Abstracts of Scientific Presentations 2008 AALAS National Meeting Indianapolis, Indiana Platform Sessions 10 times, led to seropositive recipients after embryo transfer. However, when oocytes in the cumulus-oocyte complex (COCs) were fertilized in vitro with cryopreserved spermatozoa (IVF) PS1 Decrease in Helicobacter hepaticus Titers over Time in in the presence of 104 TCID50/ml MMVp and the 2-cell embryos BALB/c Mice were washed 10 times, recipients were seronegative. The lack of CM Nagamine*1,2, Z Ge2, ME McBee3, KA Knox3, AB Rogers2, transmission of infectious MMVp by IVF embryos may be due JG Fox2, DB Schauer2,3 to the cumulus cells on the oocytes, which may have prevented 1Comparative Medicine, Stanford University, Stanford, CA; 2Com- entry of MMVp to the zona pellucida or they may have absorbed the virus. To test this hypothesis, COCs or cumulus-free oocytes parative Medicine, 3Biological Engineering, MIT, Cambridge, MA (CFOs) and spermatozoa were exposed to 104 TCID50 MMVp/ Helicobacter hepaticus induces clinical disease and confounds ml in the fertilization medium (HTF). The resulting embryos the interpretation of data in certain mouse strains. We recently were washed 10 times in virus-free KSOM and kept in holding reported that H. hepaticus appeared to be lost in BALB/cJ and media (M2). Media and groups of 20 embryos were analysed BALB-IL10 null mice starting at 13 wk postinoculation (wpi) based using PCR and in vitro infectivity assay (IVia). Of 5 replications on helicobacter-specific PCR on fecal pellets. We hypothesize that with COCs, 1/5 holding drop and 2/5 groups of embryos were the BALB/c genetic background inhibits long-term, high-titer MMV-positive using PCR, while neither holding drops nor colonization by H. hepaticus. To test this hypothesis, we gav- embryos were positive using IVia. From IVF with CFOs, PCR aged 35 BALB/cJ (BALB) females with H. hepaticus. Five 129S6/ revealed that 3/5 holding drops and 4/5 groups of embryos SvEv-Rag2tm1Fwa (129RAG2 null) females were treated similarly were MMV-positive, while 1/5 holding drops and no embryos as positive controls. Negative control BALB females received were positive for MMV using IVia. The present data show that media only. Fecal pellets were collected from individual mice at 2-cell embryos from COCs did not transmit infectious virus to 2, 6, 10, 14, 18, 22, 26, and 30 wpi. Fecal DNAs were extracted and the holding drops, most likely due to the presence of cumulus used in an all-helicobacter, all-lactobacillus duplex PCR assay. cells during fertilization. All 129RAG2 null positive controls were helicobacter-positive, and all BALB negative controls were helicobacter-negative. PS3 Absence of Germline Transmission after Blastocyst Injec- Inoculated BALB mice were helicobacter-positive at 2, 6, and 10 tion with Murine Embryonic Stem Cells Cultured with Mouse wpi. However, at 14 wpi, 2 mice were helicobacter-negative. At Hepatitis Virus and Mouse Minute Virus 18 wpi, 6 mice were helicobacter-negative and 1 of the 2 mice that were initially negative reverted to being helicobacter-positive. E Mahabir*1, K Reindl1, J Mysliwietz2, J Needham3, D Bulian1, K Overall, 56% of inoculated BALB mice were helicobacter-negative Markoullis1, J Schmidt1 at 1 or more time points. Quantitative PCR (qPCR) was per- 1Department of Comparative Medicine, 2Institute for Molecular formed on the cecal DNA of 3 129RAG2 null positive controls, Immunology, Helmholtz Center Munich—German Research 3 BALB mice that were helicobacter-positive at all time points, Center for Environmental Health, Neuherberg, Germany; 3 The and 7 BALB mice that were helicobacter-negative at 2 or more Microbiology Laboratories, North Harrow, United Kingdom time points. 129RAG2 null mice averaged 13 x 106 copies of H. hepaticus in their ceca. BALB mice that were helicobacter-positive The aim of this study was to determine the susceptibility of at all time points had an average of 2.5 x 106 copies. BALB that murine embryonic stem cells (mESCs) to infection with mouse were helicobacter-negative at 2 or more time points had the least hepatitis virus (MHV-A59) and mouse minute virus (MMVp) number of H. hepaticus in their ceca (0.1 x 106 copies). This was and the effect of these viruses on germline transmission (GLT), 24-fold fewer copies relative to BALB mice that were helicobacter- and the serological status of the recipients and pups. The mESCs positive at all time points. The data suggest that some BALB were inoculated with 10-1 TCID50/cell MHV-A59 or MMVp and mice develop low titers of H. hepaticus in their ceca, resulting in cultured for 6 h or over 4 to 5 passages, with an interval of 2 d H. hepaticus not being easily detected by fecal PCR. These data between passages. mESCs were productively infected with MHV- suggest caution in the interpretation of fecal PCR results used A59 whereas infection was restricted in mESCs after exposure to to screen for the presence of H. hepaticus. MMVp. Also, MHV-A59, but not MMVp, was cytolytic for the mESCs. Exposure of mESCs to the viruses over 4 to 5 passages, but not for 6 h prior to blastocyst injection and embryo transfer, PS2 Lower Transmission of Mouse Minute Virus by In Vitro- affected GLT to the extent that no chimeras were born. In contrast, produced Embryos Due to Cumulus Cells control mESCs led to 6.2 pups per germline chimera. Recipients E Mahabir*, D Bulian, R Schmöller, J Schmidt and pups were seropositive for MHV-A59, but not for MMVp, when mESCs were cultured with the virus over 4 or 5 passages. Department of Comparative Medicine, Helmholtz Zentrum The present data show that mESCs can transmit viruses during München—German Research Center for Environmental Health, blastocyst injection and embryo transfer, depending on the virus Neuherberg, Germany itself, and that germline transmission is dramatically affected by In vivo-produced embryos, which had been exposed in vitro virus-contaminated mESCs. to 104 TCID50/ml of mouse minute virus (MMVp) and washed Abstracts of scientific papers 2008 AALAS National Meeting PS4 Mycoplasmas Affect Cell Parameters of Murine Embry- detectable through 56 DPI. The mean peak concentration of MPV onic Stem Cells, Germline Transmission, and the Resulting DNA in BALB/c feces of approximately 1.5 million copies per mg Chimeric Progeny of feces was 3 times higher than in B6 feces. The apex of viral DNA and mRNA transcript expression in tissues largely corresponded K Markoullis1, D Bulian1, G Hölzlwimmer2, L Quintanilla-Fend2, to the pattern described for fecal shedding. In addition to being H Zitzelsberger3, K Heiliger3, J Mysliwietz4, CC Uphoff5, HG Drexler5, T Adler6, DH Busch6, J Schmidt1, E Mahabir*1 delayed and at a lower level, MPV infection, as measured by qPCR, waned sooner in B6 than in BALB/c mice. Concentrations of viral 1Department of Comparative Medicine, 2Institute of Pathology, DNA and mRNA transcripts in tissue specimens were highest in 3Institute of Molecular Radiobiology, 4Institute of Molecular Im- mesenteric lymph nodes and lowest in spleens. At 56 DPI, viral munology, Helmholtz Center Munich—German Research Center DNA concentrations in tissues remained high, whereas viral mRNA for Environmental Health, Neuherberg, Germany; 5Department transcripts were no longer detectable. MPV-1 seroconversion was of Human and Animal Cell Cultures, DSMZ—German Collection first detected in a majority of BALB/c and B6 mice at 7 and 10 DPI, of Microorganisms and Cell Cultures, Braunschweig, Germany; respectively. In summary, the resistance of B6 mice to MPV infec- 6Institute of Medical Microbiology, Immunology and Hygiene, tion correlates with lower levels of virus replication and shedding, Technical University Munich, Munich, Germany which can complicate detection and confirmation of MPV infection Production of genetically modified mice implies frequent exchange in mice that are B6 or on a B6 background. of murine embryonic stem cells (mESCs) between laboratories, which increases the risk of contaminations of cell cultures and PS6 Comparison of Side Effects between Buprenorphine and mice, thereby jeopardizing experimental protocols. To determine Meloxicam Used Postoperatively in Dutch Belted Rabbits the susceptibility of mESCs to mycoplasmas and the effects of (Oryctolagus cuniculus) mycoplasmas on cell parameters, the germline transmission CS Cooper*1, KA Pate1, C Barat2, L Swarthout1, J Anastasio1, D (GLT), and the resulting chimeric progeny, 129/SvPas mESCs Scorpio1 were inoculated at passage P13 with M. hominis, M. fermentans, 1Molecular and Comparative Pathobiology, Johns Hopkins Uni- and M. orale and cultured over 20 passages (P13 + 20). Com- pared to controls, there was a 7-log reduction in the growth of versity SOM, Baltimore, MD; 2Mathematics, Villa Julie College, mycoplasma-contaminated mESCs. Karyotypic analysis revealed Baltimore, MD non-clonal chromosomal aberrations, which increased with the One of the challenges faced by veterinarians and researchers who duration of infection. At passage P13 + 6, where mycoplasma use rabbits (Oryctolagus cuniculus) as a surgical model is choos- infection was strongest, 46.3% of mycoplasma-infected mESCs ing an appropriate postoperative analgesic that is efficacious at expressed POU5F1 (OCT-4) and SSEA-1 markers, in contrast to relieving pain without unwanted side effects. Buprenorphine 84.8% of control mESCs, indicating a decrease in the expression and other opioid derivatives are common analgesics used to of stem cell differentiation markers. The GLT of mycoplasma- alleviate postsurgical pain in rabbits. However, this class of infected mESCs at passage P13+5 decreased from 90%, as found analgesics has been noted to cause anorexia and gut stasis in in controls, to 50%. Similarly, the number of litters and the number various mammalian species, including humans. The objective of pups born decreased from 100% to 50%, and from 45% to 22%, of this study was to evaluate the occurrence of buprenorphine respectively. Some chimeras obtained from blastocyst injection side effects in Dutch Belted rabbits when used postoperatively. with mycoplasma-infected mES cells showed nasal discharge, We evaluated meloxicam as an equal alternative to opioid use. osteoarthropathia, and cachexia. The present data indicate that We expected that meloxicam would provide the same quality mycoplasmas affect cell parameters, germline transmission of of pain relief for most surgical interventions in rabbits without mESCs, and postnatal development, underlining the importance unwanted side effects such as anorexia and gut stasis. Baseline of screening mES cells for mycoplasmas prior to their use. parameters including activity, pain assessment, food intake/fecal output (g), urine output, body weight (kg), physical examination, PS5 Comparing Mouse Parvovirus Infection of Susceptible abdominal palpation and auscultation, CBC/serum biochemis- BALB/c and Resistant C57BL/6 Mice Using Quantitative Poly- try, and fecal cultures were collected for 30 female Dutch Belted merase Chain Reaction and Serologic Assays rabbits. A routine ovariohysterectomy was performed on each rabbit. Postsurgically, rabbits were randomly assigned to groups KS Henderson, CL Perkins, LA Banu, SM Jennings, WR Shek* receiving either postoperative buprenorphine (n = 10), meloxi- Research Animal Diagnostic Services, Charles River, Wilming- cam (n = 10), or bupivicaine incision infiltration (n = 10). Rabbits ton, MA in the meloxicam group demonstrated the fastest recovery to In a previous study, we demonstrated that the dose of the MPV-1e baseline parameters compared to rabbits treated with either field isolate required to infect C57BL/6NCrl (B6) mice was 10 to buprenorphine or bupivacaine. Normal eating and voiding be- 100 times higher than the infectious dose for BALB/cAnNCrl mice, haviors of rabbits treated with buprenorphine gradually returned whether the virus was inoculated by gavage or intraperitoneally. to baseline; however, rabbits treated with bupivicaine needed To investigate this susceptibility difference further, we inoculated medical intervention to fully recover to baseline. Meloxicam is B6 and BALB/c mice by gavage with approximately 1000 and 100 an appropriate alternative to buprenorphine for alleviation of BALB/c infectious doses (ID50) of MPV-1e, respectively. Feces, postoperative pain in the rabbit. Rabbits can be given meloxicam mesenteric lymph nodes, ileum, spleen, and blood were collected safely without onset of anorexia or gut stasis, which are docu- at 11 time points, from 1 to 56 d postinoculation (DPI). Viral DNA mented side effects associated with buprenorphine use. We also and reverse-transcribed messenger (m) RNA copies were measured recommend strong consideration and justification when using in feces (DNA only) and tissue specimens by fluorogenic nuclease bupivicaine infiltration as a sole method of pain relief. qPCR assays. MPV seroconversion was assayed by several meth- ods, including ELISA, IFA and MFIA. MPV DNA was detected PS7 Identification of a Novel Low-pathogenicity Rabbit Calici- in the feces of both B6 and BALB/c mice at 1 DPI; fecal shedding virus in a Spontaneous Disease Outbreak in the United States peaked in BALB/c mice at 5 DPI and 2 d later in B6 mice, and was I Bergin*, SR Bolin, M Kiupel, RK Maes, TP Mullaney, AG Wise Abstracts of scientific papers 2008 AALAS National Meeting Michigan State University, Ypsilanti, MI with Taconic mice (0.850 ± 0.130; P < 0.01). These data indicate that the environment in which the mice are reared is more im- We report a spontaneous disease outbreak in a privately owned portant than genetic relatedness. The notable difference between Michigan rabbitry caused by a novel rabbit calicivirus related Taconic and Jax husbandry is that Taconic rederives their mice to but distinct from rabbit hemorrhagic disease virus (RHDV) into the germ-free state with subsequent ASF colonization. In and having low-level pathogenicity. At this facility, 35% of ap- contrast, Jax maintains their mice in SPF without rederivation and proximately 200 New Zealand White rabbits died over a period colonization with ASF. This study suggests that the indigenous of 1 wk. Pregnant does were predominantly affected; principal gastrointestinal microbiome plays a significant role in murine lesions consisted of pulmonary hemorrhage and hepatocellular necrosis. Calicivirus-like particles were detected in liver ho- gallstone formation. mogenate by electron microscopy. Initial RT-PCR amplification of viral sequence using RHDV-specific primers was positive; PS9 Single Nucleotide Polymorphisms That Differentiate however, partial genomic sequencing demonstrated an overall Common Inbred Strains and Substrains of Mice 79% nucleic acid identity with RHDV. Phylogenetic compari- GW Bothe*1, JL Seymour1, JL Moran2, SM Festin1 sons of the partial viral genomic and amino acid sequences 1 were consistent with a unique calicivirus in the Lagovirus genus Taconic, Rensselaer, NY; 2Brigham and Women’s Hospital, that was related to but distinct from RHDV and other reported Harvard Medical School, Boston, MA lagomorph caliciviruses. We provisionally describe this virus While microsatellite markers can be used to verify the identity of as Michigan rabbit calicivirus (MRCV). To our knowledge, this mice, single nucleotide polymorphisms (SNPs) are more easily is the first report of a low-pathogenicity rabbit calicivirus in available in current databases, and are more amenable to high- the Lagovirus genus in the U.S., although low or nonpathogenic throughput analytic technologies. SNP data on some mouse rabbit caliciviruses have been reported in other countries. We strains have been widely reported. However, the almost complete performed an infection study using tissue homogenate from absence of data on certain substrains made the development of affected rabbits inoculated into specific pathogen-free rabbits. a genetic quality control program difficult. Therefore, we first This resulted in individual hepatocellular necrosis and positive analyzed a panel of 768 mouse SNPs that contained a number in situ hybridization within hepatocytes, but no overt clinical of markers that differentiate C57BL strains. Markers were de- disease or widespread histological changes. We propose that tected using BeadArrays. Among major strains and substrains, MRCV primarily causes subclinical infection of healthy rabbits there were 19 differences between C57BL/6NTac (B6NTac) and and may induce overt, RHD-like disease under certain field C57BL/6J (B6J), 11 B6NTac and C57BL/6JBom (B6JBOM), and 8 conditions. In addition to the possibility of clinical disease, this B6J and B6JBom, placing these strains into the expected positions report calls attention to the potential diagnostic confusion of a in the C57BL family tree. We found 459 differences between B6N- low-pathogenicity calicivirus such as MRCV with the reportable Tac and BALB/c, and no difference among BALB/c substrains foreign animal disease agent RHDV. AnNTac, ABomTac, J, and JBomTac. This result is unexpected given the long separation time of the substrains. However, it may PS8 Role of the Gastrointestinal Microbiome in Cholesterol be explained by a “marker identification bias” caused by nearly Gallstone Pathogenesis all SNPs having been identified in screens of B6J compared with other strains. We used a 140,000 SNP DNA chip to search for JJ Fremont*1, JG Fox1, MC Carey2, KJ Maurer1,2 SNP markers differentiating BALB/c substrains. We were able 1Division of Comparative Medicine, MIT, Cambridge, MA; 2Divi- to identify 8 candidate SNPs in the chip data, and have verified 3 sion of Gastroenterology, Department of Medicine, Brigham and SNPs by PCR amplification and sequencing. We then developed a Women’s Hospital, Harvard Medical School, Boston, MA genetic quality control program that would detect any mix-up of inbred mice after transport from one facility to another. For each Cholesterol gallstone formation is a common costly disease with strain-specific panel, at least 2 markers were selected for each no definitive nonsurgical treatment. Recently, we demonstrated that BALB/c mice from different commercial sources exhibit strain that could distinguish that strain from any other strain, and at least 2 additional markers were added to each panel to markedly different prevalences in cholesterol gallstone forma- tion. Specifically, BALB/c mice from The Jackson Laboratory enable differentiation of substrains where necessary, resulting in (BALB/cJ) demonstrated an increased prevalence compared panels of 4 to 8 SNPs. This testing program combines maximum to those from Taconic Farms. To determine if these differences clarity of results with minimum cost. All SNP assays in the qual- resulted from variations in the indigenous microbiota or genetic ity control panels were verified by fluorogenic PCR. differences between the substrains, we performed a prospective study analyzing biliary phenotype in closely related BALB/c PS10 Mouse Parvovirus Infection of Immunodeficient Mice mice. Three groups (n = 10/group) of mice were analyzed for J Macy*, PC Smith, J Wilson, SR Compton gallstone phenotype: Group 1 was BALB/cJ mice; group 2 was BALB/cByJ mice, which are from the same genetic substrain as Comparative Medicine, Yale University, New Haven, CT BALB/cAnNTac mice, but are housed at Jax; and group 3 was The interplay of mouse parvovirus (MPV) and the host immune BALB/cAnNTac. Eight-week-old mice were fed a lithogenic system is not well understood. In order to determine how the diet for 8 wk. At the conclusion of the study, gallbladder bile immune system impacts MPV infection shedding, transmission, was examined to determine biliary phenotype. BABL/cByJ and immune cell tropism 10 SCID (CB-17/Icr-Prkdcscid/Crl), 5 and BALB/cJ mice displayed similar prevalences of cholesterol SCID-BG (CB-17/Icr.Cg-Prkdcscid Lystbg/Crl), 5 Igh (STOCK- monohydrate crystals (70% and 90%) and cholesterol gallstones Igh-Jtm1Dhu N?+N2), 5 CB17 (C.B-Igh1b/IcrTac) and 5 BALB (50% and 70%). In contrast, BALB/cAnNTac mice developed (BALB/cAnNTac) mice were infected with MPV. Fecal samples cholesterol monohydrate crystals and cholesterol gallstones at were collected weekly from each mouse for PCR. One SW sentinel a significantly lower prevalence (10% for each; P ≤ 0.05). Mucin was added to each cage at postinfection day (PID) 0 and replaced gel score were increased in Jackson mice of either substrain at PID 14 and PID 28. Sentinel mice were held for an additional 2 (BALB/cJ: 2.250 ± 0.367; BALB/cByJ: 2.150 ± 0.317) compared wk and tested for seroconversion and fecal shedding. Index mice Abstracts of scientific papers 2008 AALAS National Meeting were euthanized on PID 14 (5 SCID) or PID 42 (all other mice) tively allocate limited resources to multiple projects and requests and mesenteric lymph nodes (MLN) were harvested. PCR and from diverse end users. By using task and study time analysis FACS were performed on MLNs to detect the presence of MPV in methods during the scheduling process, we can track available CD4, CD8, B220, and NK cells. The results showed that all BALB, study support time compared to resources previously alloted. CB17 and Igh, 8/10 SCID and 1/5 SCID-BG mice were shedding Technicians supply actual time-in-motion data at the comple- MPV in the feces on PID 7. At PID 14, feces from SCID, SCID-BG, tion of each day, permitting the development of a database on and Igh mice averaged 4.2 pg/µl of MPV DNA, whereas CB17 task performance. This practice has revealed a valuable “team and BALB feces contained 1.6 and 2.8 pg/µl of MPV DNA, re- effect”—the efficiency that occurs when 2 or more technicians spectively. MLNs from SCID and CB17 mice contained 66.5 and complete a specified task in less time than predicted by adding 7.4 pg/µl of MPV DNA, respectively. Seroconversion and fecal time spent alone. We then assign daily schedules and study shedding occurred in contact sentinels placed with SCID-BG or work based on data collected from one technician performing Igh mice on PID 14, but not with SCID, CB17 or BALB mice. By each task, allowing for the team effect to provide flexibility in PID 21, CB17 and BALB mice ceased shedding MPV, whereas accommodating last-minute demands and increase the capac- the immunodeficient mice were still shedding on PID 28, 35, and ity of the team’s workload without strain to any individual. It 42. Sentinels placed on PID 28 with SCID-BG mice but not with is driven by increasing participation from the technicians and other mouse strains seroconverted and shed virus. At PID 42, directly influences how the calendars are managed. PCR detected 0.77 (Igh) to 6.7 (SCID-BG) pg/µl of MPV DNA in feces and detected from 1.9 (CB17) to 86.4 (SCID) pg/µl of PS13 Developing Meaningful Training in Laboratory Animal MPV DNA in MLNs. FACS analysis indicated a low number of Science: An Educational Methodology B cells and NK cells contained MPV antigen on PID 42 (SCID > SCID-BG > Igh). These results indicate that fecal shedding is S Pack, P Clifford* delayed in SCID and SCID-BG but virus levels in the feces and ULAR, University of Pennsylvania, Philadelphia, PA MLNs were high at PID 42. Although SCID and SCID-BG had A new concept to bring to the laboratory animal community is high levels of virus in the feces and MLN, only the SCID-BG designing learning environments to promote understanding and transmitted infection after PID28. This suggests a role for NK reason. There are a few basic instructional design techniques that cells in controlling infection. can immediately be applied in training development to promote understanding. Applying a “backwards design” method can help PS11 New Solutions for Management and Husbandry: RFID in developing effective training by focusing on goals, evidence, Census and Wash Cycle Tracking and activities. These 3 simple words can make a difference for B Scher* those needing to deliver training sessions to support or research staff. Before developing training, the goals of the session must Dynasys, Clearwater, FL first be clearly defined. Next, determine the acceptable evidence. The process of arriving at an accurate census for large rodent How are you going to assess the learners and be confident that populations is time consuming, labor intensive, and prone to they understand what is being taught? Lastly, develop the ac- many errors. Principal investigators often challenge the ac- tivities, which include presentations. Typically those delivering curacy of their billed services and census reports. Tracking the training sessions or presentations start with their activities first current location of every cage housed within a large animal (such as a PowerPoint presentation). This step in development care facility requires continuous monitoring and extensive should be last, hence the “backwards design” method. Assess- recordkeeping. Radio Frequency Identification (RFID) tagging ment and evidence are key and need to be clearly determined of each cage allows for real-time, error-free electronic detection prior to activity development. The 6 facets of understanding need of hundreds of cages per second. The Electronic Product Code to be considered to truly teach and train learners so that they gain (EPC) is an open global standard designated for RFID labeling an enduring understanding that is applied beyond the training analogous to the Universal Product Code (UPC) barcode that has session. These facets of understanding that are needed to effec- been identifying all consumer goods for the past 30 y. EPC tags tively assess knowledge building are as follows: can explain, can can be directly attached to animal cages, racks, and cage card interpret, can apply, have perspective, can empathize, and have holders. These passive tags can survive autoclave sterilization self-knowledge. Particularly, when working in our industry with and can be directly attached to metal. EPC technology features live animals, a deep and meaningful workplace understanding anti-collision properties such that hundreds of tags can be read is paramount. Therefore, by applying the above methodology in seconds using a portable RFID reader, and the animal census and building an awareness of the 6 facets of understanding, of a room can be reconciled immediately. In addition to taking trainers can gain an active awareness of their learners’ progress, animal census, these tagged cages can be individually identified while being assured that their learners are gaining practical and as they pass through cage washers. RFID readers attached to the meaningful knowledge. washers identify each cage and integrate the process control data with the cage tag identifiers, providing an accurate data record PS14 Refining Rodent Paralysis Models through Animal Care of each wash cycle. RFID technology will find many new ap- Staff Involvement plications within the laboratory facility. A Heiser*, E Westhoven, RJ Hurley, DM Jarrell PS12 The Team Effect and Technician Efficiency at Genzyme CCM, Massachusetts General Hospital, Charlestown, MA M Scribner* A recent advance in laboratory animal care is to leverage the daily interaction that front-line animal care staff already has Department of Comparative Medicine, Genzyme Corporation, with their assigned animals to notify appropriate parties when Framingham, MA an animal has health or welfare problems. Delegation of such When effectively scheduling animal work for a technical support tasks can result in a faster and equally effective response rather team, it is necessary to monitor the team’s workload to effec- than waiting for higher-level employees (veterinary technicians, Abstracts of scientific papers 2008 AALAS National Meeting veterinarians, investigators) to discover or confirm the same PS16 Retaining Your Top Animal Care and Cage Wash Per- problem. But IACUC-approved endpoints in potentially painful or formers distressful protocols may be unclearly defined or not adequately C Kilcullen-Steiner* understood by animal care staff to permit appropriate and timely intervention. Murine models of amyotrophic lateral sclerosis (Lou University Animal Care, University of Arizona, Tucson, AZ Gehrig’s Disease) and Huntington’s Disease have helped identify At academic institutions, keeping top performers in cage wash new drug strategies, an important advance for these and other and direct animal care positions is becoming more difficult. neurodegenerative conditions for which there are no effective These employees usually have the lowest remuneration, are treatments. Such models routinely involve severe clinical signs required to be on their feet during their whole shift, experience that should require euthanasia rather than allow the animal’s little variation in routine, and have no loyalty to the depart- condition to decline further. IACUC-approved protocols at our ment/institution. As research funding and other sources of institution mandate euthanasia following hind limb paralysis, revenue become harder to acquire, more principal investigators the most common severe clinical sign in these models, if the are recruiting for new employees from the animal care staff. animal cannot right itself within 15 s. Our animal care staff used The PI knows he will obtain a highly trained animal technician multiple variations of the “15-s” rule, resulting in recommended familiar with the laws, regulations, policies, and procedures euthanasia at varying stages of disease. To standardize monitor- that govern the care and use of research animals. He just trains ing of an animal’s condition, we developed a precise and simple them on his actual research. Usually the PI can offer the animal decision tree for more consistent reporting and intervention. A technician more money per hour and give them the opportunity combination of pictorial guides, clearly defined clinical signs, to conduct actual research in addition to direct animal care. In and possible outcomes provided in an SOP format resulted in order to slow or stop this outflow of trained staff, the animal more effective training, simple interpretation, and reduction in care facility had to move toward incentive methodologies, as variability. Benefits of this approach include improved animal well as good business practices to increase employee retention. welfare without compromising research and increased awareness As a start, new positions were created to provide more career and involvement by a caring workforce. paths. One job was created for an environmental enrichment specialist, two instructional specialist positions were created for PS15 Development of Team Leaders in a Team-oriented Or- upward mobility in cage wash, back-ups were designated for the ganization supervisors, and technician senior leads were created to give top employees experience in team leadership. In addition, monetary C Gibbons*, J Fuller, DM Jarrell incentives were established for achieving certification, excelling CCM, Massachusetts General Hospital, Charlestown, MA in job performance, etc. The outflow of staff has decreased, but Today’s laboratory animal care workspace mirrors recent evolu- we know not every employee can be promoted to a new position tion in biomedical research by being more complex, expensive, and money is limited. We continue to work on more novel ways and potentially hazardous. However, the standard approach to to retain valuable staff. organizational structure and workforce development remains unchanged. Conventional responses are to either increase su- PS17 Developing and Implementing a Program for Standard- pervisory oversight (greater control) or narrow the scope of each izing and Harmonizing Training Requirements and Records worker stratum (less risk) to avoid mistakes of greater impact. Both for a Global Organization approaches increase total labor expense while research funding is M Butler*1, I Garrod3, G Ma4, M Defeo1, SA Drinen2, SE Deboer1, tighter and other costs (fuel, raw materials, supplies, equipment) R Burke1, KJ Rogers3, CF Fox3 are growing. Furthermore, employees are forced to wait for pro- 1Worldwide Comparative Medicine, Groton, CT; 2Worldwide motions or lateral transfers before they attain additional skills and enjoy career growth without leaving. By contrast, in line with using Comparative Medicine, St. Louis, MO; 3Worldwide Compara- Toyota Production Systems (TPS) as our main operational and tive Medicine, Sandwich, Kent, UK; 4Worldwide Comparative management strategy for improving work quality, we established Medicine, Pfizer, La Jolla, CA 18 multi-functional teams of research animal specialists divided Developing and implementing an effective training program to amongst 6 facilities. Team responsibilities include all husbandry, support quality animal care and use is critically important. Our initial health assessment and basic veterinary care, operational global organization, consisting of three sites in the United States maintenance, and research services within their designated area. and 1 site in the United Kingdom, was faced with the challenge Strong team leadership is paramount in sustaining a highly func- of developing such a program. To address this challenge, our tional team. Team leaders are responsible for coordination of daily organization formed a team consisting of select personnel from tasks, team-directed evaluation, and improvement of work flow each site. This team was given the task of developing a train- processes, and assurance that team members work in a productive ing program that was aligned across sites, creating a learning and collaborative manner. An intensive team leadership training environment with shared and measured accountability, and program developed in-house includes a comprehensive review developing and implementing an innovative learning approach of problem-solving tools and principles of TPS, an emphasis on that would save the organization both time and money. The mature and clear communication skills for a strong researcher/ approach developed by this team, modeled after a successful customer service focus, and building work style-focused interper- approach used at our U.K. site, consists of four components. sonal skills to support a highly functional team-environment. One A master training index is used to identify all training require- significant impact seen at MGH of improved team leaders and ments specific to our global training program. Each item in this highly functional teams has been our ability to increase staffing index is listed under an appropriate training category and is levels only 30%, while increasing our overall rodent cage census supported by a corresponding training module. Each training by 60% over the last 3 y. Financial impacts in less overtime and module contains harmonized learning objectives agreed to by increases in non per-diem revenues (such as research services all sites and identifies the minimum requirements that an indi- support) were also observed. vidual must meet before they are considered proficient for that Abstracts of scientific papers 2008 AALAS National Meeting module. Qualified trainers use these training modules to ensure hands-on classes were provided to learn postural modifications that training is provided at an appropriate level of detail and and exercises used to prevent injury when performing tasks such that all required information is addressed during the training. as rodent cage changing, dog cage cleaning, and moving heavy Training schedules are used by managers at each site as a check- equipment in feed and bedding storage areas and washrooms. A list and guide to ensure that new personnel complete assigned review of job specific exercises is presented here, as well as staff training requirements within established timelines. Standardized members’ comments by survey of the success of the program training records mirror the contents of the master training index in decreasing areas of discomfort and fatigue in the workplace. and require that individuals are assessed while working under Our departmental goal of decreasing injury rates by at least 10% supervision, assessed as competent, and assessed as a competent was met after implementing these changes and our future goals trainer, if appropriate, for each assigned training module. This related to safety will be addressed. session details the approach taken to develop our global training program and describes the tools developed to support it. PS20 Development and Implementation of a Quality Control Program for Animal Care Activities PS18 Ergonomic Improvements through Equipment Updates N O’Callaghan*, R Palmisano in a Laboratory Animal Facility Yale University, New Haven, CT R Starr*, N Montero, C Fuzzi, J Nolan, A Kopec, M Hargaden, L Singer Monitoring the care and use of laboratory animals is an integral component of humane biomedical research and a responsibility Department of Laboratory Animal Resources, Roche, Nutley, of institutional oversight and evaluation. Ongoing monitoring NJ must be performed and documented to ensure humane and The study of body movement and posture in relation to work compliant direct care activities between federally mandated demands, or ergonomics, has become a major point of interest in inspections and accrediting site visits. This provides early detec- places of work, including laboratory animal facilities. In 2007, a tion and identification of noncompliance for corrective action and global initiative to decrease work-related injuries and associated quality control. To accomplish this, a position was created within costs prompted the Department of Laboratory Animal Resources the Regulatory and Safety Services (RSS) section with dedicated (LAR) to invite an ergonomist to evaluate specific job functions effort to monitoring animal care and related operations. Due to in the animal facility. An internal safety committee comprised of Yale University’s large, diverse, and geographically decentralized an operations manager, building supervisors, laboratory animal animal care and use program, a streamlined process and self- technicians, clinical veterinary associates, and veterinarians assessment tool was created to ensure that animal care activities was responsible for working with the ergonomist to alleviate are in accordance with institutional, federal, and state regulations sources of repetitive motion or overuse injury and fatigue in and policies. This assessment is conducted in every animal room our employees. The committee evaluated new equipment and and support space twice a year. The findings from each space variations on the use of space to enhance safety in work areas, monitored are immediately reported to the facility manager so including cagewash rooms, large and small animal housing that corrective action can be taken. Monthly reports with global rooms, and food and bedding storage areas. Our evaluation findings are given to all facility managers, and quarterly global of several pieces of equipment, including hydraulic lifts, tote reports are presented to all managers at a convened meeting. tippers, table lifts, scooters, and ergonomic brushes and mop Corrective action plans are then identified and implemented. handles, will be discussed, as well as an overview of related This quality control monitoring program promotes timely cor- costs. Our departmental goal of decreasing injury rates by at rective action and provides a global view of the entire program. least 10% was met after implementing these changes. Our future Additionally, this internal assessment provides a link to both goals related to safety will be addressed. the refinement of the training program and the creation of and revision to the internal standard operating procedures (SOPs). PS19 No-cost Solutions for Ergonomic Improvements in a The institution’s investment in quality control monitoring affirms Laboratory Animal Facility its commitment to improving the quality of humane animal care while heightening the regulatory awareness of all levels of animal C Fuzzi*, M Hargaden, L Singer care staff and ultimately reducing noncompliance. Department of Laboratory Animal Resources, Roche, Nutley, NJ PS31 Spontaneous Acute Tumor Lysis Syndrome in Swiss Webster The study of body movement and posture in relation to work Mice with Spontaneous Leukemias and Lymphomas demands, or ergonomics, has become a major point of interest in S Rasmussen*1,3, K LaPerle1,3, S Couto1,3, J White1,3, V Gillespie1,2, places of work, including laboratory animal facilities. In 2007, a FR Wolf1,4 global initiative to decrease work-related injuries and associated 1Postdoctoral Research, Tri-Institutional Postdoctoral Training costs prompted the Department of Laboratory Animal Resources (LAR) to invite an ergonomist to evaluate specific job functions Program in Laboratory Animal Medicine and Science, New York, NY; 2Pathology, Animal Medical Center, New York, NY; 3RARC/ in the animal facility. An internal safety committee comprised CBC, Memorial Sloan-Kettering Cancer Center, The Rockefeller of an operations manager, building supervisors, laboratory University, and The Weill Medical College of Cornell Univer- animal technicians, clinical veterinary associates, and veterinar- sity, New York, NY; 4RARC, Memorial Sloan-Kettering Cancer ians was responsible for working along with the ergonomist to locate potential sources of repetitive motion injury or fatigue Center and The Weill Medical College of Cornell University, New York, NY and discomfort in our employees. In addition to providing our employees with training in ergonomics, we consulted an exercise We have observed a high incidence of spontaneous acute tumor physiologist to address staff members’ individual concerns with lysis syndrome (ATLS) in our colony of Swiss Webster mice used their specific daily tasks. An exercise program was implemented as dirty bedding sentinels. ATLS is thought to be rare in mice, to increase awareness in posture and stretching, and small group with a single reported case in a DBA/1J mouse. ATLS results Abstracts of scientific papers 2008 AALAS National Meeting from a rapid destruction of neoplastic cells. When large volumes findings. Vascular lesions, considered the cause of death in the of byproducts from neoplastic cells are released, the body’s published reports of GVHD in mice, were not observed in our excretory mechanisms are overwhelmed, resulting in metabolic cohort. X-GVHD has not been previously described in the NOD- derangements, renal failure, and disseminated intravascular scid IL2Rγnull strain, nor has the disease been fully characterized coagulation and thrombosis. ATLS can occur spontaneously, clinically and pathologically in mice. This comprehensive study or more commonly following chemotherapy, radiation, or will further the understanding of X-GVHD in immunodeficient corticosteroid administration. In humans, ATLS has a strong mice and provide useful information for those using NOD-scid association with high-grade lymphoproliferative malignancies, IL2Rγnull mice in cancer immunobiology. although also reported to occur with other tumor types. Com- plete anatomic pathologic evaluations were performed on 146 PS33 Incidence, Evaluation of Sampling Techniques, Isolate Swiss Webster mice that were either found dead without a history Diversity, and Antibiotic Susceptibility of Corynebacterium of clinical signs or were sacrificed due to lymphadenopathy or bovis in Immunodeficient Mice varying signs of morbidity. The mice were not subject to prior experimental manipulation. Results from mice necropsied over HN Burr*1,2, NS Lipman1,3, A Toma3, FR Wolf1,3 a 12-mo period revealed that 94 of 146 (64%) mice evaluated 1Tri-Institutional Training Program in Laboratory Animal were found to have either lymphoma or leukemia. These find- Medicine and Science, New York, NY; 2 Memorial Sloan-Kettering ings are not surprising given that a high percentage of Swiss Cancer Center, The Rockefeller University, and the Weill Medical Webster mice are known to carry and express ecotropic murine College of Cornell University, New York, NY; 3 Memorial Sloan- leukemia viruses. Seven of 146 (4%) mice evaluated also had Kettering Cancer Center and the Weill Medical College of Cornell histologic lesions consistent with ATLS. Five of the 7 ATLS cases University, New York, NY died spontaneously. Of the remaining 2, 1 mouse was euthanized Corynebacterium bovis infection remains an endemic problem in due to gross abdominal distension as a result of splenomegaly athymic nude mice at academic institutions. Infected athymic and lymphadenopathy; the other due to neurologic signs. Four nude mice used in cancer research frequently exhibit signs of of the 7 mice with ATLS had pathologic findings consistent with clinical disease, including hyperkeratotic dermatitis, septicemia, both leukemia and lymphoma. The remaining 3 mice had find- and death, when experimentally manipulated. We sought to ings consistent with disseminated lymphoma. All mice were evaluate C. bovis sampling techniques in immunodeficient hirsute found to have had intravascular multifocal thrombi composed and glabrous rodents, incidence of infection, isolate diversity, and of basophilic nuclear and eosinophilic cytoplasmic debris con- antibiotic susceptibility. Rodents in 2 animal holding rooms, each sistent with ATLS. housing immunodeficient rodents including athymic nude mice and rats, were sampled and found to have a C. bovis infection PS32 Clinical and Pathological Characterization of Xenogeneic prevalence of 30% and 67%, respectively, with 95% of infections Graft-versus-host Disease in NOD-scid IL2Rγnull Mice subclinical. Skin swab and buccal swab techniques were found HN Burr*1,2, JC Markley3,4, JR White1,2, SS Couto1,2 to be comparable for isolation of C. bovis from both hirsute and 1 glabrous mice, but in rats, the buccal swab was more sensitive. In Tri-Institutional Training Program in Laboratory Animal hirsute rodents, the skin scrape technique was also evaluated and Medicine and Science, New York, NY; 2 Memorial Sloan-Kettering found to be less sensitive. C. bovis isolates demonstrated 8 differ- Cancer Center, The Rockefeller University, and the Weill Medi- ent biochemical profiles with colorimetric biochemical tests; each cal College of Cornell University, New York, NY; 3 Department profile was confirmed by colorimetric carbon source metabolic of Molecular Pharmacology, New York, NY; 4 Memorial Sloan- fingerprinting, PCR and 16S rRNA sequencing to be C. bovis. C. Kettering Cancer Center, New York, NY bovis colony size also varied. Both a large and small colony-type Immunodeficient mouse strains are commonly used to study grew on Columbia colistin-nalidixic acid agar with 5% sheep blood human tumor biology and adoptive immunotherapy. Conse- media. The Kirby-Bauer disk diffusion antibiotic susceptibility test quently, they can develop xenogeneic graft-versus-host disease revealed different sensitivities to penicillin between the large and (X-GVHD) which may confound interpretation of experimental small colony-type. Our findings demonstrate the skin and buccal results. We characterized X-GVHD in NOD-scid IL2Rγnull mice to swab sampling techniques in mice to be equally sensitive at C. bovis aid in the differentiation between the disease and cancer-related detection, contrary to published reports. Diversity within C. bovis death. Eight-week-old male mice received intravenous human isolates is newly reported and may lead to further understanding Burkitt lymphoma cells (Raji cell line) alone (n = 6) or followed of virulence and disease manifestation. Additional research that by intravenous tumor-specific human T cells 6 d later (n = 19). evaluates the efficacy of amoxicillin diet prophylaxis and treat- After approximately 2 mo, mice receiving human T cells pre- ment will be presented. sented with nonspecific signs of illness including weight loss, scruffy hair coat, and hunched posture. Histologically, a hetero- PS34 Abnormally High Serum Potassium Levels in Laboratory geneous infiltration of CD3 positive lymphocytes was observed Guinea Pigs and Rodents Caused by CO2 Euthanasia–Induced in several organs. In addition to dermatitis, colitis and hepatitis Respiratory Acidosis commonly seen in allogeneic GVHD, the mice had significant lesions in the lungs, liver, salivary glands, prostate, thyroid P Cowley, B Hayes, A Greene, WR Shek* gland, and epididymis. Mild nephritis, arthritis, and meningoen- R&D, Charles River, Wilmington, MA cephalitis were also observed. Raji tumor cells were completely The carbonate/bicarbonate buffer system is essential in main- eliminated in mice receiving human T cells. Mice receiving Raji taining pH homeostasis in animal physiology. CO2 euthanasia cells alone presented with hindlimb paralysis and histological overwhelms the buffering capacity of the system, causing a evidence of spinal cord and liver infiltration by Raji cells, but no respiratory acidosis that may lead to abnormal serum chem- lesions compatible with X-GVHD. Mice suffering from X-GVHD istry values. We performed serum chemistry on specimens likely die of multiorgan dysfunction with thyroid destruction collected from 16 specific pathogen-free Hartley guinea pigs and pulmonary impairment being the most clinically relevant (Crl:HA ORIGIN), following ketamine/xylazine (40 to 80 mg/ Abstracts of scientific papers 2008 AALAS National Meeting kg; 2 to 5mg/kg) anesthesia and after CO2 euthanasia. Serum progression of disease have not been clearly identified. Glutamic potassium (K+) levels were normal in specimens collected from acid decarboxylase (GAD), found in mouse and human pancreatic anesthetized animals (4.41 mmol/l). Following CO2 euthanasia, beta cells, is one potential candidate autoantigen. We constructed however, the serum K+ concentration increased 3-fold on average mice with a human:mouse chimeric TCR recognizing GAD65 (12.95 mmol/l); moreover, less marked increases in glucose and (555-567) which were crossed with murine MHC Class II knockout inorganic phosphate levels were noted. Increases in K+ above mice expressing human diabetes-associated MHC Class II alleles normal concentrations (3.5 to 5.1 mmol/L) were also found in (C57BL/6 background), with and without a recombination- sera from CO2-euthanized mice (n = 182; K+ = 12.81 mmol/l) activating gene deletion. Unexpectedly, we observed spontaneous and rats (n = 572; K+ = 8.88 mmol/l). lymphoblastic lymphoma in 2 lines of GAD TCR transgenic mice (4.13αβ and 164αβ). Health records were examined over a PS35 Poliomyelitis in SCID Mice Following Injection of 2-y period; mice presenting with clinical signs compatible with Basement Membrane Matrix Contaminated with Lactate lymphoma such as combinations of abdominal or subcutaneous Dehydrogenase-elevating Virus masses, anemia, hunched, and ruffled fur plus cases confirmed by necropsy or histopathology were counted. Forty-four mice met J Carlson*1, R Garg2, SR Compton1, C Zeiss1, E Uchio2 these criteria. Age ranges of affected 4.13αβ and 164αβ mice were 1Section of Comparative Medicine, 2Department of Surgery, 17.1 to 54.6 wk (average = 28 wk) and 18.7 to 53 wk (average = Section of Urology, Yale University School of Medicine, New 34.5 wk), respectively, with no sex bias. The incidence of disease Haven, CT in 4.13αβ mice was 19.7% and 13.9% in 164αβmice. Interestingly, mice transgenic only for the 164β chain of the TCR had a disease Five 30- to 40-wk-old female ICR/SCID mice presented for incidence of 0.03% based on the above criteria. A large thymus, para- or quadriplegia. The mice were part of a 50-mouse study splenomegaly, and lymphadenopathy were the most common involving subcutaneous injection of carcinoma cells suspended gross findings. Histopathology performed on 6 animals revealed in basement membrane matrix. Ruleouts included mouse extensive lymphoma in multiple organs with bone marrow in- hepatitis virus, mouse encephalomyelitis virus, lactate dehy- volvement where examined. The necropsy findings combined with drogenase-elevating virus (LDV), and CNS metastasis. Upon characteristic lymphoblastic cells strongly support a T cell origin; initial investigation, it was discovered that the matrix was con- however, immunophenotyping will aid in further characterization. taminated with LDV. Twenty-one mice were analyzed to confirm Clinicians and investigators are reminded that lymphoma may be LDV as a cause of clinical findings; tissues from 14/21 mice were an unintended consequence in genetically engineered mice. positive for LDV by RT-PCR. Histology was performed on 18 mice, of which 4 presented with paralysis; 12/18 mice had CNS lesions consisting of neuronal degeneration, marked astrocytic PS37 Development and Implementation of Multimedia Con- hypertrophy and gliosis and myelin loss. The spinal cord was tent for Web-based Rodent Microsurgery Courses most commonly affected, with less consistent involvement of the S Baran*1, J Kehler2, E Johnson3, F Hankenson4 brainstem and caudal midbrain. Cord lesions co-localized with 1Department of Surgery, Veterinary Bioscience Institute, Har- focal marked astrogliosis on GFAP immunohistochemistry. Wal- leysville, PA; 2Department of Clinical Studies, University of lerian degeneration of sciatic nerves was detected in 4/18 mice. Pennsylvania, Kennet Square, PA; 3 Harleysville Veterinary Lymphoma was present in 5/18 mice. Additionally, 21/21 mice Hospital, Harleysville, PA; 4ULAR, University of Pennsylvania, were positive for murine leukemia virus (MuLV) by RT-PCR. Cell lines were negative by PCR for viral contaminants, while Philadelphia, PA matrix was positive for LDV. These results strongly implicated Microsurgery is commonly used in the human, veterinary and LDV-contaminated matrix as a cause for clinical findings. LDV is biomedical fields. Advances in microsurgical equipment and a mouse arterivirus that is generally asymptomatic. MuLVs are rodent technologies have increased the demand for biomedical endogenous retroviruses found in all laboratory mice as provi- microsurgeons. While a limited number of microsurgical training ruses integrated in the chromosome, although only strains that courses are available, ultimately many rodent biomedical proce- carry the Fv-1n/n allele express infectious virus. Co-infection dures are self-taught, introducing a bias of technical proficiency with MuLV and LDV in immunosuppressed Fv-1n/n mice results into experimental outcomes. Electronic-learning (e-learning) has in the development of age-dependent poliomyelitis (ADPM), a been successfully incorporated into the human surgical curricula. model of human amyotrophic lateral sclerosis. In this case, clini- E-learning allows practitioners to study and improve their skills cal disease consistent with ADPM developed due to incidental at their convenience. Translating this model to the biomedical inoculation of LDV into immunocompromised mice. This report research field provides users with an opportunity to learn rodent highlights the importance of screening biological materials before surgical techniques through electronic resources like interac- inoculation into laboratory animals. tive and non-interactive videos, animations, presentations, and standard operating procedures. This training can be completed PS36 Spontaneous Lymphoma in Glutamic Acid Decarboxylase prior to using live animals, which promotes the principle of re- T-cell Receptor Transgenic Mice duction since animals are not used and enhances the experience of hands-on courses. The goals of the project were to develop AJ Burich*1,2, PM Treuting2, JA Gebe1, TL Brabb2 web-based interactive learning for rodent surgical techniques. 1Benaroya Research Institute at Virginia Mason, Seattle, WA; Unique e-learning microsurgical modules were incorporated into 2Department of Comparative Medicine, University of Washing- a training class of undergraduate laboratory animal students ton, Seattle, WA (n = 60). The tested modules included laws, regulations, and guidelines; surgical patient preparation; and proper tissue han- T-cell receptor (TCR) transgenic mice are used to understand dling and dissection. The project was evaluated using postcourse thymocyte development, peripheral tolerance, and T cell:antigen survey data. Within the training class, all students responded interactions in infectious and autoimmune diseases. Type 1 favorably to videos, step-by-step photographs of procedures, and diabetes is thought to be a T-cell mediated autoimmune process, 24-h availability of the course. Voiceovers with the videos were although the autoantigens responsible for the initiation and Abstracts of scientific papers 2008 AALAS National Meeting preferred by 65% of students, and short video clips embedded been previously described in these animals; however, there are in the text over compilation videos were preferred by 87% of currently no reports pertaining to other clinical findings when students. Also, 95% responded positively to review questions these rabbits are placed on a high-cholesterol diet. In the current provided after each section of the course. Critiques included study, Watanabe rabbits (n = 14) were placed on a diet contain- improving video resolution and the quality of the voiceover ing 0.5% cholesterol. In addition to the high-cholesterol diet, performance. These preliminary results support the continued 2 groups of these rabbits also received atherosclerotic plaque- development and implementation of electronic and interactive reducing drugs. Bloodwork, serial cholesterol, and triglyceride rodent surgical technique courses to the growing laboratory levels were monitored throughout the study. After 2 mo, the animal and biomedical research communities. rabbits began to develop some unusual lesions. Complete oph- thalmic examinations revealed that all rabbits had a significant PS38 Cesarean-derived SPF Sheep for Biomedical Research lipid arcus and uveitis, and 2 out of the 14 had scleral and uveal eruptive xanthomas. Six rabbits developed complete bullous RJ Hurley*1, JJ Hurley2 retinal detachments and glaucoma secondary to the uveitis. The 1CCM, Massachusetts General Hospital, Charlestown, MA; 2 retinal detachments and secondary glaucoma led to vision loss New England Ovis, LLC, Rollinsford, NH in this population. In addition, integumentary xanthomas and weight loss were seen clinically. At necropsy, affected animals Great strides have been made in recent decades to minimize or had granulomatous inflammation in the pyloric region lead- eliminate unintended environmental and biological variables in ing to partial gastric obstruction. Histological analysis of the research animals. Pathogens have been eliminated from many ocular lesions revealed that the sub-retinal space was markedly commonly used species. This has resulted in healthier animals, expanded by lipid-filled macrophages and cholesterol clefts. of which fewer are needed to generate reproducible and sta- Additional histopathological findings included severe diffuse tistically significant data. Many of these advantages, however, hepatic lipidosis and lipogranulomatous inflammation of the have not been realized in research using sheep because their meninges, ependymis, renal papilla and choroid. Although clini- commercial production does not employ the same “barrier” ap- cal complications are expected in these rabbits, especially when proach applied to smaller mammals. In particular, it has been they are fed a high-cholesterol diet, the severity and extent of the experience at our facility that pulmonary and cardiovascular systemic involvement seen in these animals was unprecedented studies using sheep are very dependent on the health status of and warrants disclosure to the scientific community. the animal subject. Sheep from commercial flocks would ar- rive clinically normal, but, following shipment-related stress, changes in housing and diet, and experimental manipulation, PS40 Use of Tricaine Methanesulfonate for Euthanasia of they often developed respiratory signs. In addition, some were Reptiles discovered during thoracotomy to have bacterial or mycoplasmal NE Hahn* pneumonia that excluded them from sensitive physiological studies. Other occasional infections such as contagious ecthyma Office of Laboratory Animal Care, University of California and clinical coccidiosis imposed additional health management Berkeley, Berkeley, CA burdens in a vivarium setting. In response, a flock of cesarean- Tricaine methanesulfonate (MS-222) injected into the intrac- derived Polypay sheep was established. Lambs were aseptically oelomic (ICm) cavity of reptiles was evaluated as a chemical and immediately removed from their ewes and hand-raised at a euthanasia method. This euthanasia method was developed separate facility isolated from other ruminants. Extensive test- as an alternative to sodium pentobarbital, which due to Drug ing (serology, PCR, culture) has shown these animals to be free Enforcement Agency control is problematic to use, particularly of bacterial, viral, and parasitic organisms commonly found in for international fieldwork. Three western fence lizards, 2 des- conventional flocks. Monitored and excluded agents include all ert iguanas, 4 garter snakes and 6 geckos were successfully intestinal parasites (helminthes, cryptosporidia, giardia), lung euthanized using this protocol. The reptiles were given an ICm worms, ectoparasites, bluetongue virus, bovine viral diarrhea injection of 250 to 500 mg/kg of 0.7 to 1% sodium bicarbonate- virus, contagious ecthyma, ovine progressive pleuropneumonia, buffered MS-222. Loss of response to toe or skin pinch occurred Toxoplasma, Brucella, chlamydia, Q-fever, Mycobacterium paratu- for most reptiles within 2 to 3 min, at which point a second ICm berculosis, Leptospira, and all respiratory bacterial pathogens injection of 0.1 to 1.0 ml unbuffered 50% (v/v) MS-222 solu- (culture negative). Coccidia were detected 1 y into the project, tion was administered. Respiratory and cardiac arrest occurred but no clinical cases have been seen. When used in research pro- within seconds in all animals, and was confirmed visually after tocols, these animals have been free of clinical signs and lesions, opening the coelom. This 2-stage protocol for the euthanasia of providing investigators with consistently reliable quality along reptiles MS-222 can be easily followed in the laboratory or field. with an improvement in animal welfare. In conclusion, MS-222 offers an acceptable alternative to sodium pentobarbital for euthanasia of reptiles. PS39 Clinical Complications in Watanabe Heritable Hyper- lipidemic Rabbits on a High-cholesterol Diet PS41 Does Sex Matter in a Rhesus Macaque (Macaca mulatta) Model of Focal Stroke? LGallaugher*1, C Forrider1, S Stone2, L Wancket2, D DaCunha1, S Lewis1, B Farrar3, M Kariisa3, S Rajagopalan3 SJ Murphy*1, JR Kirsch1, W Zhang1, M Grafe1,2, PD Hurn1 1UniversityLaboratory Animal Resources, 2College of Veteri- 1Anesthesiology and Peri-operative Medicine, 2Pathology, Or- nary Medicine, 3College of Medicine, The Ohio State University, egon Health and Science University, Portland, OR Columbus, OH Gender differences, sex steroid effects, and sex-specific candi- The Watanabe heritable hyperlipidemic (WHHL) rabbits are the date therapeutics in experimental ischemic stroke have been most commonly used genetically altered strain of rabbits used studied primarily in rodents. Many preclinical stroke studies today for atherosclerosis research. Clinical complications such as involving nonhuman primates use only males, do not indicate lipid keratopathy, eruptive xanthomas and hepatic lipidosis have sex, or do not stratify outcomes relative to sex. The lack of data Abstracts of scientific papers 2008 AALAS National Meeting in female higher order species severely limits the translation cord, spinal nerve roots, and peripheral nerves were submitted of experimental data to women requiring treatment for stroke for pathologic evaluation. Light and electron microscopy demon- or neuroprotection during stroke. In this feasibility study (n = strated severe, diffuse spongiform changes in the central nervous 3 per group), we developed a model of transient focal cerebral system only. Widespread intracellular vacuolation of the astro- ischemia in adult male and female rhesus macaques that consis- cytes was seen predominantly in the gray matter of the brain and tently includes white matter injury. We also questioned whether spinal cord. The brain stem (medulla oblongata), cerebellum, and gender-linked differences in histopathological outcomes could be midbrain were more severely affected than the cerebral cortex. evaluated in this model for future, larger preclinical trials. Male Future characterization will focus on differentiating a primary and female rhesus macaques aged 7 to 11 y were subjected to 90 astrocyte membrane dysfunction from vacuolation secondary to min of reversible middle cerebral artery occlusion (MCAO) using a metabolic disorder. These ataxic OFA rats constitute a novel the transorbital approach with temporary aneurysm clip place- model for studying the etiology, pathology, and treatment of ment. Histological evaluations of the caudate nucleus, putamen, astrocyte associated neurodegenerative diseases. globus pallidus, claustrum, internal capsule, external capsule and hemisphere were done 4 d following MCAO using cresyl violet PS43 Normal Feeding and Body Weight in Fischer 344 Rats staining. Intra-ischemic and reperfusion blood glucose, blood Lacking the Cholecystokinin-1 Receptor Gene pressure, heart rate, oxygen saturation, and rectal temperatures were determined and found equivalent between females and J Blevins3, J Overduin3, J Fuller2, D Cummings3, K Matsumoto4, males. Although there were no differences in regional infarction DH Moralejo*1 volumes between male and female subjects in this small sample, 1Comparative Medicine, 2Medicine, University of Washington, females showed greater variability in infarction volume (mm3; Seattle, WA; 3Division of Endocrinology/Metabolism, VA Puget % contralateral structure) than did males in the caudate nucleus Sound Health Care System, Seattle, WA; 4Division for Animal (125 ± 109 mm3; 45 ± 38% compared with 217 ± 63 mm3; 97 ± Research Resources, The University of Tokushima, Tokushima, 16%), putamen (283 ± 184 mm3; 62 ± 42% compared with 499 ± Japan 58 mm3; 106 ± 11%), and hemisphere (781 ± 437 mm3; 10 ± 8% Cholecystokinin (CCK) is hypothesized to inhibit food intake by compared with 1261 ± 383 mm3; 18 ± 6%). This is the first report acting at CCK1 receptors (CCK1R) on vagal afferent neurons that to compare cerebral ischemic outcomes in female nonhuman innervate the gastrointestinal tract and project to the hindbrain. primates to that of their male counterparts. Infarction volume Recent studies have shown that OLETF rats, which carry a spon- variability in male compared with female macaques was com- taneous null mutation of the CCK1R, are hyperphagic, obese, and parable to what is observed in human focal cerebral ischemia predispositioned to type 2 diabetes. In contrast, we have gener- and in other nonhuman primate proximal MCAO models. The ated a congenic strain in which the OLETF-derived CCK1R gene female macaque ischemic stroke model could be used follow- identified by the QTL (Niddm10/of) on chromosome 14 was in- ing rodent studies in providing vital preclinical data for clinical trogressed into the normoglycemic and nonobese Fischer 344 rat trials in women. genetic background, producing a CCK1R-deficient Fischer 344 rat (F344.Cck1r-/-) with a lean and normoglycemic phenotype. Here PS42 Characterizing the Neurological Phenotype of the Spon- we characterized the behavioral and neurobiological phenotype taneous Mutant Ataxic OFA Rats of this rat strain more fully. F344.Cck1r-/- and F344.Cck1r+/+ ST Chou1, KL Walter*2, E Ahearn1, A Harper1, R Driscoll1, LA rats did not differ in body weight and showed comparable weight Conour1 gain when maintained on Ensure Plus for 3 wk. No difference 1 Charles River, Wilmington, MA; 2 was found in 24-h food intake and dark-phase meal frequency Harleysville Veterinary or meal size between F344.Cck1r+/+ and F344.Cck1r-/- rats. Hospital, Harleysville, PA As expected, intraperitoneal injections CCK-8 inhibited intake A spontaneous autosomal recessive mutation in a colony of out- of chow and Ensure Plus in F344.Cck1r+/+ rats but not in F344. bred Oncins France Strain A rats (Crl:OFA[SD] resulted in a novel Cck1r-/- rats. CCK-8 failed to induce Fos in the area postrema neurological phenotype. A clinical scoring system was used to and the gelatinosus, commissural and medial nucleus tractus establish a timeline for onset and progression of phenotype. The solitarius of F344.Cck1r-/- rats, whereas robust Fos responses average body weight of the mutants is higher at weaning, but were seen in these areas in the F344.Cck1r+/+ rats. Together, falls behind in growth curve by 6 to 7 wk of age. There is no gross these results confirm that CCK-1R mediates CCK-8-induced difference in clinical disease progression by sex. Whole body inhibition of food intake and Fos activation in the hindbrain, but tremors are noted at weaning and progress into an exaggerated we find that in rats with a F344 background, genetic ablation of “bouncing” of hindlimbs during ambulation. This is evident in CCK1R is not associated with obesity or hyperphagia. >90% of the mutants by 7 wk of age. Gait analysis at 4 wk of age shows the affected rats to have a shorter stride length and wider PS44 A Novel Neonatal Non-invasive Motor Function Test with base stance than the unaffected rats (P ≤ 0.01). By 8 wk of age, Potential Application for Neuromuscular Diseases 100% of the affected rats exhibit ataxia and incoordination. Print interpretation from gait analysis becomes increasingly difficult C Pater*1, S Ramboz1, M Winberg2, A Chen1, N Edgar1, B El- as their gait leaves behind multiple sets of hindlimb prints with Khodor1 each step taken. Despite abnormal gait, open field assessment 1PsychoGenics, Inc., Tarrytown, NY; 2SMA Foundation, New showed a significantly higher number of rearing events (P ≤ 0.01) York, NY and no difference in distance traveled between rat groups within Comprehensive motor and sensorimotor studies have been per- 10-min test sessions. The neurological phenotype fluctuates in formed on a variety of neonatal neuromuscular disease models. progression rate and severity as animals age. By 6 mo of age, We realized that the existing neonatal neuromuscular tests do not clinical signs include loss of body condition and righting reflex, adequately measure muscle strength and possible subtle drug ef- recumbency, and absence seizures. The onsets of these clinical fects. Such a test would ideally be a scored, reflex-based motor test signs are used as humane endpoint criterions. The brain, spinal that could be easily and reliably administered to rodent infants as Abstracts of scientific papers 2008 AALAS National Meeting early as the day of birth and would remain valid for at least the first Most experimental approaches in stroke research are carried out 2 wk of life. The hind-limb suspension test (the tube test) evaluates in rodents, but given differences between rodents and primates, the proximal hindlimb muscle strength, weakness, and fatigue in nonhuman primate stroke models may provide a valuable tool mouse or rat neonates. It uses simple equipment found around the to study therapeutic approaches in stroke. We have developed lab, such as 50 ml conical tubes or beakers for larger mice. There a reproducible surgical method for cerebral infarction of the are 2 consecutive trials. The mouse is placed head down, hanging M1 branch of the middle cerebral artery (MCA) using 7 African by its hind limbs in a 50 ml conical tube with a cotton ball at the green monkeys (Chlorocebus aethiops). A left fronto-temporal bottom to cushion its fall. Three parameters are evaluated: latency craniotomy was performed and after dural exposure, the M1 to fall from edge of tubes; number of pulls that animal performs branch was surgically exposed. A vascular clip was applied to the during its attempt to escape the tube; and hindlimb score (HLS), distal portion of M1 for 2 h. After releasing the clip, cranioplasty which assesses the positioning of the legs and tail. Observations and repair of the temporalis muscle were carried out. Neuro- with neonates exposed to the tube test showed that WT mice readily logical examinations and behavioral testing were developed held on with their hind limbs from postnatal day (P) 2 until P12. The for functional assessments. Postsurgically, 4 animals showed effects of various factors known to influence motor performance typical right hemiparesis, similar to human patients with M1 in neonates such as hypothermia have been evaluated in the tube infarction. Magnetic resonance imaging (MRI) demonstrated test. In addition, using a neonatal mouse model of spinal muscular extensive infarction throughout the left MCA territory (complete atrophy (SMA), we showed that the performance of the KO was occlusion group). Three animals showed mild hemiparesis, and significantly different from WT in the 3 parameters of the tube test MRI demonstrated smaller areas of infarction (partial occlusion as early as P4. This test would enable anyone to obtain a reliable group). During M1 clipping, we observed darkening of cerebral quantitative assessment of motor function in mice as early as the venous blood, small vessel collapse, and blood pooling of corti- day of birth. We would like to acknowledge the SMA foundation cal veins, evidenced by back-and-forth movement of stagnant for their help and support in this project. blood during brain pulsation. In animals with partial occlusion, we did not observe blood pooling of cortical veins. These data PS45 Validation of Chronic Mild Stress in Rodents as a Trans- indicate that a highly reproducible MCA occlusion model with lational Model of Clinical Depression consistent functional deficits in the nonhuman primate can be obtained with the clipping of the distal portion of M1 for 2 h, TA Swanson*1, G Lessard1, K Muravnick2, RP Giovanelli2, L if the above visual criteria of the infarcted cortex are met. The Badura3, PA Seymour4, S McLean4, M Moalli1 reproducibility of the model is important for future experimental 1Comparative Medicine, 2Pathology, 3Translational Medicine, therapeutic strategies. 4Neuroscience, Pfizer Global Research and Development, Gro- ton, CT PS47 The Role of Positive Reinforcement Training in Creating Chronic mild stress (CMS) is considered to be a predictive ro- a Reproducible Surgical Stroke Model dent model that recreates some of the pathophysiology present S Rubino*1,2, M Sasaki2, J Kocsis2 in human depression. Anhedonia, a core symptom of clinical 1Comparative Medicine, Pfizer, Inc., Griswold, CT; 2Neurosci- depression, is characterized by the loss of interest in normally ence, Yale University, New Haven, CT rewarding stimuli. This can be quantitatively assessed in the rodent by measuring decreases in consumption of sucrose During the development of a reproducible surgical stroke solution. Clinical depression has also been associated with a model, it was necessary to develop neurological examinations disruption in circadian rhythm. These two outcome measures and behavioral tests for functional assessments. As part of the were used to validate chronic mild stress as a translational model neurological exam, African green monkeys (Chlorocebus aethiops) of clinical depression. Experiments focused on optimization of were trained to touch a target stick with their left hand or right the stress paradigm, adding the use of telemetry to monitor ac- hand and to track an object through horizontal and vertical tivity rhythms, and validation of biomarkers. Approved by the movements. Animals were also exposed to a puzzle feeder that Pfizer IACUC, this model consists of exposure to a series of mild, required the use of 2 hands to remove food rewards. For func- random, unpredictable stressors which result in a state of anhe- tional assessments, animals were trained to perform fine motor donia, disruption of normal circadian rhythms, and weight loss. grasping tasks including a well/reach task and a staircase task. Stressors include periods of cage tilt, exposure to stroboscopic These tests were timed and assessed prior to surgery as a baseline light, white noise, and wet bedding. Strain and sex differences for postoperative data collection. Behaviors were taught using in response to CMS were examined in both Sprague-Dawley positive reinforcement. The training plans developed for these and Wistar rats. Biomarkers such as serum and urinary corti- tasks allowed us to successfully train all animals selected for the costerone and lymphocyte counts were assessed. As predicted, preliminary project. sucrose consumption was decreased in response to the chronic stress paradigm. However, reversal by antidepressant therapy PS48 Establishment of a Nonhuman Primate Cognition Labora- was inconsistent with published data and warrants further in- tory vestigation of this model. JA Vivian*1, JL Basile1, MR Tinsley1, R Morris1, N Poy2, D Brandt2, C Waugh2, L Santarelli1 PS46 Establishment of a Reproducible Surgical Stroke Model 1Neuroscience, 2Comparative Medicine, Roche Pharmaceuticals, in the Nonhuman Primate Palo Alto, CA M Sasaki1, J Kocsis1, S Rubino*2,1 1Neuroscience, Due to their expanded social and behavioral repertoire, and exten- Yale University, New Haven, CT; 2Comparative sive frontal cortical development, the establishment of a cognition Medicine, Pfizer, Inc., Groton, CT facility using nonhuman primates would appear relevant and an Stroke is a leading cause of death in adult humans, and extensive important endeavor for basic, pharmaceutical, and translational research is needed to develop new therapies for stroke patients. (preclinical to clinical) science. However, issues including the 3 Rs, Abstracts of scientific papers 2008 AALAS National Meeting and the special requirements for monkey research, have proven (9 mitral, 2 aortic). We originally assumed that the main cause of challenging. In this presentation, an overview will be provided for mortality was due to surgeon error and immediately instituted more the development of a research effort in neuroscience (specifically, conservative measures for de-airing the heart and assuring hemo- cognition) using nonhuman primates, explaining why monkeys are stasis. After analysis of the data, we saw that while this was true used, and providing some detail for key decision points, includ- for aortic valves, for mitral implants the key cause of mortality was ing the choice of monkey strain and gender; handling, housing sheep-related. Since introducing the procedure changes we have and enrichment concerns; veterinary and staff oversight; and reduced surgeon-related mortality, and surprisingly also reduced test model development (training and testing strategies). Briefly, the incidence of fatal GI dysfunction and pulmonary edema. The socially housed adult male cynomolgus monkeys were chair- and exact mechanism for this improvement is unclear. This study shows test system-trained using a modified Wisconsin General Test that it is important to review records to determine the actual causes Apparatus, operant chambers (lever boxes) and computer touch- of mortality and that solving one problem can actually improve sensitive screens (CANTAB). Subsequently, they were tested with outcomes in other areas. prototypic drugs effective in the treatment of Alzheimer disease and schizophrenia. Monkeys readily learned the various tasks, and PS51 Primate Tuberculosis Testing Comparison: Gamma In- demonstrated cognition-enhancing effects after drug treatment. terferon Assay and Mammalian Tuberculin Taken together, these results marshal support for the unique value of monkeys in the assessment of complex cognitive behavior, and ST Shipley*1, T Coksaygan1, C McLeod1, DK Johnson2, LJ De- hopefully, their predictive and translational value for the clinic. Tolla1 1Comparative Medicine/Veterinary Resources, University of PS49 Nasal Intubation in the Rabbit: A New Twist on a Dif- Maryland School of Medicine, Baltimore, MD; 2Scientific Advi- ficult Procedure sor, Harlan, Chester Springs, PA J Stephens-DeValle* Tuberculosis (TB) is a highly infectious disease in nonhuman pri- mates (NHPs) causing high morbidity and mortality that is also Department of Clinical Investigations, DD Eisenhower Army important for its zoonotic potential. A comprehensive diagnostic Medical Center, Fort Gordon, GA program is essential for primate colony health management. The Although nasal intubation has been described in the rabbit, there gold-standard assay for TB screening in NHP colonies is the delayed is limited literature concerning this technique. At our facility we hypersensitivity test using mammalian old tuberculin (MOT) to needed to intubate rabbits for a protocol that required the animals marshal a delayed dermal hypersensitivity response over a 72-h to be placed under general anesthesia twice, with a 2-min time span period. A new primate diagnostic test is available measuring gamma between the initial intubation and the final intubation. Initially, the interferon release from sensitized lymphocytes in response to ex- animals were orally intubated; however, after encountering difficul- posure to TB antigens. This interferon release is then measured by ties with oral intubation, nasal intubation was performed. During ELISA assay. This gamma interferon assay patterns itself after an the development of this technique, we learned several essential key analogous test of human tuberculosis testing. The antigens used points: total relaxation, modification of the classic blind position, for the primate gamma interferon assay are 2 strains of myco- the proper approach, and securing the tube. Contraindications for bacterium (M. bovis and M. avium complex), whereas the human nasal intubation include the possibility of introducing pathogens assay uses purified protein derivative of human mycobacterium into the lungs and the need for high oxygen flow rates. However, (M. tuberculosis). A 2-y comparative evaluation of tuberculosis test- clinically, neither item was an issue. At our facility, after a few ing in primates was performed by alternating between MOT and modifications and practice, nasal intubation was preferred as an primate gamma interferon assays during both primate quarantine easy, less traumatic method of rabbit intubation. and semi-annual facility TB testing postquarantine. Results from both types of testing were found to be equivalent, and both tests PS50 Analysis of Early Postoperative Mortality in a Sheep (Ovis correctly identified animals positive for naturally acquired TB aries) Model of Mitral and Aortic Valve Replacement (culture-confirmed infections). The advantage of the interferon as- say is the ability to have results within 24 h of blood draw, which JM Olin*, A Okos, S Meyer is 48 h faster than the MOT assay. However, the initial steps of the Biological Resource Center, Edwards Lifesciences, Irvine, CA interferon assay must be performed in-house within 6 to 12 h of blood collection using validated lymphocyte culturing procedures Higher than expected mortality in early 2008 prompted us to de- by highly trained technicians. The interferon assay has limits based velop a plan to improve early postoperative outcomes for surgical on cost of reagents, technician labor time, and intermittent delays of valve replacement in sheep. The records of 99 consecutive animals test kit availability. Although the USDA has licensed this interferon from Jan 2007 through Apr 2008 were reviewed to analyze the cause test in several species of NHPs, the MOT (delayed dermal hyper- of death. Early mortality was considered to be any death or illness sensitivity) assay remains the principal recognized diagnostic test requiring euthanasia within the first 7 d postimplant. Mortality was by regulatory agencies such as CDC and USDA. For these reasons categorized as surgeon error, device complication, or spontaneous or the gamma interferon assay cannot be used alone in absence of the sheep-related. Surgeon error included hemorrhage, emboli, or injury gold standard mammalian intradermal test. to other cardiac structures. Some of the devices were prototypes or atypical configurations; any mortality that was linked to a unique device feature was categorized as a device complication. Spontane- PS52 Methicillin-resistant Staphylococcus aureus Infection ous or sheep-related mortality included untreatable GI dysfunction and Cranial Abscessation in a Rhesus Macaque (Macaca and pulmonary edema, and deaths for which no other cause could mulatta) be assigned. All implants were performed by the same surgical BH Jasmin*1, JI Gold2, CH Vite3, SC Rankin4, J Baxter1, F Han- team using sheep from the same source. In the 16-mo period, 57 kenson1,4 mitral and 42 aortic valve replacements were performed, with early 1University Laboratory Animal Resources, 2Department of Neu- mortality distributed across surgeon error (4 mitral, 6 aortic), device complication (1 mitral, 3 aortic) and spontaneous or sheep-related roscience, 3Department of Clinical Studies-Philadelphia, Section Abstracts of scientific papers 2008 AALAS National Meeting Chief of Neurology, 4Department of Pathobiology, University of that the quinolone resistance for this isolate was plasmid-mediated Pennsylvania, Philadelphia, PA and transferable to a quinolone-susceptible E. coli. PMQR is clini- cally important when treating macaques prophylactically and for A 7-y-old female rhesus macaque (Macaca mulatta) presented for bacterial infections, especially given the common clinical usage clinically abnormal behavior within 24 h of surgery to relocate a of fluoroquinolones. Of the antibiotics tested, the shigella isolates cylinder in a pre-existing head stage implant. This animal was used were only susceptible to trimethoprim/sulfamethoxazole and that in an IACUC-approved behavioral study exploring mechanisms antibiotic was used to eradicate S. flexneri from the macaques. To of learning and visual discrimination. Clinically, the animal was our knowledge, this is the first report of PMQR in S. flexneri isolated intermittently depressed over the first 3 postsurgical weeks despite from monkeys. antibiotic therapy and pain management. Serum chemistry and complete blood count were within normal limits. During routine cleaning and maintenance of the implant cylinders, abnormal tissue PS54 Corynebacterium spp in Ferrets (Mustela putorius furo) growth within the caudal cyclinder was noted. Cultures from the and Macaques (Macaca mulatta) cylinders grew methicillin-resistant Staphylococcus aureus (MRSA). RP Marini*1, EM Buckley1, Y Peters1, P Cassiday2, JG Fox1 Paired samples were collected and submitted to a contract diag- 1Comparative nostic laboratory specializing in culturing MRSA for confirmation Medicine, Massachusetts Institute of Technology, and bacterial typing. Topical flushing treatments of the dura around Cambridge, MA; 2pertussis and Diphtheria Laboratory, Centers the implant were attempted with sterile saline, 1% chlorhexidine, for Disease Control and Prevention, Atlanta, GA and appropriate antibiotics without successful MRSA eradication. The genus Corynebacterium contains organisms responsible for A routine MRI for assessment of cylinder placement unexpectedly diphtheria of humans (C diphtheriae and, sporadically, C ulcerans identified a 1.7-cm space-occupying lesion in the left occipital lobe. and C pseudotuberculosis) and a number of diseases of veterinary A repeat MRI with contrast was then performed and the lesion importance. Recent publications reporting the zoonootic trans- was classified as an abscess with evidence of hemorrhage and mission of the diphtheroid bacterium Corynebacterium ulcerans meningitis. Approximately 5 mo after initial MRSA diagnosis, the from cats and dogs to people motivated a survey of bacteriologi- animal’s cranial implant was removed. During removal, purulent cal isolates from cephalic implants (n = 73) and oropharynges (n = material found at the implant site yielded MRSA. MRSA was also 78) in a colony of macaques (Macaca mulatta) and ferrets (Mustela isolated from a sample of the actual hardware pieces. Following putorius furo; n = 3 and n = 1, respectively). Bacteria identified implant removal, the skull and dural surface were thoroughly from cephalic implants by the apiCoryne system included C flushed with alternating 1% chlorhexidine and saline. Cultures of ulcerans (n = 40), C renale (n = 3), C pseudotuberculosis (n = 6), these disinfected tissues were negative for MRSA. Due to its loca- and C striatum (n = 1) in macaques and C ulcerans (n = 3) and C tion, the cranial abscess could not be readily accessed for culture pseudotuberculosis (n = 1)in ferrets. Oropharyngeal recovery of but was presumed to be related to the chronic bacterial infection coryneforms from macaques included C ulcerans (n = 7), C propin- in this animal; no neurological abnormalities have been observed quum (n = 2) and C auris (n = 1). C pseudotuberculosis was cultured in this animal. This case exemplifies the difficulty in eradication of from the oropharynx from the ferret harboring that organism in MRSA, an emerging infectious disease in laboratory animal species, its implant. Mixed infections of Corynebacterium spp and other from nonhuman primates with chronic cranial implants. microorganisms were common in both macaques and ferrets. Human contacts of these animals (n = 34) were subjected to PS53 Plasmid-mediated Quinolone Resistance in Shigella oropharyngeal culture to evaluate carriage and zoonotic spread flexneri Isolated from Macaques (Macaca mulatta) of Corynebacterium spp. No humans shared organisms with ani- mals with which they had direct contact, although carriage of C HR Martin*, Z Shen, EM Buckley, JG Fox pseudotuberculosis (n = 1), C jejeukium (n = 3) and C striatum (n = 1) Division of Comparative Medicine, Massachusetts Institute of was demonstrated. Diphtheria toxin activity of select C ulcerans, Technology, Cambridge, MA C renale, and C pseudotuberculosis isolates were negative in Elek tests. Sequence analysis of 16s ribosomal RNA genes of select Plasmid-mediated quinolone resistance (PMQR) is an emerging is- macaque isolates revealed over 99% homology with reference sue with bacterial species in the family Enterobacteriaceae. Typically, sequences for C ulcerans (n = 5) and 98.9% homology for C renale quinolone resistance is chromosomally mediated; however, PMQR was first reported in 1998 in Klebsiella pneumoniae. The first PMQR (n = 1). To our knowledge, this is the first demonstration of puta- determinant, QnrA, was a 218 amino acid protein protecting DNA tive C ulcerans and C pseudotuberculosis infection of ferrets and from quinolone binding. Two other Qnr-type determinants, QnrB the first demonstration of cephalic implant infection of macaques and QnrS, confer resistance to the quinolone, nalidixic acid, and with C renale and C pseudotuberculosis. Molecular epizootiology, increase the mean inhibitory concentrations of fluoroquinolones. evaluation for virulence factors, and confirmation of isolate Five different isolates of multi-drug resistant Shigella flexneri were identity with 16s ribosomal RNA gene sequencing is ongoing. isolated from the feces of 3 macaques. Antibiotic susceptibility test- Personal protective practices used in the vivarium appear to have ing was performed using disc diffusion and resistance to ampicillin, prevented zoonotic transmission of these organisms. amoxicillin + clavulanic acid, cephalothin, gentamicin, tetracycline, enrofloxacin and nalidixic acid was noted. PCR was performed on PS55 Diabetic Ketoacidosis in a Pregnant Rhesus Macaque the shigella isolates for the presence of the qnrA, qnrB, and qnrS (Macaca mulatta) genes. Genomic DNA from all isolates was positive for qnrS. Plas- E Paster*, L Colgin, J Wilk, G Maginnis mid DNA was extracted and 4/5 isolates tested positive for qnrS. A standard transformation protocol was followed using plasmid Portland VA Medical Center, Portland, OR DNA and E. coli competent cells. E. coli competent cells were used A 13-y-old pregnant female rhesus macaque presented to the as a control and were susceptible to nalidixic acid by disc diffusion. clinic for partial anorexia. A fetal ultrasound had been performed A transformed E. coli colony was isolated in the zone of susceptibil- under sedation earlier that week. She had previously delivered ity for nalidixic acid and confirmed to be resistant to nalidixic acid. live infants vaginally in February of 2005 and 2006 and the infants’ The zone of enrofloxacin sensitivity was decreased, demonstrating birth weights were 0.54 kg and 0.60 kg, respectively. Ultrasound Abstracts of scientific papers 2008 AALAS National Meeting revealed that the placental attachments were normal, there was no R Duran-Struuck*1, PS Cho1, SG Moran1, A Teague1, K Fergu- evidence of fetal distress, and the estimated gestational age was son1, BV Lutton1, I Hanekamp1, KJ Wikiel1, B Horner1, DP Lo1, 118 d, based on biparietal diameter measurements. Two days later, M Duggan1, SJ Arn1, B Billiter1, S Houser1, SV Westmoreland2, there was suspected vomitus in the cage and the patient developed DH Sachs1, CA Huang1 diarrhea. A follow-up ultrasound was performed and no fetal 1Surgery, Transplantation Biology Research Center, Massachu- heartbeat was detected. A cesarean section was performed and setts General Hospital—Harvard Medical School, Boston, MA; a nonviable 0.273 kg fetus was removed. An in-house chemistry 2Pathology, New England Primate Center, Harvard Medical analysis performed during surgery revealed metabolic acidosis. School, Southborough, MA Blood chemistry results revealed hyperglycemia, azotemia, hypertriglyceridemia, hyperphosphatemia, hyperkalemia, and The Massachusetts General Hospital (MGH) miniature swine panhypoproteinemia. Hematologic abnormalities demonstrated characterized at the major histocompatibility complex (MHC) a mild leukocytosis with a minimal left shift and a glycosylated serve as a model system for human clinical transplantation. hemoglobin of >13%. Urinalysis showed a specific gravity of 1.011, Sublines of these animals have been successfully inbred and now mild glucosuria, and significant ketonuria. Gestational diabetic ke- permit adoptive transfer studies in a large animal preclinical toacidosis was suspected; however, as previous blood profiles were model. We report here 5 cases of spontaneous chronic myeloid lacking in this patient, pre-existing diabetes mellitus could not be leukemia with similarities to human disease that have occurred ruled out. Fetal macrosomia, a consequence of gestational diabetes in our inbred miniature swine herd. Four of these cases occurred in humans, was not present in this case. Despite aggressive fluid within the highly inbred histocompatible sublines. All animals therapy, the patient continued to decline and was euthanized due presented with symptoms of weight loss, lethargy, and marked to poor prognosis. Necropsy findings of the mother included acute leukocytosis. Samples were obtained for complete blood count, tubular necrosis, hepatic lipidosis, lymphatic ectasia and fibrin peripheral blood smear, and flow cytometry analysis. Animals thrombi in the heart and liver, along with placental infarcts. A that were confirmed to have neoplasms were then euthanized. At review of the current literature reveals that this is the first reported necropsy, all animals had diffuse disease involvement of vascu- case of gestational DKA in a rhesus macaque. To improve maternal larized organs and severe hepatosplenomegaly. Peripheral blood and fetal outcome, screening for diabetes should be considered mononuclear cells and tissues were phenotyped by fluorescence in obstetric cases that have a history of large fetuses or in those activated cell sorting (FACS). In addition, all affected tissues were patients that develop signs of morbidity. assessed histopathologically. Taking under consideration the clini- cal presentation, FACS, gross pathology and histopathology we diagnosed these cases as chronic myelogenous leukemias. Some PS56 Acute Respiratory Distress Syndrome in Two Rhesus of these tumors have been successfully established as in vitro cell Macaques (Macaca mulatta) lines and are candidates to become the first leukemias to be used as JJ Fremont*, RP Marini, JG Fox, AB Rogers transplantable tumors in inbred and MHC-defined large animals. Department of Comparative Medicine, MIT, Cambridge, MA Thus, MGH MHC-inbred miniature swine and established porcine tumors may provide useful preclinical translational models to Acute respiratory distress syndrome (ARDS) is an important study immunotherapeutic approaches to treat human leukemia. and potentially life-threatening complication in humans subse- quent to a variety of primary insults, including noxious fume PS58 Epidemiologic Investigation of a Multidrug-resistant inhalation, infection, and trauma. Clinically, animals with ARDS Staphylococcus aureus Outbreak in Two Macaque Colonies present with progressive dyspnea and pulmonary edema. Physi- ologically, ARDS is characterized by severe hypoxemia (partial JR Baxter*1, BH Jasmin1, F Hankenson1, A Behrman2, S Rankin3 pressure of oxygen [PaO2]/fractional concentration of oxygen 1University Laboratory Animal Resources, University of Penn- in inspired air [FIO2]<200). Histopathologically, the disease is sylvania, Philadelphia, PA; 2Occupational Medicine, Hospital of characterized by 2 phases. In the initial phase, the lungs are University of Pennsylvania, Philadelphia, PA; 3Clinical Microbi- edematous admixed with variable numbers of neutrophils, ology, Ryan Veterinary Hospital, Philadelphia, PA alveolar hemorrhage, and atelectasis. The disease progresses to the fibrinoproliferative phase 7 to 10 d postinsult. Here we Methicillin-resistant Staphylococcus aureus (MRSA) is a multi-drug describe for the first time ARDS in 2 rhesus macaques associated resistant bacterium that has become a prevalent pathogen among with complications following experimental surgical craniotomy. people and is now emerging among the veterinary patient popula- ARDS in 1 monkey was attributed to sepsis resulting from tion, underscoring its potential as a zoonotic. This microorganism meningeal infection with Staphylococcus aureus, group A Strepto- has not been reported in nonhuman primates (NHP). MRSA was coccus pyogenes, and Corynebacterium ulcerans. ARDS in the other diagnosed in 3 clinically compromised animals within 2 macaque monkey was ascribed to direct neurogenic injury. Despite the colonies, Colony A and Colony B, in one of our facilities. The 2 different etiologies, both monkeys developed clinical features rhesus macaques in Colony-A were used for IACUC-approved of ARDS and syndrome-defining histopathologic criteria includ- neurobehavioral studies requiring invasive cranial implants, while ing edema with intra-alveolar neutrophils, fibrinohemorrhagic the cynomologus macaque in Colony B was used for IACUC- effusions with crescentic membranes, and interstitial vascular approved transplant research requiring immunosuppression. degeneration. ARDS may be underreported in macaques and Containment and eradication planning was critical because these other nonhuman primates. Recognition and aggressive treatment chronic studies involve numerous animals. Additional samples of ARDS at an early stage may improve survival rates in these were analyzed from colony animals using chrome agar enrichment highly valuable animals. techniques specific for MRSA. Colony B had no additional positive animals, while Colony A had 2 additional implants culture posi- tive from a pool of 5 clinically healthy animals, yielding an overall PS57 Immunophenotypic Characterization of Spontaneous colonization rate of 57% in Colony A. Body swabs were obtained Myeloid Leukemias in Adult MHC-defined and Histocompat- from the individuals with colonized implants to determine if ible Miniature Swine: Towards the Development of a Large Animal Transplantable Tumor Model NHP can be colonized in a pattern similar to humans colonized with MRSA. Nasal swabbing is the gold standard test for human Abstracts of scientific papers 2008 AALAS National Meeting colonization; however, NHP with colonized axillary and perineal structurally complex housing configuration with an exercise areas did not consistently have positive nasal cultures. Our vet- program. In the initial housing system, cats were housed either erinary and husbandry team reviewed personnel and equipment singly (males) or in isosexual (females) dyads or triads in pens trafficking and animal practices that may have contributed to the with minimal PVC furniture for climbing. Two new housing rooms outbreak. An interdepartmental collaborative effort was made to with 6 pens in each room were designed from recycled dog pens screen all personnel with NHP contact (n = 44) for MRSA. DNA so the cats could be divided into 2 groups. In each housing room fingerprint analysis of the single human isolate and all NHP the 6 pens were contiguous by wall-mounted tunnels and side isolates were of the healthcare-associated (HA) strain of MRSA. doors to allow the cats continuous free access among all of the Our investigation led to improved hand washing, sanitization, pens. Vertical complexity was achieved through placement of wall- aseptic technique, and heightened institutional awareness of the mounted staggered perches, varying height tunnels between pens, importance of consistent use of personal protective equipment to and commercially available climbing furniture with hammocks for prevent the potential transmission of zoonotic agents. resting and sleeping. Eventually all 16 cats were combined into 1 of the 2 rooms. The other room was converted into an exercise/ PS59 Animal Health and History: Electronic System for Captur- play room with an area rug, floor toys, and suspended toys. The ing Medical and Husbandry-related Data housing room and exercise/play room were connected to one another through a walled off ante-room to allow cats direct access J Sims*2, SA Mischler1, J Ashline1 to the exercise/play room 4 d per wk for 4 to 6 h per d. The incor- 1Drug Safety, 2Information Services, Wyeth, Chazy, NY poration of the exercise/play room into the enrichment program resulted in the desired weight loss to a healthy weight range and Our facility’s husbandry and veterinary activities in drug safety body condition score. Additionally, the response of care staff and were originally built around a paper-based system. All informa- regulatory agencies to this program was positive. tion for animals was recorded and maintained on paper, including medical records, scheduled tasks (physical exams, body weights, vaccinations, and so forth), and training activities. The paper PS61 Cage-level Barrier System for Primates and Other Non- system was an effective way to maintain compliance with the rodent Species Application for Biodefense Select Agent and Food and Drug Administration’s (FDA) 21 CFR Part 58 (Good Infectious Disease Studies (ABSL-3) Laboratory Practice for Non-clinical Laboratory Studies) and 21 DK Johnson*1, D Allen1, LJ DeTolla1, Y Gal1, J Huss1, SD Reyn- CFR Part 11 (Electronic Records and Electronic Signature) guide- olds2 lines. The paper system, however, was time consuming and made 1Harlan Sprague Dawley Inc, Indianapolis, IN; 2CAES Group, it difficult to perform certain tasks, including analyzing trends, M/E Engineering P.C., Syracuse, NY inspecting data for internal quality control, reviewing data for poststudy activities, and analyzing animal care staffing require- A cage-level barrier system for primates that is also suitable ments. Data for animals was also stored in disconnected systems for other large laboratory animals was engineered with reason- that required duplicate data entry and complicated data searches. able adaptations of flexible film isolators, along with minor Wyeth formed a team of veterinary, husbandry, and information modifications of conventional cages. The rapid growth of funded services staff to gather requirements and develop a custom ap- laboratory animal research for biodefense studies, as well as in- plication to replace the paper system. The project had a broad fectious disease investigations, require the use of special agents, scope so the agile software development framework was used to highly contagious infectious organisms, aerosols, toxins, and new incrementally deliver functionality. The result was a web-based vaccine testing in primates and non-rodent species. This com- application called Animal Health and History (AHH), which is bined flexible film isolator and cage system establishes a barrier currently at version 2.1. Wireless tablet PCs are used to record at the cage level for these larger laboratory animals. It allows the data. Other validated systems were also integrated with AHH to safe conduct of infectious disease studies and meets occupational minimize duplicate data entry and facilitate searches. AHH has health requirements for containment of infected animals. The been in production in a validated state since January 31, 2005. Data safety protection to personnel and the environment meet insti- and reports from AHH have been used to meet both USDA and tutional and governmental guidelines. A validation engineering FDA requirements. Quality control activities have been facilitated study was conducted to verify the operational effectiveness by by making the data available electronically. Retrospective pain and using the technology of Computational Fluid Dynamics (CFD). census information is easily recorded and reported to the USDA. This credible engineering program accurately determines airflow, Husbandry and veterinary activities required in FDA submissions velocity, distribution, thermal loads, and related aerodynamic are easily analyzed and retrieved. AHH continues to evolve as new parameters. Multiple examples of leak scenarios, including van- functionality is added. The ultimate goal is to provide a completely dalism, were evaluated. The CFD data provides documentation paperless system to capture husbandry and veterinary data, while for the successful approval of this cage-level barrier system. In leveraging related data in other systems and maintaining compli- addition, the system has application for large laboratory animals ance with applicable FDA and USDA guidelines. assigned to immunodeficiency studies. PS60 Development of a Complex Housing Arrangement with PS62 Working with State Regulating Agencies: The Value of Integration of an Exercise Program for Laboratory Cats State Biomedical Research Organizations M Wilkinson*3, L Wood1, A Boyd1, S Watson2, A Wilkinson1 JS Ellis* 1 Animal Resources Unit, Alcon Labs, Fort Worth, TX; 2De- PA Society for Biomedical Research, Camp Hill, PA partment of Psychology, University of Southern Mississippi, In addition to federal laws and regulations, in many states Hattiesburg, MS; 3Veterinary Sciences, Tulane National Primate research facilities must also comply with state laws and regula- Research Center, Covington, LA tions. Most often research facilities focus on the federal laws and Over the course of 36 mo, 16 domestic shorthair cats were transi- regulations, which are well known and discussed at national tioned from a standard feline housing system to a group-housed, meetings. State laws and regulations may not be as well known Abstracts of scientific papers 2008 AALAS National Meeting throughout the research community, and state agencies may not Animal Resources Center, University of Texas Medical Branch, fully understand the research process. Research facilities may Galveston, TX not even be aware that certain state agencies have regulations The Galveston National Laboratory (GNL) at the University of that apply to them. The state biomedical research associations Texas Medical Branch in Galveston Texas is 1 of 2 state-of-the-art can play a key role in educating both its membership and state national biocontainment laboratories constructed under grants agencies. This is only one of the many benefits of membership awarded by the National Institute of Allergy and Infectious in the state biomedical research organizations. This presenta- Diseases/National Institutes of Health (NIAID/NIH) that will tion will discuss 2 such examples that have recently taken place support Biosafety Level 2 through Biosafety Level 4 research. in Pennsylvania. The first example is the education of the PA While the GNL will provide much-needed research space for Department of Agriculture on why federally registered research naturally occurring emerging diseases, as well as for viral and facilities should be exempt from legislation designed to improve bacterial agents, attracting an exceptionally talented and mul- the conditions of dogs living in commercial kennels. The second tidisciplinary team of techs for high-biocontainment work is a case involves efforts to educate the PA Game Commission on daunting endeavor. Our initial criteria for a Animal Resources research needs, as well as to educate research facilities on the Biohazard Technologist required AALAS certification at the LAT Game Commission’s regulations. Since many voices working or LATG level, a minimum of 3 y experience in research, and 1 together are stronger than a single voice, working collectively as y experience in biocontainment. Finding ideal candidates that a research community makes sense. The value of state biomedi- possessed our requirements was found to be nearly impossible. cal research organizations is exemplified by the coordination of After seeing many applications that had met 1 or 2, but not all, this research community. of the requirements, we modified our criteria to fit the applicants we were seeing. These modifications opened a larger pool of PS63 A Novel Web-based Approach to Ethical Committee applicants for us, and we used novel recruiting strategies. We (IACUC) Functions: Managing Protocols, Meeting Minutes, took our show on the road, using platform sessions at AALAS and Inspections branch meetings and attending job fairs at regional universi- CM Pate*1, B Bell1, R Fosse2, A Staubi3 ties. Our plan was to hire the traditional applicants, give them 1Laboratory intensive biocontainment training in animal husbandry and Animal Science & Welfare, US, Sanofi-Aventis, procedures, provide AALAS certification “boot camp,” and pro- Bridgewater, NJ; 2 Laboratory Animal Science & Welfare, vide concurrent training in quality system and GLP standards. Northern Europe, Sanofi-Aventis, Montpellier, France; 3 a-Tune AALAS certification at the LAT level within 1 y is a condition Software AG, Darmstadt, Germany of employment. The proliferation of Animal Biosafety Level 3 The Consolidated Laboratory Animal Database (COLADA) is a and 4 labs throughout the country has left an increased gap of novel web-based solution we developed to provide our organiza- demand outweighing the supply of biocontainment technicians. tion with a flexible system that automates the office workflow in The most realistic solution is to hire traditional animal techni- a completely transparent manner. COLADA provides automated cians and train internally while time is not on your side. Doing reminders to researchers, management, and the ethical committee this entails an entirely new problem with retention. on required actions such as notices of annual reviews or inspec- tion findings that may require action. The system creates protocol PS65 Handling during Routine Care Affects Responses of documents using a “dictionary” system populated with techniques Laboratory Rats toward Humans and methods that are approved by the ethical committee. The researcher is able to create new protocols using a plug-and-play S Cloutier*, J Panksepp, RC Newberry system of adding items listed within the dictionary. Because the VCAPP, Washington State University, Pullman, WA system is web based, each protocol is easily tracked through the We hypothesised that the type of handling used by caretakers workflow by researchers, management, and the ethical committee. during routine care of laboratory rats affects fear and anxiety The program checks training records of individuals by technique responses in behavior tests used in biomedical research. We as- and immediately shows if an individual has the training required sessed caretaker handling effects on male Sprague-Dawley rats to perform the task. This feature provides the principal investi- (Rattus norvegicus; Sim:[SD]fBR Albino; found free of internal gator immediate feedback regarding the training qualifications and external parasites and disease by quarterly serology and without having to cross-reference outside paper or electronic files. parasitology, and yearly necropsy, evaluations) in 4 treatments COLADA is an improvement on the current internally written pro- (n = 8 rats/treatment) between 57 and 74 d of age. The standard tocol review software, which only provides a platform for protocol (control) treatment involved rats being handled on cage cleaning document generation and review but does not manage workflow days (2 times/wk) only (picked up by tail and placed in clean or any other administrative aspects of the ethical committee. CO- cage). The passive hand treatment involved rats being exposed LADA is an integrated system that also provides a platform for to a motionless hand for 2 min daily. Pinned rats were restrained managing meeting minutes and inspections, generating reports, on their backs for 2 min daily. Tickled rats were tickled for 2 min and documenting workflow of review and approval of the re- daily. Behavior was assessed between 78 and 80 d of age in a hu- ports. Our company continually seeks to improve animal welfare man approach test (2 min without hand [HA], followed by 3 min compliance and ethical committee review. A global initiative of with a familiar or an unfamiliar hand); an elevated plus maze harmonizing ethical review of research protocols across multiple (EPM), and a cat odor (CAT) test. Ultrasonic vocalizations (50 countries has been initiated. COLADA was developed as a web- kHz, interpreted as evidence for positive affective states) were based tool to manage compliance for the German, American, and recorded during each test. Tickled and pinned rats emitted more French sites using animals for research. 50-kHz calls with a familiar hand than rats from the passive hand and standard treatments (P = 0.01). Tickled rats reared more than PS64 Recruiting Qualified Animal Technicians for a High- rats from the pinned and standard treatments in the HA (P = biocontainment Animal Facility 0.05) and with a familiar hand (P = 0.01). Tickled rats spent more NS Linde*, MK Tate, M Maccallum time near, and made more contact with, the familiar hand than Abstracts of scientific papers 2008 AALAS National Meeting rats from the standard and passive hand treatments (P ≤ 0.01). contract. The members could explain findings and corrective Rats in the standard treatment made the fewest entries into the action in terms that coworkers would relate to and would be area close to the unfamiliar hand (P = 0.02). Treatment did not more likely to institute since they were not being directed from affect behavior in the EPM and CAT. In conclusion, incorporat- a supervisor or management personnel. The goal was to try to ing playful handling (such as tickling) into the routine care of bridge the gap between staff and management by including laboratory rats reduces fear of familiar and unfamiliar humans staff in the resolution process, teach staff to identify and correct but not general anxiety. problem areas, improve communication between management and staff regarding improvements to processes, and ultimately PS66 Benefits of Postapproval Monitoring reduce the number of findings for each department. CM Malinowski* PS68 Extracellular Matrix Adjuvant for Cancer Vaccines Center for Comparative Medicine, Northwestern University, Chicago, IL MA Suckow*1, P Hall2, W Wolter1, V Sailes1, M Hiles2 1Biological Sciences, University of Notre Dame, Notre Dame, IN; Monitoring of animal use protocols for compliance is a require- 2Cook ment of the Animal Welfare Act, the Guide for the Care and Use of Biotech, Inc., West Lafayette, IN Laboratory Animals, the Public Health Service Policy on Humane Most vaccines include an adjuvant of some type, a compound Care and Use of Laboratory Animals, and the United States which enhances the immune response to the immunogens. Government Principles for the Utilization and Care of Vertebrate Currently, most commercial vaccines use adjuvants based on Animals Used in Testing, Research and Training. However, the aluminum salts or combinations of such salts, such as alum. methods used to accomplish this monitoring are not specified. Some studies have used specific cytokines as adjuvants, though Postapproval monitoring can therefore be a major hurdle for cytokines typically enhance specific features of the immune re- institutions to face. In order to accomplish this task, a postap- sponse and may be unstable outside of very controlled storage proval monitoring program was developed and implemented at conditions. We have noted that an extracellular matrix material, our institution. Postapproval monitoring is a program through small intestinal submucosa (SIS), attracts large numbers of an- which an IACUC and the institution itself can ensure and tigen processing cells (macrophages) as it is incorporated into document program integrity, compliance with the regulations surrounding tissue; this feature suggests that SIS could serve and guidelines, and adherence to protocols. At our institution, as an effective vaccine adjuvant. To test the hypothesis that SIS postapproval monitoring is conducted on individual protocols. can enhance the immune response to cancer antigens, groups of Procedures are observed with focus on the following areas of 5 Lobund-Wistar rats were administered 1 × 106 PAIII prostate the protocol: personnel, study procedures, anesthesia, surgery, cancer cells subcutaneously over the flank. The animals under- postsurgical care, euthanasia, recordkeeping, and the labora- went aseptic surgical debulking of the tumor mass 14 d later. At tory environment. The benefits of postapproval monitoring are the time of tumor resection, animals were either left untreated, numerous and include the ability to identify any problems that vaccinated with 5 × 106 glutaraldehyde-fixed tumor (GFT) cells may occur and deal with them before they get to a point where harvested from a subcutaneous PAIII tumor; vaccinated with they need to be reported to regulatory agencies. The program GFT cells that had been grown on SIS for 3 d prior to fixation, also helps foster an environment of openness, encourages the or vaccinated with fixed SIS without GFT cells. In cases where investigative staff to engage the animal facility and IACUC per- animals were vaccinated, vaccines were administered directly sonnel when they need assistance, and provides an educational onto the tumor bed. Animals were euthanized 21 d later, and and training forum for the research staff. This program has met regrown tumors weighed. While there were no significant dif- with much success at our institution, and to date 39 visits have ferences in rats left untreated or vaccinated with GFT cells or SIS been conducted. alone, tumor weight was reduced by greater than 50% in rats vaccinated with GFT cells on SIS, significantly less (P < 0.005) PS67 New Approach to Quality Assurance than controls. Further, only 40% of rats vaccinated with GFT cells on SIS had pulmonary metastases, compared to 100% of controls. K DeMuro* These data demonstrate that cells and immunogens affixed to 1 SoBran, Bethesda, MD; 2 NIH/DVR, Bethesda, MD extracellular matrices such as SIS offer an outstanding platform for vaccine adjuvant development. In an animal facility, quality assurance (QA) is a part of each individual’s responsibility and can be demonstrated as review- ing one’s own work, supervisory checks at the end of the day, PS69 Clinical Monitoring Study of Rhesus Macaques (Macaca management walk-throughs, and a formal QA program. It can mulatta) Given Humanized Monoclonal Antibody be difficult to incorporate a QA mentality into staffs that are W Porter*1, A Wickham1, C Johnson1, A Stadheim2, J Drummond3, not familiar with the QA process. In order to improve upon our B Doerning1, W Strohl3 performance and decrease the number of QA findings, we imple- 1Laboratory mented QA committees that were made up of the QA manager, Animal Rescources, Merck Research Laboratories, Rahway, NJ; 2GlycoFi, Lebanon, NH; 3Biologics Research, Merck program manager, training staff, and a representative from each Research Laboratories, West Point, PA building within the contract. There was a separate committee for the different areas of the facilities (cage wash, animal caretaker, When humans are intravenously dosed with humanized and veterinary technician). During these meetings, the repre- monoclonal antibody (huMab), any resulting infusion reactions sentatives discussed routine findings from the buildings and are usually minor; however, on rare occasions fatalities have the corrective actions and recommendations to prevent their occurred. Due to this phenomenon, the short- and long-term occurrences. The representatives were then asked to present the health status of nonhuman primates on huMab research studies information from the meeting to their peers at their building’s could be at risk and make their use on future studies prob- next weekly training session. This provided a more consistent lematic. These considerations led us to closely monitor a very corrective action plan amongst the 6 facilities that are under this comprehensive range of clinical parameters in unanesthetized Abstracts of scientific papers 2008 AALAS National Meeting adult, male rhesus macaques (Macaca mulatta) during and after of tissues prior to spontaneous death without significantly im- dosing with huMab. Twelve animals were intravenously dosed pacting longevity outcomes. with huMab; repeat dosing occurred 21 d later. Four animals were dosed once with vehicle control. Animals were monitored PS71 Refinement of Mouse Telemetry Transmitter Implantation extensively for 24 h after each dosing, and were followed for using 3D Microultrasound-guided Surgery up to 135 d for changes in general health and for potential on- set of immune-mediated complications such as anaphylaxis or X Shen*1, C Ahn2, B Campbell3, P Zhou2, M Bunzel3, J Mane1, C immune-complex disease. We monitored and tracked the follow- Zhang3, P Cheng2, R Sophie2, D Cully2, B Doerning1, M Walker ing; skin/mucous membrane coloration, physical examination III3, T Cai2, C Johnson1 including cardio-pulmonary auscultation and heart/respiration 1Laboratory Animal Resources, 2Cardiovascular Diseases, 3Im- rate, body weight/temperature, behavior, blood cell count and age, Merck & Co., Inc., Rahway, NJ chemistries, total serum complement activity (CH 50 assay), Previously, we reported implementation and characterization of erythrocyte sedimentation rate, urinalysis, and other aspects a radiotelemetry system measuring blood pressure with C57BL/6 of health status. Only 2 parameters were altered with statistical mice. In the process of expanding the system to other strains of significance (decreased lymphocyte counts and increased CH mice, we found that the length of the carotid artery varies from 50 serum activity) in the huMab group compared with the con- strain to strain of mice, or even animal to animal within the trol group, and these transient physiological changes occurred same strain, making consistent catheter placement a challenge. only during the first 24 h after the initial dosing. From this we Surgical placement of the PA-C10 transmitter catheter resulted conclude that huMab administration did not adversely affect in a high variability in the blood pressure readout, and a sub- the health of the animals in the study, through at least 135 d. We sequent lower success rate of the telemetry modeling system. consider these animals to be clinically normal and suitable for Ultrasound-guided surgical techniques have been successfully use in other types of studies. To our knowledge, this is the first used to minimize invasiveness and to increase effectiveness, lead- report of a comprehensive monitoring of rhesus macaques for ing to a reduced procedure and recovery time in animals. Our acute and chronic health effects after dosing with therapeutic group introduced a quantitative 3D-microultrasound (Q3DMU) levels of huMab. to improve the surgical catheter placement accuracy and blood pressure recording stability in several strains of mice. Standard PS70 Endpoints Related to Imminent Death in Mice Used for surgical procedure was followed in implanting a PA-C10 trans- Longevity Research mitter to the branch level of the aorta by way of the left carotid MA Ray, N Johnston, S Verhulst, LA Toth* artery. Real-time adjustments were then applied while imaging with the Q3DMU such that the probe tip is precisely placed at Southern Illinois University School of Medicine, Springfield, the aortic arch region of the left carotid. Following a standard IL postsurgical recovery process (2 wk), the blood pressures were Despite anticipated health problems, aged mice can be found recorded to document successful placement. We have applied dead without preceding signs of illness. The goal of this study this method with 3 strains of mice (C57, BPH and BALB/c), and was to identify objective criteria for prediction of imminent death a success rate of over 90% was achieved with ultrasound-guided in mice used in longevity research. ICR mice (retired breeders; technique, based on quality of blood pressure measurements. 58 females and 47 males) were subcutaneously implanted with This modeling system addresses the 3Rs by refining the radiote- an identification chip that also allowed remote measurement lemetry placement procedure and reducing the experiment time of body temperature. Male mice were housed individually and number of animals required. to avoid injury due to fighting; female mice were assigned to either group (10 mice per cage) or individual housing. Mice PS72 Sustained Delivery of GLP-1 to Treat a Rat Model of Type were evaluated weekly for body temperature, weight, body 2 Diabetes and Obesity condition, respiratory rate and character, dehydration, posture, hair coat condition, spontaneous ambulation, and response to DH Moralejo*1, O Yanay3, J Fuller2, R Tatake4, R Barton4, A Lern- manipulation. Twenty-four mice underwent euthanasia based mark2, W Osborne3 on clinical signs (inability or unwillingness to walk, lack of overt 1Comparative Medicine, 2Medicine, 3Pediatrics, University of response to manipulation, large or ulcerated tumors, or palpable Washington, Seattle, WA; 4Boehringer Ingelheim Pharmaceuti- hypothermia), and 81 were monitored until spontaneous death cals, Inc., Ridgefield, CT occurred. Mice that underwent euthanasia were killed at a mean Type 2 diabetes (T2D), a common disorder with a prevalence age of 25.3 ± 1.3 mo, whereas the mean age at death among mice of 4% to 5% that is increasing worldwide, is associated with that died spontaneously was 24.5 ± 0.7 mo. Housing and gender shortened life expectancy due to late diabetes complications; did not significantly influence outcome measures. Weight loss thus, novel gene therapy treatments are warranted. We treated was the most common and earliest sign of imminent death. Mice a leptin receptor-deficient rat model (DR.lepr/lepr) of T2D and lost about 20% of their weight during the 3-mo period that pre- obesity with encapsulated allogeneic vascular smooth muscle cells ceded death, with about half of that loss occurring during the last (VSMC) genetically modified to secrete glucagon like-peptide-1 week of life. Body temperatures fell by over 1 °C during the last (GLP-1). GLP-1 exhibits biological and clinical effects that include week of life. Within 24 h of death, 43% showed slow or labored stimulation of glucose-dependent insulin secretion, insulin biosyn- respiration. Thus, monitoring weight as a preliminary marker, thesis, beta cell proliferation, neogenesis and induction of a satiety with hypothermia, respiratory difficulty, and/or precipitous signal. Rat VSMC transduced with a retrovirus encoding GLP-1 weight loss as penultimate markers, appears to allow prediction and selected in G418 secreted 294.4 pg GLP-1 per 106 cells per 24 of death to within 1 wk. Among the mice used in this study, 1 wk h of GLP-1. We seeded encapsulating devices with 107 transduced is about 1% of the median life span. Closer evaluation of high- cells and implanted a single device SQ into random fed male risk mice could potentially allow even more accurate prediction. DR.lepr/lepr rats and monitored blood glucose, weight, and islets. Euthanasia of mice based on these criteria may allow collection Blood glucose levels decreased after surgery and fell within the Abstracts of scientific papers 2008 AALAS National Meeting normal range (90 to 120 mg/dl) 5 d after cell implantation. For the macaques. Currently, there are 5 well-described serotypes of SRV 27-d period before devices were removed, the blood glucose level that are all genetically related to the prototypic type D retrovi- mean was 112.8 ± 41.7 mg/dl (n = 2) without hypoglycemia. After rus, Mason-Pfizer monkey virus (MPMV or SRV3). Retroviruses 27 d we removed the encapsulation devices, and although blood are enveloped, single-stranded RNA viruses that can reverse glucose levels rose to a mean of 160.7 ± 46.1 mg/dl (n = 2), they transcribe their RNA genome to DNA and then integrate into did not return to the hyperglycemic levels recorded before surgery the host’s genomic DNA, resulting in a persistent, lifelong virus (>350 mg/dl). In rats treated with GLP-1 delivery, pancreatic sec- infection. Depending upon various host, virus, and environmen- tions stained for insulin and H&E showed enlarged islets without tal factors, SRV infection can vary from nonpathogenic to an multifocal vacuolization and with normal morphology, and an- immunosuppressive AIDS-like disease that results in death from giogenesis with islet proliferation and neogenesis, respectively. opportunistic infections. Diagnosis of SRV infection is difficult, These data indicate we achieved sustained delivery of GLP-1 to since not all SRV-infected macaques mount a robust humoral im- treat diabetes in our rat model of T2D and obesity. mune response to the virus and SRV-infected macaques are often negative by polymerase chain reaction (PCR). This presentation PS73 Comparison of Two Euthanasia Methods for Zebrafish will describe the development of a truncated, recombinant SRV (Danio rerio) glycoprotein 20 (rgp20) antigen for use in SRV serodiagnosis. The antigen was produced as a chimeric fusion protein with a JM Wilson*, AJ Carty polyhistidine tag in a baculovirus expression system, and was University Laboratory Animal Resources, University of Penn- partially purified by metal affinity chromatography. The rgp20 sylvania, Philadelphia, PA antigen was then used to develop a Luminex xMAP-based as- say for SRV1-5. Diagnostic results from a single accession of 45 The 2007 “AVMA Guidelines on Euthanasia” and the ILAR report cynomolgus macaques (Macaca fascicularis) from an SRV endemi- Recognition and Alleviation of Pain and Distress in Laboratory Animals cally infected colony revealed that 42 of 45 animals tested were state that hypothermia (also referred to as rapid cooling) is an positive for SRV by serology, PCR, or both. Of the 42 animals unacceptable method of euthanasia in fish. Although no scientific that were positive for SRV, 36 were positive by serology using justification is provided as to why rapid cooling is unacceptable, SRV2 whole virus lysate as antigen and 41 were positive by there is speculation that ice crystal formation occurs in the tissue serology using rgp20 as antigen. All 5 samples that were serol- during rapid cooling, which is presumably painful. In order to ogy positive using rgp20 but negative using SRV2 whole virus assist Institutional Animal Care and Use Committees about the lysate were confirmed as SRV-infected by PCR. When compared most humane method of euthanasia in zebrafish (Danio rerio), we to assays that use partially purified SRV2 whole virus lysates as hypothesized that rapid cooling in zebrafish is a humane method antigen, the rgp20 assay has enhanced diagnostic performance of euthanasia, due to a more rapid time between cold exposure and characteristics, including improved diagnostic sensitivity and death, with minimal signs of distress. Zebrafish were individu- specificity. ally euthanized by immersion in 1 of 2 solutions. Twenty-three zebrafish were placed in an ice-water bath (<4 °C) and 23 were placed in MS-222 solution (250 mg/l; 25 to 30 °C). The time from PS75 Intracardiac Pacing Lead in Telemeterized Cynomolgus exposure to the solution until the fish were unable to swim and Monkey (Macaca fascicularis): Evaluation of Refinements right themselves, as well as the cessation of gill movement (death), from 2005 to 2008 were noted. Specific signs of distress were also recorded. Two M Volberg*, N Poy, P Harris, M Engwall minutes following the last opercular movement, animals were placed in a “recovery tank” of system water to determine the Pfizer Global Research & Development, Groton, CT effectiveness of each euthanasia solution. Tissues were then col- Telemetry is widely used as a noninvasive method of collecting lected for histologic analysis. Time to cessation of gill movement in continuous blood pressure and ECG data in canine and nonhu- animals euthanized by rapid cooling ranged from 5 to 9 s (average man primates particularly for safety pharmacology studies. The = 7.13 s). Time to death with MS-222 was significantly longer (P = use of an intracardiac bipolar pacing lead in conjunction with 0.0005) and ranged from 29 to 124 s (average = 53.52 s). Signs of a telemetry transmitter has many advantages over traditional distress occurred in 39% of animals euthanized by rapid cooling subcutaneous lead placement. Advantages include improve- as opposed to 100% of fish euthanized with MS-222 (P < 0.0001). ment of signal quality, higher signal-to-noise ratio and minimal Of the zebrafish euthanized with MS-222, 17.4% animals regained movement artifacts, all resulting in more consistent waveforms consciousness in the recovery tank after exposure to MS-222 solu- to enhance data analysis. During the early development of the tion for 2 min past the last observable opercular movement. This model, the bipolar pacing lead with 3 cm distance between phenomenon did not occur with any animals euthanized by the electrodes was used. In the course of study use, inconsistency rapid cooling method. Based upon the more rapid, less stressful, in size and morphology of the P-wave was noted, and the inter- more effective death of zebrafish euthanized by rapid cooling, we val between electrodes became progressively longer or wider. advocate this method as a humane practice. Data suggested that the most favorable placement consisted of the terminal/tip electrode in the right ventricular apex and the PS74 Development of a Recombinant Serodiagnostic Assay proximal/ring electrode inside or just above the right atrium. for Simian Type D Retrovirus Infection in Macaques (Macaca Refinement for capturing optimal waveforms has resulted in 5- fascicularis) or 6-cm–spaced electrodes for cynomolgus weighing between 4 to 7 kg. As an additional refinement of the surgical procedure JH Simmons*, J Cowley, ML Wunderlich, BA Bronson, RK Dha- with no impact on data capture, the transmitter body is located wan in an intramuscular pocket placed by way of laparotomy. Eighty- Research Animal Diagnostic Services, Charles River, Wilming- five cynomolgus monkeys have been instrumented to date with ton, MA these methods and have experienced minimal or no postopera- tive complications. A review of the surgical procedure and ECG Exogenous simian type D retroviruses (SRV) are a closely re- enhancements will be presented. lated group of viruses that infect free-ranging and laboratory Abstracts of scientific papers 2008 AALAS National Meeting PS76 Circadian Disruption Induced by Dark-phase Light for measurement of AFP, a biomarker that reflects presence and Contamination in Laboratory Animal Facilities growth of human tumor cells. Tumors were collected at various times throughout the study for histological analysis of vessel RT Dauchy*, EM Dauchy, RP Tirrell, LK Davidson, DT Lynch, density using Factor VIII staining. Following 7 d of treatment PC Tirrell, DE Blask with an antiangiogenic drug, uptake of microbubble contrast Bassett Research Institute, Cooperstown, NY was assessed and showed a 35% decrease (P = <0.05) when Diurnal light cycles and circadian rhythms are important to compared to vehicle-treated tumors. After 10 d there was a 30% humans and laboratory animals alike, and have been investi- difference (P = <0.1) between treatment and control groups. gated for many years. Previous studies from this laboratory Blood pressure showed no significant change between treatment demonstrated that minimal light contamination (0.25 lux) during groups throughout the study, indicating that the changes seen in an otherwise normal dark phase resulted in suppression of the tumor blood flow are independent of anesthetic effects. We have circadian neurohormone melatonin. Here, 36 adult male Sprague demonstrated a correlation between changes in tumor vascular- Dawley were randomized into 6 designated groups (n = 6/ ity and response to compound treatment. While power Doppler group), housed and maintained in light exposure chambers and imaging is somewhat limited to large vessels with high flow, the provided free access to food and water. Animals were maintained use of microbubble contrast enhancement provides much greater on either control (I) 12 L (345 lux; 141.5 µW/cm2; lights on 0600 detail about flow and microvessels within a tumor. h):12 D or experimental light-at-night (LAN) lighting cycles as follows: (II) 12 L:12 D–0.02 µW/cm2; (III) 12 L:12 D –0.05 µW/cm2; PS78 Facility Design Requirements for Integrating Gaseous (IV) 12 L:12 D–0.06 µW/cm2; (V) 12 L:12 D–0.08 µW/cm2; and Chlorine Dioxide Decontamination Systems constant light (24L). Following 2 wk on these lighting regimens P Lorcheim, M Czarneski* animals were subjected to a series of 6 low-volume blood draws by cardiac puncture at 4-h intervals (beginning at 0400 h). Plasma Clordisys Solutions Inc, Lebanon, NJ samples were measured for total fatty acids (TFA), triglycerides This presentation provides detailed information on designing (TGA), free fatty acids (FFA), phospholipids (PL), cholesterol and integrating a gaseous chlorine dioxide decontamination esters (CE), glucose, lactic acid, corticosterone, and melatonin system into a new animal research facility. Topics to be covered levels. Plasma TFA and LA, respectively, for groups I–V revealed include reasons for choosing gaseous chlorine dioxide, facility an identical diurnal rhythm, highest during the dark phase (4.66 HVAC design requirements, and room construction require- and 1.61 mg/ml, 0400 h) and lowest during light phase (0.91 and ments, as well as ways to interface the decontamination system 0.30 mg/ml, 1600 h), but remained at the same level throughout to rooms, pass-through chambers, and entire suites such as that the day in group VI (2.72 and 0.92 mg/ml, constant light; TGA > of a BSL facility. A distributed installation configuration layout, FFA > PL > CE). Diurnal plasma glucose levels, similar for groups where the decontamination system could remotely decontami- I–IV (peak, 2000 h; nadir, 0400 h), were significantly elevated (P nate any room of a BSL-3 suite area, will be presented, and the < 0.05) and phase-delayed in groups V (peak, 1200 h) and VI use of decontamination chambers to decontaminate equipment (peak, 2400 h), respectively; melatonin rhythms were normal for and supplies entering a barrier facility or exiting a BSL facility groups I–IV (130.8 pg/ml, dark phase; 6.1 pg/ml, light phase), will be explored. The benefits of a complete suite or facility but disrupted and low diurnally for groups V and VI (6.1 pg/ decontamination system will be compared to a room-by-room ml, light phase). These findings argue that even minimal light decontamination system. contamination during an otherwise normal dark phase in the laboratory animal facility causes disruptions in normal physi- PS79 Circadian Disruption Induced by Dark-Phase Light ologic and metabolic circadian rhythms. Contamination in Animal Facilities Stimulates Human Tumor Growth and Metabolism in Nude Rats PS77 Application of High-frequency Ultrasound to Assess Tumor Neovascularization and Compound Response RT Dauchy*, EM Dauchy, LK Davidson, PC Tirrell, DT Lynch, RP Tirrell, MW Greene, LA Sauer, DE Blask TA Swanson*, KJ Patterson, CJ Andresen, C Bagi, MR Moalli Bassett Research Institute, Cooperstown, NY Comparative Medicine, Pfizer Global Research and Develop- ment, Groton, CT Laboratory animal facility diurnal lighting protocols significantly impact the chronobiologic nature and outcome of scientific inves- The use of high-frequency ultrasound (HFUS) in preclinical drug tigations, and strict adherence to such protocols, as outlined in the development has many applications. Ultrasound measures of Guide, is often overlooked. Previously, our laboratory demonstrated blood flow and vascularity are directly translatable to the clinic that animal facility dark-phase “light contamination” with as little and are comparable to clinical ultrasound. Our current studies as 0.25 lux (0.08 μW/cm2) suppressed production of the circadian involve the validation of high-frequency ultrasound power neurohormone melatonin and stimulated rodent and human breast Doppler and contrast-enhanced imaging on xenograft tumor tumor growth and metabolism. This occurs by way of an MT1 models to assess growth kinetics and response to drugs that are melatonin receptor-mediated suppression of signal transduction, known to effect neovascularization. Our objective was to assess resulting in inhibition of linoleic acid (LA) uptake and conversion to neo-vascularization events in mouse and rat tumor xenografts, the mitogen 13-hydroxyoctadecadienoic acid (13-HODE). Here we quantify changes in vascularization detected by HFUS, and cor- tested whether dark-phase light contamination disrupts temporal relate those changes with biomarkers such as alpha-feto protein coordination of human breast tumor–host metabolic mechanisms. (AFP) and tumor histochemistry. Nude rats were implanted with Female nude rats (170 g; n = 36/group) bearing “tissue-isolated” Huh7.5 hepatocarcinoma cells injected subcutaneously into the human breast cancer xenografts were maintained on either control flank. Tumor size and vascularity were assessed using 3D gray- (I) 12 L(345 lux; 141.5 μW/cm2):12 D or experimental (II) 12 L:12 scale volume imaging, power Doppler and microbubble contrast D–0.08 μW/cm2 lighting regimen (lights on 0500 h). Tumor latency bolus injections. Blood pressure and body temperature were to onset (first palpable mass) and growth rates for groups I and II monitored during these studies. Blood samples were collected were, respectively, 16 and 11 d, and 0.26 ± 0.03 and 0.58 ± 0.04 g/d. Abstracts of scientific papers 2008 AALAS National Meeting When tumors reached an estimated weight of 5 to 6 g, animals animal care staff and facility managers overseeing time manage- were prepared for arteriovenous measurements over 6 circadian ment and per diem rates. Our objective was to determine the timepoints in a 24-h period (n = 6/timepoint). Results showed impact of microisolation cage bedding volume and cage change plasma melatonin levels in group I were high in dark-phase, and interval on microenvironmental conditions, mouse health, and low in light-phase (I) or group II animals. Diurnal tumor glucose behavior. Fifteen cages housing ICR mice female (n = 5/cage) uptake, lactic acid release, cAMP levels, LA-uptake, 13-HODE were monitored for 17 d. The mice were obtained at 12 wk of age release, and DNA [3H]thymidine incorporation were elevated and allowed a 2-wk acclimation period. Three different volumes significantly (P < 0.001) in daytime (I) and group II by over 60%, of autoclaved aspen bedding chips were used: minimal volume 400%, 700%, 1000%, and 900%, respectively, compared to dark- (approximately 250 ml/40 g), large volume (approximately 550 phase (I). Tumor Akt, MEK, ERK 1/2 activation was elevated for ml/100 g), or normal volume (approximately 400 ml/70 g). Mice groups I (light-phase) and II. These findings are the first to show were evaluated (physical exam, weight) on d 7, 10, 14, and 17, in vivo that temporally coordinated and integrated metabolic and and a qualitative assessment of cage soiling was photographed signal transduction mechanisms underlying human breast cancer at these timepoints. Pooled feces were collected for corticos- growth can be disrupted by dark-phase light contamination in the terone analysis, and bedding mass was also measured at these animal facility. timepoints. On d 17, 1 mouse from each cage was euthanized and bronchoalveoloar lavage was performed for cytologic analysis. PS80 A Comprehensive Rodent Quarantine System That Jump- Mice were videotaped for approximately 10 min during the active starts Research periods on d 1, 8, and 15. Atmospheric analysis was performed to determine intra-cage ammonia, particulate matter concen- J Watson* tration, and cage humidity/temperature on a daily basis. The MCP, Johns Hopkins, Baltimore, MD atmosphere was measured using a photoionization detector and particulate size and number was recorded by an aerodynamic This presentation outlines the logistics of setting up a full-service particle sizer. There were no indications that the animals were rodent quarantine program geared for research support. Statistics stressed, experienced adverse health effects, or exposed to higher from our first year of implementation will be presented. Rodent than recommended levels of particulates or adverse environ- quarantine is a necessary component of most research facilities to mental conditions during the 14-d period. Significant findings prevent introduction of unwanted pathogens. Interinstitutional included variations in humidity depending upon cage location rodent shipments are increasingly common due to NIH sharing and ammonia concentration among cage bedding levels. Cages agreements, research collaborations, faculty recruitment, and were visibly soiled beyond what would be considered accept- international vendor sources. However, there is no universal able to most observers by d 14. This study demonstrates that a standard regarding which organisms should be excluded from 2-wk interval between cage changes did not impact measurable rodent colonies, and the varied rodent housing arrangements animal welfare indices, despite the appearance of excessive soil- and systems for colony health surveillance make it hard to assess ing by this timepoint. infection risk. Thus, it is usually necessary to quarantine incom- ing animals to evaluate their infection status, which can take 8 to 12 wk. In some cases, this process may impede or even prevent PS82 Ammonia Concentrations at the Cage Level for Mice research. Colony founders may be too old to breed by the time and Rats: Effects of Strain, Housing Density, and Ongoing they are released, and age-sensitive phenotypes may occur during Breeding the quarantine period. Some shipments may be rejected because of A Ostdiek*2, A Frazier1, N Johnston1,4, T Liberati1,4, R Trammell4, infections in the source colony or incur additional delays and costs LA Toth3,5 for surgical rederivation. We developed a comprehensive quar- 1Division of Laboratory Animal Medicine, Southern Illinois antine program to address these problems. Research delays were University School of Medicine, Springfield, IL; 2College of mitigated by providing technical services in quarantine, including breeding, identification, sample collection, genotyping, and basic Veterinary Medicine, University of Illinois, Urbana, IL; 3Phar- macology, 4Internal Medicine, 5Associate Dean for Research and experimental manipulations. Shipments that would previously Faculty Affairs, Southern Illinois University School of Medicine, have been rejected or delayed because of infection were instead Springfield, IL placed in high-risk quarantine and tested individually. Infected animals were rederived by neonatal transfer. Our first year, 59 High ammonia in the microenvironment can adversely impact of 230 shipments were placed in high-risk quarantine, and 58 of rodent health. Many factors influence ammonia concentrations them tested free of infection. Thirty-four shipments used technical in the cage, including housing density, air flow rate, room tem- services, 118 shipments were bred, and 17 genetic lines were red- perature and humidity, bedding type, type of caging system, erived to eliminate Helicobacter sp. Our experience demonstrates and the frequency of cage changes. The NIOSH-established that rodent quarantine, far from impeding research, can actively exposure limit for people is a time-weighted average of 25 ppm promote an efficient start to research following interinstitutional over an 8-h period. Levels above this are reported to have det- rodent transfers. rimental effects in people and rodents. We performed this study to examine the relationship between ammonia levels and strain PS81 Bedding Change Interval Influence on the Microenviron- differences, housing density, and breeding animals. To verify ment of Mice in Ventilated Microisolation Housing that our standard housing conditions did not generate damag- ing concentrations of ammonia in the cage, we used a hand-held M Rosenbaum*1, S VandeWoude1, T Johnson2 piston pump and ammonia indicator tubes to monitor ammonia 1Microbiology, Immunology, and Pathology, 2Environmental of 2 strains of rats (GEPR-9 and Sprague Dawley) and 3 strains and Radiation Health Sciences, Colorado State University, Fort of mice (Maf/D2, Ames dwarf, and PEPCK-BGH1). These strains Collins, CO were selected due to their frequent use in our facility or to their apparent tendencies to produce relatively high amounts of urine. The frequency of mouse cage bedding changing has significant The animals were housed in static micro-isolator cages that were implications not only for the animals themselves, but also for the Abstracts of scientific papers 2008 AALAS National Meeting changed weekly. Breeding cages contained 1 to 3 adults with as The fast increase in the production of new lines of genetically many as 17 pups. Single-sex cages contained 1 to 5 animals. For modified mice has raised concern about how gene manipula- each strain, 2 breeding cages and 2 non-breeding cages were tion techniques affect the welfare and normal development of evaluated over a 2-wk cycle, providing a total of 4 cages per the resulting animals. The phenotype of the mice obtained by strain that were sampled over 2 cage changes. For both rats and those techniques is influenced by the background strains, the mice, ammonia concentrations rose above 25 ppm between 4 environment, and possibly by the occurrence of compensatory and 6 d after a cage change. To determine whether this degree processes. Unexpected phenotypes may result in welfare prob- of exposure caused a pulmonary inflammatory response, we lems and excessive suffering, which justify the establishment compared concentrations of 10 cytokines in the lungs of mice of humane endpoints. While the need for the development of maintained in a constant low ammonia environment by more monitoring routines at an early age has been discussed, no pro- frequent cage changes to those in lungs of mice that were exposed tocols for preweaning characterization of genetically modified to a high-ammonia environment for 2 d. Concentrations of 4 of and mutant mice have so far been validated. This study com- the 10 cytokines were below limits of assay detection. Significant prises a first assessment of a characterization routine that has differences between groups were not detected. We conclude that been developed in our laboratory. A total of 56 mouse pups of 3 the practice of weekly cage changes does not cause inflammatory inbred strains and 1 hybrid commonly used as backgrounds for changes in the lungs of mice, despite their exposure to levels of the construction of transgenic and knockout mice (C57BL/6Bkl, ammonia over 25 ppm for periods of up to 2 d. BALB/cBkl, 129S6Bkl, and B6CBAF1Bkl) were monitored from birth to weaning on key days of morphological, physiological, PS83 Radical New Approach to IVC Housing for Rats and behavioral development. The results presented show that the developed characterization routine is efficient in detect- M Jackson* ing differences in physical and sensory development between BSF, University of Manchester, Manchester, United Kingdom strains, namely concerning weight gain, reaction to handling, physical condition at weaning, and emotional reactivity. A future A rat focus group featuring experienced animal technologists study will address its efficiency in detecting differences in the from academia, contract research, and the pharmaceutical sectors development of genetically modified pups and their wild-type was formed to address the need for a new rat IVC cage. The key litter mates. considerations were to produce a cage which is compliant with UK and European legislation. It should hold up to 5 adult rats or a breeding pair in an enriched environment that allows rats to PS85 Predicting the Impact of Noise from Construction on express a full range of behaviors including bi-pedal posture. The Mouse Gestation and Nursing Behavior cage must also have good manual handling characteristics and S Rasmussen*1,3, GM Glickman2, FW Quimby1,3, R Tolwani1,3 be as light and well balanced as possible. Flexibility is important 1CBC, The Rockefeller University, New York, NY; 2 Wilson, so as to be acceptable to a range of research environments. The cage should be operational at positive or negative pressure and Ihrig & Associates, Inc., New York, NY; 3Postdoctoral Research, allow use of various water delivery systems. As automation is Tri-Institutional Postdoctoral Training Program in Laboratory increasingly used for washing and bedding replacement in many Animal Medicine and Science, New York, NY institutes, the cage must withstand being handled by robotics. A comprehensive study was undertaken to identify the impact The cage design includes a base and a high, domed lid. A horizon- of noise from a planned expansion of our animal facility on tal U-shaped divider provides an extra story, effectively doubling mouse gestation and nursing behavior. Ambient noise levels floor space in the same footprint. Two test cages were fabricated were analyzed first in order to gain an understanding of daily using stereolithography and connected to an IVC rack system noise exposure. We then compared the ambient noise levels to for 4 wk. Three male S/D rats were allocated to each test cage. that predicted from construction activities to see if noise from Animals were weighed daily, and food and water consumption construction would impact mouse gestation or maternal nursing were noted. Growth rates and diet consumption were normal behavior. We hypothesized that high noise levels would result in for this strain. Video monitoring was used to study activity both fewer live births and higher pup mortality as a result of disrupted in light and dark phases of the daily cycle and after weighing or nursing behavior. To study the impact of noise on mouse gesta- cleaning of cages. Observations revealed rats using all areas of tion, 70 8-wk-old female ICR mice underwent embryo transfer. the cage. Sleeping was confined mainly to the lower half of the Mice were divided into groups of 10, with each group receiving cage, while play occurred on both levels. Exploratory behavior exposure to a single intensity of simulated noise analogous to and climbing are seen as positive welfare aspects of the design. concrete saw cutting at 70 dBA, 80 dBA, or 90 dBA for 1 h daily. Animal technologists of varying experience were encouraged to The frequency of the simulated noise ranged from 2 to 4 kHz. use the cage and complete a questionnaire to elicit their opinions. Thirty mice received noise exposure d 1 through 7 following The paper shows that the design brief was met and an enriched, embryo transfer, 30 were exposed d 8 through 14, and 10 mice radical new rat cage is ready to go to mass production. received ambient noise only. The number of viable pups born was recorded. An additional 20 mice were used for a second study to PS84 Characterization Routine for Improved Animal Welfare examine the effects of construction noise on nursing behavior. A in Preweaning Mice control group of 5 mice received normal ambient daily noise. The remaining 15 mice were divided into groups of 5. Each group was JM Marques1,2, H Augustsson*3, S Ögren4, K Dahlborn1 exposed to a single intensity of noise simulated from concrete 1Department of Clinical Sciences, Swedish University of Ag- saw cutting at 70 dBA, 80 dBA, or 90 dBA for 1 h daily during ricultural Sciences, Uppsala, Sweden; 2Laboratory Animal the first 7 d following parturition. Female mice were monitored Science, Institute for Molecular & Cell Biology, Porto, Portugal; for cessation of nursing behavior. Pups were weighed daily 3Department for Animal Welfare and Health, Swedish Board of and monitored for increased mortality. Based on these studies, Agriculture, Jönköping, Sweden; 4Department of Neuroscience, specific construction noise limits were established to minimize Karolinska Institute, Stockholm, Sweden any negative impact on mouse reproduction. Abstracts of scientific papers 2008 AALAS National Meeting PS86 Novel Means of Continuously Monitoring Laboratory packaging and transport, production and distribution of IVCs, Rodents and use and disposal of IVCs throughout the life cycles of the alternative systems. The analysis was performed using a system SM Niemi*1, M Dempsey2, N Sims2, M Wollowitz3 perspective and the LCA methodology. The functional unit was 1Center for Comparative Medicine, 2Center for Integration of set to the use of 1 IVC (disposable or multiuse) for a period of Medicine and Innovative Technology, Massachusetts General 1 y. The yearly number of replacements and the disposal of the Hospital, Boston, MA; 3 Tangible Design, Inc., Chatham, NY single-use PET IVCs compared with the yearly number of wash- The dramatic growth of genetically engineered mice in biomedi- ing and sterilization cycles of the PSU IVC were considered. The cal research has resulted in more complex breeding programs, results of the LCA study are presented both in terms of direct sophisticated endpoints, subtly different phenotypes, occasional inventory data, including mJ of primary energy from nature, by difficult breeders, and consequently greater financial losses in the fuel type, and in terms of impact assessment, meaning tons of event of husbandry failures. But because mice are nocturnal, they each greenhouse gas emitted to the atmosphere by all life cycle are more likely to display abnormal behavior or clinical signs of processes, and estimates of human health consequences and disease or injury at night. All of these considerations demand ecological consequences of the pollutant releases to air, water, better observation of cages than a perfunctory visual check once and soils over the life cycle. In addition, a sensitivity analysis daily. However, there may be difficulties or inconsistencies in was performed considering particularly different IVC disposal observing every mouse on a routine basis because the bottom scenarios so as to understand how to reduce the environmental and top rows of high-density cage racks are not easily viewed. To burden at the end of an IVC’s life. address these problems, a novel approach was devised in which a series of gantry-mounted infrared cameras scan all cages on a rack; video (at programmable intervals and of programmable durations) images are recorded and stored for later viewing. Poster Sessions Each gantry communicates to an on-site or remote web server. With appropriate authentication, live video feeds or stored data P1 Model for Clinical Evaluation of Perioperative Analgesia can be viewed through any web browser. Features include ob- in the Rabbit servation of mice 24/7, digital documentation of cage location AZ Karas*, LA Weaver, KA Andrutis, CA Blaze and animal signalment, storage of retrievable images, restricted Clinical Sciences, Tufts University Cummings School of Veteri- access to images for each user, and remote override of scanning schedules. Software enhancements will include automated alerts nary Medicine, North Grafton, MA in the event of excessive activity (such as fighting, stereotypy) Assessment of pain and distress in rabbits is problematic, and or absence of activity (moribundity or death), flooded cages, studies of effective surgical analgesia are lacking for this species. empty feed hoppers and so forth. Advantages of this solution Seeking potential indicators of postoperative pain or distress, with respect to discovery research and GLP testing include we performed ovariohysterectomy and telemeter placement as continuous observation for subtle changes in behavior or unan- a form of moderate surgical injury in 20 female rabbits. Rabbits ticipated phenotypes in neuroscience protocols, more precise were randomly assigned to 1 of 4 treatment groups (n = 5 per determination of impending morbidity for toxicology, and safe group): buprenorphine (0.03 mg/kg SC, every 12 h for 3 d); fen- (remote) observation for select agent protocols and other highly tanyl (25 mcg patch placed 24 h prior to surgery); ketoprofen (1 hazardous studies. In addition, this solution ensures that every mg/kg, SC, every 24 h for 3 d) and control (no treatment given). animal is, indeed, observed daily per regulatory requirements Various physiologic and behavioral parameters were recorded or and requires fewer staff to do so. scored by blinded observers, including heart rate and blood pres- sure, temperature, food and water consumption, fecal output, PS87 A Comparison of Single-use and Multi-use Indi- and remotely recorded video-recorded behaviors during daily vidually Ventilated Cages: CO 2 and Energy Environmental exercise in 1.8 × 2.4 m floor pens. Compared to preoperative Performance values, declines in behaviors occurred on postoperative day 1: mean travel distance (70%), vertical exploring (94%), decrease GA Norris*1, V D’Incognito2 in food consumption (87.6%). Many values had not returned to 1School of Public Health, Harvard University, Boston, MA; 2Take baseline levels after 7 d. Fecal output decreased significantly as Care, Milano, Italy well. Although no single treatment proved significantly better than another, some analgesic treated rabbits did suffer a change Individually ventilated cages (IVCs) have been under close in behavior values the day after cessation of therapy. Our results scrutiny recently, especially the choice of plastic materials for demonstrate substantial inappetance and reduction of normal single-use or multi-use cages. Many different requirements activity levels in rabbits after surgery. Although empirical doses such as medical safety, rodent care, treatment functionality, (those typically recommended) of analgesics did not appear costs and efficiency, and environmental performance have to be substantially better than placebo, comparison of other doses met. For several years now, the most commonly used materials and multimodal analgesic techniques using these monitoring for IVC manufacturing have been polysulfone (PSU) for multi- strategies may prove useful in future studies aimed at optimiz- use cages and, more recently, polyethilene terphthalate (PET) ing postoperative analgesia in rabbits. for single-use cages. A life cycle assessment (LCA) study was used to compare the environmental performance of the 2 cage types, with focus on the energy consumption and global warm- P2 Evaluation of Supplemental Warming Methods for Preven- ing contribution due to greenhouse gas (GHG) emissions. The tion of Hypothermia in Rats LCA study is based primarily on the use of publicly available A Koch*2,1, DG Scorpio1, D Ruben1 life cycle inventory databases that quantify raw material input 1 Molecular and Comparative Pathobiology, Johns Hopkins from nature and releases of pollution to air, water, and land as- University, Baltimore, MD; 2Cummings School of Veterinary sociated with the input of primary and intermediate materials, Medicine, Tufts University, North Grafton, MA Abstracts of scientific papers 2008 AALAS National Meeting Prolonged anesthesia and hypothermia is a common problem in mg/dl). The macaques that received Telazol for induction had an rodents undergoing experimental procedures. Previous studies average temperature decrease of 3.5°F (preoperative values = 100.5 conducted with Mylar-backed drapes in anesthetized rats revealed ± 1.3 °F (x¯ ± SD); range 98.7 to 102.2 °F)(postoperative values = 99 reduced hypothermia compared to conventionally draped and ± 2.1 °F (x¯ ± SD); range 96.1 to 101.5 °F) and blood glucose levels undraped rats. Mylar reduces radiant heat loss in rodents under- increased an average of 30.7 mg/dl (preoperative values = 62.4 going procedures in ventilated hoods or atop laboratory benches. mg/dl ± 6.9 mg/dl (x¯ ± SD); range 55 to 75 mg/dl; postoperative This study was performed to compare Mylar-backed drapes to values = 93 mg/dl ± 23.1 mg/dl (x¯ ± SD); range 69 to 127 mg/ other warming methods in anesthetized rats undergoing sham dl). In conclusion, although the group premedicated with Telazol laparotomy. Conventional surgical drapes and Mylar drapes were showed increased postoperative glucose concentrations compared compared, both with and without a heat-emitting gel pad. A control to preoperative values relative to the ketamine group, the change group included no drapes or gel pad. Rats were anesthetized with was not statistically significant; thus the choice of one sedative intraperitoneal ketamine/xylazine. A ventral midline incision was over the other does not impact surgical recovery. created through a drape fenestration covering each rat’s ventrum. Temperature was monitored with digital rectal probes for 60 min. P4 Suppurative Osteomyelitis, Meningoencephalitis, and Rats were euthanized at conclusion of experiments. All groups Multifocal Suppurative Bronchopneumonia in the Gambian developed hypothermia except for the group with the isothermal Rat (Cricetomys gambianus) pad combined with the Mylar drape. These rats exceeded nor- motherrmia with temperatures between 107 to 109 °F, due to gel BP Herrod*, N Bryant, D Carroll, E Jackson, N Powell pad isothermal release of 108 °F+ reflecting off the Mylar radiant Animal Resources Branch, Pathology Laboratory, Centers for drape. The standard drape with gel pad allowed an average 2.19% Disease Control, Atlanta, GA heat loss from baseline; conventional and mylar drapes used alone The African giant pouch or Gambian rat (Cricetomys gambianus) allowed 8.44% and 9.65% heat loss from baseline, respectively, is a nocturnal rodent from central and southern Africa, where it though far less than control rats (14.39%). Heat loss likely occurred is used as a meat source by natives. Gambian rats were popular from the abdominal incision, thus both drape types yielded similar pets in the US until June of 2003, when they were suspected to results. Rats completely wrapped in the Mylar drape without gel be a source of a monkeypox outbreak. Health officials suspected pad sustained 4% to 5% heat loss from baseline. Mylar drapes and that they were the source of infection because tests have shown heat-emitting pads serve to prevent hypothermia during anesthesia that these rats have antibodies to monkeypox. Presently there in rodents. Used together, however, rat core body temperature rises are no licensed therapies for human monkeypox; therefore, the quickly. Using both methods together is beneficial for hypothermic human population is at risk to this zoonotic agent. Due to this rats. Other supplemental heat modalities such as warm water recir- susceptibility, there are fears that the monkeypox virus could be culation or forced air should be ideal when used with Mylar-backed used as a bioterrorism agent. Gambian rats, in addition to other drapes because they generate less heat than the gel pad; both will animal species, are being studied to investigate the pathogenesis be further evaluated. and treatment of orthopoxvirus infections. We received wild- caught, female Gambian rat, which died suddenly, for necropsy. P3 Perioperative Effect of Ketamine Compared with Telazol The animal was negative for the monkeypox virus and had not on Body Temperature and Blood Glucose Concentration with been used in a study. At gross necropsy, a few abnormalities were Rhesus Macaques (Macaca mulatta) noted. An abscess was found within the right temporomandibu- BC Davisson*, M Patterson, RP Marini lar region which extended into the right temporomandibular joint and the base of the skull. We also found multifocal white Division of Comparative Medicine, Massachusetts Institute for nodules along the surface of the both lungs. Aerobic bacterial Technology, Cambridge, MA cultures were taken in an effort to identify the microbial agents Variables likely to impact intraoperative and postoperative man- involved. The results from the mandibular region revealed a agement of macaques include anesthetic induction regimen, body heavy growth of Pasteurella multocida, light growth of Escherichia temperature, and blood glucose concentration. A sample group of coli, and scant growth of alpha-hemolytic streptococci. Bacterial macaques (n = 30) undergoing extensive cranial surgeries (rang- cultures taken from the base of the skull revealed moderate ing from 2 to 11 h) was examined to compare these parameters growth of Pasteurella multocida and scant growth of a Coryne- and determine what impact induction agent had in maintaining bacterium spp. Histopathology revealed evidence of glossitis, normothermia and blood glucose concentrations. All macaques pneumonia, meningoencephalitis, osteomyelitis, and myositis. In underwent at least a 12-h fast and then received either intramus- this presentation, we will further characterize the histopathologic cular ketamine for induction (n = 23) or intramuscular Telazol lesions through special stains and other molecular diagnostic (n = 7). After sedation, rectal temperatures were recorded and procedures. We will provide histopathologic evidence of an in- blood glucose concentrations were measured using a handheld fection that we encountered in a Gambian rat. After an extensive glucometer. Animals were intubated and maintained on 1% to 3% literature review, we have concluded that this is the first case isoflurane in balance oxygen and were intravenously administered report of this type of infection in a research Gambian rat. a lactated Ringers solution containing 5% dextrose. Immediately postoperatively, a second blood glucose sample was tested from P5 Uterine Epithelioid Trophoblastic Tumor in a Cynomolgus each animal and postoperative rectal temperatures were recorded. Macaque (Macaca fascicularis) In the group that received ketamine for induction, body tempera- tures decreased an average of 3.7 °F (preoperative values = 101.2 ± CL Emerson*1, J Hutt1, TK Cooper2 1.6 °F (x¯ ± SD); range 97.3 to 102.6 °F; postoperative values = 97.9 1Lovelace Respiratory Research Institute, Albuquerque, NM; ± 1.7 °F [x¯ ± SD]; range 94.3 to 100.8 °F), while blood glucose levels 2Comparative Medicine, Penn State Milton S. Hershey Medical increased an average of 22.9 mg/dl (preoperative values = 61.3 Center, Hershey, PA mg/dl ± 12.7 mg/dl [x¯ ± SD]; range 41 to 97 mg/dl; postopera- An adult female cynomolgus macaque was evaluated for lethargy tive values = 84.1 mg/dl ± 21.4 mg/dl [x¯ ± SD]; range 52 to 126 and dehydration. Results of blood work were unremarkable, Abstracts of scientific papers 2008 AALAS National Meeting and the animal was rehydrated. The animal’s clinical signs im- 1Microbiology, Immunology and Pathology, 2LAR, Colorado proved the day after admission, but declined dramatically by State University, Fort Collins, CO the following day and the animal was euthanized. At necropsy, a A wildlife sanctuary presented a postmortem adult female cot- sessile mass was identified in the wall of the uterine body, with a tontail (Sylvilagus spp.) rabbit, age unknown, to the Colorado portion of the mass extending into the lumen. The mass was ap- State University Pathology service for necropsy. The available proximately 3 cm in diameter, yellowish-white, and friable in the history on this animal was limited, but it was reported that at center. There were numerous multifocal to coalescing metastatic least 1 other Sylvilagus spp. rabbit in the same area was reported nodules of similar appearance present in the lungs, with efface- to exhibit similar gross cutaneous lesions. On gross exam, the ment of approximately 80% of the normal lung parenchyma. animal had numerous pigmented wartlike lesions arising from Scattered metastases were also present in the liver and spleen. the skin of the head surrounding the ears, eyes, nares, mouth, and Based upon the histological appearance of the uterine mass and a few areas along the dorsum. Masses were firm, greasy, friable, select immunohistochemical stains, the tumor was identified and easily detached from the underlying skin. Affected cutaneous as an epithelioid trophoblastic tumor (ETT). ETT is one type of tissue was fixed in formalin, embedded in paraffin, and sectioned gestational trophoblastic disease derived from the placental tro- by microtome. The slides were stained with hematoxylin and phoblast. Gestational trophoblastic diseases present with a wide eosin for histopathology. Histologic evaluation revealed multiple range of clinical behaviors and histological appearances, some of papillary masses lined by stratified squamous epithelial cells which are aggressive and malignant. These tumors are distinct with central cores of fibrovascular connective tissue arising from from all other tumors in that they are fetal tumors (products of the epidermis. Cells of the stratum spinosum were frequently conception) arising in the mother’s body. ETT was first described swollen with abundant perinuclear cytoplasmic clearing and in women in 1998 and has features that are similar to carcinoma. occasional intranuclear amphophilic glassy spherical inclusions ETT has been diagnosed in nonhuman primates and found in up to 3 µm in diameter. The lesions were consistent with rabbit the uteri of a red-tailed guenon (Cercopithecus ascanius) and an Shope papilloma virus. Differential diagnoses included Shope African green monkey (Chlorocebus aethiops). A germ cell tumor fibroma virus and myxoma virus. To definitively confirm the with features of ETT (nongestational ETT) has been described in Shope papilloma virus diagnosis, histopathology slides were the ovary of a cynomolgus macaque (Macaca fascicularis). submitted for immunohistochemistry; results were positive. Affected tissue was fixed in glutaraldehyde and submitted for P6 Comprehensive Dental Health Assessment and Treatment electron microscopy, through which the virus was identified. This Program case presents an opportunity for review of the numerous virally CL Gai*1, T Trinh1, A Parlapiano2, GO Voronin1 induced cutaneous lesions due to papillomavirus and poxvirus 1Laboratory Animal infection in rabbit species, and it illustrates an important model of Resources, 2Lead Optimization, Merck and viral carcinogenesis as well as a disease that represents different Co., Inc., Rahway, NJ disease spectrums following cross-species transmission between Dental health and hygiene are important components of the over- Sylvilagus and Oryctolagus spp. A detailed explanation of this all health and well-being of animals. Undiagnosed or overlooked differential disease spectrum is included in the discussion. dental disease can lead to poor patient health and discomfort, disease in other body systems, and may be a confounding factor P8 Use of Behavior Conditioning to Facilitate Venipuncture in research. Providing adequate dental care to a large number of animals presents a logistical challenge. Mandating dental clean- C Lanman* ings for all animals on a predetermined schedule could lead to Pet Care, Proctor and Gamble, Lewisburg, OH animals receiving unnecessary treatment and exposure to anes- In every capacity of research and in every facility, it can be as- thesia. It would lead to the expenditure of animal care resources sumed that methodology improvements come to the minds of and manpower that would be better used elsewhere. Tailoring technicians quite often. In the case of feline restraint for blood care to the individual needs of an animal would require a greater collections, new ways to reduce the stressful nature of these amount of effort for each animal, but would also ensure that each procedures for both the felines and the technicians have been animal receives the appropriate level of care in a timely manner. difficult to uncover. This innovative venipuncture method not In order to deliver animal-specific and -appropriate dental health only decreases the stress sometimes associated with restraint and care, a comprehensive dental health assessment and treatment venipuncture, but also allows for blood collection by a single program was developed, tested and implemented in our canine technician. The method involves first conditioning felines at a colony. Initially, a simplified dental health scoring system was younger age to be comfortable with venipuncture, toenail trims, developed that was used to grade the current dental health status and veterinary treatments. Training begins at 2 d of age. At 3 wk of the colony. A spreadsheet was designed that could not only of age, the kittens are acclimated to being placed dorsally in a be used to keep track of dental health, but would also generate technician’s lap with the sound of clippers running. From this the appropriate dental care recommendation based on the data point, the technicians continue the dorsal placement training entered. Trained observers examined100 dogs, and their recom- while adding different collection techniques such as stretching mendations were evaluated for interobserver reliability by use of the neck, stroking the jugular furrow, and clipping the hair along a kappa statistic. Veterinarians and veterinary associates shared the furrow. This training is ongoing to ensure the safety of the the spreadsheet to generate a plan to deliver dental health care felines and technicians. These training protocols and venipunc- to individual animals during the course of 6 mo. This poster ture method have proven to be instrumental in improving feline presents the development, implementation, and results of our well-being and technician aptitude in this research facility. comprehensive dental health plan during this time period. P9 Research Opossum (Monodelphis domestica) with Perior- P7 Cutaneous Lesions in a Cottontail Rabbit (Sylvilagus bital Swelling spp.) D da Cunha*1,2, LM Wancket2, C Freed1, L Gallaugher1 C Weiner*1,2, M Rosenbaum1,2, K Fox1, S VandeWoude2,1 Abstracts of scientific papers 2008 AALAS National Meeting 1 University of Laboratory Animal Resources, 2 College of tion program. Newly replaced lines developed the same level of Veterinary Medicine—Veterinary Biosciences, The Ohio State ATP as used lines after the first week of use. Although the ATP University, Columbus, OH levels in lines associated with clinical signs were lower than those without, the sanitation procedure reduced the overall incidence There are around 1,500 publications in the PubMed database of clinical signs to 0.4%. This study demonstrates that use of pertaining to the use of marsupials in experimental, biomedical- ABS systems in rats is associated with development of resident oriented research. Monodelphis domestica are naturally found in bacteria in the lines that can result in clinical infections. The oc- arboreal habitats in South America. Here we report a case of a currence of infection is not predicted by bacterial load alone, but 3-y-old breeding female, part of an endocrinology study, that presented with right enophthalmia, moderate periorbital swell- is likely related to other factors. However, controlling the overall level of contamination through weekly sanitation reduces clinical ing, prolapsed and inflamed third eyelid, blepharospasm, and infections, but complete sterility cannot be achieved, even when blepharitis with mild purulent discharge causing irritation and medial canthus alopecia. Triple antibiotic ophthalmic ointment new lines are used. A luminescent bacterial-ATP assay used in (TAO) with hydrocortisone was initiated, and systemic treat- conjunction with bacterial culture and identification is a useful ment with ibuprofen (15 mg/kg) provided for 7 d, after which tool for monitoring and managing ABS line infections. no signs of inflammation remained. Two months later (January 11th) the same animal presented with ventral periorbital swell- P11 Regimens and Results for Treating Myobia musculis ing, mildly prolapsed third eyelid, and buphthalmia in the right D Newsom*1, IM Washington1, K Chase2, TL Brabb1, L Maggio- eye. Oral cavity examination under isoflurane anesthesia ruled Price1, S Dowling1 out tooth root abscess. Systemic treatment with enrofloxacin (5 1 Comparative mg/kg), ibuprofen (5 mg/kg) and topical TAO was initiated. Medicine, University of Washington, Seattle, On re-evaluation 3 d later, the animal presented with severe WA; 2Veterinary Services, California Institute of Technology, buphthalmia, blepharoedema, third eyelid thickening, and hy- Pasadena, CA peremia. Self-trauma to the periorbital area resulted in alopecia The University of Washington (UW) was notified that a mouse and hyperemia of the surrounding tissue. Based on the clinical sent to another university tested positive for fur mites. Upon presentation with poor prognosis and the age of the female, the notification, the UW colony from which the positive mouse origi- investigator consented to euthanasia and postmortem evalua- nated was tested by tape test and confirmed positive for Myobia tion. Blood was collected for CBC analysis. Grossly, the surface musculis. Additionally, over a 1000 tape tests were performed of the spleen appeared irregular, but otherwise within normal on mice from 7 different rooms, 5 investigators, and 13 racks; 15 limits. The liver appeared muddy-brown in color with a smooth positive animals were identified. The positive animals were from surface. CBC findings included neutrophilia without bands, 2 investigative groups, and were of 8 different strains, some im- accompanied by slight lymphopenia resulting in normal total munocompetent and some immunodeficient. All sentinels were white cell count. These results were supportive of transient negative for mites by pelt testing. All mice from the 2 positive stress most likely due to disease. H&E-stained sections showed investigative groups (approximately 3400) were treated with severe unilateral suppurative maxillary cellulitis with granula- a nestlet impregnated with fipronil and methoprene (1.0 mg/ tion tissue that displaced the right globe. The lesion contained nestlet). The treated nestlet was replaced 2 wk later, for a total both Gram-positive cocci (consistent with Staphylococcus or Strep- exposure of 28 d. A single dose of topical selamectin (10 mg/kg) tococcus sp.) and filamentous bacteria (consistent with Nocardia was applied to the intrascapular region of mice in any cage with or Actinomyces sp.). an incompletely torn nestlet and to all mice in cages containing the 15 positive animals. No side effects from treatment were P10 Management of Line Infections in an Automated Blood observed. Culled animals were submitted posttreatment for pelt Sampling System for Rats examination. The treated mice remained fur mite negative for 5 mo. In January 2008, live Myobia musculis mites were noted on K Carpenter3, DM Cooper*1, K Heinz-Taheney2, J Sullivan2 1 mouse that had been treated with impregnated nestlets alone. 1Veterinary Resources, 2Discovery Pathology, 3Animal Studies, Based on this finding, it was assumed that treatment with only Eli Lilly and Company, Greenfield, IN fipronil/methoprene neslets failed to eliminate all mites. A sec- ond treatment was carried out, in which all haired mice were We observed a 6.8% incidence of rear limb paralysis in femoral treated topically with a combination of selamectin (10 mg/kg) artery– and vein–cannulated rats within several days of being and fipronil/methoprene (8 mg/kg) applied to the intrascapular placed on an automated blood sampling (ABS) system. This prob- region. Mice were treated twice, 30 d apart. Pelt examination lem was not seen with a previously used system and was not seen continues. No mice have tested positive for fur mites at 5.5 mo in animals from the same shipment not used on study. Necropsy of affected animals showed catheter-associated phlebitis or cel- posttreatment. All sentinels remained negative throughout this lulitis of the lumbar epaxial muscles or spinal cord. Cultures of outbreak. lesions and the pump lines detected Stenotrophomonas maltophilia, Escherichia coli, Staphylococcus aureus, Burkholderia cepacia group, P12 Effects of Altitude on Red Blood Cell Count, Hemoglo- Klebsiella oxytoca, and K pneumoniae. A luminescent assay for bin, and Hematocrit in African Green Monkeys (Chlorocebus bacterial ATP in the pump lines had average readings of 98,635 aethiops) relative light units (RLUs) from all pumps tested, even ones that DC O’Donnell*, CL Emerson were not associated with clinical signs. As the lines for the ABS were not designed for replacement between studies, we started a Animal Resources, Lovelace Respiratory Research Institute, weekly sanitation cycle in addition to that recommended by the Albuquerque, NM manufacturer. ATP levels were markedly reduced immediately Lovelace Respiratory Research Institute is in Albuquerque, NM following sanitation, and further decreased after the unit was idle with an altitude of approximately 5,300 ft. African green monkeys over a weekend (average of 6,135 RLUs.) Lines did not develop (AGMs) arrived at the facility in November 2006 from St. Kitts, an increased baseline in ATP after several months on the sanita- West Indies, Eastern Caribbean (n = 16), and February 2008 from Abstracts of scientific papers 2008 AALAS National Meeting Barbados (n = 20). It was noted that results of hematologic tests Portland VA Medical Center, Portland, OR performed on AGMs indicated that a few red blood cell (RBC) B6.129-Slc6a4tm1KplN10 strains of mice have a mutation resulting indices were significantly increased, compared with published in reduced serotonin-transporter function, which causes phe- reference intervals obtained at sea level. Analysis of hematologic notypic changes such as gut dysfunction, bone weakness, and tests performed on AGMs received in February 2008 and sampled stress-related behaviors. They are used at our facility for osteopo- approximately 2 wk after arriving at the facility revealed RBC rosis studies. We have received clinical reports in heterozygous counts were high in 13 of 20 AGMs and hemoglobin and hema- mice at about 100 d of age describing lethargy, dehydration, and tocrit values were high in 17 of 20 AGMs. Additionally, analysis hypersalivation. These clinical reports represented over 60% of of hematologic tests performed on AGMs received in November the heterozygous mice of this age. Bloodwork was consistent 2006 and sampled approximately 1.5 y after arriving at the facil- with dehydration. Necropsy revealed a flaccid, dilated distal ity revealed similar increases in RBC counts (12 of 16 AGMs), esophagus impacted with ingesta, which are findings consistent hemoglobin (14 of 16), and hematocrit (13 of 16). In most AGMs with achalasia. Literature has suggested small intestinal motility from both sources, values for mean corpuscular volume, mean is decreased in these strains, but this is the first time alteration corpuscular hemoglobin, and mean corpuscular hemoglobin of esophageal motility or lower esophageal function has been concentration were within published reference intervals. Simi- described. Microscopic evaluation of the esophagus around the lar changes in RBC indices have been observed in humans and impaction in these mice has been largely unremarkable to date. laboratory rodents at altitude. These results may be important Esophageal achalasia is primary motility disorder in which peri- to consider during acclimation of research animals, as well as in stalsis is lacking in the esophageal body and there is abnormal studies that would potentially be affected by RBC mass. relaxation in the lower esophageal sphincter. In humans, the etiology of esophageal achalasia is ill-defined, although an au- P13 Use of Dietary Fiber in the Management of Chronic and Re- toimmune component is suspected. The diagnostic test of choice occurring Diarrhea in Pigtail Macaques (Macaca nemestrina) for human esophageal achalasia is manometry. K McGee2, D Patel*2, J McKenna2, F Banks2, S Sekou2, K Prevost1, D Barnard1, B Skopets1 P15 Sublingual Blood Collection in Rats: An Alternative to 1DVR, 2SoBran, Tail Vein and Orbital Sinus Collection Inc., NIH, Bethesda, MD EM Wolak*, A Freeman Chronic diarrhea is a common syndrome in cage-housed nonhu- man primates. Six 3- to 12-y-old male pigtail macaques (Macaca Toxicology, Novartis Pharmaceuticals, East Hanover, NJ nemestrina) with chronic or reoccurring diarrhea tested negative Nonterminal blood collection for rats has routinely been accom- for common bacterial and parasitic intestinal agents. All animals’ plished using the tail vein and orbital sinus puncture (primary) diarrhea had been successfully managed for several months procedures. These techniques require more skill to master and by various repeated treatments with exception of dietary fiber may lead to tail lesions as well as lesions in the orbital plexus, supplementation. Two animals were fed a commercial labora- resulting in damage to the eyes. Ophthalmoscopic examinations tory primate diet containing 5% crude fiber. Four animals were and obtaining quality blood samples for use in clinical pathology, fed a commercial laboratory primate diet containing 8.5% crude toxicokinetics, and genomics are important in all toxicity studies. fiber. Studied animals were not involved in ongoing research. Currently, we are collecting blood sublingually from rats on our We hypothesized that higher fiber content may be curative for toxicity studies. Sublingual tongue vein collection has proven chronic diarrhea. To test this hypothesis, we arbitrarily supple- to be a successful and suitable method for collection and yields mented 6 animals’ diet with 1 tsp of Metamucil once daily and sufficient blood volume for quality clinical pathology and toxi- gradually discontinued other treatments. As a result, 4 out of 6 cokinetics analyses. In addition, the site can be drawn from more monkeys still had diarrhea, which was addressed by increas- than once in a day and the animals recover more quickly. The use ing the dose of Metamucil to 1 tsp twice daily. The increase of of this procedure has no impact on the animal’s health or ability Metamucil resolved diarrhea in 3 out of 4 animals. Animals to eat and the site of collection heals quickly. The technique is were maintained on Metamucil for a period of several weeks, relatively easy learn and can be applied for all types of blood after which the decision was made to switch them to a higher collections on research and toxicity studies. fiber (approximately 15.5%) commercial diet. Initially, the ani- mals were unwilling to eat the high-fiber diet. The problem was overcome by gradually transitioning the monkeys from the low- P16 Development of a Species-specific Giardia PCR Assay fiber diet to the high-fiber diet and moistening the biscuits with EK O’Connor*, LK Riley, CL Franklin, R Livingston orange juice. During the transition, 2 animals had 1 to 2 episodes Pathobiology, University of Missouri, Columbia, MO of diarrhea, which resolved without treatment or was quickly reversed by administering Pepto-Bismol. In conclusion, chronic Giardia muris is a flagellated protozoan that colonizes the small or reoccurring diarrhea of unclear etiology in pigtail macaques intestine of rodents and is generally considered nonpathogenic. can be successfully managed by daily supplementation of 1 to However, immunodeficient mice or mice with large parasite bur- 2 tsp of Metamucil or by feeding a diet containing 15.5% crude dens may develop clinical signs associated with malabsorption fiber. However, the study showed that primate diets containing and maldigestion, including a rough haircoat, weight loss, and 15.5% crude fiber are not highly palatable to macaques and re- abdominal distention. Current methods used to identify Giardia quire a very gradual transition period and possibly a temporary infections in rodents include histopathologic evaluation of the flavoring of the new diet (such as orange juice). small intestine or visualization of Giardia trophozoites on direct smears of small intestinal contents. Drawbacks to these methods P14 Dehydration and Excessive Salivation in B6.129-Slc6a4t- include their inability to accurately speciate Giardia organisms m1KplN10 Mice and the requirement for postmortem sample collection. Our hypothesis is Giardia PCR assays performed on feces will allow E Paster*, D Hickman, R Baldwin antemortem detection of infected mice and provide more sensi- Abstracts of scientific papers 2008 AALAS National Meeting tive and specific identification of Giardia infections compared Tissues from a 5-y-old privately owned, male guinea pig that to current detection methods. In this study, we developed PCR died suddenly were evaluated histologically. The stomach con- assays that target rRNA gene sequences to detect all Giardia spp. tained a spherical (approximately 7 mm diameter) expansile or to specifically identify G. muris. Feces and duodenal tissue mass originating within the muscularis externa and elevating from 10 mice from a colony with a history of Giardia infection the submucosa and mucosa. The mass was densely cellular and were tested using the developed PCR assays. All 10 mice tested consisted of spindle-shaped cells with ovoid to flattened nuclei positive on both PCR assays, while trophozoites were only arranged in tightly interwoven bundles and whorls within a scant visualized in 5/10 mice on direct smear. Additional testing of collagenous stroma. There were foci of central and superficial samples from pet store hamsters and mice with the G. muris necrosis and a low mitotic index. The initial differential diag- primers yielded positive results in 13/13 animals. The sensi- nosis was leiomyoma; however, immunohistochemistry using tivity of the G. muris primers was further evaluated by testing antibody against the tyrosine kinase receptor KIT demonstrated serial dilutions of plasmids containing cloned amplicons. The strongly positive cytoplasmic labeling. This was consistent with G. muris primers had a detection limit of 55 template copies. a diagnosis of a gastrointestinal stromal tumor (GIST). GISTs are The specificity of both PCR assays was confirmed by testing 17 mesenchymal tumors derived from the interstitial cells of Cajal in fecal samples from uninfected mice as well as through sequence the gastrointestinal tract, omentum, and mesentery. In humans, analysis of amplicons generated from infected mice. These data these tumors typically have activating mutations in KIT isoforms. demonstrate that testing rodent feces with the developed PCR Spontaneously occurring GISTs have been reported in dogs, assays provides sensitive and specific antemortem detection of horses, and as a case report in a rat. Because of their histologi- G. muris infections in mice. cal resemblance to leiomyomas, they may be underdiagnosed. KIT immunohistochemistry is generally necessary to confirm P17 Incidence of Mycoplasma Contaminants in Cell Cultures: the diagnosis. Differentiating these tumors from leiomyomas Detection and Growth on Trypticase Soy Agar Containing 5% is significant in human medicine because of the possibility of Sheep Blood metastasis and the use of tyrosine kinase inhibitors for therapy. To our knowledge, this is the first report of a spontaneous GIST GF Caviness*, JE Thigpen, J Locklear, TE Whiteside, MG Grant, in a small laboratory animal species other than the rat. DB Forsythe National Institutes of Health, Research Triangle Park, NC P19 Arthropod Detection during Screening of Sentinel Mice Mycoplasma contaminants in cell cultures, tumors, tissues, and J Locklear*, GF Caviness, JE Thigpen, TE Whiteside, MG Grant, animal products can significantly impact in vitro and in vivo DB Forsythe studies. The Quality Assurance Laboratory routinely tests all cell National Institutes of Health, NIEHS, Research Triangle Park, cultures, tumors, and animal products for viral, mycoplasmal, NC and bacterial contaminants prior to animal inoculation. This report describes the results from 2,241 specimens assayed for Our health surveillance program is designed to detect pathogenic microbial contaminants during the past 12 y and describes the bacteria, mycoplasmas, viruses, ectoparasites and endoparasites unexpected growth of some mycoplasma species on trypticase in our laboratory animals. Sentinel CD-1 mice and F344 rats soy agar containing 5% sheep blood (blood agar). The murine housed in polypropylene cages with autoclaved hardwood bed- viruses were detected using the conventional mouse antibody ding and autoclaved NIH-31 diet were submitted for a complete production test, serological assays, or polymerase chain reaction microbial work-up. On physical examination, no abnormal clini- (PCR) assay. Mycoplasmal and bacterial contaminants were cal signs were noted. All sentinels were euthanized using carbon detected using PCR assays or by plating the specimens on con- dioxide. Blood, oropharyngeal swabs, and fecal samples were ventional mycoplasma agar and blood agar. Microbial isolates collected for microbial assays. Direct cecal preps and perianal were identified using conventional microbial methods. Known impression tapes were examined for endoparsites. Pelts were mycoplasma control species were purchased from the American examined for ectoparasites. Upon observation of the perianal Type Culture Collection for comparative testing. The average impression slides, 2 types of mites were detected. Murine mites incidence of positive cell cultures was approximately 11% (255 of were ruled out based on gross anatomical features. The first mite 2,241) during the past 12 y. The yearly incidence of mycoplasmal species possessed a distinctive enlarged pair of pincer-like palps contaminants in cell cultures varied from 5% (17/376) in 2000 on the dorsal claws. The second mite, which was 420 µm long to 20% (39/196) in 1998. Surprisingly, no murine viruses were and 320 µm wide, was creamy white, globular in shape, and had detected in any of the cell cultures tested. An unexpected con- a striated cuticle. The mites were diagnosed as the Cheyletus mite firmed finding was that M. arginini, M. laidlawii, and M. pulmonis and the American house dust mite. Mites were observed in the formed small, visible, shiny colonies within 5 to 7 d on blood feed bin fines, the suspected route of exposure. These species agar. Our results confirm that mycoplasma contaminants are a of mites are commonly found in feed warehouses where their major problem for researchers using cell cultures; mycoplasma food sources, such as grains, oils, and paper products, are stored. species continue to infect a high percentage of cell cultures; These arthropods have not been reported to cause any known mycoplasma contaminants in cell cultures and animal products harm to rodent colonies or research studies. It is important to can significantly impact both in vitro and in vivo studies; and distinguish them from pathogenic mites. Personal protective mycoplasma species such as M. arginini, M. laidlawii, and M. equipment should be used to protect against potential mite al- pulmonis will grow on blood agar, which may be a useful tool in lergens. The mite’s lifecycle requires a humid, warm environment the detection and isolation of some mycoplasma species. for survival. Therefore, feed storage environmental conditions should ensure relative humidity is below 50% and temperature P18 Gastric Tumor in a Guinea Pig (Cavia porcellus) is maintained within 55 to 65 °F. Autoclaving the diet and inspec- tions and cleaning of feed bins and feed storage areas should be I Bergin*, M Kiupel conducted routinely to prevent any pest infestation of laboratory Michigan State University, Ypsilanti, MI animal feed products. Abstracts of scientific papers 2008 AALAS National Meeting P20 Mycoplasma arginini Associated with Swollen Joints in exposure to the rederived mice or soiled bedding. Lesions ob- Severe Combined Immunodeficient Mice Implanted with served in nu/+ sentinel mice included bronchoalveolar adenoma Tumor Cells (15/646) and epicardial mineralization (15/646). Lesions most commonly encountered in nu/nu mice were splenic lymphoid J Story, J Thigpen*, K Meshaw, M Grant, L Cheatham, B Hol- lister hyperplasia (18/646), splenic plasmacytosis (8/646), cystic change of lymph nodes (12/646), bronchoalveolar carcinoma Piedmont Research Center, Morrisville, NC (12/646) and acidophilic macrophage pneumonia (6/646). In Severe combined immunodeficient (SCID) mice are more sus- conclusion, both NU/NU and CD-1 Nude mice are acceptable ceptible to microbial pathogens than immunocompetent mice. sentinels for monitoring the health profile of research animals. C.B.-17lcr Hsd-Prkcdscid mice were implanted subcutaneously The strain-related lesions summarized in this report emphasizes with human tumor cells and subsequently developed swollen the importance of a thorough knowledge of the background le- joints and hyperkeratotic dermatitis. The objective of this report sions associated with the sentinel stock/strain used in a rodent is to describe the microbial and histological findings observed health program. in these mice. Specimens of skin, intact tumors, affected hocks, forelimbs and toes, and tumor cells were plated on mycoplasma P22 Chronic Experimental High-fat Diet Exposure Leads to agar and trypticase soy agar containing 5% sheep blood. M. Coagulopathy in African Green Monkeys (Chlorocebus aethiops arginini was isolated in pure culture on blood agar, and myco- saebeus) plasma agar and was confirmed by PCR assays from the intact K Ethun*, K Kavanagh, JR Kaplan tumors and from purulent fluid from the swollen limbs and toes. The skin from 5 mice was culture and PCR positive for C. Wake Forest University, Winston-Salem, NC bovis. The tumor cell line was PCR positive for M. hyorhinis and High-fat diets (HDF) are often used experimentally in animal M. arginini. Surprisingly, small, shiny, irregular colonies of M. models to study human disease. We hypothesized that prolonged arginini grew within 5 to 7 d on the blood agar plates inoculated feeding of HFD can result in abnormal hemostasis after observ- with the purulent fluid from the swollen joints and toes. These ing 3 cases of fatal hemorrhage in nonhuman primates following colonies were confirmed to be M. arginini by PCR procedures routine clinical procedures. A 19-y-old male African green mon- and sequence analysis. Swollen hocks revealed a moderate to key (AGM) was euthanized after observing severe hemorrhage severe pyogranulomatous inflammation involving the entire (hematocrit [Hct] 13%) following routine venipuncture. A 13-y- joint. There was fibrosis, destruction of bone, and pockets of old male AGM and a 7-y-old female cynomolgus macaque died neutrophils. Basophilic stringy material was observed in areas due to acute anemia (Hct 17% and 18%, respectively) after initial of inflammation of forelimbs and toes. It was concluded that presentations of bleeding plantar xanthomas and coagulopathy Mycoplasma arginini was associated with the swollen joints, and characterized by melena and hematuria, respectively. All animals was isolated in pure culture from most swollen joints. Mycoplasma were healthy prior to presentations, but were maintained on contaminants present a major problem to researchers using cell HFDs containing ≥30% calories supplied by fat and supplemental cultures and should be tested for microbial contaminants prior to cholesterol for >1 y. The aim of this study was to screen in-house injection into animals. This is the first report to describe swollen AGMs (Chlorocebus aethiops saebeus) for abnormal hemostasis joints in SCID mice due to M. arginini. under variable conditions of diet, disease, and age. Diabetic and non-diabetic AGMs (n = 21 [10 males, 11 females], 3 to 21 y old) P21 Background Lesions in Homozygous and Heterozygous were divided into 2 experimental groups, including those con- Mice of Two Stocks, Crl:NU-Foxn1nu and Crl:CD1-Foxn1nu, suming a high-fat diet (HFD; 30% calories supplied by fat) and Used as Sentinels those consuming a commercial low-fat chow diet (13% calories provided by fat) for >6 mo. There were 5 diabetics in the control JE Thigpen*1, BA Hobbs1, CB Clifford2, MG Grant1, MB Myers1, group and 3 diabetics in the HFD group. Buccal mucosal bleeding DB Forsythe1 times (BMBT) were performed once per animal and evaluated by 1Comparative Medicine Branch, NIEHS, Research Triangle Park, a single blind observer. Results revealed that the consumption of NC; 2 Charles River, Wilmington, DE a HFD was associated with a 32% increase in BMBT compared Ideally, animals used as sentinels in health surveillance programs to those consuming a chow diet (P = 0.05). The mean BMBT for should be free of microbial pathogens as well as background those animals consuming a chow diet was 62.78 ± 11.016 s and lesions which might confound results from health surveillance 99.83 ± 11.06 s for those consuming a HFD. There was a signifi- programs or research studies. Therefore, knowledge of the cant positive association of age with bleeding times (r = 0.56, P = prevalence of spontaneous lesions in the age, gender, and strain 0.008). Hyperglycemic status did not affect BMBT (P = 0.8). The selected for sentinels is critical. This report describes the spon- duration of HFD consumption was positively associated with taneous lesions observed over a 3-y-period in over 1200 NU/ a prolonged BMBT (r = 0.48, P = 0.03). Our results demonstrate NU and CD-1 Nude mice. Of each stock, half of the mice were that HFD prolongs bleeding times particularly in older animals homozygous athymic nu/nu; the remainder were immunocom- or following prolonged (years) of consumption. petent heterozygous nu/+. These mice were used as sentinels in 43 isolators housing rederived mice at Charles River. Specific P23 Ependymal Cyst Involving the Occipital Cortex in an pathogen-free sentinel mice were reared in isolators, then were Asymptomatic, Adult Cynomolgus Macaque (Macaca fas- transferred to other isolators and housed in direct contact with cicularis) colony mice for 2 wk, followed by 4 to 6 wk of exposure to soiled KP Yamada*1, SJ Popilskis2, A Garate3, M Oglesbee4, K LaPerle5 bedding. After 6 to 8 wk, sentinel mice were necropsied and ap- 1Tri-Institutional Training Program in Laboratory Animal Medi- propriate specimens collected for microbiology, serology, PCR, and histopathology. No microbial pathogens were detected. All cine and Science, Memorial Sloan-Kettering Cancer Center, The lesions observed in the nu/nu and nu/+ sentinel mice of both Rockefeller University, and Weill Cornell Medical College, New stocks appeared to be strain-related, rather than resulting from York, NY; 2Research Animal Resource Center, Memorial Sloan- Abstracts of scientific papers 2008 AALAS National Meeting Kettering Cancer Center, New York, NY; 3 The Animal Medical Reducing time and manpower are also concerns, for research Center, New York, NY; 4Department of Veterinary Biosciences, technicians and investigators alike. A novel approach to rat re- The Ohio State University, Columbus, OH; 5Laboratory of Com- straint has been employed by our staff for such tasks. The rats parative Pathology, Memorial Sloan-Kettering Cancer Center, are guided into a conical-shaped terry cloth and held in restraint New York, NY by rolling the edges and securing them with 2 large binder clips. The device has a small opening at the cranial end, which allows An experimentally naïve, adult, male cynomolgus macaque for unrestricted breathing for the animal. It is also opaque and (Macaca fascicularis) underwent a routine prestudy MRI of the brain. The MRI revealed a diffuse, bilateral cystic lesion, in- soft and gently molds to the animal to provide secure restraint. volving the occipital cortex, whose content was isointense with Most rats quickly become trained and enter the restraint readily. cerebrospinal fluid (CSF). Clinically, the animal had normal be- They can then be manipulated while lying quietly on one side, havior and mentation. No observable visual deficits were noted allowing the handler to have both hands free for equipment or on tracking tasks, and the animal had no deficiencies in fine animal manipulation. Holes near the rear limbs allow for access motor control. On necropsy, the cystic structures communicated to the saphenous veins, and the entire tail is exposed for access with the caudal aspect of the lateral ventricles and contained to vessels. The cloth is both inexpensive and sanitizable. Regular clear fluid compatible with CSF by fluid analysis. Histopathol- application of this technique by our staff has helped to decrease ogy showed ciliated ependymal cells lining the cyst with focal the amount of time and manpower needed to perform injection areas of hemosiderophages within the cyst wall. This lesion is or blood collection. In comparing 3 types of restraint methods, 9 compatible with an ependymal cyst, previously reported only female rats were bled from the saphenous vein. Three rats were in humans and generally considered an asymptomatic devel- placed in each of 3 different forms of restraint: the plastic bag opmental anomaly. decapitation cone, the bootie/shoe cover, and the terry cloth sewn into a cone shape. The venipunctures using the terry cloth restraint were completed more quickly than the other methods; P24 Impact of Selective Alpha-2 Agonists on Blood Glucose finishing 1 min and 7 s ahead of the bootie method and more than Levels in the Laboratory Rat 5 min ahead of the decapitation cone method. The technician re- JT Larsen, E Reardon, K Grayson* ported that the cloth restraint was more secure, in that there was little opportunity for escape or injury to the handler or the rats. Clinical Investigation Facility, David Grant USAF Medical Center, The rats demonstrated very little resistance or vocalization while Travis AFB, CA in the cloth restraint, and less manipulation was necessary. We randomly assigned 40 female Sprague-Dawley rats to 4 groups and anesthetized each with either: xylazine (5 mg/kg) + P26 Anaplastic Large Cell Lymphoma in a Sooty Mangabey ketamine (90 mg/kg); yohimbine (4 mg/kg) followed in 15 min (Cercocebus atys atys) by xylazine + ketamine; medetomidine (0.5 mg/kg) + ketamine (90 mg/kg); or atipamezole (2.5 mg/kg) followed in 15 min by L Collura*1, D Anderson2, JK Cohen1, P Sharma2 medetomidine + ketamine. We measured blood glucose levels 1Division of Animal Resources, 2Division of Pathology, Yerkes (BGL) at baseline and at 15-min intervals for the 1st h, then at 2, National Primate Research Center, Atlanta, GA 4, 8, and 24 h. In a separate experiment, 9 groups of 4 rats were A 13-y-old female sooty mangabey (Cercocebus atys atys) was anesthetized with medetomidine + ketamine and had serum evaluated due to a large submandibular mass and decreased analyzed for insulin and glucagon concentrations and liver appetite. The mangabey was serologically positive for simian biopsies taken to determine glycogen content. In the xylazine- T-cell leukemia virus type 1 (STLV-1) and simian immunode- ketamine group, BGL rose to 260 mg/dl by 1 h and fell to baseline ficiency virus (SIV). Generalized peripheral lymphadenopathy by 8 h. Yohimbine partially blocked the post-xylazine blood glucose elevation. BGL showed a much slower progression in was also observed on physical examination. A fine needle aspirate of the mass was poorly cellular and did not grow any the medetomidine-ketamine group, peaking at 2 h and remain- significant pathogens by aerobic bacterial culture. Biopsies from ing elevated through 8 h. Atipamezole pretreatment prevented medetomidine BGL increases. Serum insulin levels decreased the submandibular mass and an inguinal lymph node revealed a poorly differentiated tumor, with initial consideration given and glucagon levels increased in parallel with BGL, reaching 1 to a diagnosis of possible sarcoma. The animal had a severe ng/ml and 250 pg/ml, respectively. Similarly, the percentage of leukocytosis (79,900 with 55% lymphocytes). Due to poor prog- liver cells stained with periodic acid-Schiff reagent that contained glycogen fell sharply during the first 4 h after anesthesia, but nosis, the mangabey was euthanized. Subsequently, based on the histologic evaluation and immunohistochemical analysis had returned to baseline levels by 24 h. Our findings confirmed (vimentin positive, CD3 positive, cytokeratin negative) of the a previous study that reported xylazine administration causes lymph nodes, a diagnosis of anaplastic large cell lymphoma, hyperglycemia in rats. Medetomidine administration resulted a type of non-Hodgkin lymphoma, was made. This tumor has in nearly the same BGL elevation; however, the hyperglycemia been reported in humans, baboons, and a chimpanzee but has lasted much longer. BGL quickly returned to normal once the not been documented in a sooty mangabey. In addition, reports rats recovered from anesthesia and resumed eating. of neoplasia in sooty mangabeys are infrequent. P25 Alternative Method of Rat Restraint P27 Hind Limb Weakness in a Common Marmoset (Callithrix LL Roberts*, M Dyson jacchus) Unit for Laboratory Animal Medicine, University of Michigan, M Batchelder*, M Sauer Ann Arbor, MI Veterinary Sciences, Bristol-Myers Squibb, Wallingford, CT Creating a comfortable confinement for animals that allows ac- A 3.5-y-old pair-housed male common marmoset (Callithrix complishing tasks such as blood collection and injections while jacchus) presented with lameness and swelling of the left hind minimizing stress is integral to the success of a research project. leg. The animal’s history included multiple minimally invasive Abstracts of scientific papers 2008 AALAS National Meeting studies under isoflurane anesthesia. Physical exam revealed P29 Characterizing a Body Condition Scoring Technique and poor grasping ability in the left hind foot, weak reflexes, and Scale for Rats swelling of the left foot and ankle. Radiographs obtained the DL Hickman1, M Swan*1, R Luksic2,1, B Bustamante1, E Paster1 following day revealed multiple abnormalities in the left tibia, 1VA Medical Ctr, VA Medical Ctr, Portland, OR; 2College of Vet- femur, tarsus, and phalanges consisting of radiolucencies within the cortex and bullous periosteal densities. No lesions were erinary Medicine, Oregon State University, Corvallis, OR detected in other limbs, ribs, or pelvis. Subsequent radiographs Evaluation of weight loss or gain in rodents is often problematic revealed progression of the radiographic lesions in the left hind for multiple reasons. Rodents are often group-housed, making leg and the development of similar lesions in the right hind leg. feed intake difficult to assess. Weights can vary depending on The animal remained clinically stable during this time except time of day, scale used, and even observer. Also, for changes for weakness in the left hind leg. The animal did not appear in weight to be clinically meaningful, it is necessary to have a distressed or in pain, and was eating well. Ten months follow- baseline weight or age-matched control rodent for comparison. ing the initial presentation, the animal became depressed and Body condition score (BCS) scales have been described for mul- spent more time inside the nest box. Both hind legs were weak, tiple species including domestic (dogs, cats, livestock), mice, and and the animal was losing weight. The animal was euthanized, African claw toed frogs. In this study, we used a rat model of and both tibias were noted to be grossly abnormal at necropsy. inherited polycystic kidney disease to characterize the applicabil- Histopathology revealed bone marrow hyperplasia consisting ity of the published mouse BCS scale to the rat. We have found predominantly of granulocytic precursors and megakaryocytes, that minor modifications to the mouse BCS scale are required widespread cortical bone absorption, and periosteal bone for- in overweight (BCS4) and obese (BCS5) animals as fat does not mation. Extramedullary hematopoesis was evident in multiple reliably deposit over the lumbar vertebrae in the rat. Specifically, organs, and colitis was also noted. The histologic interpreta- palpation of the dorsal fat pads over the pelvic bones is a more tion was suggestive of several chronic myeloproliferative bone reliable assessment of the rat BCS than palpation of the fat over disorders in humans, but was not characteristic for any single the vertebral column, as described for mice. The BCS technique syndrome. This is believed to be only the second case of spon- and scale presented here will be of use for personnel assessing taneous chronic myeloproliferative bone disease reported in the health of rats used in biomedical research. nonhuman primates. P30 Comparison of Phlebotomy Techniques for Serial Bleeds P28 Successful Treatment of C57Bl/6 Ulcerative Dermatitis in Common Marmosets (Callithrix jacchus jacchus) with Caladryl Lotion MB Moyer*, J Barclay, J Devorak, AM Schiavetta ME Crowley*1, ML Delano2, SM Kirchain1 Comparative Medicine, USAMRICD, Gunpowder, MD 1Laboratory Animal Science Center, Boston University Medical Center, Boston, MA; 2Animal Resources at Children’s Hospital, Veterinary technicians were assigned to collect blood samples Children’s Hospital Boston, Boston, MA for 3 time points, ranging from 10 to 45 min apart, in 16 common marmosets (200 to 500 g). The goal was to obtain 400 µm of whole Ulcerative dermatitis (UD) in C57BL/6 mice and related strains blood at each time point from the saphenous vein. Several methods is a frustrating condition to manage in the laboratory animal of saphenous vein blood collection were compared: use of a 1 cc facility. Mice often become extremely pruritic ,with scratch- tuberculin syringe, use of a cutoff butterfly catheter, and use of a ing leading to several complications. Most of the pathology of blood collection needle into a microvette tube or capillary tube. UD results from secondary infections caused by the relentless Collection devices were prepared using either liquid heparin or scratching. Many topical and parenteral products and therapeutic EDTA. One or more of these techniques were used on one or both protocols have been used for years with varying success. We legs of each marmoset. Femoral vein or tail vein phlebotomy were have been using a therapeutic protocol for UD incorporating conducted when all other techniques did not yield an appropriate camphor/pramoxine/zinc (Caladryl) lotion for 2.5 y and have sample size. Femoral phlebotomy increased the chance of large seen a dramatic improvement in 65% of the UD cases. The first hematomas; when used, that leg was only sampled once. The component of the protocol is attentiveness by animal care staff, marmosets were anesthetized using an isoflurane system (initial who are the first to identify health cases and are critical to the induction in an inhalant anesthesia chamber, followed by nosecone success of this therapy. After early detection, the UD site is gently maintenance) on a heated surface. Procedures were conducted cleaned with chlorhexidine solution using a clean gauze and on 4 animals simultaneously. Two technicians were assigned to then dried with clean gauze. Caladryl is then applied in a thin each animal: one monitored anesthesia and positioned the leg; the layer once daily for 5 to 7 d using either a clean cotton swab or a other performed the phlebotomy. Marmosets were prepared by clean gloved finger. Caladryl dries the UD lesions effectively and clipping the venipuncture site, then wiping the area with alcohol relieves the itching. Improvement is seen in most cases within 2 to maximize vasodilation. The assistant occluded the vessel, and to 3 d with a range up to 10 d. Healing ensues sequentially after the sample was collected. Pressure and ice pack were then ap- cessation of scratching, with resolution of erythema and drying plied to the site. During recovery, technicians held marmosets of the lesion. Treatment must continue after lesions have healed until awake, then orally administered up to 1 ml of corn syrup. grossly and must continue or be reinstituted if any scratching is Upon return to their home cages, animals were provided a heat noted. Some cases do recur several months after the initial di- padrock in their hammock and a fleece pad on the floor. Different agnosis. Therapy has been less successful in mice with multiple techniques yielded different results for each technician; however, UD lesions or advanced full skin thickness lesions. Pruritis is the tuberculin syringe method or cutoff butterfly catheter method a significant inciting event in many allergic and inflammatory worked best, based on length of time to achieve blood flow skin diseases. A major advantage of Caladryl is mitigating the and total sample size collected. By using different phlebotomy scratching to allow lesions to dry and heal. Many valuable mice techniques, veterinary technicians met the study requirement have been saved using this therapy. challenges of conducting concurrent serial bleeds on marmosets with no adverse effects on the animals. Abstracts of scientific papers 2008 AALAS National Meeting P31 Topically Administered Recombinant Leptin for the Treat- similar muscle contractions were observed in only some frogs ment of Ulcerative Dermatitis in Zucker Lean Rats (2/6) with acetic acid at 15 min postinduction in the absence of a wiping response. AAT did not differ from baseline at 60 and NV Gopee*, J Leakey, J Carraway 120 min. Most animals had a depressed RR at 0, 30 and 60 min National Center for Toxicological Research, Jefferson, AR (100%, 83%, and 66%, respectively), and a depressed WR at 0 Zucker lean rats developed ulcerative dermatitis predominantly and 30 min (83% and 66%, respectively). RR and WR nearly behind the forelimbs, with isolated lesions on the head and fully recovered 60 and 120 min, respectively. Propofol did not behind the ears, several weeks following procurement from the show any significant effects on HR (P > 0.05). On the other hand, vendor. Histopathology of the skin lesions revealed moderate propofol did induce respiratory depression (P < 0.01). The mean to marked epidermal ulceration with neutrophilic infiltrates in respiratory rate significantly decreased from baseline (27.8 ± 6.2 the presence of bacterial colonies, dermal necrosis of variable breaths/min) to an average of 0.7 ± 1.6 and 0 breaths/min at 0 degrees of severity and numerous inflammatory polymorphonu- and 15 min, respectively, and progressively returned to 19.5 ± 7.7 clear and mononuclear infiltrates, with several animals having breaths/min at 2 h postinduction. AUC0-t was 24.07 µg min/ml microabscesses or granulomas. Skin culture showed the pres- and AUC to infinity (AUCinf) was 24.71 µg min/ml. The elimina- ence of Staphylococcus aureus. Treatment with broad-spectrum tion half-life (T1/2) was 1.18 h. In conclusion, when administered amoxicillin/clavulanic acid per os did not resolve the skin lesions as a single-bath immersion (88 mg/L) for 15 min, propofol does despite bacterial sensitivity and previously reported success in not appear to be a safe and effective anesthetic for Xenopus laevis obese mice. Leptin is a 16 kDa circulating hormonal product of frogs due to the narrow dose-effective window, as well as the the ob gene and is reported to be higher in Zucker obese rats than short duration and light anesthesia level obtained. in lean rats. Review of the literature has reported that following exogenous administration of leptin, enhanced re-epithelialization P33 NHP Biodata Chart and accelerated wound-healing were observed in obese mice. R Ford* In addition, leptin alleviated the severity of the inflammation in S. aureus-induced arthritis in rodent models. As such, a treat- Veterinary Services, Primate Products Inc. Panther Tracks Learn- ment regimen consisting of topical administration of 1 and 5 μg ing Center, Immokalee, FL recombinant rat leptin applied to the affected areas once daily Data is very important in the world of biomedical research. It every other day was initiated and maintained for several months. will always determine your next step. Nonhuman primates are Significant healing of the lesions, as evidenced by a reduction in an important model in the biomedical industry. They are mainly wound size and severity, was observed in the rats treated with used for human diseases when lower-rank animals will not suf- the higher (5 μg) dose of leptin compared with saline controls fice, so we greatly rely on their data to determine our next step. and lower dose (1 μg) animals. In addition, histology of the As a technician who works with primates daily, I’ve noticed that healing lesions was characterized by epithelial hyperplasia and there are few pieces of information that contain important data dermal fibroplasia adjacent to the ulcerated areas. However, due about primates all in one place. If you have a question, you often to the extensive nature of some of the ulcerations, healing was find yourself riffling through pages and pages of references to slow and prolonged and did not completely resolve in some find an answer. I’ve designed and developed a chart that will of the treated rats. No adverse effect on weight was observed help answer your most common questions in one place. This throughout the treatment period, and S. aureus was cultured chart includes information on specific primates such as rhesus, from the lesions following termination of leptin treatment. The cynomolgus, African green, baboons, and chimpanzees, and results further substantiate the potential for leptin to provide date including hematology, chemistry, arterial blood gas, blood beneficial therapeutic effects on the management of moderate groups, physiology, growth curve, and dentition. ulcerative dermatitis in Zucker rats. P34 Mouse Immunocomb Kit: A Quick and Easy Way to Si- P32 Anesthetic Properties of Propofol in African Clawed Frogs multaneously Detect Antibodies against Mouse Parvovirus, (Xenopus laevis) Mouse Minute Virus, Murine Norovirus, and Mouse Hepatitis S Guénette, F Beaudry, P Vachon* Virus in Laboratory Mice Veterinary Biomedicine, University of Montreal, St-Hyacinthe, ML Wunderlich1, S Mazar2, L Campbell1, WR Shek1, RK Dha- QC, Canada wan*1 1Diagnostic Reagents, Charles River, Wilmington, MA; 2Biogal The objective of this study was to determine the level of an- esthesia attained in Xenopus laevis frogs using a propofol bath Galed Labs, Kibbutz, Israel administration. Thirty-three Xenopus laevis nonbreeding female Immunocomb kit has been shown to be an effective tool to test frogs were used. Following the determination of an appropriate mouse sera for several important infectious agents of labora- anesthetic dose (pilot study to show effectiveness of bath immer- tory mice, including Sendai virus, coronavirus, and Mycoplasma sion 88 mg/l for 15 min), the acetic acid test (AAT), withdrawal pulmonis. Our hypothesis was that this same technology could reflex (WR), righting reflex (WR), heart rate (HR), and respiratory be used to detect serum antibodies against currently prevalent frequency (RF) were used to evaluate central nervous system infectious agents mouse parvovirus (MPV), minute virus of depression. Pharmacokinetics of propofol were calculated fol- mince (MMV), mouse norovirus (MNV), and mouse hepatitis lowing blood concentration determination by tandem liquid virus (MHV) in laboratory mice, and that this tool would perform chromatography/mass spectrometry analyses. AAT results with equal efficacy as the microplate ELISA and multiplexed show that loss of the nociceptive response occurred between the fluorometric immunoassay (MFIA) technologies. Serial dilu- moment frogs were removed from the bath and during the fol- tions of previously characterized standard mouse antisera and lowing 30 min (P < 0.0001). Only at 15 min postimmersion was nonimmune mouse serum were tested to compare the new kit the wiping response absent. Muscle fasciculations following the and ELISA detection limits and to assess the kit selectivity and application of the acetic acid were observed (4/6 animals), and diagnostic accuracy. The standard dilution at which sera are Abstracts of scientific papers 2008 AALAS National Meeting tested in the new kit is 36-fold. The ratio of the kit antiserum Ten blood samples from 3 different colonies of mice were tested at titers to this dilution ranged from 22 to >178. This proves that random. Three samples were from mice known to be multiplexed the kit sensitivity is ample for demonstrating viral seroconver- fluorometric immunoassay (MFIA) negative for all viruses; 2 sion by mice from adventitiously infected colonies. Diagnostic samples were from mice positive for mouse parvoviruses but performance of the kit was assessed with serum samples from negative for MNV; and 5 samples were from mice known to be barrier-reared retired breeder and young adult SPF mice and positive for MNV. We instilled 40 μl of blood from a heparin- sera from experimentally infected mice. No false-positive or microcapillary tube into the filter lid. Plasma was allowed to false-negative results were obtained in 33 tests of known ELISA- filter to the test membrane for 60 s. The lid was opened, and 150 positive sera and 128 tests of known ELISA-negative sera. The kit μl of chase buffer was added to activate the test mechanism. The assays detected seroconversion at the same days postinoculation buffer saturated the test membrane, causing the plasma to flow (DPI) as did ELISA and MFIA. These results show that the mouse unidirectionally to a test port. The buffer resuspended a colloidal immunocomb kit performance characteristics are equivalent to gold marker that followed the plasma and conjugated with any those of established and validated microplate ELISA and MFIA antibody captured on the test or control lines. Captured colloidal for serosurveillance of SPF colonies. gold marker appeared as a red stripe of varying intensity (Rann scale). Tests gave positive results at the test or control lines within P35 Biosecurity Lockout System to Influence Human Traf- 10 min. All 5 devices detected the presence of MNV in the MNV ficking: Ingress Control as a Component of the Prevention of positive samples. The remaining five devices were negative for Rodent Infectious Agent Transmission MNV. Because no color developed at the test line with plasma positive for mouse parvoviruses, cross-reactivity did not occur. RP Reynolds*1, SJ Pomeroy1, M Elks2, JN Norton1 Control lines for all 10 tests were distinctly positive. Test results 1DLAR, 2Engineering and Operations, Duke University Medical were 100% accurate for selectivity and specificity. Comparison of Center, Durham, NC Rann scale intensity and MFIA optical density showed correlation in sensitivity. Results with the Fenway Direct Test demonstrated Although there are many causes of disease transmission between a rapid test for MNV was feasible and could be used to detect rodent areas in the research environment, one of the most diffi- infected mice and eliminate them from colonies. cult to control is the traffic flow patterns of humans. Movement of personnel and associated fomites from animal areas (whether a module/suite or facility) of a lesser health status to a superior P37 Spontaneous Case of Spina Bifida on a Mouse health status can spread infectious agents. An application was S Duarte-Vogel*, GW Lawson developed where the Division of Laboratory Animal Resources DLAM, UCLA, Los Angeles, CA codes the existing card access readers with a numerical level cor- responding to the health status of the rodents in the specific area. A spontaneous case of spina bifida occulta was found in a mouse Individuals that enter a facility or area with rodents of a level at our facilities. The mouse was a 22-d-old male, CD1-lpr/lpr corresponding to lesser health status are locked out of areas of with a “bulldog” tail that was short, thick, and had a 180-degree a higher health status until the system automatically resets at a curve to the right that began immediately caudal to the sacral predetermined time. Unimpeded traffic flow can continue to occur vertebrae. The mouse had complete bilateral rear limb paralysis within the same level or to a level corresponding to a lesser health with absence of superficial and deep pain as well as lack of anal status. Currently, any restrictions incurred on one day will not tone and severe generalized muscular atrophy. The differential carry over to the next day due to the system reset during the early diagnoses at this point were trauma, genetic malformation, and morning hours, although this period of restricted access can be spina bifida. Necropsy revealed the pelvic canal of this mouse extended. This application also has the ability to generate reports was extremely wide and deep because of a dorsal curve in the of individuals who attempt to violate the prescribed traffic flow. sacral vertebrae that began directly following the sacroiliac joint. Training and proper notification of institutional personnel is criti- A CT scan showed the defect in the vertebral column began at the cal in mitigating concerns of the research staff prior to initiating the level of the proximal thorax and continued to the coccyges. The use of this bio-security lockout system. The support of institutional thoracic vertebrae had partial development of the lamina and administration is important in implementing this system and in absence of the spinous process with exposure of the spinal cord. setting penalties for those who violate the appropriate traffic flow. Lumbar, sacral, and caudal vertebrae were abnormal on shape This presentation will describe the system and its specifications, and distribution, small, compressed, and difficult to differentiate. other traffic flow considerations, and implementation of the system The clinical signs and the gross and CT scan findings revealed into the overall rodent health program. spina bifida. Spina bifida is defined as failure of the neural tube to close during embryonic development. Different types of spina P36 Detection of Mouse Norovirus in Less Than 30 Minutes bifida have been described and include spina bifida occulta, Using a Novel Lateral Flow Test spina bifida cystica (myelomeningocele), and meningocele. The neurological damage as well as the disability degree depends N O’Luanaigh, CC Otte, WT Sauls, MP Goggin, BA Keniston, on the area of the lesion and how far down the spine it extends. RE Hopkins* Spina bifida is commonly associated with animals with short 1BBI Research, British Biocell, Inc., Madison, WI; 2LACF, Boston tails such as the Manx cat and the bulldog. University, Boston, MA; 3Research-Development, Fenway Biosci- ence, Boston, MA P38 Method for Repeated Blood Sampling from the Medial Ten novel lateral flow test devices for detecting mouse norovirus Saphenous Vein of Conscious Rats in a Long-term Study (MNV) were developed to rapidly detect MNV in mouse blood K Koszdin1, S Frayo*2, D Cummings2 without sample preparation or instrumentation. Each device 1Animal Research Facility, VA Puget Sound Health Care System, had a filter lid for fractionating plasma from blood and a hous- Seattle, WA; 2Endocrinology and Metabolism, Veterans Affairs ing containing a lateral flow test membrane. The membrane was Puget Sound Health Care System, Seattle, WA striped with MNV antigen and anti-mouse antibody as a control. Abstracts of scientific papers 2008 AALAS National Meeting We developed a method for withdrawing up to 1 ml of blood from 1Division of Animal Resources, Emory University School of the medial saphenous vein because we needed to consistently col- Medicine, Atlanta, GA; 2Department of Pathology, Yerkes Na- lect at least 400 µl of blood from rats without assistance. The rat tional Primate Research Center-Emory University, Atlanta, GA; is restrained in a 1.5-in. diameter, 12-in. long cloth stockinet tube 3Department of Pathology and Laboratory Medicine, Emory with one end partially closed with tape, such that the rat can push University, Atlanta, GA its nose out to breathe but cannot put its head through the hole. A high incidence of gingival overgrowth occurred in a group During placement, the front legs are pushed back to decrease the of New Zealand White rabbits receiving daily cyclosporine possibility of a toenail catching in the fabric. The stockinet is rolled (15 mg/kg, IM) while on a retinoblastoma study. Initially, up to expose the lower half of the body, and the rat is placed gently 4 rabbits presented with hypersalivation and inappetence. on its back and restrained with the palm of one hand. One hind Physical exam and accompanying necropsy revealed varying limb is exposed, and the thumb and index finger of the restraining degrees of hyperemia and moist dermatitis of the facial skin, hand grasps this limb above the knee, straightening the knee and and thickened gingiva in the maxillary and mandibular molar occluding the vessel. Petroleum jelly is spread on the medial side area. Over the next 9 mo, 10 of 18 additional rabbits presented of the leg to visualize the vein and to prevent blood from soaking with similar clinical signs (hypersalivation: 4/18, inappetence: into the fur. A bright light helps with visualization. The medial 3/18, both: 3/18), prompting attempts at treatment and a clini- saphenous vein runs along the inner thigh and branches just distal cal investigation. In an effort to alleviate symptoms, the dose to the knee in the shape of a wishbone. The vein is pierced 2 mm of cyclosporine was reduced to 10 mg/kg. The dose reduction proximal to this branch point with a 20-gauge needle. Blood can resulted in complete resolution of clinical signs in all but 2 rab- be collected in a capillary tube. After blood collection, the vein is bits. These rabbits were administered azithromycin (62.5 mg PO compressed to minimize hematoma formation. The stockinet is once daily for 7 d), a common treatment for cyclosporine-induced gently rolled off the rat. We were able to rapidly collect up to 1 gingival overgrowth in other species. Following dose reduction ml of blood from a single vein in rats ranging from 350 to 660 g. or azithromycin treatment, clinical signs resolved and rabbits The same animals were sampled every 3 to 4 wk for over 1 y. The remained free of symptoms throughout the remainder of the minimum time between collections was 17 d. Using this method, study. Fourteen rabbits were necropsied at the end of the study, one person can reliably and repeatedly collect at least 1 ml of blood and gingival width was measured. Although clinically normal, from a conscious rat. the gingiva in these rabbits was grossly thickened. Rabbits on cyclosporine for at least 2 wk had gingiva around the molars P39 Ensuring Normothermia during General Anesthesia that was significantly thicker (4.8 mm; P < 0.05) than untreated S Luboyeski* controls (2.5 mm). Histologic analysis revealed mild to moder- ate gingival epithelial hyperplasia and mild inflammation of CCM, Massachusetts General Hospital, Charlestown, MA the molar gingiva. Mild inflammation but not hyperplasia was Animals undergoing prolonged imaging studies or invasive observed in the incisor gingiva. Gingival overgrowth is a known surgery, or having a low percentage of body fat or a high ratio side effect of cyclosporine administration in other species but, of surface area to body weight may exhibit up to 8% loss of core to the authors’ knowledge, this is the first report in rabbits. Be- body heat during just 90 min of general anesthesia or prolonged cause rabbits are commonly used in studies which may involve sedation in the absence of appropriate supportive care. The resul- systemic cyclosporine administration, clinicians are advised to tant hypothermia routinely reduces heart and respiratory rates, include this possibility on their differential list in cases involving blood pressure, and cardiac output, and prolongs recovery times. hypersalivation, facial dermatitis or inappetence. Widely used methods to avoid hypothermia, such as circulating warm water pads under the animal or heat lamps overhead, can P41 Effect of Automated Cross-over Serial Blood Sampling on be inadequate or interfere with MRI and other scanners. The effi- Urinary Corticosterone in Rats cacy of covering the animal’s torso and extremities with pediatric hand and foot booties and using padded woolly blankets with SE DeBoer*1, E Kadyszewski2, MS Kovacs1 warm water bottles during sedation and transport, was assessed 1Comparative Medicine, 2Non-Clinical Statistics, Pfizer, Inc., in 4 3-y-old rhesus macaques in the course of routine veterinary Groton, CT support during general anesthesia for MRI scans. Animals were Automated blood sampling provides a method for sequential provided a circulating warm water pad only, hand and feet sampling of small amounts of blood in freely moving animals. booties only, or a water pad in conjunction with booties. Some We wanted to evaluate the utility of performing crossover pK animals were also provided blankets and warm water bottles studies on the same animal using an automated blood sampler. during sedated transport to the imaging room. All animals were The objective of this study was to compare the effect of 1 and 2 subjected to each combination over 5 scans. Rectal temperature serial blood collection sessions on the physiologic stress hormone was obtained at the onset of anesthesia and at extubation. When corticosterone in rats. A total of 15, male Sprague-Dawley rats these novel patient comforts were applied, animals experienced were used in this study. Five precannulated rats were assigned less than an average 0.5 °F body temperature difference between to each treatment group. Five rats received a standard pK blood onset and end of anesthesia, compared with an average 2 to 3 collection protocol over a 24-h period, involving collection of °F drop in body temperature with only a circulating water pad approximately 10% of their blood volume. Five animals received provided. In addition, recoveries were smoother and recovery 2 blood collection protocols separated by a 24-h wash-out, in- times were reduced by 20 to 30 min. The addition of these items volving a total collection of approximately of 20% of their blood has a greater impact on maintaining normothermia than current volume. Five control animals were tethered in the automated conventional methods alone. blood sampling units but were not subjected to automated sam- pling. Urine was collected twice daily from each rat to measure P40 Cyclosporine-induced Gingival Overgrowth in New Zea- urinary corticosterone and creatine. For single and double serial land White Rabbits (Oryctolagus cuniculus) blood sampling protocols, there was a statistically significant S Jean*1, P Sharma2, DK Taylor1,3, DM Mook1,3 elevation in the urinary corticosterone/creatine ratio during Abstracts of scientific papers 2008 AALAS National Meeting each of the blood collection sessions in the automated sampler tered a single subcutaneous dose of 1 of 4 different PEG-leptin when compared to controls. Despite significant decreases in variants, native human leptin, or vehicle control at study initia- hematocrit in both the single and double serial blood sampling tion. Body weight and food consumption were recorded daily for sessions, urine corticosterone/creatine ratios returned to normal 7 d after treatment. Over that period, we saw increased weight within 24 h of each sampling session. We determined that rats loss and decreased food consumption in mice treated with the recover quickly from the stress of blood collection and despite PEG-leptin variants. Mice treated with native human leptin or lower than normal hematocrits, evaluation of the stress hormone vehicle control did not lose weight and had slightly increased corticosterone indicates no residual stress effects. food consumption. In conclusion, we have shown that a single subcutaneous administration of a PEG-leptin variant in ob/ob P42 Ambulatory Approach to Long-term Bile Collection in mice caused increased weight loss and decreased food consump- the Canine tion when compared to mice treated with native human leptin, which indicates a therapeutic advantage of these compounds S Vessie, T Ziegelhofer* when compared to commercially available compounds. Cardiovascular/Infusion Toxicology, Huntingdon Life Sciences, East Millstone, NJ P43 Critical Analysis of a Biosecurity Program After presurgery bloods and jacket acclimation, each canine is AR Santiago* prepared for an aseptic surgical procedure during which a T- DVMS, UT—MD Anderson Cancer Center, Houston, TX catheter is secured within the lumen of the common bile duct. The terminal ends of the catheter are tunnelled through the With the exponential growth of animal research in the Texas abdominal wall and connected subcutaneously to 3 vascular Medical Center comes increased concerns about biosecurity. access ports (VAPs). For 6 wk postsurgery, animals are carefully Maintaining the integrity of research and assuring the health monitored for signs of pain, distress, and jaundice. Catheters are and welfare of laboratory animal colonies require an up-to-date, frequently flushed to ensure patency, and liver function is as- feasible, and cost-effective biosecurity program in every animal sessed through weekly blood collection. The occluding balloon facility. The following major areas at our facility are described: within the catheter is inflated, temporarily obstructing the flow traffic control, pest control, cleaning and disinfection, host sus- of bile into the duodenum. Animals are placed in jackets, with ceptibility, and communication. The biosecurity program of M.D. a bile sampling line and a cooled bile collection bulb within the Anderson’s vivarium is current and cost effective. Occasional jacket pocket. Simultaneously, an ambulatory pump in the op- biosecurity breaches are attributed mainly to lack of training or posite jacket pocket infuses a bile salts solution into the duodenal language barrier. Human error and economic constraints are the end of the catheter. Upon study termination, jackets are removed, major limiting factors to an ideal program. The recommenda- catheters are flushed with saline and the occluding balloon is tions are inclusion of biosecurity in orientation of personnel and deflated to allow bile flow from the gall bladder to the intestines. investigators; translation of signs into 2 other major languages The animals are flushed at least once every 14 d to ensure proper spoken in the facility; enrollment in English as a second language catheter function. Bloods are collected once per month to evalu- courses offered at the institional level; incentive program for ate liver values. We have successfully performed surgery on 10 compliance; termination for top 3 biosecurity measure breaches; canines and executed several studies to completion. We believe and the formation of an interinstitutional committee for regional our refined, ambulatory method of long-term bile sampling is perspective. the most optimal. Since the model does not require tethering, the animals are not only paired housed during non-sampling P44 Contamination Control Mat: Solving a Hairy Situation periods, but are also readily exercised during collection intervals, a significant advantage to this procedure. A Dryman* University Research Animal Resources Life Sciences, University P42.5 Increased Weight Loss and Decreased Food Consump- of Georgia, Athens, GA tion in Ob/ob Mice Administered PEG-leptin Variants when Working with New Zealand White rabbits results in large Compared to Mice Administered Human Leptin amounts of rabbit fur floating in the air. This fur coats the work- M Cook*, V Kraynov, H Cho, B Kimmel, D Axelrod, J Pinkstaff, space and personnel, aggravating allergies and any personnel L Sullivan trying to clean the workspace after procedures using rabbits are complete. We run a polyclonal antibody production service Ambrx, La Jolla, California that uses New Zealand White rabbits, and sometimes an entire This study was designed to evaluate the efficacy of 4 different workday is spent handling rabbits in our small procedure room. PEG-modified human leptin compounds when compared to After multiple procedures, the rabbit fur adheres to all horizontal native human leptin in ob/ob mutant mice. Leptin, a 10-kDa surfaces, along with scrubs and lab coats. Previously, our sani- protein, is produced by adipose tissue and is thought to play a tation method was to carefully vacuum the workspace, trying critical role in body weight regulation. Due to leptin effects on to suction up the fur without blowing it throughout the room, appetite satiety, energy expenditure, and metabolism, leptin is which met with limited success. About a year ago, we started a viable therapeutic option for weight loss and weight manage- using a mat on our exam tables, similar to the contamination ment in obese patients. Initial studies evaluating the therapeutic control flooring mats we use outside of our barrier and quar- potential of leptin have been performed in mutant ob/ob mice. antine rooms. Initially, the idea was to see if it would improve These mice lack functional leptin and, as a result, have reduced the rabbits’ welfare by preventing distress due to sliding on the activity, decreased metabolism, and are genetically obese. We slick metal exam table. We were excited to find a different and hypothesized that administration of PEG-leptin would cause entirely helpful use for the mat. The fur that once roamed our increased weight loss and decreased food consumption when procedure room would instead collect on the mat. We found that compared to animals administered native human leptin. Female those members of our staff with rabbit allergies had reduced ob/ob mutant mice (approximately 8 wk of age) were adminis- symptoms when we started using the mat. The mat is easy to Abstracts of scientific papers 2008 AALAS National Meeting care for and shortens cleanup time immensely. We would rec- weight was measured. Average food wasted was significantly ommend using this mat for rabbit work, and hope to try it with (P = 1.94e-05) reduced from 26% to 0.5% with the modification. other species soon. Food left in the ball did not vary significantly between the groups (P = 0.37). Each tray cost $10 in materials and took 1 h to build. P45 Effect of Cage Change Frequency on Rodent Breeding Daily cleaning with running water and scheduled cage sanita- Performance tion in the rackwasher were both easily accomplished with balls in situ. This simple and inexpensive modification significantly AE Sanderson*, HM Castor, J Lohmiller, S Boutin reduced food waste, with the added benefit of reduced cleanup Veterinary Services, Taconic Farms Inc., Hudson, NY time following use. The Guide states that “in some instances, frequent bedding changes are contraindicated, such as during some portions of P47 Online Inventory Equipment Database System the prepartum or postpartum period.” Consequently, the pro- B Liu*, KA Wendler, CA Osina, L Sun, JJ Sharp fessional judgment of many investigators, facility managers, Center of Comparative Medicine, Baylor College of Medicine, and animal care technicians is that disrupting certain sensitive Houston, TX rodents will result in breeding failures and loss of newborn pups. We assessed breeding performance of 1 presumed sensitive rat Equipment used in vivaria includes hundreds of racks, numer- line and 1 mouse line to determine what effect cage change had ous hoods, and many pieces of other equipment types. Due to on the survivability of new litters. We selected C57BL/6NTac frequent movement of many types of equipment among animal mice and NTac:NIH-Whn rats based on their presumed high facilities in geographically separate locations, it is important to sensitivity to parental neglect or cannibalization in response to track the location of equipment and maintain updated informa- cage manipulation. The study consisted of approximately 250 tion about the condition of the equipment. We have introduced a breeding rat females and 5,700 breeding mouse females per wk web-based online inventory database system that keeps records and was conducted in multiple barriers at 2 geographic locations. of equipment for animal facilities. The system is compatible with Breeding performance was evaluated over a 22- to 25-wk period. different computer platforms and different browsers. Each piece Study animals were placed in 1 of 2 study groups, “change” or of equipment is identified by an applied barcode; this barcode “no change.” The change group had their cages changed on a technology ensures the integrity of data records. Data entry is weekly basis regardless of the presence of newborn pups, and facilitated for single items, multiples of the same item, or a large the no change group was not changed until the following week quantity of items. An implemented web interface allows batch if new pups were observed. The study design was such that all data entry. For convenience, a formatted spreadsheet can also be environmental and caretaker factors were rigorously controlled. uploaded from the website, which allows facility users to record The primary factor for evaluating breeding performance was the information quickly for racks and cages. Users may search comparing change with no-change pup survivability at the the database for equipment not in use and submit a request to time of weaning. Data was collected on a weekly basis, captur- have it moved to their facility or request a transfer of equipment ing the total number of female breeders and the total number from one facility to another. Alternatively, users may request that of pups weaned. The production efficiency was calculated by equipment be removed from their vivarium to the warehouse. dividing the number of pups weaned by the total number of ac- Equipment needing repair or to be discarded is also noted. Users tive breeder females. The data demonstrated that there was no can search the inventory items in the database, review the im- statistically significant difference in the breeding performance ages of the items, and make a request for the specific items. The or efficiency of C57BL/6NTac mice or NTac:NIH-Whn rats based warehouse manager will then be notified and deliver the items on cage change alone. This outcome is contrary to the general to the correct site. The database keeps warranty information, belief of rodent breeding and would not support modifying which will be helpful in the event that service is needed for any recommendations of the Guide stated as “There is no absolute equipment. Equipment usage and performance information can minimum frequency of changing bedding, but it typically varies be obtained from archived data which can inform equipment from daily to weekly.” replacement decisions. The database provides accessibility for future mobile systems such pocket PCs. P46 Simple Modification of a Commercially Available Primate Foraging Device Significantly Reduces Food Waste P48Web-based Animal Internal Transfer Systems AS Glenn*, J Watson B Liu*, L Sun, JM Simonet, KA Wendler, JJ Sharp Animal Resources, Johns Hopkins University, Baltimore, MD Center of Comparative Medicine, Baylor College of Medicine, Houston, TX Commercially available perforated foraging balls that hang on the outside of the cage are used at our institution for nonhuman Internal animal transfers (room to room, PI to PI, protocol to primate enrichment. Food treats are placed in the balls and pri- protocol, and so forth) are one of the most important activities mates forage for them through holes in the balls. However, 15 of an animal program. Prior to our web-based animal internal to 55% of the treats were wasted when they fell onto the floor transfer system, all transfer requests were submitted on paper, and out of reach. We addressed this problem by constructing which was slow, and there was no central database to reference a polycarbonate tray to catch dropped food and permanently for status of transfers. Coordinated by the transfer agent, a series attaching it to the bottom of each ball with 2 3-in stainless steel of actions was required to ensure that the transfers were done cor- bolts. Eight macaques were used in a study to assess the effect rectly and promptly. For example, when a PI submitted a transfer of the modification on wasted food. Balls were placed on cages request, the transfer agent had to verify the animal protocols, for a fixed period each day (1.5 to 5 h) in a crossover design. retrieve health information from veterinarians, check with the Four macaques were given the modified ball for 3 d then the IACUC for possible compliance issues, and ask the receiving unmodified ball for 3 d, and 4 were given the balls in reverse facility manager to verify space availability. Once the request order. Initial, final (left in the ball), and wasted (on the floor) food was fully approved, the transfer agent had to contact the PI, the Abstracts of scientific papers 2008 AALAS National Meeting manager of the sending facility, and the manager responsible for were counted following 48 h of incubation at 37 °C. Mean CFU the actual move. After the transfer was completed, the receiving counts for clean and dirty samples from unsealed barn concrete PI had to be informed. In most cases, the parties involved in this floor were significantly lower than corresponding counts from process were in separate locations. Our new web-based system the sealed barn concrete floor. Mean CFU counts obtained after allows all of the management steps to be handled with just a few cleaning and disinfection were acceptable in all barn locations clicks. Completion of an upstream step automatically triggers a tested. Pooling of disinfectant in cracks, crevices, and pores of notification email for next downstream steps. Notification emails unsealed floor may allow for longer contact time compared to for each step are automatically sent to the appropriate persons. sealed flooring, where residual disinfectant dries more quickly Investigators benefit from this automated system because they or runs off into drains. can view the real-time status of their requests. Completed transfer requests are archived, with reporting capability available if de- P51 Simple Environmental Enrichment Technique for Am- sired. Validation elements have also been implemented to ensure phibian Species the security and integrity of data. Early use of this system shows it to be efficient and easy to use by all parties with a high level of CA Bluhm*, PA Stiles, A Johnson acceptance among researchers and animal facility managers. Animal Care, Ferris State University, Big Rapids, MI A common misconception is that captive reptiles are stoic, P49 Novel Cage Flag System for Identifying Special Husbandry adaptable, and highly tolerant of the abnormal conditions in Requests captivity. However, research suggests that reptiles may be less BA Davis*, H Graninger, ND Hitt tolerant and less adaptable to captive environments than was suspected. Because there has been limited information on ap- NINDS/AHCS, Charles River, Germantown, MD propriate enriched artificial habitats, many of the current reptile Rodent facilities are often faced with special husbandry requests husbandry and enrichment techniques have been derived from because of phenotype- or protocol-related issues. These requests empirical evidence. We developed a lab-tested, simple, sustain- can range from special feed for an entire room to varying indi- able environmental enrichment technique for an amphibian vidual cage care instructions. This degree of customization of species habitat. Our previous habitat consisted of a water-filled care can result in miscommunication and lost research data if an 36 by 36 by 24 in. square polystyrene container raised at one efficient, accurate method to alert the animal care staff to changes end. Rocks were placed throughout to allow the frogs rest areas. in husbandry is not used. After trying numerous methods to While this habitat was in use, we experienced a high incidence identify cages where special husbandry was required, a system of redleg and as a result a high mortality rate. A decision was was designed where small laminated flags are clipped to cage made to design and construct a habitat that would more closely card holders on the individual cages. These color-coded flags resemble that of the frog’s and turtle’s natural habitat. The end display special instructions using a combination of the general result was the use of a 100-gal rugged plastic livestock tank that description of the change with specific details noted in perma- was divided using a polycarbonate resin sheet. One side of the nent marker. When special care is no longer required, cards are tank was filled with river rock and pea gravel; large stones were easily detached and wiped clear with ethanol for reuse. To make placed on the bottom of the tank on the opposite side and the identification of cages convenient for animal care and research tank filled with water. In doing this, we created a dry side and staff, several sets of each type of flag are provided in each animal swimming hole that is approximately 12 in. deep. The frogs rest holding room. After several revisions, we have streamlined our on the rock-filled side and eat crickets, and swim in the water- flag system so that it consists of 5 flags, each with our most com- filled side. A heat lamp on a timer was placed above the tank to mon special care and feed options, which can be checked by the allow the frogs to bathe in the artificial sunlight. A base and lid researcher using a permanent marker. This system has greatly were constructed for the tank using composite decking material. improved our ability to provide the customized care required The base of the tank is on casters to allow easy movement of by so many of the research projects in our facility with minimal the tank. The lid was designed to remain in place until the frogs additional expense. are removed or the tank is cleaned. A drain in the bottom side of the tank with a shut-off valve allows for easy cleaning. We P50 Microbiological Monitoring of Sealed and Unsealed Barn have housed up to 75 frogs at a time in this habitat with little Floors using RODAC Bacterial Culture Plates or no death and have not had an incidence of redleg since. We also extended the same concept to housing our turtles. The tank BF Grasperge*, J London, R Stout, D Baker is a 50-gal plastic stock tank, though it lacks a base and lid. A Division of Laboratory Animal Medicine, Louisiana State Uni- simple ramp was designed using polycarbonate sheeting and versity School of Veterinary Medicine, Baton Rouge, LA indoor/outdoor carpeting secured with carpet tape to allow the turtles easy access to the dry area. Sealed flooring is considered ideal for animal holding facilities due to its impermeability to contaminants. The objective of this study was to determine the cleanliness of sealed and unsealed P52 Making the Transition to Biometrics concrete floors within the Louisiana State University School of C Kilcullen-Steiner* Veterinary Medicine client barn before and after cleaning and University Animal Care, University of Arizona, Tucson, AZ disinfection using replicate organism detection and counting (RODAC) bacterial culture plates. The floor locations sampled Security is paramount in biomedical research facilities. Keys and included unsealed concrete barn aisle, sealed concrete dump- security cards are commonly used to restrict access to the animal ster storage area, sealed equine intensive care unit, and sealed facility to only authorized personnel. However, these items can equine surgery floors. For each of the 4 locations, 10 sites were be easily shared among research personnel, and some may write sampled immediately before and 10 min after cleaning and dis- the access code on the card, jeopardizing facility security. To infection. Floor sites were sampled with RODAC plates, and the increase security, the university decided to go biometric. Optic total numbers of bacterial colony-forming units (CFU) per plate scanners, finger scanners, and palm scanners were investigated. Abstracts of scientific papers 2008 AALAS National Meeting Optic scanners were ruled out because many felt the scanners Laboratory Animal Services, Novartis Pharmaceuticals Corpora- could damage their eyes or make them go blind. We selected tion, East Hanover, NJ fingerprint scanners, despite some employees’ fears of having The common marmoset (Callithrix jacchus) can be an invaluable their fingerprints captured; the idea that the fingerprint was research model. Its small size makes it particularly useful when converted to a logarithmic equation was hard to accept. The compound is in short supply or expensive to produce, and safety change to biometrics took several months. New biometric cards concerns such as those encountered when working with old had to be purchased and issued. Fingerprint equations had to be world nonhuman primates are relatively nonexistent. We have downloaded on the biometric cards and the new readers had to used this species for approximately 10 y with success in several be in place and functioning prior to the changeover. Having the types of research in both discovery and preclinical safety. The new biometric system makes it impossible for an access card to small size of the marmoset allows its use with approximately be used by someone other than the approved card holder. Usage 1/10 the amount of compound that would be necessary when can be tracked better with this technology, thus creating security working with a more commonly used model, such as the cyno- levels to meet the needs of our users. The new system has not molgus macaque. Unfortunately, two aspects of the marmoset’s eliminated all access problems; a very small percentage of us- physiology can present significant challenges for the researcher. ers do not have readable fingerprints, and the system does not It has a high metabolism rate that makes post–blood collection prevent “piggyback” entry. We are looking to new technology hemostasis difficult even though it has small blood vessels. to remedy those problems. Once blood is drawn, it seems to clot quickly, often causing blood samples to need to be redrawn. The methods of blood P53 The Effects of Social Housing on Diabetic Laboratory collection and intravenous injection used at our site have been Canines: Food Consumption, Weight, Behavior, Glucose Toler- continually modified over the years in an effort to minimize ance, and Insulin Levels these challenges. Modifications in restraint methods, equipment, C Coke Murphy*1, KJ Salleng1, M Scott2 and post-collection hemostasis procedures have progressed to 1Division of Animal Care, 2Molecular Physiology & Biophysics, continually improve to what are considered best practices from an efficiency and animal welfare aspect. Today, the techniques Vanderbilt University Medical Center, Nashville, TN used for these procedures with marmosets are proven to be suc- Obesity and diabetes research models depend on monitoring cessful; the result of many years of working with this species are many factors, such as weight maintenance, daily food consump- presented in this poster. Researchers will find these best practices tion, and glucose and insulin measurement. This has historically for injection and bleeding to be helpful in their work with the resulted in the single housing of canines. Long-term compatible common marmoset. social housing has been shown to be beneficial to the physical and psychological well-being of laboratory canines and further P55 Quantitative Monitoring for Laboratory Animal Aller- led to a reduction in the incidence of stereotypical behaviors. gens The aim of this study was to determine if social housing initi- ated mid-study would affect experimental variables of weight, C Zumbaugh*, C Davis food consumption, stereotypic behavior expression, oral glucose Eli Lilly and Company, Indianapolis, IN tolerance test (OGTT), and fasting plasma insulin. Three adult Managing potential exposures to laboratory animal allergens male hound dogs (Class B) that were part of a study on the ef- (LAA) is vital for maintaining a healthy work environment. fects of a chronic high-fat diet and pancreatic insufficiency of Quantitative air monitoring studies can be used to characterize glucose metabolism were tested for compatibility. Dogs were employee exposure to LAA and assign effective control measures. socially housed in the mornings after food bowls were pulled A series of studies were conducted within a large research facil- and then separated in the afternoon to be fed. Two of the dogs ity to investigate the airborne levels of LAA, specifically mouse were not compatible so 1 of the dogs alternated being paired with urinary protein (MUP) and rat urinary protein (RUP). A new the other two dogs daily. After 12 mo of pair housing, 2 of the 3 analytical technique, multiplex array technology, was established dogs had gained weight (+2.04 kg and +1.66 kg), and the third to analyze the samples. After the successful completion of a pilot had maintained its weight. Food consumption 60 d before and study, a comprehensive monitoring strategy was implemented to 60 d after pairing showed similar results, with 2 of the 3 dogs collect exposure data for common rodent handling tasks. More increasing food consumption post-pairing (+5% and +12.5%) than 150 personal air samples and over 200 area samples were and 1 maintaining. The dog that maintained its weight and food collected. The resulting data was categorized into exposure control consumption was the control dog, so the results were as expected. bands of high (>50 ng/m3), medium (5 to 50 ng/m3), and low (<5 One of the 3 dogs had exhibited an atypical behavior prior to pair- ng/m3) based upon relative risk. The exposure control banding ing, as evidenced by a lick granuloma. Following social housing, system provided a data-driven framework for prioritization of no atypical behaviors were noted and the granuloma resolved activities intended to minimize employee exposure potential. within 31 d of pairing. OGTT and fasting plasma insulin results The quantitative data illustrated that tasks involving significant were within expected range and do not appear to have been af- bedding disturbance, such as cage changing and bedding disposal fected by the pair housing. Compatible pair housing of captive without local exhaust ventilation (LEV), were categorized within canines optimizes animal welfare by reducing stress levels as the high-exposure band. The disposal of bedding with LEV re- shown through weight maintenance, daily food consumption, vealed data that placed the task in the medium-exposure band, decreased presence of abnormal behaviors, and no assignable which confirmed the importance of source ventilation. All of the negative effects on research, thereby refining and reducing the area sample results taken in the hallways and corridors were below number of dogs needed for research. the limit of detection. These results assisted in the verification of our building ventilation capability. Current control measures, P54 Blood Collection and Intravenous Injection in the Common such as respiratory protection and ventilation containment, were Marmoset (Callithrix jacchus) evaluated and adjusted to align with the exposure band require- C Maurus*, M Simkins, A Lange ments. Now that tasks can be banded according to the associated Abstracts of scientific papers 2008 AALAS National Meeting airborne concentration of LAA, future metrics around employee manufacture date. During the 5-mo assessment, approximately sensitization can be cross-referenced in order to evaluate the ef- 17,000 valves were assessed, of which, 2,300 failed (6.1% low-flow, fectiveness of the control strategy for the reduction of LAA cases 1.4% no-flow, 3.5% leaky). The manufacturer’s target total failure in vivarium staff. rate is 0.1%. Working together with the vendor, we determined that flow rate was impacted by the total number of autoclave cycles. P56 Colors of Communication in the Veterinary Care Manage- Failures were impacted by several factors, including residual metal ment of a Large Rodent Population shavings, silicone flashing, introduced debris or bedding and, most commonly, a distortion of the autoclave-rated internal diaphragm. L Schleenbaker, DI Del Castillo*, HE Ramirez It was determined that the combination of heat, pressure, and Animal Care Services, University of Florida, Gainesville, FL steam caused the diaphragm to become water-permeable and led to formation of a blister. Normal flow rate often returned following Veterinary medical management of a rodent population in a a period of drying out, only to reabsorb water at the next autoclave decentralized facility requires organization and an effective cycle. We also established a process for rebuilding valves with the method of communication between research, husbandry, and 3 components susceptible to wear (shield, diaphragm, and O-ring). veterinary staff. The proper identification of clinically sick To aid with future tracking of valve age, the manufacturer has cre- and postsurgical animals, in addition to the documentation of ated an annual color scheme for the valve shield. A preventative husbandry procedures performed by investigators, is of utmost program for water valve maintenance should include autoclave importance in providing quality veterinary care for a larger ro- recalibration, prescreening of valves, and continual assessment all dent population. Hence, the veterinary care staff has developed floods and clinical dehydration cases. a silent communication system using colored cards and symbols as the basic communication language. The cards are used by husbandry, research, and veterinary staff to maintain an efficient P58 Advancing and Refining Animal Welfare within an Estab- flow of information. The following cards are primarily used in lished Group-housed Primate Facility our facility to document veterinary related concerns. A yellow D Philip*, WJ Archibald, M Zubiaurre “sick animal” card is used by the husbandry and research staff Preclinical Technologies, Aptuit Ltd, Edinburgh, United King- to identify newly observed sick animals in the colony. A pink dom “under veterinary care” card replaces the yellow card when the veterinary care staff assess the animal and formulate a plan Nonhuman primates have been used in drug metabolism, pharma- of action. This pink “under veterinary care” card acts as the cology, and regulatory toxicology studies for many years. In 1992, animal’s clinical record, with all observations, treatments, and we opened a new facility, specifically designed to meet the objec- outcome recorded by either research or veterinary staff and is tives of these studies, while housing the animals in social groups to removed after resolution of the problem. A peel-off “surgery” provide for their physiological and behavioral needs. Some of the sticker is used to identify surgically manipulated animals and challenges we faced were ensuring the quality and integrity of the to indicate a postoperative check has been performed. A purple study data, avoiding cross-contamination, and positively identify- “under PI care” card identifies those animals in which some or all ing individuals when in a gang cage. Simultaneously, we wanted to husbandry procedures (food, water, cage change) are performed further improve the animals’ environment while modifying study by the research staff. In addition, other colored identification designs to take advantage of the spacious living accommodation. We and notification cards are used for identifying overcrowded group-house all primates in large gang cages incorporating perches, cages and notification of dead animals. As a result, the colors of swings, puzzles, and toys. Deep litter substrate allows foraging for documentation have made animal health–related communication food, fruit, and supplementary mixes. Single cages are accessed from among research, husbandry, and veterinary staff more efficient, the gang cage by way of a hatch to allow precise post-dose observa- therefore dramatically improving the level of veterinary care tions. We have added acrylic sheet dividers and verandas between provided in our rodent population. cages, enabling enhanced visual contact between animals. Colored dyes applied on the animals’ eyebrows enable visual identification P57 Watering Valve Life Cycle Evaluation when socially housed, with minimal disturbance to the animals. Novel methods of environmental enrichment, including swim- DM Gonzalez*, SJ Gibson, M Leblanc, J Reuter ming pools, have also been introduced over time. Test compound Animal Resources Department, The Salk Institute, La Jolla, CA cross-contamination is avoided by careful physical separation of dose groups. Study procedures have been refined and adapted to Automated watering systems provide a reliable source of ad libitum complement social housing, including use of vascular access ports water to animal cages. Our facility uses reverse osmosis automated for infusion studies, enabling animals to remain in pairs or groups watering for 95% of mice (approximately 14,000 cages). Racks and throughout dosing. We have found that we can meet the needs of cages were brought online in 2002 and featured a quick-release a range of studies while housing animals in social groups. Animals water valve that allows its removal for easy sanitation. Valves were are generally calm and cooperative during procedures, thereby routinely sanitized monthly through a tunnel washer (alkaline de- contributing to reliable, quality study data. Later additions such as tergent and a 180 °F rinse) and autoclaved at 250 °F for 15 min. In verandas and swimming pools have proved successful. This poster the fall of 2007, we observed an increase in clinical dehydration in discusses our conclusion that new refinements can be incorporated a subset of the mouse population, which prompted an investigation into existing facilities, further enhancing good environmental en- of the water delivery system. Two major contributing factors were richment and welfare. indentified: phenotype and decreased water flow. We sought further guidance from the vendor and implemented an assessment plan for measuring water output. Using manufacturer’s specifications for a P59 Evaluation of Disposable Rodent Cage Inserts water flow rate of 25 to 50 ml/min, we prescreened all valves. Leaky DM Hull* flows were identified by continual release of water after toggling. Research Animal Resources, Johns Hopkins University, Balti- Low-flow, no-flow, and leaky valves were removed from circula- tion. Unfortunately, no methods were available to identify the valve more, MD Abstracts of scientific papers 2008 AALAS National Meeting Every animal facility develops occasional malfunctions of its and contacting references and past employers can be enlightening, cage wash equipment. This equipment can be out of service but often only represents the aspects that the candidate wants the anywhere from hours to weeks, depending on the severity of potential employer to see. Interviewing the candidate can also be the problem. A product has been developed that allows for an helpful, but it is sometimes difficult to evaluate responses and appropriate intracage environment to be maintained when cage predict how the candidate will fit with the existing team. For 3 y, wash equipment is broken for long periods of time and there is we have been using teams and performance-based interviewing to a need for alternative cage changing methods. Trials were con- successfully screen potential candidates. This method of interview- ducted in 5 ventilated mice cages. The same procedures were ing involves asking the candidates to respond to a predetermined followed for each cage change. These procedures consisted of panel of questions selected to ask them to describe actions they having the mice transferred into the disposable cage inserts, then took with past events in their lives that meet certain criteria. having the insert applied to the home cage, with care taken to For example, “Tell me about a time when you had to work with ensure proper alignment and exposure of the grommet supplying someone you did not like. How did you handle the situation?” air and water. The same home cage was used for 10 wk, during The panel includes questions that require the candidates to share which time an insert was changed every other week. These cages positive and negative memories and experiences, with the expec- had a variety of different numbers and strains of mice. During tation that past behavior is predictive of future behavior. Our use this 10-wk interval, these cages were evaluated for ease of use, of this interview technique has allowed us to identify individuals effects on food, bedding, grommet and wire bars, as well as the who share our commitment to teamwork and customer service, condition of the mice. By the end of the 10-wk test, we found decreasing staffing turnover and increasing job satisfaction. that the cage insert was easy to use and had no adverse affect on the bedding, food, or health of the mice. These inserts were an P62 Comparison of Costs Associated with the Preparation of effective and convenient way to keep mice in a clean environ- Nutritional Support to Multiple Rodents ment until clean autoclaved cages are available. DL Hickman* P60 A Novel Biocontainment Signage System Veterinary Medical Unit, VA Medical Ctr, Portland, OR DM Mook*1,2, W Thompson1, MJ Huerkamp1,2 Providing nutritional support to debilitated rodents is instrumental 1Division of Animal Resources, 2Department of Pathology and in minimizing pain and distress. Provision of nutritional support can be in the form of oral gavage, moistening of rodent block chow, or Laboratory Medicine, Emory University, Atlanta, GA through the provision of commercial soft diets that require minimal The use of biohazardous agents in research increased sharply to no preparation. In this study, we evaluated the labor and material in the past few years, as indicated in our facility by a tripling of costs associated with the provision of moistened rodent block chow, the number of persons oriented to our biocontainment program powdered diet that is reconstituted with water, and a preformulated between fiscal year 2006–2007 and 2007–2008. As is typically nutritional supplement. Technicians recorded the time required to the case in laboratory animal facilities, this increase in use was prepare and provide 1 of these 3 diets in the facility animal rooms. associated with no concurrent increase in space. In a multiple The time per serving was estimated by dividing the total time by the investigator room, the possibility existed for several investiga- number of cages serviced. Four animal rooms, averaging 20 cages tors to use various agents, and for these agents to be in use at of animals requiring nutritional support per room, were included different times. Users of the biocontainment suite had the right in the study, and time was recorded over 31 d. The mean time to be appraised of the specific risks associated with their work required to supply moistened rodent block chow was 1.25 min/ environment, and this function was accomplished by placing cage; the mean time required to supply reconstituted powdered signage on animal room doors listing the agents used therein. diet was 1.96 min/cage; and the mean time required to supply With heavy usage, it became increasingly difficult for the animal the preformulated diet was 0.49 min/cage. With an hourly labor resources program to keep signage up to date. We created a novel charge of $27/h and a cost of $0.08/disposable petri dish (required magnetic biohazard signage system in which users changed the for the moistened rodent block chow and reconstituted powdered sign themselves as they added or deleted agents. A laminated diet, but optional for the preformulated diet), we concluded that biohazard sign was affixed to a magnetic backing, and another the preformulated diet was the least expensive option overall, with sign, identified as “approved agents,” was also created. Remov- a per cage cost of $0.54 per cage. The per cage costs for moistened able magnetic labels were generated with investigator names rodent block chow and reconstituted powdered diet were $0.66 and and agent names. When using an agent in a room, laboratory $1.13, respectively. This cost differential was primarily due to the staff retrieved the agent and PI name from the approved agents fact that the preformulated diet did not require daily refreshment. sign and affixed it to the biohazard sign on the door. Staff then As our facility provides nutritional supplementation for an average removed the magnet when they were finished using the agent. of 200 cages per d due to experimental manipulation, selecting the The signage system was easy to use and intuitive, was well- preformulated diet over the moistened rodent block chow or the received by both laboratory and animal care staff, and received a reconstituted powdered diet represents an annual cost savings of specific commendation from the Association for the Assessment $8,760 and $43,070, respectively. Additionally, as the use of a petri and Accreditation of Laboratory Animal Care, International. This dish is not required, less waste is produced by the animal facility. poster will share the signs themselves with our community, and describe their rationale and use. P63 Mock IACUC Meeting: A Novel Approach to Animal Care Staff Training P61 Use of Performance-based Interviewing DM Ostmeyer*1, L Martin2, M Rosenbaum3,4 DL Hickman* 1Officeof the Vice President for Research, 2Research Integrity Veterinary Medical Unit, VA Medical Ctr, Portland, OR and Compliance Review Office, 3Microbiology, Immunology and Pathology, 4Laboratory Animal Resources, Colorado State Selection of employees who are a good fit with your animal care University, Fort Collins, CO and use program can be quite challenging. Review of applications Abstracts of scientific papers 2008 AALAS National Meeting Animal care technicians are an integral part of the dynamic re- are hairless, their eyes and ears are closed, the hind limbs and search team. As primary caretakers, they must be knowledgeable tail are stumps, and bone development is extremely immature. regarding animal welfare regulations and governing agencies Hind limbs begin to form between d 9 to 12.The ears opened including the Institutional Animal Care and Use Committee between d 12 and 15, and hair appears on the body around d (IACUC). Technicians often receive classroom instruction about 18. The eyes begin to open on d 33. There was little variance in the committee while preparing to take certification exams, but the rate of development between opossum pups within a litter they rarely have the opportunity to gain firsthand knowledge or among litters. Additionally, male and female opossum pups by interacting with the committee. A mock IACUC meeting do not appear to vary greatly in size between the ages of 0 and was designed to give the animal care staff at our institution an 90 d. We believe that these images will provide a useful guide interactive, first-hand experience. Numerous members of the for staging developing M. domestica pups. animal research community enthusiastically volunteered to take part in this exercise. The faculty for the mock IACUC training P65 Measurement of Isoflurane Waste Anesthetic Gas Concen- included the committee chair, 3 additional committee members, 2 tration When Using the Open-drop Technique comparative medicine resident veterinarians, the senior commit- tee coordinator, and the animal program coordinator. The mock DK Taylor* meeting was designed as a 2-part activity. The first part included Division of Animal Resources, Emory University, Atlanta, GA a presentation about the committee’s make-up and function, The open-drop technique is a commonly used method of an- distribution of mock animal use protocols, and solicitation of esthetic delivery to small rodents. Operator exposure to waste staff volunteers to serve as “student” committee members. After anesthetic gas (WAG) is a potential occupational hazard if this the presentation, a mock committee of 3 technician volunteers method is used without a control such as a chemical fume hood. and 7 “faculty” committee members were seated at a confer- This study was designed to determine if using the open-drop ence table, with the remaining technicians in the audience. The technique on a surface without exposure controls generates committee coordinator and chair each gave brief explanations significant WAG concentrations. Liquid isoflurane (0.1, 0.2, or about their respective roles, and then deliberation began. The 0.3 ml) was placed into screw-top 500-ml (3.5 in. tall) or 1000-ml committee generated a great deal of discussion about the mock (6.5 in. tall) glass containers. WAG concentration was measured protocols, and the audience interjected with several comments 1, 2, or 3 min after liquid placement at the vessel opening and and questions, resulting in a highly interactive and educational at 8 and 16 in. from the opening (operator height) using a por- experience. The mock meeting also created an atmosphere that table infrared gas analyzer. A 2-factor ANOVA showed that, at fostered positive relationships between the committee members the vessel opening, time was only significant factor (P < 0.0001) and the animal care technicians. Positive feedback was received impacting WAG concentration when the 500-ml jar was used, from both the students and the faculty after this training event. while neither time nor liquid volume were significant factors A course evaluation was completed by the technicians in atten- when the 1000-ml jar was used. The WAG concentration using dance, and they made excellent suggestions for improvement the 500-ml and 1000-ml containers were compared using the which will be implemented at future mock IACUC meetings. Student t-test. At all liquid volumes and time points, the WAG concentration using the 500 ml container was marginally (P = P64 Photographic Staging Guide for the Normal Development 0.057) to significantly (P < 0.05) greater than that for the 1000 ml of Short-tailed Opossums (Monodelphis domestica) container. At operator level, WAG concentrations were at or near D Paulus*1, SJ Karlen2, L Krubitzer1,2, CJ Clayton1 0. Importantly, WAG concentrations at the vessel opening were 1Department of Psychology, 2Center for Neuroscience, University transiently as high as 662 (+168) ppm using the 500-ml jar and 122 (+87) ppm using the 1000-ml jar, much greater than the NIOSH of California, Davis, CA 60-min recommended exposure level of 2 ppm; concentrations The grey short-tailed opossum (Monodelphis domestica) is a com- were well below this ceiling at operator level. These results show mon research model because it is one of the few marsupials that that operator exposure to WAG using an open-drop technique is can be kept and bred relatively easily in captivity. These opos- minimal and below levels considered hazardous; however, the sums have been used for a variety of studies, including spinal risk of significant exposure increases as the distance between cord and brain development, spinal cord injury and repair, skin the operator and the jar diminishes. It also remains important and eye cancer, genetics, olfactory communication and spatial to properly handle the liquid agent and avoid contact with skin navigation. M. domestica is a pouchless marsupial whose young or mucous membranes. are born after a 14- to 15-d gestation period. The pups migrate to the nipples and attach for approximately 2 wk, and the pups P66 Light Dark Studies Meeting the Demands are weaned from the mother at 8 wk of age. The purpose of this study was to develop a staging guide for opossums to use for EJ Delaney*1, KL Krueger1, M Thompson2 developmental studies and animal husbandry. Photographs 1Animal Research Children’s Hospital, Children’s Hospital Bos- were taken of the litters from 6 female opossums. Litters were ton, Boston, MA; 2Plastic Concepts, INC., Billerica, MA of varying size and date of birth. Imaging was done every 3rd Neuroscientists at our institution are evaluating the developing d from d 0 (day of birth) to 30, every day from d 30 to 35 (the brain and how it is affected by environmental changes, such as time of eye opening), every 5th d from d 35 to 60 (the age of alteration in light cycle. While it would be simplest to alter the weaning), and on d 90 (prepuberty). From d 0 to 21, isofluorine light cycle of an animal room, the small number of animals needed was used to anesthetize the mother so that the pups could be for these studies was a problem where space is at a premium in imaged. After d 21, pups were large enough to remove from the our facility. In addition, the coordination of altered light cycles mother, so one individual pup was selected at random for imag- between several protocols was a concern. It was determined the ing. All photographs were adjusted for brightness and contrast best solution would be to design a chamber which was light-tight, and are presented in chronological order. We found that pups in which light cycles could be determined by the investigator, and changed dramatically between d 0 and d 90. At birth, the animals that would fit up to 8 small mouse cages. A plastics manufacturing Abstracts of scientific papers 2008 AALAS National Meeting firm was identified, and together with animal resources manage- Instituto de Ciencias Básicas y Medicina Experimental Hospital ment staff and the investigators, a design was developed. Two Italiano de Buenos Aires, Ciudad Autónoma de Buenos Aires, chambers per unit were created, each holding approximately 8 Argentina small mouse cages or 6 large mouse cages. To ensure complete Exposure and training to laboratory animal science is scarce darkness of the interior enclosure, the doors had an overlapping for most medical students and future researchers. One possible gasket seal design. Small air blowers in the back of the unit pro- reason for this is that in most biomedical careers prior training is vided ambient air to the chambers. Temperature and humidity not obligatory when beginning laboratory work. Our facility re- probes were installed, with digital readout displays to ensure quires that people who use animals in their projects first undergo appropriate environmental parameters were maintained for the training with basic contents. We designed and implemented a animals. Air supply and return, along with the electrical con- program with the purpose of enhancing the adequate care and nections, were designed with a double-wall unit to prevent any use of laboratory animals and thus the achievement of quality reflective light from entering the interior chamber. It was critical and reliability in the expected results. It consists of an optional that these animals experience no exposure to light, and so a very 36-h course for 2nd-y students called Science of the Laboratory sensitive light detection device (4 lumis of light) was placed inside Animal. Our aim is to sensitize the student to the harmonious the chambers to alert users of any internal lighting by way of both care and use of laboratory animals, with a special emphasis on a visual and audible alarm. A timer was installed in each chamber the welfare of the animal in regards to biomedical activity. The for properly determining the duration cycle of the study. Finally, main idea is to educate implementing the principle of the 3 Rs of the units were designed to keep outside noise to a minimum, Russell and Burch. Teachers evaluated the knowledge acquired which is critical for most neuroscience studies. These units have with partial and final exams and a monograph. Through observa- now been in place, and in constant use, for more than 6 mo. The tion, teachers also evaluated the practical abilities and attitudes design ensures the animals are not exposed to even small amounts towards animals acquired by students. We conclude that the of light. Investigators are extremely pleased with these units; they proposed objectives were satisfactorily acquired since the results are easy to open and close even in total darkness, and can accom- showed that students understand the importance of a previous modate multiple cages, allowing investigators to perform entire training, as much for themselves as for future collaborators. experiments under the same environmental conditions. P69 Murine Retro-orbital Bleeding Under Anesthesia: An P67 Single or Social Housing for Research Animals Alternative Method E Scott*, K Ryder, DB Anderson R Scott, E Villalobos, A Gaudiel, R Garcia-Gonzalez, E Chua* Animals Scientific Procedures Inspectorate, Home Office, Laboratory Animal Resources, Genentech, Inc., South San Fran- Dundee, United Kingdom cisco, CA Social housing is promulgated in virtually all legislation or guide- This poster will outline the procedure as an alternative to using lines for species used in scientific work. Traditional approaches glass capillary tube or anesthesia chamber retro-orbital bleeding using single housing should be discouraged where social housing methods. Sharper glass capillary tubes have been replaced with is advantageous and improves animal welfare, yet is not detrimen- sterilized plastic pipette tips. Anesthesia chamber use has been tal to the scientific research outcomes. In the United Kingdom, replaced by use of sterile, flip-top plastic vials for induction by inspectors play a role in encouraging group housing of animals open-drop method. The animals are also securely restrained by and ensuring that a clear scientific, veterinary, or husbandry case is the procedure operator at all times, minimizing head movement made by researchers for single housing of social animals. There are of the animal during anesthesia induction, which can cause examples where single housing may be justified, but social housing trauma. All materials in this procedure can be sterilized, which options need to be explored, as they are often possible and can be helps prevent cross-contamination between animals. In our use, beneficial. In neuroscience, when chronic implants are used, there this method has also aided in providing faster, more successful is concern that interaction between animals may lead to infection training of general animal users, because of the ease of restraint, or implant disturbance. In the immediate postoperative period anesthesia induction, and avoidance of sharp objects in the ocular single housing may be a good option, but thereafter experience area of the animals. This method works well in larger murine in the United Kingdom shows that social housing is possible with facilities, while minimizing animal stress, trauma, and time spent reduced incidence of stereotypic behavior. Telemetry animals can by the procedure operator. There has also been a significant de- be housed singly for short monitoring periods and then restored crease in hemolyzed or clotted samples due to the more conical to their groups without disruption of social hierarchies. When shape of the pipette tip in comparison to a capillary tube. severe illness is expected with bacterial challenge models it may be best to single house during the acute phase only, to allow close and frequent monitoring. Such periods are likely to be short P70 eCertificate: A Fully Automated Web-based Health Certi- (hours or days) and the animal is likely to be sufficiently ill that fication System for Rodents interaction with peers would be unlikely. The measurement of FR Wolf*1,2, Q Zhang2 individual feed intake or monitoring postdosing may necessitate 1Weill Cornell Medical College, New York, NY; 2 Memorial Sloan- single housing. Husbandry measures of short-term separation at Kettering Cancer Center, New York, NY feeding and dosing times, feeding systems with individual animal recognition systems, or alternative measures of food intake, such Sharing of unique genetically modified rodents among scientists as body weight, have been used to reduce social isolation time is important in biomedical research. To properly assess the risks with no impact on study data acceptability. associated with the receipt of the animals at the receiving facility, it is essential to receive a comprehensive health report from the P68 Laboratory Animal Science Education for Medical Stu- facility of origin. To facilitate this process, we developed eCertifi- dents cate, a web-based application that creates and distributes health certificates within seconds based on user input and preloaded E Herrero*, G Dabrowski, PF Argibay Abstracts of scientific papers 2008 AALAS National Meeting data. Each eCertificate consists of a PDF file with 4 pages. A cover the innate behaviors of burrowing, tunneling, chamber grouping, page provides general information as well as a summary of the and a tower for them to see around their “town.” The configura- colony’s current and historical health status, including counter- tion of tunnels, burrows, and towers can be altered to fit any size measures taken in cases of pathogen detection. The second page, room and almost any number of prairie dogs. The hope was that a pro forma invoice and health certificate, is used by customs the prairie dogs would use the burrows for different purposes officials when shipping internationally. The third page details the (eating, sleeping, defecating). The tower will provide them the op- health monitoring program and husbandry procedures used in portunity to climb above their town and perhaps give “warning” the room and facility of origin. The final page is a FELASA-style or “all clear” vocalizations. We constructed the “den” from 100-gal health report providing current as well as historic results for each livestock troughs. Each trough is fitted with 4 6-in. wheels to allow pathogen tested. eCertificate incorporates a number of features. for easier sanitation and reconfigurations. The troughs are con- Cover letters and health monitoring profiles can be custom- nected to each other using CPVC unthreaded 6-in. diameter pipe tailored to individual rooms, to groups of rooms, or to a specific that serves as the tunnels. The pipe is attached to the troughs using vivarium. Investigators have access to previous eCertificates and CPVC unthreaded 6-in. diameter pipe fitting. The tower of each can resend the certificate with current results with a single click. CHEU is a cat gang-housed cage and is attached to the CPVC pipe A search function allows the biosecurity veterinarian to find all with CPVC unthreaded 6-in. pipe fitting without hex in center and certificates issued for animals shipped from a specific room, CPVC unthreaded 6-in. pipe fitting, 90 degree elbow. All exits from facility, investigator, and so forth within a specified time frame, the dens can be capped to prevent movement of prairie dogs. The allowing a follow-up e-mail to be sent to selected recipients if a troughs are covered with 48 by 96 by 0.25 in. polypropylene perfo- change in health status is detected. The next release will collect rated sheets which are 20% open area to allow air flow and light into test results from a laboratory information system automatically the troughs. It was determined that PVC would not withstand cage updating the health report in eCertificate. wash temperature, and the prototype was redesigned with CPVC to withstand temperatures of 180 ° F. New shipments of prairie dogs P71 How Safe Is Your Facility? are scheduled to arrive within the next few months. These will be the first groups ever housed in the new CHEU. F Warren* Lab Animal Medicine, University of Delaware, Newark, DE P73 Animal Enrichment and Socialization in an Indoor Re- Animal facilities have the advantage of providing everything that search Facility an animal needs, which includes food, water, sanitation, air, and G Jeans*1, KD Corcoran1, JL Wheatley2, AS Fix2 lighting. The disadvantage of providing lighting is that animal 1Pharmaceuticals, 2Central Product Safety, Procter & Gamble, facilities are constructed with no windows that allow natural Cincinnati, OH light to enter. If there was a power failure, an animal facility can be an extremely dark place. Navigating around a dark facility Environmental enrichment is an important element of any animal can be not only intimidating, but dangerous. Large equipment research program. The goals of the Procter & Gamble program would become hazardous if personnel were trying to find their include providing comfort and decreasing stress for the animal, way out. In our facility, we have instituted many safety features promoting species-specific behaviors, creating a more complex to be used in the event of a power failure. Placing flashlights in living environment, and enhancing animal welfare standards. All corridors is one way. We have also found a simple, inexpensive animals are provided enrichment unless exempted due to health method of providing an extra measure of safety. All rooms and reasons or scientific justification approved by the IACUC. Environ- support areas have luminescent materials on the door frames. mental enrichment items have been chosen for each species based Next to the doorframe is a pen-style flashlight that can help on actual observations of animal interactions with the items. Study guide someone out of an animal room. We also put in place other personnel choose species-specific enrichment items, which are safety features, including large battery-operated flashlights in the rotated to provide stimulation and variety. For example, rodents office area and hallway maps showing exit doors. The hallway are provided a variety of enrichment items (nestlets, isopads, huts, flashlights have automatic night lights built in that light when igloos, gumabones, nylabones, tunnels, marbles, stainless steel power is off. Another safety feature of the hallway flashlights bells) on a rotating basis. Dog enrichment includes a variety of toys, is that they make a beeping sound after 3 min when power has treats, and various socialization activities (grooming, brushing, been cut off, which aids in locating them. nail trimming, person-to-dog and dog-to-dog socialization, and clicker training). In addition, we provide a large indoor play area P72 Cynomys Housing and Enrichment Units using child-based playground structures and setup a mock living room using common household furnishings. Dogs quickly learn to G Langham*1, K Kelly1, C Walls2, N Powell1 anticipate their socialization experience and these experiences have 1Animal Resources Branch, Centers for Disease Control and greatly facilitated the adoption of dogs from our research programs. Prevention, Atlanta, GA; 2 Four Seasons Environmental, Inc, In addition, these rooms have become a center of activity for the Monroe, OH animal care and veterinary staff, providing an additional positive benefit to facility personnel. Based on our experiences, a broadly The use of prairie dogs (Cynomys spp.) in infectious disease research applied environmental enrichment program across all species im- has increased due to their susceptibility to monkeypox, bubonic proves the welfare of the research animals, the quality of the research plague, and hepatitis. Cynomys have a complex social system and environment, and job satisfaction for employees. housing has posed a difficult challenge. Other facilities have tried housing them in storage bins or on the floor. We were dissatisfied with these housing methods, and decided to construct a Cyonmys P74 Enhanced Indoor Housing for Research Dogs housing and enrichment unit (CHEU) that is easily sanitizable, GL Jeans*1, DA Fryer1, KW Casey1, CR Bohne1, GL Calihan1, mobile, and promotes as many species typical behaviors as possible AS Fix2 in a captive environment. A group housing scheme was developed 1Pharmaceuticals, 2Pet that provides quarantined prairie dogs the opportunity to exhibit Care, Procter & Gamble, Lewisburg, OH Abstracts of scientific papers 2008 AALAS National Meeting Enhanced indoor housing for our research dogs was a key interest shows that allergen exposure can be drastically reduced by use among our personnel and stakeholders. In response, we bench- of ventilated caging, by handling the cages always with filter top marked with other research institutions and collected input from on, and by changing cages only under laminar flow conditions. our staff to determine how best to optimize our kennels and meet In the cage wash area, a 32-fold reduction was achieved using an our animal welfare goals. The goals were to improve our existing inexpensive dust extractor. Further studies will be necessary to kennel wards; enhance our animal welfare standards; provide op- elucidate whether this allergen reduction is sufficient to prevent portunities for the dogs to be group-housed, yet maintain ability mouse-related allergic sensitizations. A striking result was that to be periodically single-housed; create a more visually appealing relevant Mus m 1 levels were detectable even at an indirectly and open kennel environment; and promote increased dog-to-dog exposed work area like a PC work desk. and human-to-dog interaction. After researching kennel options with various vendors, we decided upon a modification of our P76 Quantitative Culture and ATP Detection Used to Validate existing design. The new panels were shorter and included side- Rodent Water Bottle Change Intervals access gates, visual barriers, and raised benches. Additionally, we added sound baffling panels to the wards, which dramatically H Tobin*, NH Shomer reduced the reverberant noise levels. After installation of the Merck Research Laboratories—Boston, Boston, MA modified kennels, the dogs quickly adapted to the new design Changing water bottles is labor intensive, poses an ergonomic and are now predominantly pair-housed. The combination of hazard for husbandry staff, and increases biosafety risks to the the new kennel design and installed sound baffling has resulted mice as it requires handling the bottles more frequently. We in quieter dogs who appear more content and no longer exhibit investigated the feasibility of extending the water bottle change undesirable behaviors. The dogs all use their raised platforms and interval from 7 to 14 d for singly housed, specific pathogen-free can easily see what is happening throughout the ward because of mice. We hypothesized that water quality in 14-d-old water the increased visibility. The flexibility of the side access gates allow bottles from singly housed (SH) cages would be at least as good us to house dogs based on compatibility and research study needs. as the quality in 7-d-old cages from group-housed (2 to 5 per In addition, shorter kennel walls invite human-to-dog interaction. cage) mice. Based on the current accepted 7-d change interval for In our experience, these redesigned kennels met our research group-housed (GH) mice, we set the GH contamination level at 7 needs, provided optimal housing for our dogs, and increased job d as our acceptance criterion. Water in our facility was autoclaved satisfaction for employees. municipal tap water with undetectable levels of free chlorine after autoclaving. Mice tested were free of pathogens and common P75 Effective Strategies to Diminish Critical Allergen Exposure opportunistic pathogens, including Pseudomonas. Water bottles while Handling Laboratory Mice of GH and SH mice were tested at predetermined intervals by H Riedesel*2, J Krause1, E Hallier3, S Turowski3, A Heutelbeck3 aseptically removing a 1-ml water sample, vortexing, and testing 1Animal Facility, Max-Planck-Institute of Experimental Medicine, using a luminometer to estimate organic contamination levels. Quantitative cultures were performed using nutrient agar plates Goettingen, Germany; 2Animal Facility, Helmholtz-Centre for incubated for 3 d at 37 °C. At 24 h postplacement, water bottles Infection Research, Braunschweig, Germany; 3Occupational from SH and GH mice averaged 95 ± 210 colony-forming units Medicine, Georg-August-University, Goettingen, Germany (CFU)/ml (n = 19) and 489 ± 1778 CFU/ml (n = 14), respectively. Allergy caused by laboratory mice is a common occupational At 7 d, SH and GH mice averaged 35518 ± 152843 CFU/ml (n = health hazard for personnel. We evaluated the exposure of mouse 19) and 96412 ± 241625 CFU/ml (n = 14), respectively. At 14 d, allergens in the air under different working conditions when SH mice averaged 126320 ± 257137 CFU/ml (n = 19). Average handling mice. With on-the-person and stationary air sampling luminometer counts for SH and GH mice at 24 h postplacement methods (flow: 2.5 to 3 l/min, 8 µm cellulose nitrate filter), inhal- were 1 ± 0 (n = 19) and 62 ± 230 (n = 14), respectively. At 7 d, SH able and respirable dust was collected over several day shifts and and GH mice had counts of 992 ± 2020 (n = 19) and 12612 ± 22870 during night in animal rooms, cage wash, and areas with a PC (n = 14), respectively, and at d 14 SH mice had an average count work desk. Work areas with different conditions were included, of 11639 ± 35127 (n = 19). Plate counts and organic contamina- such as open cages compared with individually ventilated cages tion levels at 14 d for SH mice were not significantly different (with positive or negative pressure) and technical equipment (P = 0.73 and 0.92, respectively) from our acceptance criterion (changing cages with or without a laminar-flow hood). After (contamination levels in GH mice at d). Based on this data, we extraction of the dust samples the level of mouse allergen Mus extended our water bottle change interval for SH mice to 14 d. m 1 was determined using a commercially available ELISA. In the laboratory rooms with individually ventilated cages, over P77 Work-related Mouse Allergens in the Home Environment the measuring time mean levels of Mus m 1 were detectable be- of Animal Technicians tween 0.55 ng/m3 (positive pressure) and 0.31 ng/m3 (negative pressure), in rooms with open cages up to 12.97 ng/m3. Without J Krause*1, H Riedesel2, E Hallier3, S Turowski3, A Heutelbeck3 using a laminar flow hood for cage change, Mus m 1 levels up to 1Animal Facility, Max-Planck-Institute of Experimental Medicine, 17.55 ng/m3 were measured compared to 0.67 ng/m3 when using Goettingen, Germany; 2Animal Facility, Helmholtz-Centre for a laminar flow hood. Higher allergen levels were detected in the Infection Research, Braunschweig, Germany; 3Occupational cage wash area near the disposal station for soiled bedding. The Medicine, Georg-August-University, Goettingen, Germany air here showed 84.93 ng/m3 Mus m 1. By installing a commercial Laboratory animal allergy is a common occupational health dust extractor, levels of Mus m 1 could be lowered to 2.59 ng/ hazard for research scientists and animal technicians. Risk fac- m3. Levels up to 7.16 ng/m3 were detectable on indirect exposed tors include genetic and environmental components. The aim of work areas, such as a PC work desk. Under most working con- this study was to evaluate the airborne mobility characteristics ditions, relevant mouse allergen levels were detectable. Up to of mice allergens and their transmission into the homes of ani- 20 times higher Mus m 1 level could be found in animal rooms mal technicians. Dust samples were vacuumed from car seats, with open cages compared to rooms with ventilated caging. This carpets, kitchen chairs, and mattresses of animal technicians’ Abstracts of scientific papers 2008 AALAS National Meeting private dwellings. Eight technicians working in 4 workplaces P79 Creating a Safer Environment for Laboratory Animal Han- of different allergen exposure levels (specific pathogen-free dlers Anesthetizing Rodents in Induction Chambers [SPF] and conventional rooms equipped with open compared JB Hoff* with individually ventilated cages) participated in this study. Depending on the housing unit different types of personal pro- Center for Intgrative Genomics, University of Michigan, Ann tective equipment was used, including sterile gowns with or Arbor, MI without face mask, hair cover, and gloves. Technicians working Recent research indicates laboratory animal personnel using in the SPF barrier unit passed a shower lock and changed their induction chambers to anesthetize rodents may be placing clothes twice. Technicians of the conventional unit did not shower themselves at risk of exposure to unknown levels of volatile and changed their gowns only once. Showering after work was anesthetics. We hypothesize risk can be greatly reduced by per- optional; however, every employee used 2 separate lockers for forming a simple modification to an existing rodent anesthesia private and for work clothing. All animal rooms were ventilated station. An isoflurane vaporizing system was used to deliver a with HEPA-filtered, conditioned air at a temperature of 22 ± 1 °C measured amount of isoflurane and oxygen into an induction and 55 ± 5% relative humidity, with 14 air changes per hour. No chamber. Rats were anesthetized in the chamber by administer- study participant had pet rodents at home. After extraction of ing 5% isoflurane and oxygen at a rate of 1 l/min continuously the dust samples, the level of the mouse allergen Mus m 1 was for 60 to 90 s. Isoflurane and oxygen were turned off before determined using a commercially available ELISA. In the settled opening the chamber to retrieve the animal. With additional dust of the private dwellings of the animal handlers, the Mus tubing and a Y connect, we attached the blower end of a small m 1 levels ranged between 0.29 and 2.46 µg/g (mean 1.3 µg/g) hand-held vacuum in the gas line directed towards the chamber. in cars, 0.006 to 0.18 µg/g (mean 0.081 µg/g) in carpets, lower A stopcock was used to stop the anesthetic gas from backing up than detection limit up to 0.81 µg/g in mattresses, and 0.028 to into the line or to allow the air from the blower to flow into the 0.19 (mean 0.07 µg/g) on kitchen chairs. Our results indicate induction chamber, blowing most of the volatile gas out of the that the private home may be an important location for exposure chamber before opening. We compared isoflurane concentration to work-related allergens from mice. This should be taken into levels in the environment surrounding the induction chamber consideration when developing an overall occupational health with and without the benefit of the blower. A halogenated gas intervention strategy. We observe the tendency of mouse al- monitoring badge was placed near the opening of the induction lergens to stick to skin, hair, and clothes, thus giving allergens chamber for a period of 150 min. In that time, 6 rats were anes- the opportunity to be carried from work into private areas. This thetized, as described above. Isoflurane concentration reached can only be prevented by strict separation of the working and 6.7 ppm. The following day, a new halogenated gas monitoring living environments. badge was placed in the same vicinity as the first for 135 min. Six rats were anesthetized. With the added benefit of blowing the P78 Fur Mite Treatment 101: Labor, Time Management, and volatile anesthetic out of the induction chamber before opening, Project Coordination concentrations were reduced to 1.7 ppm. The system described here reduces exposure to potentially harmful volatile anesthet- J Pinkney1,2, D Smith2, T Burkholder*2 ics by 75%. Our system is unique because few rodent anesthetic 1 SoBran, Bethesda, MD; 2 NIH/DVR, Bethesda, MD stations allow the operator to flush isoflurane from the induction Fur mite eradication is known to be a challenge in large trans- chamber before opening. It is easy to obtain, affordable, and genic rodent colonies. Considerations when making a plan of adaptable to permanent and portable anesthetic systems. action include the size of the colony or number of cages to be treated, ease of administration, choice of chemical or drug for P80 Washroom Efficiency: A Pilot Study treatment, and the coordination of animal care staff. Our past JM Traylor*, GR Robinson facility treatment protocols required a time commitment of 10 to 20 h per wk from the technical staff for up to a year per room. Comparative Medicine, Pfizer Inc., Groton, CT When developing our treatment plan for fur mite eradication, Washrooms are essential to the operation of an animal research efforts were taken to ensure the plan would provide the most facility and are busy places that have to meet the needs of the effective care while striving to minimize the amount of labor and scientific and husbandry staff. Historically, the washroom staff time involved in providing the treatment. Animals were screened was sometimes comprised of contract personnel with limited to confirm suspected fur mite presence based on 1 mouse that experience in laboratory animal science. To mitigate this lack had been exported from the colony. Once confirmed positive, the of experience, more experienced husbandry technicians would room was treated with ivermectin in water at specific intervals. help when time was available. They did so by assisting with Results from retesting indicated eradication of mites was possible equipment cleaning and preparing caging for their assigned with 2 mo of treatments on alternate weeks and follow-up testing rooms. This process created numerous trips in and out of the for 6 mo after treatment. This presentation will explain several washroom, which led to decreased efficiency and potentially factors that were considered in the development of the treatment compromised biosecurity. A pilot study was undertaken at one plan initiated in this facility. Additionally, it stresses the impor- of our facilities to improve the washroom traffic patterns, equip- tance of strategic planning, teamwork, and cooperation across ment maintenance, inventory management, and the reduction departmental lines. It also further discusses the background of biosecurity risk. We staffed the washroom with 4 experienced and support information, the advantages and disadvantages of husbandry technicians and implemented a menu system to ad- the program, diagnostic techniques and frequency of testing, dress the organization, inventory, and biosecurity issues. This and the overall outcome of the program used to combat such system is a service provided by the washroom personnel to fill a novel topic. Lastly, this presentation can also be useful when the scientists’ and husbandry technicians’ caging requirements. combating other treatable common laboratory animal conditions The use of the menu system has decreased the daily clean-side when looking into treatment timelines and strategic planning entries of husbandry staff by a minimum of 64 times. The system according to testing results. has taken 14 husbandry technicians’ hands off of the equipment Abstracts of scientific papers 2008 AALAS National Meeting stocks, enabling better resource management. This tool has also attention behaviors (being at the front of their cages and watch- allowed for proper equipment rotation by the washroom staff ing the monitor) when that video was played; some NHPs also and has improved accountability and ownership. Organizational displayed lip smacking when monkeys in a similar laboratory maintenance and biosecurity has also improved and continue environment appeared on the screen. The other 3 videos elicited to be preserved. much smaller percentages of monkeys watching them (range: 10% to 40%). One conclusion drawn is that NHPs may respond P81Neonate Scooper to video scenes with which they are more familiar. J Burgess* P83 Castration Eliminates Conspecific Aggression in Group- Animal Facility—E.A.M, Trudeau Institute, Saranac Lake, NY housed Male Surveillance Mice Neonatal mortality can be a problem in mouse production fa- JL Lofgren*, SE Erdman, C Wong, R King, T Chavarria, A Discua, cilities. In breeding colonies, one of the goals is to keep stress JG Fox, K Maurer levels as low as possible. After cage changing, females with Division of Comparative Medicine, Massachusetts Institute of young pups show high stress levels. These females will pick up Technology, Cambridge, MA their pups and move around the cage franticly. Neonates will be left randomly about the cage, some left uncared for and oth- Microbiological surveillance is an essential component of murine ers cannibalized. The stress of the female is seen immediately health maintenance. To adequately ensure the pathogen status after cage changing and can continue for up to 1 h. With some of mice used in our health-surveillance program, we use in- modification to a simple food scoop, we are able to easily and house–bred CD-1 mice. To make optimal use of mice from this quickly move an entire nest of pups during cage changing. A colony, thereby reducing overall animal use, we use the female 200-ml polypropylene scoop was modified to place a 90-degree progeny for recipient animals in our core transgenic facility bend in the open portion of the scoop. The bend was made by and the male progeny in our surveillance program. Veterinary gently heating the scoop with a propane torch, creating the 90 technicians noted that there was a great deal of fighting among degree bend by pressing the scoop against a solid surface and cohoused male surveillance mice that was not remediated then quickly cooling the scoop in cold water. The modified scoop through environmental enrichment. We hypothesized that early withstands autoclaving to 121 °C. The neonate scooper is used castration may eliminate fighting in male mice. As such, male where neonates (0 to 12 d) are present. Moving the entire nest mice were castrated prior to 2 mo of age and then released into at once eliminates handling of individual pups and appears to the surveillance program. Castration was performed using an produce less stress on the female. By moving the nest, the scent open technique under general chemical anesthesia, and mice stays on the pups, there is no manual pressure on the pups, and were given buprenorphine for postoperative pain. Veterinary less time is spent in breeder cage changing. Use of the neonate technicians recorded fighting in castrated and non-castrated male scooper decreases the time needed for cage cleaning and pro- surveillance mice during their weekly rounds to determine any duces higher neonate survival. benefit. Castrated mice recovered from the surgery within 24 h with no noted negative effect. Over a 3-mo period, the overall P82 Video Enrichment Preferences of Laboratory Macaques prevalence of fighting in intact male cages was 54% (12 of 22 (Macaca mulatta) cages) compared with 0% (0 of 16 cages; P = 0.0019) in castrated cages. Additionally, 11 intact male mice were separated and 7 JN Camacho*, J Tsui were euthanized due to fighting. Castration served as both a CCM, Massachusetts General Hospital, Charlestown, MA refinement by eradicating pain and stress resulting from fight- ing and a reduction in animal numbers by allowing the males Video is a common means of environmental enrichment for from the breeding colony to be used as surveillance mice. If laboratory nonhuman primates (NHPs). Other studies indicated only females from the colony were to be used, the size of the that NHPs appear to prefer to watch something on a screen as breeding colony would need to be doubled and the majority of opposed to nothing at all. However, there is still much unknown male progeny would be euthanized. In conclusion, castration is a about which video topics may be more effective in promoting minimally invasive, safe, humane, and rapid method to eliminate species-specific behaviors and minimizing apparent boredom conspefic aggression in male CD-1 surveillance mice. in captivity. We performed basic observational assessments to determine if adult M. mulatta and M. fasicularis exhibited prefer- ences for different video subjects, involving a total of 14 monkeys. P84 Canine Home Environment Acclimation Program: A Each animal’s normal behavior was established during routine Conditioning Program to Better Prepare Research Dogs for husbandry procedures and interaction with animal care staff over Adoption several days in the absence of video enrichment; other enrich- JL Wheatley*, GL Jeans, KD Corcoran, AS Fix ment modalities (for example, contact with conspecifics, audio Procter & Gamble Company, Cincinnati, OH enrichment, nutritional variety, manipulanda) were unchanged. Four videos were played: NHPs in a naturalistic setting, a nature Many canines retiring from research service are deemed eligible video involving no NHPs, a children’s education video, and an for adoption. However, without the proper intervention, a dog AALAS video tutorial on proper occupational safety practices raised solely in a research institution may have difficulty adjusting when working with NHPs. Each video was played 10 to 12 times to life outside the facility. In an effort to better prepare these dogs between 0800 and 1600 on a given day, while NHP behavior was for a successful transition to a home environment, we describe an observed for 20-min increments 4 times throughout the day. A acclimation program implemented at the Procter and Gamble Co. different video was played on each of 4 consecutive d. Time and This program includes a specific acclimation and training plan for reaction were compared for each animal directly observing each each dog with exposure to environments and stimuli likely to be video to determine if different behavior resulted. Nonhuman encountered postadoption. Our goal is to address any existing primates appeared to prefer the human tutorial video more behavioral or socialization issues, introduce housebreaking prin- than the other videos because 70% of macaques displayed direct cipals, and familiarize dogs to common household items prior to Abstracts of scientific papers 2008 AALAS National Meeting adoption. The artificial home environment resides in an unoccu- staff management and performance. Bioluminescence systems im- pied lab within our facility and includes a television, refrigerator, mediately calculate adenosine triphosphate (ATP) levels, indicating radio, telephone, floor fan, couch, dinette, mirror, and area rugs. the amount of organic material present on surfaces. At our facility, Acclimation to the home environment, along with other interven- we used a bioluminescence device to identify which management tions such as leash walking and obedience training, are provided factors contribute to enhanced work performance. We assessed based on individual animal needs. We have found this program sanitization of animal room floors, fencing, and pens under 4 condi- to be beneficial and enriching not only for the canine subjects, but tions: blinded (baseline), non-blinded (supervision), non-blinded also for the animal care and veterinary staff members who have with 48 h delayed feedback and delayed accountability (required participated. The program has been used for many dogs that have to resanitize and retest), and non-blinded with immediate feedback been placed in adoptive homes. The success of these adoptions and immediate accountability. Our results indicate that without staff has been increased by exposing the dogs to the home environ- knowledge of testing (blinded), there was no significant difference ment and household items prior to their leaving the facility. We between pre- and postsanitization values. This suggests that without advocate an acclimation program for canine adoption candidates supervision, the staff were not effectively performing sanitization as a way to improve the transition to a home environment and to procedures. When the staff were aware of testing (supervised), increase job satisfaction for employees working in a laboratory pre- and postcleaning values were significantly different (t = 1.90, animal research setting. P < 0.05), but only 60% of the measured areas reached our pass cri- teria of greater than 75% reduction in surface ATP levels. Delayed P85 Supplemental Care for ERCC-1–deficient Mice feedback and delayed accountability had little effect on sanitation efficacy. However, immediate feedback and immediate accountabil- J Mensch* ity cohorts demonstrated significantly improved performance (t = SoBran, Inc., Rockville, MD; OLAS, NIAAA/FLAC, Rockville, 2.95, P < 0.01). Also, in the immediate feedback and accountability MD condition, 100% of the tested areas reached a pass score of greater than 75% reduction in ATP. In conclusion, bioluminescence can be ERCC-1–deficient mice tend to be runted at birth and die before both an effective and inexpensive vivarium management tool. By weaning. Knockout of the ERCC-1 gene in mice results in a shorter using bioluminescence to collect quantitative assessments, and by life span and the mice display signs of premature aging. At our providing real-time feedback and accountability to staff, supervisors facility, these strains accurately portray these problems, along can improve employee performance. Furthermore, because surface with degeneration of the hind limbs. The litters born from ERCC-1 ATP values are obtained before and after sanitization procedures, -/- deficient females were broken down into 3 categories based on bioluminescence reduces the amount of direct supervision required genotype. The wild type mice resembled normal mouse weights at during sanitization, increasing supervisors’ time management and weaning (15 g at 21 d). The heterozygous mice tend to be slightly efficiency. smaller at weaning age (7 to 8 g), and the knockout mice were very small (3 to 5 g) at weaning age. Our weaning policy indicates that 7 d post weaning, the animals are given extra supplements such a P87 Double-mouse Perch Designed as an Enrichment and fruits (apples and oranges) and gelatin containing whey protein. Life-saving Device Incidentally, the heterozygous and knockout mice struggled to KA Phillips*, G King, A Ibanez, L Altholtz, CV Johnson mature, and survival was not expected. Researchers required these Laboratory Animal Resources, Merck & Co., Rahway, NJ animals to be weaned early (14 d) and performed an AAV1 injection on the brain. Postsurgical knockouts recovered much slower than Rodents housed on racks with automatic watering are widely the heterozygous animals subjected to the same procedure. In order used in the research community. Although there are many ben- to increase their survival probability, the animals were provided efits to this type of housing, occasionally cage flooding due to a with additional surveillance and extra nutritional supplements leaky valve occurs. A novel double-mouse perch was designed beyond the 7 d postweaning period. These supplements consisted as an enrichment device and provides elevated shelter in the of fruits, gelatin fortified with whey protein, a nutritional drink, event of a cage flooding. It is completely constructed of stain- bacon-flavored soft pellets, and a special-needs dough diet. With the less steel and is suspended from the wire grid of a mouse cage supplemental care in place, we found that the animals gained weight lid without compromising floor space. The upper level indents more rapidly and their overall body condition did not degenerate over the bottom level giving it a stair-step, climbing functional- as quickly. One particular mouse was given a supplementary brain ity that provides 2 levels of activity for enrichment opportunity. injection of AAV1 and ERCC-1 gene which posed to reverse the The perch allows for full viewing of the mice through the cage effects of the knockout genotype. The mouse’s presurgical weight during daily observations and can easily be sanitized through was 4.30 g; it continued losing weight postsurgery, until reaching the rack washer. It also provides aid for small and aged mice in 3.25 g. After our postsurgical treatment, its weight increased to accessing food and water within the cage, as well as increased 3.30 g and it appeared much healthier than the previous knockout vertical space. The functionality of this novel device was tested in animals injected only with AAV1 injection. Combined with the an observational study. A total of 10 mice, 5 per box, were housed constant supplemental care and ERCC-1 injection our test animal’s in a small ventilated mouse box (7.5 by 13.75 by 5.5 in.) and were condition remained stable and healthy. given 1 wk to acclimate to the perch. During the acclimation pe- riod, mice were observed interacting with the perch by actively P86 Use of Bioluminescent Technology as a Novel Husbandry climbing on both levels. After the acclimation period, the mice Staff Management Tool underwent 2 separate simulations of an actual cage flooding. The first simulation was a pilot study in which warm water was KM McDonald*, PA Chalmers, JT Newsome slowly introduced into the cage using a flexible tube. This was Division of Laboratory Animal Resources, University of Pitts- done to monitor their response to the rising water and interaction burgh, Pittsburgh, PA with the double mouse perch. The second simulation study was done to mimic an actual cage flooding from a leaking sipper tube. Bioluminescent technology is often used in vivariums to verify the In both simulations, all 10 mice safely climbed to elevated shelter. efficacy of santiation procedures, but it can also be used to enhance Abstracts of scientific papers 2008 AALAS National Meeting They were moderately wet and huddled together on the upper over a 12-mo period. As a control, biofilm from rack manifolds level of the perch. From these studies, we have determined that in a facility with no additional water treatment (municipal wa- this device is a well-used form of enrichment and an effective ter passed through a 5-μm filter) was also sampled. At periodic life-saving platform during accidental cage flooding. intervals, biofilm samples were obtained from the inside of rack manifold piping using sterile swabs. Biofilm samples were as- P88 Use of a Remote, Real-time Infusion Monitoring System sayed for total bacterial population by a rapid assay for microbial ATP. Results showed minimal biofilm growth in 4 racks sampled K Pickel*, J Resendez from the facility with RO-DI-UV–treated water for up to 12 mo Infusion, MPI Research, Mattawan, MI of use. In contrast, the 3 racks from the control facility showed substantial biofilm growth between 3 and 6 mo of use. Median The use of percutaneous indwelling catheters is a widely ac- RLU/swab values for the RO-DI-UV facility were 11.8, 24.0, and cepted method of administering pharmaceutical compounds 19.3 for mo 6, 9, and 12, respectively, compared to 90.0, 203.0, into systemic circulation. Vascular access is readily established 515.0, and 759.0 for mo 3, 4, 5, and 6 in the control facility. These using this type of catheter, but long-term patency is often dif- results suggest that the combined processes of reverse osmosis, ficult to maintain. Consequently, surgically implanted vascular deionization, and ultraviolet filtration are effective in limiting catheters are often used when longer duration administrations biofilm growth in AWS for up to 12 mo. Facilities that provide are required. As this infusion model has become more established drinking water conditioned in this manner may be able to reduce in the preclinical arena, so has the acknowledgment that continu- the frequency of sanitization and sterilization to once per year, ous intravascular administration can present unique challenges which would mitigate the cost of purchase and installation of compared with other more standard routes. Consideration must the water treatment system. now be given to the technical requirements and equipment necessary to successfully conduct this type of study (such as tethers, animal jackets, infusion pumps, surgical capabilities, P90 Comparison of Two Foraging Devices for the Psycho- and aseptic technique). Longer administration durations are logical Well-being of Caged Cynomolgus Primates (Macaca accomplished by using syringe or ambulatory infusion pumps fascicularis) either intermittently for repeated doses over the course of a day J Howard*, K Pyle, M Liddle, C Noyes, E Kirchner, H Byrnes, C or weeks, or continuously over a period of consecutive days. The Grant, C King, K Trotta logistics required to maintain and monitor the infusion over the course of a typical study often result in increased labor hours Veterinary Science Technology, SUNY Delhi, Delhi, NY or staffing. This operational impact may result in a less efficient In maintaining compliance for the psychological well-being of overall infusion program. To address this issue, MPI Research caged cynomolgus monkeys (Macaca fascicularis), environmental has developed a monitoring system capable of continuously enrichment through the use of foraging devices has been accepted evaluating the performance status of each infusion pump. An in our industry. This study compares a widely used, commercially in-house software program was created that communicates di- available foraging device that is hung outside the cage with a rectly with each pump; receives specific, real-time operational new prototype foraging/enrichment device that is offered inside information regarding the infusion progress; and remotely alerts the cage. The prototype is made of polypropylene plastic with a technical staff of situations or interruptions, such as catheter line 2.5-in. hollow spout connected to a 2.75-in. hollow ball. Peeled occlusions or a low battery, that may compromise the infusion. baby carrots were used as a treat in this device. This study used 9 Use of this system allows for a more timely response to pump monkeys ages 1 to 27 y old, individually and pair housed. For a alerts, which is critical to the ultimate success of the planned 4-wk period, 10-min direct observations of the animal’s interaction administration. Since the monitoring system was implemented, with the foraging devices were performed 3 times per d, 3 d per the number of technical staff and time required to maintain an wk. Interactions were recorded as discrete events on an ethogram infusion study has decreased, and the ability to successfully ad- to evaluate the animals behavior in the categories of agonistic, dress infusion interruptions has significantly improved. human-directed, affiliative, non-social, exploratory (including environment and foraging device) and enrichment. At 1100, the P89 Effect of Reverse Osmosis, Deionization, and Ultraviolet foraging devices were loaded and presented to the monkeys. The Filtration on Biofilm Development in Automated Watering test objects were left in or on the cages and were not refilled for the Manifolds 1300 and 1500 10-min ethogram evaluations. Treat removal from the available device was significantly easier than treat removal KM Hugunin*, CJ Maute, RC Dysko from the prototype because the treat was visible and could be eas- Unit for Laboratory Animal Medicine, University of Michigan, ily grasped through the holes. The prototype required substantial Ann Arbor, MI manipulation to get the carrot lined up with the spout in order to obtain the treat. The opacity of the material also made it dif- Providing drinking water to laboratory rodents through auto- ficult for the primate to see the treat inside, therefore providing mated watering systems (AWS) has become commonplace over extended enrichment. The new prototype foraging/enrichment the last decade. Previously, our laboratory found that biofilm device performed statistically better (1-tailed t tests were used with increased significantly over a 6-mo period within automated α = 0.05) than the widely used, commercially available foraging watering manifolds provided municipal water (2.8 to 3.3 ppm device in the areas described. monochloramine) passed through a 5-μm filter. The effects of various water purification treatments on biofilm growth within AWS have not been independently examined. The newest vi- P91 Commercial Modular Housing Facilities for Ossabaw varium at our institution provides municipal water that has been Miniature Swine with Metabolic Syndrome treated by reverse osmosis (RO), deionization (DI), and ultra- KM Pohle*, LD Jefferson, JW Wenzel, M Sturek violet filtration (UV) using AWS. The combined effect of these 3 Cellular and Integrative Physiology, Indiana University Purdue processes on biofilm growth in AWS over time was investigated by sampling biofilm from rack manifolds in use in this facility University Indianapolis, Indianapolis, IN Abstracts of scientific papers 2008 AALAS National Meeting This presentation describes the husbandry facilities used suc- replacing toe clipping, toe tattooing can be used as a reliable cessfully for Ossabaw miniature swine over the past 4 y. Main back-up to ear tagging or notching. It is also significantly less guidelines for care include easy-to-clean surfaces, daily pen expensive than installing a transponder. washing and excrement removal, space, temperature, and light:dark cycle. Ossabaw miniature swine are housed in 3 P93 Disposable Nonhuman Primate Puzzle for High-contain- separate commercially produced modular swine housing units ment Labs located near Purdue University. Each building is equipped with 2 60,000 btu natural gas heaters and 2 12-in. and 2 24-in. fans for K Kelly*, N Powell convective cooling. The units house the entire Ossabaw miniature Animal Resources Branch, Centers for Disease Control and swine colony, including breeding boars; open, gestating, and Prevention, Atlanta, GA lactating sows; suckling piglets; weaned piglets; young adult pigs Some primates are used for the advancement of public health that have not yet been assigned to research projects; and pigs that with regards to infectious diseases. A few of these are located are on research protocols being fed excess calorie, atherogenic in high-containment laboratories that deal with emerging in- diets to induce obesity, metabolic syndrome, and heart disease. fectious diseases at BSL3/4. Enrichment items that enter these Stainless steel gating, fiberglass outer walls, and slatted flooring laboratories do not come back out once the study has ended; enable rapid, efficient daily sanitation. All buildings maintained they are discarded. The disposable enrichment puzzle costs animal holding room average monthly temperatures between pennies to construct, does not require cleaning, and allows 61 °F (low limit) and 81 °F (high limit) guidelines over a 1-y primates located in BSL3/4 the opportunity to express species- data collection period, despite the variable Indiana temperature specific behaviors that they would otherwise not be able to. The fluctuations, which exceeded the limits on all days. The modular bulk of the device is made from cardboard plotter paper tubes. housing units exceeded the low-temperature limit on a mean of One tube is approximately 3.5 in. in diameter and cut to a 6-in. only 9 (8%) of the days that the exterior temperature was below length. The other tube is 2.5 in. in diameter and cut to a 10-in. the low limit (111 d). The high-temperature limit was exceeded length. The smaller diameter tube comprises the inside of the a mean of only 17 (6%) of the d that the exterior temperature puzzle and nestles inside the larger tube. Holes are cut into both was above the high limit (288 d). Effective sanitation, space, and tubes to allow access to treats that will be later hidden within. temperature maintenance of the animal housing area makes these Two lengths of plastic chain are cut to around 10 to 11 in. in commercially produced modular swine housing units acceptable length. These pieces of chain will be attached at the end of the housing for biomedical research swine. tubes through holes cut on both sides of the tube. Another link of larger plastic chain is used to hold the length together. This P92 Benefits of Toe Tattooing as a Method of Identification single link is used by cutting a small gap on its side and holds KJ Diven* the entire device to the front of the primate cage. The inner tube is filled with shredded paper and treats. The primates must align Animal Care and Use Committee, John Hopkins University, the holes to have access and then have a fun time shredding the Baltimore, MD cardboard. The puzzle device was tested on a room of 8 singly Small rodent identification can be problematic when using vari- housed adult male rhesus macaques at BSL2. These primates ous popular techniques. When rodent pups are young they are had used a similar PVC device and were aware of the necessary too immature to ear tag, ear notch, or implant a transponder. mechanics required. Most of the contents were removed within Historically, the solution to this problem was to toe clip. This 15 to 30 min, and the tubes themselves were a continued source involves the removal of the first bone of certain toes. The Guide of interest for several days as the cardboard could be shredded. for the Care and Use of Laboratory Animals limits its use and states, Some primates had destroyed the device by the next morning; “toe-clipping, as a method of identification of small rodents, others were still working on theirs 2 d later. These devices could should be used only when no other individual identification be used for any level of containment, but were specifically de- method is feasible and should be performed only on altricial signed to be used in a high-containment lab. These labs will soon neonates.” Other rodent identification methods also have their be coming online at CDC, and this device will be given its first drawbacks. Ear tags, if not properly sterilized and placed, can true trial run in the coming months. cause irritation to the ear pinna and be pulled out. Ear notching uses a numbering code that is limited to the hundreds. When P94 Wild-caught Virginia Opossums (Didelphis virginiana) animals fight, the ear can be ripped, resulting in a loss of the tag or Thriving in a Research Environment disruption of the pattern. Even excessive grooming can cause the pattern to be unreadable or the tag to be removed. Toe tattooing L Andrews*, W Beckman, KJ Thompson, E Paster, DL Hickman is accomplished with simple tools and requires limited practice Veterenary Medical Unit, VA Medical Center, Portland, OR time. The tools needed include a sterile lancet, green tattoo paste, This presentation will describe our experiences in developing and something suitable to hold a dab of the paste, and an alcohol implementing a regiment to support sustained health, provide wipe. Pups (1 to 14 d old) are cupped in the hand; older animals environmental enrichment, and acclimate wild opossums to a are put in a restrainer. The lancet tip is dipped in the tattoo paste research setting. The wild-caught opossum is widely regarded and the desired toe or combination of toes are poked with the as a difficult and possibly dangerous animal. Handling, restrain- lancet. The skin must be punctured to introduce the paste into ing, and transporting these misunderstood marsupials presents the skin, leaving a mark. The spot is then gently wiped with the a unique set of challenges to the animal care staff and investiga- alcohol pad to remove excess paste, and the pup is returned to tors. These essential techniques, if not done properly, can result in its dam. A green dot is visible on the toe that has been tattooed. great physical harm to the handler and the opossum. Using food This dot will persist for the life of the animal. Toe tattooing is an as enrichment helps determine each individual opossum’s palate, excellent replacement for toe clipping because it is less invasive helps maintain health and promote weight gain, and builds a re- and uses the same coding system. Therefore, transitioning from lationship with the technician. The demeanor and personality of one to the other does not disrupt recordkeeping. In addition to each animal can be determined through these daily interactions. Abstracts of scientific papers 2008 AALAS National Meeting Through the use of proper handling techniques and positive en- (P = 0.001), were observed between the means of the samples richment, some opossums are able to be tamed to the point where that were taken after the installation of the HEPA filtration (15 procedures and treatment administration are considerably less CFU/m3) to that of samples taken before the installation of the stressful on the animals and technicians. Learning to understand HEPA filtration (36 CFU/m3). This poster presents the findings attitude and behavior trends through working with animals on of a study to investigate the efficacy of a portable HEPA filtration an individual basis increases a technician’s ability to provide unit installed in the operating theatre. appropriate care. By improving the animal’s environment, the productivity and quality of the research is enhanced. P97 Cage Color Preference for Maintenance and Breeding of Rats P95 Use of a Hand-held Computer Device to Increase Efficiency DR Almeida, VL Amaral, VB Rosa, M Cordini, M Frajblat* in Observations of Single-housed Nonhuman Primates Department of Health Sciences, Universidade do Vale do Itajaí, LM Mello*, KL Potter UNIVALI, Itajaí, Brazil Animal Resources, Lovelace Respiratory Research Institute, Animal well-being is a fundamental aspect of laboratory animal Albuquerque, NM science. The conditions under which animals are maintained have As part of our enrichment program, all single-housed nonhuman an impact in the scientific data collected. Light is essential for the primates (NHPs) have their object enrichment rotated weekly. rat circadian rhythm; it also regulates reproduction and other The enrichment technicians, or a trained designee, monitor the systems. The objective of this study was to test rats’ preferences NHP’s reaction to the object as well as making behavioral ob- between white or black cages and to investigate if the black cage servations. Previously, these observations were made on printed has an effect on the reproduction and growth rate of these animals. forms that included animal identification and location, with the The preference study was performed with 8 male and 8 female observer filling out by hand the object offered, the NHP’s reac- Wistar rats divided in groups of 4 and maintained in 2 propyl- tion to the object, and the animal’s behavior. Even though the ene cages (black and white, 49 by 34 by 16 cm) connected with a object enrichment and the animal’s reaction to the enrichment transparent tube. A total of 8 observations were performed during had preestablished codes, the process was very time consuming, a 48-h period when the location of the animals and the nest was with observations of approximately 150 animals requiring 6 to 8 recorded. The cages were cleaned, inverted, and new set of data h. There were also issues of contamination when the enrichment/ was collected. Feed and water consumption and fecal production observation paper forms were brought out of the animal rooms. were recorded. Four groups of 6 males and 6 females (total of 48 Legibility was also a problem for this handwritten recorded data. animals) were maintained in a white or black cage for 13 wk from To help increase efficiency of all observations department wide, weaning and body weight was recorded weekly. Fifty breeding the decision was made to begin using a computer-based program pairs were divided into white and black cages and reproductive for recording and reviewing data using hand-held computer data was recorded for 6 mo. This study as approved by the institu- devices and wall-mounted computers in the animal areas. The tion ethical committee. Nests were built in the black cages in all enrichment observation forms on the computer listed the NHP’s tests. During the 8 observations, an average of 89.5 ± 1,1% of the identification and location, as well as gave the user the ability to time the animals were in the black cages. The water consumption use a default object item for the room, with drop-down menus for and fecal production was higher in the white cage (201.5 ± 15.5 the reaction and behavior codes, and a space to type in comments. compared with 145.5 ± 6.5 ml and 331.5 ± 16.5 compared with The use of the hand-held device decreased the time required to 159 ± 5.0 fecal pellets for white and black cages, respectively). complete these observations by almost half, now taking only 3 However, a higher food consumption was observed in the black to 5 h. Also, the device can be sprayed with disinfectant when cage (81.0 ± 21.1 compared with 154.3 ± 11.8 gr for white and leaving the animal room, reducing the risk of contamination. black cages, respectively). No differences were observed in the This device has increased animal observation efficiency, reduced reproductive parameters observed (8.6 ± 0.3 compared with 8.1 the potential for contamination, and can be viewed by anyone ± 0.3 pups/litter and 5.7 ± 0.2 compared with 5.8 ± 0.3 births/6 who has access to our web-based data system, which reduces mo for white and black cages, respectively). The results of this our paper usage. In addition, enrichment technicians now have study show a strong preference of rats for black cages. Also, the more time to develop and evaluate novel enrichment ideas. use of black cages did not influence the breeding performance or the growth rate. These observations suggest that these cages P96 Managing Air Quality in a Large Animal Operating The- could be used to improve the well-being of the animals without atre affecting its production. L Schofield*, D Matthews P98 Systematic Approach to Solving Breeding Colony Prob- Hercus Taieri Resource Unit, University of Otago, Dunedin, lems New Zealand MC Rosa*, D Perez, M Sabol-Jones, ND Hitt Sheep and pigs are routinely used in our large animal operating AHCS, NIH/NINDS, Bethesda, MD theatre for cardiovascular and orthopaedic survival research studies. Despite careful attention to standard aseptic techniques The increasing use of transgenic and knockout mouse models used in infection control, wound contamination can occasion- has resulted in more researchers maintaining their own breeding ally occur. Our experimental surgical facilities were originally colonies. Because many researchers are not familiar with mouse designed without the provision of HEPA filtration. With the reproductive biology and breeding colony management, vivari- advent of new technology, the use of readily portable equipment um staff members are being consulted on these topics on a more may offer significant advantages in the overall management of frequent basis. This type of consultation can be time-consuming infection control. Airborne bacterial samples were taken during for vivarium staff, but provides an opportunity to demonstrate orthopaedic surgical procedures both prior to and after the instal- the value of their unique background and expertise to directly lation of a portable HEPA filtration unit. Significant differences contribute to the success of research projects. The most common Abstracts of scientific papers 2008 AALAS National Meeting obstacles encountered when advising researchers on breeding complex and diversified environmental enrichment (EE) that colony issues are incomplete colony histories and compliance promotes well-being and alleviates stress-related abnormal with recommendations. To improve the efficiency of our breed- behaviors and health problems. Despite their positive impact ing colony consultations, we have developed a system which on research, the use of enrichment strategies in neuroscience is maximizes our ability to gather pertinent strain and breeding often limited by concerns about safety, cost, and damage to head colony background and to monitor compliance with recom- implants. In conjunction with planned facility renovations, we mendations. To obtain the necessary background information, designed independent EE cubicles that promote arboreal, object- we have developed a questionnaire consisting of breeding- and oriented, and curiosity behaviors while ensuring the safety of strain-related questions for the researcher to complete. Once we implanted macaques. We hypothesized that such units would have this information, we meet with the researcher to clarify de- improve well-being and have positive impact on stereotypic and tails and to identify the researcher’s perceptions of the problems. feeding behaviors, activity levels, obesity, and interaction with After analyzing the information, a breeding plan is provided staff. Each EE cubicle measures 36 by 8 ft, was made of stainless to the researcher and our services are offered to initiate the steel frames and panoramic 0.5-in. laminated glass windows, program. We offer to manage the colony for a trial period after and was supplied with HVAC support and floor drains. Units which a written report with our findings and recommendations were paired with a removable divider and positioned between for continuing maintenance of the colony is provided. Using 2 holding rooms to increase access and visual contact between this system we have been able to identify specific problems and animals. Each unit was equipped with multiple, sanitizable, non- improve breeding colony production for several transgenic lines. toxic enrichment devices and various destructible enrichment This system has resulted in the researchers receiving practical materials. Devices were sanitized after each use and rotated training on breeding colony management and has increased the every week to maintain novelty and increase stimulation. Each satisfaction of the vivarium staff as well. animal was given 1 to 2 rotational time slots of 2 to 3 h per wk. Animals were easily trained to transfer to and from their cages P99 Enhancements to the Canine Socialization and Training to the units in transport cages. After 1 y of operation, no injuries Program have been observed in implanted macaques. More importantly, we observed increases in time spent in species-typical activities M Huntsberry* and overall activity level, as well as decreased stereotypical SoBran, Bethesda, MD; NIH/DVR, Bethesda, MD behaviors, appetite disorders, and abnormal social behaviors. EE cubicles also improved the quality of social interaction with Dogs on long-term research studies benefit from an organized staff. Increased activity and decreased hyperphagia, combined program of socialization and obedience. Learning basic com- with implementation of a balanced diet resulted in weight loss mands increases the manageability of the dogs for care staff and in the majority of animals. The observed improvement in animal veterinarians. A formal program such as this has been executed well-being and health demonstrate that these EE cubicles offer by a dedicated behavior staff at my facility. From the beginning, significant benefits to neuroscience macaques with no obvious handlers have met once a week as a group with a behaviorist risks to head implants. for “canine class.” Classes cover positive early human social ex- periences for puppies, providing environmental enrichment for all dogs in the form of agility-related games and teaching basic P101 Use of Luminometers as an Alternative for Evaluating commands such as sit, stay, down, and loose-leash walking. In Animal Housing Sanitation the last several months, our canine socialization and training M Abernathy* program has expanded to include, among other things, talks on Borgess Medical Institute—Medical Device Research Laboratory, canine behavior. Topics taught consist of canine body language Portage, MI indicative of fear, aggression, and affiliation and the appropriate human reaction to assuage a tense situation or reward a happy The use of contact plates for evaluating animal housing sanitation dog. Additionally, handlers participate in individual monthly can be a lengthy process, especially if a positive result leads to re- meetings with the behaviorist allowing for refinement of canine- testing. Effective sanitation of animal housing is a critical function handling skills without the distraction of several other dog/ in a research laboratory and plays a role in how a facility allocates handler pairs. A systematic procedure for pairing dogs to be space for animal housing. The use of luminometers can decrease housed together has also been implemented along with a formal the time taken to verify that effective sanitation has occurred. treatment program for rehabilitating fearful animals arriving Luminometers measure the light emitted when luciferase reacts from other facilities. As a whole, the canine program at NIH with adenosine triphosphate (ATP). Luciferase is an enzyme that provides both human and dogs with supplemental enrichment, uses ATP to catalyze the oxidation of luciferin, which in turn emits positive experiences with one another, and individual atten- light. ATP is created by all living organisms and can be used to tion. Opportunities such as these contribute to alleviating and assess biological activity. A swab that contains enzymes to break possibly preventing the development of abnormal behaviors down the cell wall of organisms and release ATP can be used as the resulting in mentally, in addition to physically, healthy subjects mechanism to sample a surface for living organisms. The ATP col- for research. lected can then be coupled with the appropriate enzymes, resulting in a measureable light reaction that can be quantitated by a lumi- P100 Novel, Safe Environmental Enrichment Unit for Rhesus nometer in relative light units. The resultant relative light units Macaques (Macaca mulatta) are directly proportional to the ATP in that swabbed area. Given the level of ATP, one could assess the biological activity present M Watson*, A Ramirez, DM Gonzalez, J Reuter, M Leblanc using this technology. If a region of a dirty pen was sampled and Animal Resources Department, The Salk Institute, La Jolla, CA resultant relative light units compared to the relative light units of the same region sampled after sanitation, then a reduction in bio- Neuroscience research with macaques requires normal behavior logical activity could be calculated. A passing or failing reduction with minimal extraneous variability to produce valid, accurate, in biological activity could be assigned based on what value was and reproducible models. These animals models benefit from Abstracts of scientific papers 2008 AALAS National Meeting effective in preventing the clinical manifestation of disease due to involved water restriction protocols in which NHPs receive a bacterial contamination. The practice of using a luminometer could set volume of water each day; if the NHP removed its sipper then be used to evaluate the effectiveness of sanitation as quick tube, there was no way to know if or how much of its allotted as the sanitation could be performed. The use of luminometers as water was consumed. Lastly, this made for potential safety an alternative for evaluating animal housing sanitation is efficient hazards due to the floor being slick when wet, and also created and economical. Using this method as compared to contact plates, a general mess in the animal room. Our solution was to cut our facility obtained sanitation results 94% faster. 2-mm slits into the bottle end of the sipper tube, approximately 1 mm apart. These created sections of metal that were then bent P102 Novel Approach for Providing Rodent Soft Food Using backwards and fanned out horizontally to create a lip. When the a Semi-automated Watering Valve tube was inserted into the stopper from back to front, it could no longer be pulled all the way out. Since modifying all of our M Campbell*, K Brayton, S Nigra, KE Saunders sipper tubes, not one has been removed by an NHP, even though Comparative Medicine, Oregon Health & Science University, animals have been observed trying. This simple modification Portland, OR has helped maintain a clean, safe working environment for all staff. But more importantly, it has ensured that all NHPs have In an attempt to use resources efficiently, we looked for novel uninterrupted access to water or are receiving accurate amounts ideas to reduce labor cost. Studies at our facility determined when on restriction protocols. that providing soft food to rodents incurred significant labor costs. Although most of our rodents receive water by way of an automatic watering system, there are occasions where the P104 Refined Method of Cage Changing for Nonhuman Pri- veterinary or research staff requests that softened food be sup- mates plied to animals. Consideration was given to using city water L Teixeira, ME Graffam*, S Sleboda provided through the tap in each rodent room to soften rodent pellets. This solution was deemed unacceptable since the water CCM, Massachusetts General Hospital, Charlestown, MA supplied through the automatic watering system was treated, Nonhuman primates (NHPs) continue to serve as valuable reverse osmosis water, and the city water came directly from open animal models for biomedical research. Because animal facility surface water supplies. It was determined that providing water in space is so expensive to build and maintain, and with biomedical bottles that were filled with treated, reverse osmosis water from research funding becoming tighter, many animal care facilities a bottle-filling station, wrapped, autoclaved, and transported to are looking for ways to get more done in less space. The conven- rooms where it could be mixed with rodent pellets was an ac- tional NHP cage change procedure uses a jump box to transfer ceptable solution. We found, however, that a significant amount animals from dirty cages to clean ones. The obstacle we faced of time was spent washing the bottles, filling them, autoclaving was that the size of the rooms did not allow the jump box and a them, and transporting them. In order to reduce the amount of clean cage rack to fit in the room with the dirty cage. The solution time technicians spent preparing and transporting the water we developed proved to be a safe, easy, and efficient alternative bottles and to ensure that the animals were getting water that method to jump boxes. We intended to eliminate the jump box was consistent throughout the study, we devised a system to tap by pushing the dirty and clean cages together so their doors into the automatic watering lines at the room level. We attached lined up, open the cage doors directly across from each other air valves (the type commonly used with air compressors) to a and have the NHP jump directly into the new cage. But with quick disconnect flexible water hose. This flexible water hose the cages so close there was no way to open both doors in a way was attached to the automatic watering systems in the room with that protected the worker and was not threatening to the NHP. a quick disconnect fitting. These hoses are the same type used Therefore, we devised a pulley system that enabled one to open to provide water to the rodent racks. The installation of these both top and bottom cage doors easily from a safe distance. This air valves and hoses was simple and inexpensive and required method of cage changing proved to be quicker and resulted in little modification to the existing automatic watering system. a safer environment for the staff and an easier transition for the Importantly, the use of semi-automatic water valves provided a NHP. We recommend this method in a room of any size because significant cost savings by reducing the labor and time involved of its ease, safety, and animal acceptance. It allows animal care in the administrations of soft food to rodents. staff to change a 4-cage bank by having only to move the bank a few feet while putting less stress on the animals. P103 Solution to Sipper Tube Removal by Nonhuman Pri- mates P105 Increased Breeding Efficiency in Mice Using Miniature Treats L Teixeira, ME Graffam*, S Sleboda Y Sun, J Wang, ME Graffam* Center for Comparative Medicine, Massachusetts General Hos- pital, Charlestown, MA CCM, Massachusetts General Hospital, Charlestown, MA Nonhuman primates (NHPs) continue to be valuable animal Mice continue to be invaluable animal models in the advance- models for biomedical research. But their high level of intelli- ment of biomedical research, with unique strains maintained gence sometimes presents unique challenges in providing routine internally for individual investigator ’s study requirements. care. A common problem we faced was laboratory macaques These unique strains are often found to be difficult breeders, removing the sipper tubes from drinking water bottles, even produce small litter sizes, and experience high mortality rates though the bottles were affixed to the outside of their cages. amongst neonates. Cross-fostering is one solution, but it also An animal would pull on the sipper tube until it slid out of the leads to increased expense due to the extra cages and animals rubber stopper, resulting in emptying its contents onto the floor. maintained. Difficulties encountered at our facility with one This created several problems, the most serious being that a NHP particular breeding colony included low pregnancy rate, small could be without water for extended periods of time (up to 17 litter size, and high pup mortality rate, primarily due to aban- h if staff had already left for the day). Another major concern donment. The colony experienced only 20% of all mated females Abstracts of scientific papers 2008 AALAS National Meeting giving birth. Litters were generally small, consisting on average Hypothermic mice are generally awake; therefore, external of 4 pups, and abandonment by dams was 100%. In an effort to warmers only were considered to avoid possible ingestion of improve production, miniature treats were trialed by the animal devices placed inside the cage. Devices tested were 250W heat care staff, with each breeding female being fed 1 ball (1 g) 1 to lamps; microwaveable gel packs, electric kennel heating plates; 2 times per wk. The results were significant in both increases and reusable, chemically activated heating pads. Internal cage in pregnancy rate and litter sizes at weaning and in decreases temperature was measured with a high-low thermometer on top in neonate mortality associated with dam abandonment. Of 30 of the bedding in microisolation cages with filter-top lids, at an females, 23 gave birth (57% increase), with litter sizes averaging ambient temperature of 70 to 72 °F. Covered cages were placed 6 pups (50% increase), and only 4% of dams abandoning their directly on top of microwaveable gel packs, chemical pads, heat- offspring (96% decrease). The combination of high biomedical ing plates, or with the lid 4 in. under a heat lamp. Heat lamps research costs and industry-wide funding reductions makes it in- were the fastest, heating to 100 °F in 5 min and overheating to creasingly vital for investigators to use the most efficient practices 120 °F in less than 15 min. Lamps required constant monitor- available. With each mini-treat costing $0.02 ($20/1000-count ing to ensure they did not overheat the cage, and also required bag), this treatment proved to be a low-cost solution to docu- a free electrical outlet and significant storage space. An 8- by mented breeding difficulties. 12-in. microwavable pad could heat the cage interior to 90 °F in 10 to 20 min and maintain temperature for approximately 1 h. P106 Asexual Reproduction Rates of Box Jellyfish (Carybdea However, microwaveable pads required the most preparation marsupialis) Increase in Circular Tanks time (4 min heating at 1000 W, plus travel time), and required heat-resistant gloves to handle. The electric kennel heating plate MS Eckerd*1, VJ Martin2 could not reach 90 °F within 20 min, though after 30 min it was 1Animal Facility, 2Department of Biology, Appalachian State able to maintain 91 to 95 °F indefinitely. It required an electrical University, Boone, NC outlet for operation. Chemically activated pads (2 6- by 9-in. pads or 1 8- by 12-in. pad) warmed the cage to 90 °F within 20 Box jellyfish (Carybdea marsupialis) are currently being studied min, and maintained 90 to 97 °F for over 1 h. They require less for a variety of interests, ranging from the evolutionary history than 30 s activation time, do not require a microwave or electri- and structure of photoreceptors to anti-venom and protein- cal outlet for deployment, cannot overheat, and require minimal based toxins. The economic impact of recent increases in jellyfish storage space. The pads could be reconstituted by autoclaving in blooms worldwide underscores the need to understand the life a water-filled rat cage on a liquid cycle. Reusable chemical pads cycle and habitat preferences of the box jellyfish. In order to offered the best combination of user and animal safety, heating obtain meaningful data on growth rates, reproduction rates, speed, and convenience. and optimal living conditions, a population has been established and cultured long-term in the laboratory. The box jellyfish are housed in covered, large finger bowls, Petri dishes, and tempered P108 Harmonization of a Global Vision for Laboratory Animal glass dishes. Culture conditions include growing animals in real Medicine: The International Association of Colleges of Labora- seawater at 19 °C and feeding them brine shrimp once a week, tory Animal Medicine followed by a water change of the dishes. No attempts are made PV Turner*1, WJ White2, PJ Baneux5, K Bayne4, J MacArthur to regulate water temperature or light/dark cycles. Under these Clark3, I Alvarez Gomez de Segura6, T Kurosawa7, KJ Lee8, J conditions, animals asexually bud off polyps and jellyfish. Box Park9 jellyfish polyps were seeded into round aquaria made of clear 1Department of Pathobiology, University of Guelph, Guelph, ON, plastic with a filter system. The round design of the system Canada; 2 Charles River, Wilmington, MA; 3Animals Scientific eliminates corners which can trap jellyfish while providing more Procedures Inspectorate, The Home Office, London, United surface area for polyp attachment than do the culture dishes. Kingdom; 4 AAALAC, International, Frederick, MD; 5Center The hypothesis tested is that asexual reproduction rates of box jellyfish (polyp and jellyfish formation) will increase in the circu- for Comparative Medicine, Northwestern University, Chicago, IL; 6Veterinary Teaching Hospital, University Complutense, lar tanks with continuous water movement and water filtration Madrid, Spain; 7The Institute of Experimental Animal Sciences, when compared to budding rates of animals in tempered glass Osaka University Medical School, Osaka, Japan; 8Center for dishes. Results indicate that budding rates of polyps and jellyfish increased in the round aquaria when compared to budding rates Biomedical Research, University of North Dakota, Grand Forks, ND; 9College of Veterinary Medicine, Seoul National University, in the tempered glass dishes. As the circular tank environment Seoul, South Korea more closely resembles the natural habitat of the jellyfish than does that of the rectangular tempered glass dishes, an increased Trained, competent laboratory animal veterinarians play a critical budding rate was predicted. role in refinement of research animal care and use. Laboratory animal medicine has existed as a veterinary specialty in North P107 Comparison of Methods to Rewarm Hypothermic Mice America for more than 50 y, and several other formal colleges of laboratory animal medicine have been developed in different NH Shomer*, H Berenblit regions around the world in the past decade. Although laboratory Laboratory Animal Resources, Merck Research Labs Boston, animal medicine is an international field, it has been difficult to Boston, MA address relevant global issues with a consistent voice because of the lack of a common global platform for communication. Cage floods are a common cause of accidental morbidity and Recently, the International Association of Colleges of Laboratory mortality in research rodents. Even minor floods that do not Animal Medicine (IACLAM) was formed by ACLAM, ECLAM, result in drowning can cause death due to hypothermia. Prompt JCLAM, and KCLAM to enable the specialty to have broad in- warming is an important first aid procedure for wet mice. We ternational representation. The mission of IACLAM covers four compared methods to warm cages to a target temperature primary areas, including: certification of veterinary specialists (90 to 100 °F), considering animal and operator safety, speed (harmonizing international standards and sharing best practices of warming, convenience of use, and storage space required. Abstracts of scientific papers 2008 AALAS National Meeting among existing colleges); education of veterinarians (encourag- while no biological endpoints are available. We recently devised ing training at various levels, including training in developing a simpler and reliable approach to determine if animal care countries); dissemination of information relevant to the field staff and investigators are practicing proper cage and animal (delivering veterinary expertise in international discussions); handling techniques. Gnotobiotic mice free of aerobic bacteria and serving as research partners (promoting the veterinary were maintained in sterilized cages, provided irradiated feed role in refinement). An additional benefit of this international and acidified (sterile) drinking water, and opened or changed partnership is that demographic analyses of laboratory animal only in an appropriate laminar flow hood after decontaminating veterinary specialists may be conducted at a global level. Results gloves and forceps with chlorine dioxide before handling sterile of a recent IACLAM survey conducted within each college have equipment or mice. Fresh fecal pellets were collected from these pinpointed critical international pipeline trends of veterinary mice and cultured in tryptic soy broth for 24 h, then swabbed specialists in this field; identified employment sector statistics, onto 5% sheep blood agar plates and incubated 48 h aerobically. which may be useful for lobbying for training support at in- Any bacterial growth from these cultures indicated a break in ternational levels; and enabled identification of subspecialists technique because the intracage environment was originally free within the field who may be able to represent the community of aerobes. Such mice remained gnotobiotic for many months internationally, as needed. As globalization trends continue, IA- when proper technique was used. Conversely, aerobic culture CLAM is expected to play a key role in supporting initiatives to results were used to institute additional training in addition train veterinarians in this field, as well as in the development of to monitor cages and supplies for sterility. These gnotobiotic global standards for veterinary care of animals used in research, mice may simultaneously serve as routine pathogen sentinels, testing, and education. thereby reducing costs even further while better monitoring colony management. P109 Improving the Efficiency and Accuracy of Animal Health Reporting by Means of Personal Digital Assistant Scanners P111 Adoption as an Alternative Destiny for Retiring Research and Bar Codes Dogs R Scott, E Villalobos, A Gaudiel, R Garcia-Gonzalez*, E Chua AS Fix3, JL Borger3, RS Cooper*1, G Jeans1, B Rupert3, MA Tet- rick3, JL Wheatley2 Laboratory Animal Resources, Genentech, Inc., South San Fran- cisco, CA Product Safety, 3Pet Care, Procter and Gamble, 1BTF, 2Central In order to provide better monitoring of clinical health issues Lewisburg, OH in a large animal facility, our department has rolled out a PDA- Adoption is a viable destiny alternative for animals involved based tracking system. The in-house bioinformatics department in some, but not all, types of research service. We describe an developed a real-time application within our already existing adoption program for dogs implemented at our company. This Laboratory Animal Systems and Reports (LASAR) database, program included a set of principles for adoption, with inclusion which allows interfacing clinical case information with order- and exclusion criteria; an adoptability classification system based ing, protocol, and other sensitive information. In the past, case on thorough medical and behavioral assessments; a specific ac- information was tracked by a mixed system of paper records climation and training plan for each dog; and a screening and and spreadsheets in a shared folder, which required many points approval process for prospective owners. Adoption candidates of manual data entry. With the rapid growth of our company’s were dogs from a wide age range and service history. Dogs mouse population, this system became inadequate in handling received complete preadoption assessments and were grouped the increasing number of clinical health cases. The new system according to the adoptability classification system. This helped has significantly improved the clinical monitoring efficiency identify areas to address during preadoption acclimation and and accuracy, while also decreasing response time to urgent care training. Specific training or other interventions were provided cases. Most human data entry errors have been eliminated due to based on individual needs, including leash training and exposure the use of automated data entry through PDA barcode scanning to environments likely to be encountered postadoption. A num- and automatic exportation of data into the real-time database. ber of dogs were moved through this program and successfully This results in all data being stored in one centralized database, placed in homes. Implementation of this program required the which allows for better searching and reporting options. collaborative efforts of staff from the veterinary, animal welfare, and facility operation areas. As a way to improve the quality of P110 Using Gnotobiotic Mice as Technique Sentinels life of retired research dogs and decrease the number of animals euthanized in nonterminal research, we advocate broader con- RS Sedlacek*, DM Jarrell, SM Niemi sideration of active intervention programs and adoption as an CCM, Massachusetts General Hospital, Charlestown, MA alternate destiny for these animals. Widespread use of microisolation cages has dramatically reduced infections in rodent colonies in a cost-effective manner by shift- P112 Biohazardous Waste Disposal Cost Savings ing the effective barrier from the animal room or vivarium to RF Lariviere*, GR Robinson the cages themselves. As evidence, it has been our experience Comparative Medicine, Pfizer inc, Groton CT that unwanted infections from mice injected with contaminated cell lines or mice with previously undetected pathogens are Properly disposing of animal waste is vital to an animal research contained to only those cages, while adjacent static or ventilated vivarium. Normal animal waste and Class II biohazardous microisolation cages on separate experiments remain protected animal wastes are commonly produced in the animal research if proper technique is used. Continuous protection of rodents environment. These wastes consist of soiled bedding and ani- from infection requires strict adherence to maintaining sterility mal feces and urine, and are typically in polycarbonate animal of cages and supplies. Currently the only methods to detect poor boxes. Normal conventional animal waste is removed by dump- cage handling technique involve fluorescein powder or other ing the box contents into a vacuum bedding dump station. The chemical tracers and expensive equipment to assay samples, waste is vacuumed directly into a specialty dumpster for in Abstracts of scientific papers 2008 AALAS National Meeting state incineration. Biohazardous waste must be autoclaved in for service charges to investigators as activity coordinates with a vacuum-type sterilizer/autoclave. Once the boxes have been normal husbandry routines. sterilized, the waste is hand scraped under a protective hood and deposited into a biohazard cardboard box that is lined with 2 red P114 Use of Flashlights in Old World Nonhuman Primate biohazard bags which have a weight limit of no more than 40 Health Monitoring lbs. The biohazard boxes are then sent to a Midwestern state for disposition. The costs associated with disposal of these wastes S Savane* were significant enough to thoroughly investigate our standard SoBran, Bethesda, MD; NIH/DVR, Bethesda, MD operating procedures and determine if a change could be made. Health monitoring is a critical element in managing a nonhuman Our purpose was to determine if there would be cost savings in primate facility. Effective monitoring ensures timely identifica- diverting the sterilized biohazardous waste into the normal waste tion and reporting of animals needing veterinary care. Personnel stream. After working directly with the Environmental Health performing health checks monitor animals for illness, injuries, and Safety Department, it was determined that the biohazardous or abnormalities twice daily. This task can be challenging in an waste could in fact be diverted into the regular waste stream if environment where nonhuman primates are housed using quad- certain procedures were followed. The autoclave must be tested cage units. The Guide for the Care and Use of Laboratory Animals after every 40 h of service, the autoclave must reach 270 °F (130 recommends light at the cage level should be approximately 325 °C) at 27 lbs/in.2 gauge pressure for no less than 45 min, and it lux at 1 m above the floor, which may not provide sufficient light must be validated. With all these requirements being met, the for adequate observation of animals in the lower-level cages of autoclaved animal bedding is now dumped directly into the quad-cage units. Additionally, while some animals come to the vacuum bedding dump station along with the normal animal front of the cage during observations, others tend to hide in the waste. The result is that there is a 92.5% reduction in cost as well back of the cage where light transmission is significantly reduced. as 7 to 10 working h saved per work week. Bringing these animals to the front of the cage using a squeeze bar can be very stressful for the animal and can potentially interfere P113 Humane, Efficient, Safe, Cost-effective Rodent Euthanasia with the observation of the animal and the physical environment. Chamber Facility management evaluated the use of flashlights to observe S Gibson*, S Berry, B Lawless, M Tengco, M Leblanc, J Reuter animals in lower-level cages to determine its usefulness in al- lowing animal care staff to observe animals without the use of Animal Resources Department, The Salk Institute, La Jolla, CA squeeze bars. Tests were performed to determine the length of Population management in modern vivaria requires efficient and time required for care staff using flashlights compared to those humane methods for euthanizing large quantities of rodents. not using flashlights to find 5 ml of an experimental substance Unique solutions such as modified ventilated rack systems, placed in the cage. Results showed that the length of time to refrigerator units, or expensive commercial devices have ef- find and report the experimental substance by staff not using fectively used CO2 to meet these requirements. While greatly flashlights exceeded, by as much as 24 h, the time required for streamlining operations, each is with limitations including large those using flashlights. Our conclusion is that the use of flash- expense, obscured views, or obligation to servicing contracts. Our lights in performing daily health checks in a nonhuman primate facility has historically used single-cage CO2 euthanasia, which is facility enhances the quality of observation and the detection of time consuming, labor intensive, and cost-inefficient. We sought potential health problems. an affordable in-house method to address growing demands while optimizing cost and labor and without compromising P115 Optimizing Timed Pregnancies in Genetically Engineered animal well-being or staff safety. Working with institutional Mice facilities, we designed a novel, easy-to-maintain, transparent acrylic euthanasia chamber (17 by 27 by 33 in.) that has a total SL Mader, N Libal, R Yang, SJ Murphy* capacity of 16 standard mouse shoe box cages. It contains 2 Anesthesiology and Peri-Operative Medicine, Oregon Health rows separated by a barred shelf to allow free flow of gases, and Science University, Portland, OR and closes with an easy access doubled-latched hinge door. It’s Many departments are developing mouse breeding cores to supply equipped with a 0.5-in. CO2 entry valve, a 1.5-in. air intake valve, researchers with genetically engineered mice (GEMs). Such cores a 0.25-in. exit vent port, and a 1.5-in. exhaust valve connected to can provide other services like timed pregnant GEMs. Due to the house exhaust. The chamber is directly connected to house CO2 variety of GEMs, strategies are needed to optimize GEM-specific lines with a regulating valve, eliminating the need for portable timed pregnant protocols. Our study objective was to evaluate preg- tanks. We installed a wall-mounted CO2 room monitor/alarm nancy indices (plug rate, pregnant plugged rate, overall pregnancy for added safety. The chamber minimizes animal distress by rate, litter number and size, body weight) between a homozygous reducing animal handling, using home caging, and producing soluble epoxide hydrolase knockout (KO, n = 84 females) and a L7- rapid loss of consciousness through a steady flow of CO2. The tau-green fluorescent protein transgenic (TG, n = 69 females) mouse materials cost approximately $800. Development challenges colony in order to optimize our timed-pregnant protocols. Estrous included achieving steady CO2 dispersal, moderation of force- cycles were synchronized by exposing group-housed females to ful house exhaust, identifying port entry locations, and safety male urine-soaked bedding for 4 d, thus taking advantage of the assessment. Variables effecting cost included material, chamber Whitten effect. Timed matings were set up by placing 2 synchro- dimensions, and availability of house CO2 lines. Maintenance is nized females with a male for 48 h. Females were checked daily minimal and includes standardized cleaning and decontamina- for the presence of a vaginal plug while males were present. The tion procedures with a chlorine dioxide-based sterilant followed day a plug is found was considered to be embryonic day 1 (E1). by ethanol. Modification of work flow and husbandry practices, We weighed females upon removal of males and then weekly for 2 in conjunction with the new chamber, increased our capacity (16 wk to confirm pregnancy. Plug rates were 68% and 26%, pregnant cages), reduced required effort (10 technicians reduced to 1) and plugged rates were 49% and 78%, and overall pregnancy rates were time (10 h/d decreased to 2 h/d), and has eliminated the need 39% and 49%, respectively, for the KO and TG colonies. Average Abstracts of scientific papers 2008 AALAS National Meeting litter size for 1st (4.9 ± 0.3, n = 30) and 2nd litter (5.0 ± 1.4, n = 2) was 1Research Animal Resources, 2Research Animal Resources, Johns similar for the KO colony but was increased for the TG colony (1st Hopkins University, Baltimore, MD litter, 6.5 ± 0.3, n = 27; 2nd litter 9.6 ± 0.8, n = 7). In the KO colony, This abstract outlines the transformation of investigator-main- pregnancy (n = 33 mice) increased % baseline body weight at E7 tained satellite facilities at our facility. The chain of events that (105 ± 1%) and E14 (126 ± 1%) compared to nonpregnant mice (n = led to the transformation, the key persons that contributed to the 51; E7, 102 ± 0%; E14, 103 ± 1%). Pregnant mice (n = 34) in the TG transformation, and an outline of the actions taken to complete colony also showed an increase in % baseline body weight at E7 the transformation will be described in detail. The university (108 ± 1%) and E14 (142 ± 2%) compared to nonpregnant mice (n was confronted with the need to address inconsistencies in the = 35; E7 and E14, 102 ± 1%). Based on plug and pregnant plugged animal care practices that occurred among some of the satellite rates, we will refine our protocols by limiting male exposure to 24 animal housing facilities in our program. Key personnel within to 48 h for timing pregnancies and using weight measurements at Research Animal Resources (RAR) and the Animal Care and Use least 1 wk after male removal for early pregnancy confirmation, as Committee (ACUC) were able to devise policy and procedures plug detection was not a reliable pregnancy indicator in our KO and to improve the satellite facilities’ operation and function. These TG colonies. Further refinements are planned based on litter size policies and procedures were put in place to meet the standards and number such as using nulliparous and monoparous females outlined in the Guide for the Care and Use of Laboratory Animals, in our KO and TG colonies, respectively. as well as provide a means for maintaining these standards. To explore these events, we visited these satellite facilities and P116 Can Continuous Harem Breeding Be Successful in Small investigated the role that they played in this transformation. We Shoebox Cages and Still Meet Regulatory Expectations? examined them before and after the transformation and analyzed N Libal, SL Mader, R Yang, SJ Murphy* what it took to complete this change. The policy and standard operation procedures that were created specifically for this Anesthesiology and Peri-Operative Medicine, Oregon Health program were also explored in detail. Lastly, we explained the and Science University, Portland, OR current procedures and plans that have been devised to main- A continuous harem breeding strategy would exceed the recom- tain these standards as a long-term goal. With the cooperation mended space for group-housed mice when litters are present in a of investigators and staff, we were able to make a vast number small shoebox cage with a breeding harem. However, regulations of improvements to the satellite facilities’ operations. We were do allow space allocations to be modified to address issues like able to reduce the number of satellite facilities by moving some postnatal care and group housing of social species likely to share activities to central facilities, a clean/dirty exchange has been space. We received IACUC approval for a housing exception to created between labs and central facilities, investigators have allow continuous harem breeding in small shoebox cages (total corrected maintenance and cleanliness problems, and animal floor space 67.6 in.2 [436 cm2]) for departmental mouse breeding housing has been designated to single specific locations in the colonies so as to improve postnatal care and litter survivability. labs. Through such actions, as well as the dedication of our in- Data was collected for 10 mo using a customized breeding colony vestigators, we have enhanced the animal care and use program database from the following 6 knockout colonies: Akt1, soluble at the investigator-maintained satellite facilities. epoxide hydrolase, P450 aromatase, estrogen receptor α and/ or β. We evaluated colony performance indices (litter number/ P118 Use of Soft Hydrothermal Processing to Improve and harem, litter size, preweaning losses, incidence of death and Recycle Bedding for Laboratory Rodents euthanasia for female breeders, health and behavioral problems) as recommended by the Guide for each colony and overall to T Miyamoto*1,2, N Kasai1, Z Li2, T Kibushi1, N Yamasaki2 ensure the adequacy of continuously housing breeding harems 1 Institute for Animal Experimentation, Graduate School of and their litters in our facilities cages. Litter number/harem Medicine, Tohoku University, Sendai, Japan; 2 Graduate School was 5.6 ± 0.2 (n = 198 harems). Litter size was 6.8 ± 0.1 (n = of Environmental Studies, Tohoku University, Sendai, Japan 1103 litters). Preweaning pup loss was 28.0 ± 4.1%. Incidence Bedding is one of the common means of improving well-being of euthanasia and death for female breeders (n = 385) was 8.3 ± and the environment of laboratory animals. It also influences the 2.5%. Common health problems seen in female breeders were accuracy of the experimental results. In addition, a large amount dystocia (2.2 ± 1.0%), uterine prolapse (0.5 ± 0.4%), animals found of used bedding containing excrement is produced as the medical dead of undetermined cause (1.7 ± 1.0%), and skin problems waste from animal facilities of life science institutes. On the other (1.3 ± 0.9%). No fighting was observed in any harem cage. Litter hand, the high temperature and pressured dry steam has potential number/harem, litter size, and preweaning losses in individual for promoting organic reactions, such as extraction and drying knockout colonies are comparable to what is reported for these with soft hydrothermal processing. We developed a groundbreak- colonies in the literature or anecdotally by colleagues using pair ing system to improve fresh bedding and to recycle used bedding breeding or removal of pregnant mice from harem cages. Colony by applying a soft hydrothermal processing. The system removes performance indices for all colonies collectively are similar to or both harmful organic components and aromatic hydrocarbons at the high end of that observed in other facilities which do not that can affect animals’ metabolism. The purpose of the present use continuous harem breeding for genetically engineered mice. study is to evaluate the effects of soft hydrothermal processing on Our individual and collective colony performance indices and the beddings of laboratory rodents. Soft hydrothermal processing the lack of fighting in harem cages suggest that in our continu- removed 68% to 99% of the predominant aromatic hydrocarbons ous harem breeding system, neonatal and adult mice housed in in softwood chip bedding (spruce) treated at 0.35 MPa and 140 °C small shoebox cages have adequate space to address reproduc- for 120 min (“improved bedding”). In addition, 59.4% to 99.0% of tive, social, and individual needs. predominant harmful organic compounds derived from excrement were also removed in used softwood chip bedding (spruce) treated P117 Transformation of Investigator-maintained Satellite at 0.45 MPa and 150 °C for 60 min (“recycled bedding”). The soft Facilities hydrothermal treatment increased the number of acidic functional SP Williams*1,2, L Barnes1,2 groups on the bedding surface, thus increasing its adsorption Abstracts of scientific papers 2008 AALAS National Meeting of ammonia gas. Harmful substances such as microorganisms, consider when identifying personality traits, which include gen- heavy metals, and pesticides decreased below the detection limit. der, upbringing, education, and health. The temperament theory Then we examined effects of reproduction with these beddings in is not an exact science; however, when the staff at our facility mice. Body weight of male and female mice and reproductive data considered all the variables including each individual’s inherited (litter size and total number of offspring) showed no significant temperament, they had a good perspective of themselves and differences among different bedding groups. The results showed the people they work with. By creating an optimal atmosphere that the improved and recycled bedding are safer for laboratory in the workplace, the manager will be able to generate more rodents and has the potential to ameliorate conditions in primary teamwork and attain more productivity. and secondary enclosures (such as cages and animal rooms) used for maintaining laboratory animals. This process has the potential P121 Light Intensity Comparisons between the Macroenvi- to be one of the most advanced techniques in improving softwood ronment and Microenvironment of Various Caging Systems bedding, economizing through the recycling of used bedding and Housing Albino Laboratory Animal Species reducing by 30% the bedding waste from animal facilities. VT Lea*, SJ Young, AE Kraycirik P119 Rodent Bedding Comparison Study LASW, sanofi-aventis, Bridgewater, NJ T Putro*, PA Straeter According to the Guide for the Care and Use of Laboratory Animals, light can affect the physiology, morphology, and behavior of Veterinary Sciences, Bristol-Myers Squibb, Princeton, NJ animals used in research. Potential photostressors include in- A large percentage of the operating expenses accrued in maintain- appropriate photoperiod, photointensity, and spectral quality ing a rodent facility involves animal bedding. The predominant of light. During an AAALAC site visit, the site visitors recom- bedding used at our institution is an alpha cellulose material mended that we reevaluate light intensity levels in some of our that provides favorable benefits such as defined material com- animal holding rooms. We conducted extensive testing of the position and dust-free, sanitary environments for the rodents light intensity levels within cages housing albino rabbits, guinea and personnel. However, the alpha cellulose material is more pigs, mice and rats. All intensity levels were measured using the expensive than other bedding, which increases expenditures. Extech Instruments Model #401025 foot candle/lux meter. An ini- Other bedding alternatives, such as corn cob or a bedding blend, tial lighting intensity reading was taken in the center of an empty may be more affordable, but potentially compromise the health of animal room at approximately 1 m off the floor. Multiple types the rodent colony and possibly the integrity of the research data. of caging were tested, as were multiple cage positions through- As a result, a study was designed and executed to compare the out the room. Different types of bedding were also evaluated to microbial analysis of pre- and postautoclave bedding of 3 types: determine if bedding impacted light intensity. Measurements alpha cellulose, corn cob, and an alpha cellulose-corn cob 50/50 were taken at various positions on the rack in the microenvi- blend. Eighteen samples (9 preautoclave and 9 autoclaved), along ronment and the macroenvironment. Results were compared to with 6 control samples were evaluated. Microbial cultures were the recommended range of 130 to 325 lux at cage level, which analyzed from a 10-g homogenized slurry of each bedding type, is sufficient for animal care and does not cause clinical signs of and organism counts and identifications were performed. Results phototoxic retinopathy in albino rats. During our testing, we of the preautoclave samples indicated the corn cob bedding had found no significant changes in readings with the different types the highest quantity of bacterial and fungal organisms, while the of bedding used; however, cage positioning had a positive affect alpha cellulose bedding presented with the least. All autoclaved on the readings. Moving the cage away from direct light reduced samples yielded no microorganisms. the intensity readings. One alternative to help regulate light in- tensities in animal rooms is to rotate cage positions away from P120 Comparative Analysis of Diversity in Temperaments direct light. Data collected outside the acceptable ranges were Commonly Associated with Animal Technician Models reviewed by management. Results of this study led to corrective actions to provide a higher-quality environment for the animals VR Riojas*, JA Kundert, TK Marcotte, RE Arens, TS Frerck, DW used in drug discovery research. Harn, CM O’Rourke Animal Resources Center, Montana State University, Bozeman, P122 Effect of Directed Changes to Business Processes on Time MT to Approval for Animal Use Protocols Maintaining a team atmosphere in an animal facility with a large W Moseley*1, DM Ostmeyer2, L Martin1, J Owiny2 or diverse staff can seem daunting. The typical challenges associ- 1Research Integrity & Compliance Review, 2Vice President for ated with this type of situation can be simplified by identifying Research, Colorado State University, Fort Collins, CO each technician’s personality traits and innate characteristics. We have seen dramatic changes in our staff meetings by educating Institutional Animal Care and Use Committees (IACUCs) are ourselves on the temperaments of others in order to understand charged with oversight of the animal care program. Part of that them better. We each took a personality test based on Hippo- role involves review of proposed animal activities. Virtually all crate’s 4 basic temperaments. The Sanguine is charming, but can institutions have developed forms and processes to aid review be unproductive. The Choleric is very productive, but tends to be of animal protocols. Based on perceived problem areas, our intense and unsympathetic. Melancholies are compassionate, but IACUC and administrative office have instituted changes to moody. The Phlegmatic temperament is cool, calm, and collected, protocol application forms and review processes in a phased but can be too leisurely. Each individual possesses a blend of all manner over the past 12 mo. The changes include the addition of 4 of these traits. For this study, fabricated people conducting a university veterinarian prereview of select protocols, extensive staff meeting as models to distinguish the differences in these administrative prereview of all protocols prior to committee temperaments and identify the strengths and weaknesses of review, incremental changes to the A-100, and additional PI each were used. It is important to point out that each subject training. The goal of this study was to determine whether the is an individual and completely unique. There are variables to changes instituted over the past year have reduced the time Abstracts of scientific papers 2008 AALAS National Meeting between submission of animal use protocols and final approval. 1CCM, Massachusetts General Hospital, Charlestown, MA; 2 Using 2006 as a baseline, we compared the number of protocols Digital Paradigms, Inc, Boston, MA submitted in 2006, 2007, and the first 4 mo of 2008 to determine Per diem charges for husbandry services is the most common if our incremental changes have had any effect on protocol means of recovering lab animal care costs in academic research turnaround time. In 2006, a total of 267 protocols were reviewed institutions. Many institutions are using bar codes and scanning and approved; 219 protocols (82%) were reviewed and approved technology to more efficiently label and track cages for per diem within 30 d, while 48 (18%) took longer than 30 d to review and billing as well as for animal health, breeding, and regulatory approve. In 2007, a total of 270 protocols were reviewed and ap- compliance purposes. As lab animal care becomes more sophisti- proved; 233 (86%) of those protocols were approved within 30 d, cated and expensive while federal funding of biomedical research and 37 protocols (14%) took longer than 30 d to approve. During declines in real dollars, maximizing cost recovery through per the first 4 mo of 2008, 112 protocols were reviewed, and 97 (87%) diem revenues becomes even more important. Historically, were approved within 30 d, while 15 protocols (13%) took more our department at MGH performed cage census on Mondays, than 30 d to approve. Additionally, the mean number of days to Wednesdays, and Fridays. For billing purposes, the system was approval for each of the 3 periods has decreased from 18.49 to programmed to “fill in” for the non-census days if a cage was 14.62, to 14.22 d. Although it appears from the data on the first scanned on the 1st and the 3rd d. Taking census more frequently 4 mo of 2008 that we may have reached a point of diminishing was presumed to be an additional and costly labor expense, with returns, the overall data indicate that the incremental changes no net revenue gain. Because of the dynamic census in some of in the application form and review processes implemented thus our animal facilities, in which animals are coming and going on far resulted in improved time to approval. a daily basis, this presumption was tested. Rodent cages in one facility were scanned every weekday for 1 mo and the resultant P123 Comparison of Two Hydration Gel Products for Use in daily census and revenue were calculated. The same data were Shipping of Guinea Pigs (Cavia porcellus) then used to simulate what the census and per diem revenue Y Luo*, GB Mulder, RR Morin would have been using the traditional 3-d approach. Performing census 5 d per wk increased monthly per diem revenue 3%. On Charles River, Raleigh, NC an annual basis at our institution, this extrapolated to a $225,000 Guinea pigs are generally considered to adapt poorly to changes in increase in revenue for the department. Bar code scanning for food presentation and to novel food items. During transportation, census averages 34 cages per min, so the resultant additional it is common to provide animals with water replacement gels to labor cost for scanning cages an additional 2 d per wk consumed aid hydration. The goal of this study was to investigate preference only 10% of the additional revenue gained. This yielded a return of guinea pigs for 2 different water replacement gels. The primary on labor of 900%, creating a low cost:high yield improvement to difference between the products was the presence or absence of increase animal care revenues simply. high-fructose corn syrup (HFCS). This study also evaluated the effect of the sweetener ingredient on palatability to this specie. P125 Characterization of Western Equine Encephalitis Virus in Fifty-two male Hartley guinea pigs (Crl:HA) weighing 330 to 400 the Syrian Golden Hamster (Mesocricetus auratus) g were used. While 10 of the animals served as control for clinical chemistry and CBC, the other 36 guinea pigs were divided into 2 AL Olsen*1,2, J Morrey2 groups of 18 animals. Each group was supplied with either sweet- 1Laboratory Animal Research Center, 2Animal, Dairy, and Vet- ened or unsweetened gel product and transported within filtered erinary Sciences, Utah State University, Logan, UT crates overnight to a remote facility and subsequently held within It was hypothesized that intraperitoneal inoculation of Syrian the shipping crates for an additional 3 d. Assessment of animals Golden hamsters with Western Equine Encephalitis Virus (WEEV) after arrival included clinical chemistry, CBC, and body weights. did not routinely cause viral encephalitis, but rather resulted in In the group supplied unsweetened gel, animals consumed 99.5% a nonneuronal systemic disease, similar to a disease phenotype of supplied gel (124 g/d/animal). In contrast, the consumption reported in humans infected with similar alphaviruses. It was of HFCS-sweetened gel was 63.6% (82 g/d/animal). The average further hypothesized that virus infection produced secondary body weight losses were 2.7% and 23% for the unsweetened and effects, such as overwhelming systemic inflammation, which con- sweetened gels groups, respectively. The values for hematocrit, tributed to the high mortality and rapid death observed. Finally, and serum sodium, chloride, total protein, and albumin were sig- it was hypothesized that WEEV retained the ability to induce nificantly higher (P < 0.05) in the animals supplied sweetened gel viral encephalitis if animals survived the systemic disease, or if as compared to the animals provided unsweetened gel. Follow-up the virus was introduced directly into the central nervous system preference testing was performed by providing 6 singly housed (CNS). Hamsters inoculated intraperitoneally with WEEV exhibit male guinea pigs both water and HFCS-flavored water and moni- a unique disease phenotype as compared to that reported for mice toring the consumption of both fluids for 5 d. Animals consumed inoculated with WEEV. Hamsters displayed increased sensitivity approximately 20% more unflavored water than water sweetened to WEEV, showing high rates of mortality and death by 96 h post- with HFCS. The results of this study indicated that Crl:HA guinea virus inoculation (hpi), even when inoculated with relatively low pigs provided an unsweetened hydration gel consumed greater doses of virus. Furthermore, hamsters did not display overt signs amounts of gel, showed reduced body weight loss, and decreased of neurological disease. Experimental results found that splenic clinical pathologic indicators of dehydration than animals pro- necrosis was a consistent pathological finding in WEEV-inoculated vided a HFCS-sweetened gel product. In addition, the apparent hamsters, with an absence of neuropathological changes prior to preference for the unsweetened gel over the sweetened gel may death at 96 hpi. A potential role for the inflammatory response in have been due to decreased preference, or aversion, for HFCS. the disease process was shown by increased serum concentrations of tumor necrosis factor-alpha in WEEV-inoculated animals, and P124 Maximizing Per Diem Revenues through More Frequent significantly improved survival when virus-inoculated animals Cage Census received anti-inflammatory therapy. Evidence for the encephalitic Y Sun*1, M Moore2, M Depathy1, J Joncas1 capability of WEEV in hamsters was provided by the observation Abstracts of scientific papers 2008 AALAS National Meeting that a portion of those animals that survived beyond 96 hpi due at 0 min to 162.1 ± 14.1 mg/dl at 48 h) and isoflurane (from 72.29 to anti-inflammatory therapy displayed overt signs of neurologic ± 7.83 to 133.3 ± 17.5 mg/dl at 60 min). Both anesthetics increase disease and histopathological evidence of CNS necrosis. Intro- serum glucocorticoids and catecholamines and contribute to fat duction of WEEV directly into the CNS resulted in rapid death mobilization. However, decreases of serum cholesterol concentra- and similar neuropathological lesions. Inoculation of WEEV into tions in the group H (from 66.9 ± 5.78 to 40.4 ± 6.6 mg/dl at 24 h) Syrian Golden hamsters provides a unique disease phenotype may result from production of free radicals during biotransfor- which mimics certain characteristics of related alphavirus infec- mation of halothane. A decrease in serum albumine (from 3.88 ± tions in humans. 0.13 to 3.2 ± 0.15 g/dl at 72 h) and total protein levels (from 5.96 ± 0.12 to 4.9 ± 0.2 g/dl at 72 h) occurred in the group H. It has P126 Rabbit Model of Alzheimer’s Disease: Valid from Neu- been shown that halothane appeared to exert inhibitory effects ropathological to Cognitive Levels on synthesis of albumine and total proteins. Serum potassium levels significantly decreased in both groups (4.2 ± 0.3 mmol/L A Agelan*1, L Del Valle3, L Bright1, M April1, DS Woodruff- at 72 h in group H and 3.9 ± 0.2 mmol/l at 60 min in group I). Pak2 There was a significant decrease in serum calcium levels in the Laboratory Resources, 2Psychology, 3Neuroviology, 1University group I (12.3 ± 0.6 mg/dl at 72 h) and a significant increase in Temple University, Philadelphia, PA serum phosphorus (from 4.69 ± 0.23 to 7.6 ± 0.8 mg/dl at 120 min) and chloride concentrations (from 105.32 ± 1.33 to 110.3 ± Supplementing a rabbit’s diet with 2% cholesterol alone or with 1 mmol/l at 1 min), probably due to a transient reduced renal a trace amount of copper created neuropathological changes that function immediately after isoflurane. Caution is required in resembled those seen in Alzheimer disease (AD). AD model rab- interpreting data on serum biochemical parameters from rabbits bits were impaired in eyeblink classical conditioning, a form of anesthetized with halothane or isoflurane. learning severely impaired in AD. Our aim was to replicate AD rabbit model neuropathology, test eyeblink conditioning in this model, and determine if galantamine would ameliorate impaired P128 Heart and Respiratory Rate, Rectal Temperature, and conditioning. Rabbit chow with 2% cholesterol and drinking Glucocorticoid Concentrations after Ketamine-medetomidine water with 0.12 mg/l copper sulfate were administered for 10 Administration in New Zealand White Rabbits (Oryctolagus wk. Control rabbits received normal food and water. Rabbit cuniculus) brains were probed for neuropathology. AD model rabbits had M Martínez-Mateos1, A Gonzalez Gil*1, P Lorenzo García1, C significant neuronal loss in frontal cortex, hippocampus and cer- Pérez García2, I Díez Prieto2, J Illera1 ebellum. Changes in neurons in the hippocampus were consistent 1Fisiología Animal, Universidad Complutense de Madrid, Facul- with neurofibrillary degeneration and cytoplasmic immunore- tad de Veterinaria, Madrid, Spain; 2Patología Animal (Medicina activity for beta-amyloid and tau. Next, AD model rabbits were Veterinaria), Universidad de León, Facultad de Veterinaria, injected daily with vehicle or 3.0 mg/kg galantamine and tested León, Spain on 750 ms trace and delay eyeblink conditioning. Galantamine improved eyeblink conditioning significantly over vehicle. The The objective of this investigation was to determine the effects of AD rabbit model has validity from neuropathological to cognitive the anesthetic combination ketamine-medetomidine on heart and levels and offers a promising addition to the available animal respiratory rate, rectal temperature, and serum glucocorticoid models of AD. Galantamine ameliorated impaired eyeblink concentrations in New Zealand White rabbits. All the rabbits conditioning, extending the validity of the AD rabbit model to were clinically healthy prior the study. Ten animals maintained treatment modalities. under conventional conditions (12:12 light:dark cycle; 20 to 22 °C; 50% to 55% relative humidity; 10 to 15 air changes/h) were P127 Clinical Biochemical Response after Inhalant Anesthetics anesthetized with ketamine (15 mg/kg, IM) and medetomidine in New Zealand White Rabbits (Oryctolagus cuniculus) (0.25 mg/kg, IM). Blood samples were obtained from the central ear artery using 23-gauge needles at 6 time points: before injec- A Gonzalez Gil*1, G Silván1, C Pérez García2, I Díez Prieto2, A tion (0 min) and 10, 30, 60, and 120 min and 24 h postinjection Martínez3, J Illera1 of the anesthetics. The Departmenth of anesthesia was moni- 1Fisiología Animal, Universidad Complutense de Madrid, Facul- tored by using the pedal withdrawal, ear pinch, and righting tad de Veterinaria, Madrid, Spain; 2Patología Animal (Medicina reflexes. Serum cortisol and corticosterone concentrations were Veterinaria), Universidad de León, Facultad de Veterinaria, León, measured by competitive enzyme inmmunoassay. An adequate Spain; 3 GlaxoSmithKline R&D Spain, Madrid, Spain level of surgical anesthesia was observed for about 1 h, with no reaction to pedal and ear pinching. Heart rate was significantly The effects of anesthetics on biochemical parameters can confuse decreased at 10 to 120 min when compared to baseline (time 0) the interpretation of laboratory results. In order to characterize rate (from 255 ± 9.14 at 0 min to 197 ± 5 beats/min at 120 min) the variations of selected biochemical parameters after inhalant and respiratory rate decreased at 10 to 120 min (from 109.25 ± anesthetics, 20 New Zealand White rabbits were assigned to 2 8.73 at 0 min to 60 ± 5.88 breaths/min at 30 min), probably due to groups (n = 10): group H (halothane) and group I (isoflurane). The the medetomidine component of the mixture. It has been shown induction of the anesthesia was achieved by a face mask (3.5% that medetomidine is a respiratory and cardiovascular depres- halothane and 4.5 % isoflurane in oxygen) and were intubated sant. Rectal temperature was significantly decreased at 120 min for the maintenance of the anesthesia for 30 min (1.5% halothane when compared with the initial levels (from 38.95 ± 0.15 to 37.27 and 2.5% isoflurane in oxygen). Serum cholesterol, triglycerides, ± 0.24 °C). Serum cortisol concentrations increased at 10, 30 and albumine, total bilirubin, sodium, potasium, chloride, calcium, 120 min when compared to baseline concentrations (from 3.09 ± and phosphorus concentrations were measured by the Hitachi 0.73 to 16.91 ± 3.58 ng/ml at 10 min). It is probably that ketamine 747 autoanalyzer. Blood samples (2 ml) were obtained from the produced the inhibition of GABA receptors and consequently central ear artery before induction, 1, 10, and 30 min and 1, 2, the release of ACTH and cortisol. Although a small increase was 24, 48, and 72 h after intubation. Significant increases in serum observed at 10 to 30 min, serum corticosterone concentrations triglycerides levels occurred after halothane (from 88.94 ± 13.78 Abstracts of scientific papers 2008 AALAS National Meeting did not significantly change after injection of anesthesia when artery catheter for sampling. Following 2 recovery d, mice were compared with the baseline levels. The ketamine-medetomidine placed on an automated blood sampler with a unique movement- anesthetic combination provided an adequate level of surgical responsive caging system. Concurrently, the stylet was removed anesthesia and increase cortisol but not corticosterone secretion. from the implanted guide and replaced with a brain microdialysis Respiratory and cardiac depression effects have to be considered probe perfused with artificial CSF at a rate of 1.5 µl/min. Follow- for any experimental procedure. ing a 24-h acclimation period, 2 predose dialysate samples were collected. The mice were dosed with 1 mg/kg of carbamazepine P129 Canine Model of Relatively Sustained Atrial Fibrillation into the jugular vein catheter. Nine blood samples (5 µl) were Induced by Rapid Atrial Pacing and Phenylephrine collected from each mouse over 4 h, as well 10 dialysate samples (25 min/sample). Carbamazepine was detected in both the blood A Kijtawornrat*1,2, BM Roche2, RL Hamlin1,2 and dialysate samples. Mice remained healthy throughout the 1TheOhio State University, Columbus, OH; 2 QTest Labs, Co- study, demonstrating the efficacy of the surgical method. This lumbus, OH combination of sampling techniques within a single mouse serves as a useful model for obtaining multiple streams of information Atrial fibrillation is a common arrhythmia with important mor- while reducing interanimal variability. bidity and mortality. Limitations in studying both mechanisms and therapy arise for paucity of models that possess a high yield, are not costly, and are sustained long enough to make the P131 Effects of Doxorubicin on Renal Function in the Gerbil necessary observations. This model is based upon the thought (Meriones unguiculatus) that atrial fibrillation requires heterogeneity of repolarization, M Peña1, I Diez1, M García1, MJ Cano1, M Ríos1, AM Montes2, C that distribution of vagal fibers is heterogeneous in the atria, Gutiérrez2, CC Perez*1 and that when stimulated by elevation of systemic arterial 1Departamento de Medicina, Cirugía y Anatomía Veterinaria, pressure, atrial fibrillation will persist. We postulated that rapid atrial pacing together with phenylephrine infusion will induce Universidad de León, León, Spain; 2Departamento de Medicina relatively sustained atrial fibrillation for at least 40 min in dogs. y Cirugía Animal, Universidad de Murcia, Murcia, Spain Dogs were anesthetized with morphine-chloralose because this Doxorubicin administration in rats is a recognized model of the permits nearly intact autonomic control. Systemic arterial pres- nephrotic syndrome observed in humans, while administration sure was elevated approximately 75 mmHg and mean heart in the gerbil leads to a more moderate proteinuria in the 1st wk, rate was decreased approximately 9 beats/min with a constant which tends to settle down from d 21. The aim of this work was to infusion of phenylephrine, 2 μg/kg/min. The right atrium was establish the effect of doxorubicin administration on the different paced for 20 min at 40 Hz. Atrial fibrillation was sustained in renal function parameters in the gerbil (Meriones unguiculatus) in dogs receiving phenylephrine but terminated within seconds in a long-term study. Twenty conventionally housed gerbils, aged normotensives. In dogs receiving phenylephrine infusion, 50% 4 mo (10 males and 10 females), were used. They were housed of the dogs maintained atrial fibrillation for > 40 min, and 50% in the conventional area of the Central Laboratory Animal Facili- returned quickly (<19 s) to sinus rhythm. In conclusion, atrial ties of the University of León. A control group (n = 10) and an fibrillation may be maintained for at least 40 min following ces- experimental group (n = 10) were organized, including 5 males sation of rapid atrial pacing in dogs with hypertension produced and 5 females in both groups. In the animals belonging to the by phenylephrine infusion. experimental group, doxorubicin (a single dose of 4 mg/kg) was injected intravenously into the femoral vein. Blood and 24-h urine P130 Combinatorial Pharmacology in Mice: Integration of (collected from a metabolic cage) were obtained. Plasma calcium, Brain Microdialysis and Automated Blood Collection in a phosphorus, magnesium, urea, creatinine, and total proteins and Single Subject urinary calcium, phosphorus, magnesium, creatinine, and protein excretion were measured. Animals were sampled at 0, 14, 28, 42 C Rohde*1, R Guinn1, R DeGraw1, SN Katner1, BP Solomon2 and 56 d. Plasma creatinine remained stable, and plasma urea 1In Vivo Research, 2Bioanalytical Research, Bioanalytical Systems, increased only in the final sampling. Plasma phosphorus increased Inc, West Lafayette, IN from the second sampling, but its fractional excretion remained stable. While hypocalcemia was observed only in the final sam- In recent years, mice have become an extremely attractive model pling, calcium fractional excretion was high since d 28. Magnesium in drug research. Their small size makes them suitable candidates remained stable throughout the study, but its fractional excretion for early discovery studies where very little drug is available, and increased from d 42. Normoproteinemia was observed, but 24-h the ability to genetically modify mice adds to their appeal in the urine protein excretion increased in the final sampling. These research community. An additional trend in drug discovery has data reveal that the Mongolian gerbil is less sensitive than the rat been a major focus on central nervous system (CNS) active drugs. to the administration of doxorubicin and that the mechanisms of For this drug type, it is useful to look at drug concentration and renal adaptation are able to maintain the renal function during a effect in both the blood (systemic levels) and the brain (target reasonable period in this species. tissue levels). The ability to obtain both types of data from a single mouse will reduce the number of subjects needed for such research and improve the quality of the data by reducing inter- P132 NIH/rnu: Are They All Created Equal? A Comparison of animal variability. This study sought to develop a surgical model Taconic and Charles River Rat Strains that allowed the simultaneous collection of blood and brain CJ Andresen* dialysate samples in individual mice following an intravenous Comparative Medicine, Pfizer, Groton, CT dose of carbamazepine. Eight male CD-1 mice were anesthetized with an intraperitoneal dose of ketamine/medetomidine and Preclinical oncology research relies on nude rodent xenograft implanted with a guide cannula in the caudate putamen. After models, where human tumor cells are implanted in nude mice 4 d of recovery, mice were anesthetized using isoflurane, and or rats. Most commonly used are nude mice; however, using implanted with a jugular vein catheter for dosing and a carotid rats presents some advantages. Historically, our lab has used the Abstracts of scientific papers 2008 AALAS National Meeting NIH/rnu rat strain from 2 suppliers, Taconic (Tac) and Charles for Helicobacter, when compared to fecal pellets stored at 4.0 °C. River (CR), to create xenograft models. Some phenotypic and A comparison of multiple processing techniques demonstrated performance differences were observed between the 2 sources that nucleic acid isolated from the supernatant of low-speed which prompted us to look closer at the history and phenotype centrifuged slurries is an effective method for detecting both of the animals. The athymic hairless mutation in rats was first Helicobacter and MNV from the same sample. This study supports observed in 1953 at the Rowett Research Institute in England. that feces should be shipped and stored at 4.0 °C if used for pre- The National Institutes of Health (NIH) nude rat was developed paring slurries for Helicobacter and MNV PCR testing (although in 1979–1980. Taconic source rats were obtained from the NIH ambient temperature is also acceptable for MNV). However, all in 1981 and were derived by hysterectomy in 1987 and again in evaluated storage temperatures up to 2 mo are adequate when 1998, designation NTac:NIH-Whn. The Charles River source rats feces are placed directly into lysis buffer prior to extraction. were obtained from the NIH in 2001 and cesarean rederived, designation Crl:NIH-Foxn1rnu. The nude rat is athymic and P134 Molecular Characterization of a Newly Indentified Rat T-cell–deficient, with a normal complement of B and NK cells. Parvovirus To test our hypothesis, we performed flow cytometry analy- sis of lymphocyte subsets in 3 strains of 6- to 8-wk-old male C Wan*1, Y Liao1, Y Chang1, M Wang2 rats: Charles River CD-1, Charles River homozygote nude rat 1VeterinaryMedicine, National Taiwan University, Taipei, Tai- Crl:NIH-Foxn1rnu, and Taconic homozygote nude rat NTac: NIH- wan; 2Laboratory Animal Center, Tzu Chi University, Hualien, Whn, and tumor take-rate/growth studies in the 2 nude strains. Taiwan Data demonstrated that normal CD rats have significantly higher One new isolate of rat parvovirus was identified and char- values for NKT, T, CD8+, CD4+, CD25+ and RT1B+ than either acterized by our laboratory. The isolate was obtained from a strain of homozygote nude rats, while both strains of homozy- laboratory rat colony (RPV-1c). In this study, the DNA sequence gote nudes have higher B and NK cell values than normal CD and the predicted viral capsid protein sequence of this isolate rats. CR source nudes have significantly higher NKT, T, CD8a+, were determined and compared to previously characterized CD4+ and RT1B+ levels than Tac rnu, while Tac source nudes rodent parvoviruses. When the DNA sequence of this new RPV have significantly higher NK levels than CR. We also found that isolate was compared to other rodent parvoviruses, RPV-1a was take-rates and growth rates for tested tumor cell lines are higher the most similar to this new RPV with 98% identity. Nucleotide in Taconic source rats. In conclusion, NIH/rnu source rats differ homologies of the new RPV isolate with Kilham rat virus (RV), between vendors, and researchers should consider the applica- H-1 virus, and rat minute virus (RMV) were lower (71% to tion or particular model before choosing a supplier. 72%). The regions of the genome previously determined to be important in transcription, mRNA processing and translation of P133 Evaluation of Feces Shipping Conditions and Sample prototypic rodent parvoviruses were highly conserved between Preprocessing for PCR Detection of Helicobacter and Murine this new isolate and RPV-1a. In summary, the new RPV isolate Norovirus has been identified which are closely related to RPV-1a, but CL Perkins *1, KA Sellitto 1, JL Latchaw 1, AL Kelleher 1, ME distinctly different from other previously characterized rodent Vigliotti2, KS Henderson1 parvoviruses. 1Research Animal Diagnostic Services, 2Genetically Engineered Models and Services, Charles River, Wilmington, MA P135 Alternate Rat Pharmacokinetics Methodology Valida- tion The sensitivity of the polymerase chain reaction (PCR) is often associated with the amplification and detection portion of the C Nunes*1, X Shen1, B Wang1, S Xu2, S Zheng2, J Wang2, I Ca- process; however, detection of an agent may be affected by podanno1, W Ferguson1, BJ Doerning1, LH Cohen2, X Tong2, C shipping conditions or sample processing prior to nucleic acid Johnson1 isolation. Fecal pellets are common samples used for screening 1Lab Animal Resources, 2DMPK, Merck & Co., Rahway, NM rodents for enteric viruses and pathogens. The steps between The evaluation of novel pharmaceutical compounds using rodent sample collection and nucleic acid are not standardized, nor are pharmacokinetics screens is common within pharmaceutical the effects on agent detection clear. We investigated the impact drug discovery. Single- or double-line surgically vascular can- of storage and shipping temperature (–20 °C, 4.0 °C, ambient, nulated rats are routinely used for rodent pharmacokinetics (PK) and 37 °C) over time (2 h, 1 wk, 2 wk, 1 mo, 2 mo) on agent compound screens. Surgical preparation time, non-naïve state detection for dry feces placed directly in lysis buffer. Because of animal, and cost of surgically cannulated animals led us to fecal slurries are often used as a method for pooling samples, evaluate the feasibility of an alternate procedure for intravenous subsequent investigations were used to evaluate the impact of (IV) dosing and blood sample collections from PK rats that temperature and time on slurries and to compare different pre- would reduce our reliance upon surgically prepared animals. processing methods (direct lysis, slurry, and slurry supernatants). This study was conducted to make 2 side-by-side comparisons. Nucleic acid was evaluted by quantitative real-time fluorogenic First, we compared manual jugular vein dosing and tail blood PCR for murine norovirus (MNV) or Helicobacter. When dry sample collection with the current method of IV dosing using feces was added directly to lysis buffer, Helicobacter target copy a cannulated femoral vein and blood collection from a femoral numbers decreased by approximately 80% after 1 wk, regardless artery catheter. Second, we compared PO dosing and tail blood of storage temperature; however, the copy number remained collection with the current method of blood collection using a fairly consistent for the remaining time points. Conversely, the cannulated femoral vein. Compounds were selected for side- results for slurries indicated a 2 log10 decrease in Helicobacter by-side dosing comparisons based on known difference in PK target copy number when stored at –20 °C as compared to 4.0 °C. and physical properties to investigate if the alternate dosing and Results from a follow-up experiment demonstrated that target collection methods offered comparable PK profile data. The rats copy numbers were substantially reduced for both agents when were manually restrained, dosed IV in the jugular vein and per os feces was stored at –20 °C, –70 °C and at ambient temperature (PO) by oral gavage, and whole blood was collected by tail snip. Abstracts of scientific papers 2008 AALAS National Meeting Results of side-by-side studies showed that there was no signifi- with heat-killed ETEC on days 6 and 12 of gestation. Postparturi- cant difference (P > 0.01) in the PK profiles between alternate and tion colostrum samples were collected from each dam’s mammary conventional dosing and collection methods. The estimated cost glands using a milking device especially design for this purpose. savings in the elimination or reduction of cannulated rats in our Blood samples were drawn from each dam’s orbital sinus. The rat facility is significant, with potential reduction of up to 75% of antibody responses to ETEC were analyzed with Western blot and current expenses for vendor-supplied surgically cannulated rats. enzyme immuno-assay (EIA) to determine the specificity and titers In summary, the implementation of alternate PK methodology of antibodies in the colostrum and sera. Whole cell extracts of swine is a significant cost and time savings initiative, but feasibility at ETEC isolates were prepared for electrophoresis in 12.5% SDS PAGE this time is under consideration due to the skill set proficiencies gels. Western blots were probed with sera and colostrum from each required to reliably jugular dose the animals. dam as well as the sera from the offspring. The antibody response in rats challenged with ETEC compared with the antibody response P136 Technique Refinement for Tail Tip Biopsy Procedures for of rats challenged with saline showed an increased specificity to the Genotyping Mice swine ETEC antigens. Antibody titers measured by EIA also indicate a significant increase in the antibody titers in rats challenged with CC Sem*, S L’Italien, G Ma, H Daly, L Jackson ETEC antigens. The Sprague Dawley rat is an efficient laboratory Biogen Idec, Inc., San Diego, CA animal model for the study of the colostral antibody response to ETEC antigens of swine origin. The most commonly used technique for genotyping genetically engineered mice (GEM) is the tail tip biopsy. At many institutions, this procedure is performed by using a sterile blade to amputate P138 Effect of Selenium and Selenoprotein on Brain Function the tip of the tail, and then using direct pressure and electro- in Transgenic Rats cautery or procoagulants to ensure hemostasis. In an attempt to D Hwang*1, J Sin2, S Yim2, C Kim2, B Kim2, S Shim2, S Lee2, C refine these procedures, we used a glass bead sterilizer (275 °F) Bae2, J Woo2, J Cho3, K Chae2 to heat the end of a sterile, #21 scalpel blade for approximately 1College of Natural Resources & Life Science, Pusan National 30 s before amputating the tip of the tail. Our hypothesis was University, Miryang-si, Gyeongsangnam-do, South Korea; 2De- that simultaneously cutting and cauterizing the tissues by this partment of Laboratory Animal Resources, National Institute method would lead to rapid hemostasis, a reduction in tissue of Toxicological Research, Korea FDA, Seoul, South Korea; damage and postprocedural inflammation at the amputation 3Multidisciplinary Technology Center, Hoseo University, Asan, site, and a reduction in the technician time necessary to perform South Korea the procedure. One hundred GEM, 4 to 6 wk of age, were used for this study. All mice were anesthetized with isoflurane. Tail Selenium (Sel) is widely distributed throughout the body, but is tip biopsies were conducted on 50 mice using a sterile, room particularly well maintained in the brain. It has been reported temperature scalpel blade followed by direct pressure and elec- that changes in selenium concentration in the brain are associated trocautery to ensure hemostasis. Tail tip biopsies on the other 50 with abnormal brain functioning in various brain diseases such as mice were conducted using a sterile, heated scalpel blade only. Alzheimer disease and brain neoplasia. In this study, to investigate All tail tip biopsy procedures were performed by the same tech- whether selenium treatment would affect brain function, we ex- nician. The duration of time to complete the procedures for each amined the phosphorylation level of functional proteins including group was recorded. Daily clinical observations were conducted Tau, PKC, and MAPK in the Sel-treated SelM Tg rats. The CMV/ for 5 d following the procedure, to include scoring the degree of GFP-hSelM Tg (SelM Tg) rat used in this study were developed inflammation at the amputation sites. In the heated blade group, by microinjection of CMV/GFP-hSelM fusion gene into fertilized hemostasis was rapidly achieved in all mice, the degree of tissue rat eggs and showed a high antioxidant status in various tissues. damage at the time of amputation was noticeably reduced, and SelM Tg rats were treated with sodium selenite for 3 wk and the the incidence of postprocedural inflammation at the amputation level of related proteins were detected with western blot analyses. site was significantly reduced as compared to the group where Based on the results of the present study, we concluded that the electrocautery was used. Even though the heated blade method phosphorylation of Tau protein was significantly changed in the removed the step of using electrocautery, there was no significant ser 404, ser 396, and ser 202 site of cortex and hippocampus region. difference in the time it took to perform the two techniques. In The analysis of PKC phosphrylation showed that PKC-alpha, conclusion, we found that the heated blade technique described delta, and zeta were key proteins to phosphorylate Tau protein. here represents an animal welfare refinement. Sel treatment decreased the activity of alpha and gamma-secretase producing Abeta-42 amyloid peptides as well as increase alpha- P137 Rat Animal Model for the Study of Colostral Antibodies secretase degrading Abeta-42 amyloid peptides. The microglial cell harvested from SelM Tg rat showed the high level of cell viability C Walden*, A Byers, S Hurley, J Allen compare with that of Non-Tg rats. Therefore, our results suggested Animal Science, North Carolina A&T State University, Greens- that the SelM expression and Sel concentration play a important boro, NC role the maintain of brain function and signal pathway, and the SelM Tg rat can serve as an animal model studying the biological A laboratory animal model was developed to study the antibody function of selenoprotein and selenium on the brain disease. response of Enterotoxigenic E. coli (ETEC) of swine origin in the colostrum and sera of the Sprague Dawley rat. ETEC is an eco- nomically important bacterial infection in neonatal swine. Neonatal P139 In Vivo and In Vitro Validation of Stored Homologous swine are protected at birth from ETEC infections by passive an- Red Cell Transfusion in Pigs tibodies transferred in the mother’s colostrum. The study of these S Biagini, A Rosário, DS Camila*, C Tanikawa, L Azevedo, P antibodies and their impact on infections in neonatal swine is made Aguirre difficult by the gestation period, logistics, and overall cost. The Institute of Learning and Research, Sirio Libanês Hospital, São Sprague Dawley rat was developed as an animal model for these studies. Specific pathogen-free rats were challenged intramuscularly Paulo, Brazil Abstracts of scientific papers 2008 AALAS National Meeting This work aimed to evaluate the feasibility of using stored red basement membrane. The cause of the decrease in blood glucose blood cells in pigs in order to further validate a model of blood is still unclear but may be related to the level of Plasmodium cell transfusion. One pig (50 kg) was submitted to a controlled membrane protein GPIs (glycosylphosphatidylinositols) and hemorrhage of 1033 ml of blood. The blood collected produced the balance between Th1/Th2 responses. These results also 2 unities of red blood cells that were preserved for 13 d under corroborate the “hygiene hypothesis” which postulates that controlled temperature. Validation of these unities in vitro the increased incidence of autoimmune diseases (such as dia- was carried out at the day of the collection and at the 13th d of betes) in developed countries is result of decreased exposure storage through the evaluation of volume, level of hemoglobin to pathogens. (Hb) and hematocrit (HT), hemoglobin concentration, hemolysis determination, sodium and potassium concentrations, glycemia P141 Evaluation of Two Pachymeters for Corneal Measurement and pH. After 13 d, in vivo hemolysis was evaluated through in Rabbits (Oryctolagus cuniculus) and Guinea Pigs (Cavia chromate-marked red cells (0.7 ml of radioactive sodium chro- porcellus) mate [Na251CrO4] for 100 g of red cells). Five animals (38 to 58 kg), including the animal previously submitted to hemorrhage DN Earnest*, G Hamilton, MA McLaughlin were anesthetized and received 2.5 ml/kg of marked red cells DIS-Pharmacology Screening, Alcon Research, Ltd, Fort Worth, (0.412 MBq/kg). Blood was collected at 5 min, 10 min, 1, 3, 6, 9, TX 12, and 24 h, and these samples were counted on an automatic The link between corneal thickness (CT) and intraocular pressure gamma counter. After 24 h, the animals were sacrificed and the (IOP) has driven the need to measure CT in animal research. Pa- spleens analyzed by scintillography. Transfusion units had low chymeters measure corneal thickness, and most of those available volumes (187.1 ml and 144.7 ml at 13 d) and low concentrations of are designed for human clinical use. We evaluated 2 pachymeters total hemoglobin (41.73 g/U and 31.85 g/U at 13 d) as compared for use in the research environment and also established in-house to human standards. In vitro validation demonstrated indexes reference CT values for 2 strains of rabbits and a strain of guinea of hemolysis below 0.1% and similar electrolyte concentrations pigs. Reliability and comparability were evaluated for the Accu- when compared to human data. In vivo results demonstrated tome AccuPach V Pachymeter (measuring range 300 to 999 μm) that the percentage of chrome remaining on red blood cells was and the Sonagage Corneo-Gage Plus Pachymeter (measuring over 70% up to 24 h of observation. Spleen analysis did not show range 200 to 999 μm) in adult New Zealand albino (NZA) rabbits, significant accumulation of the radioisotope. Our results show Dutch Belted (DB) rabbits, and Hartley outbred (HOB) guinea that red blood cell storage in pigs does not induce significant pigs. Machines were evaluated for ease of use (based on portabil- hemolysis up to 14 d. These data demonstrate the feasibility of ity, set-up time, and learning time), reliability and repeatability studies that evaluate the effects of stored red cell transfusion (prolonged use for 2 h), and comparability (same eyes measured in pigs. by both machines). Conscious animals were restrained manually; 2 drops of topical anesthetic were instilled, and 3 measurements P140 Plasmodium chabaudi chabaudi CR in Non-obese Diabetic were taken on each eye and averaged. Both machines were easy (NOD) Mice to use, being portable, quick to set up, and having a sharp learn- DR Machado*1, AA Guaraldo1, BC Vidal2 ing curve. The reliability of both machines was excellent, with no 1Parasitology, 2Cell malfunctions for prolonged sessions of 2 h constant use. Corneal Biology, Unicamp, Campinas, Brazil thickness measurements were 388 μm ± 4.0 (AccuPach V) and Malaria is a parasitic disease that kills over a million people each 386 μm ± 3.8 (Corneo-Gage) in the same 44 NZA eyes; 368 μm ± year. In Brazil, 99% of malaria cases occur in the Amazon Basin. 3.4 (AccuPach V) and 362 μm ± 3.5 (Corneo-Gage) in the same Plasmodium chabaudi is an important organism for studying sev- 26 DB eyes. The AccuPach V could not measure CT in guinea eral aspects of malaria because of its usefulness as a substitute pigs since average CT was found to be less than 300 μm. With model for Plasmodium falciparum. Non-lethal P. c. chabaudi CR the Corneo-Gage, CT in 78 guinea pigs was 217 μm ± 0.9. The strain is the autocontrol in such studies. Hypoglycemia is a com- Accupach V and Corneo-Gage pachymeters are easy, accurate, mon symptom of murine malaria and of P. falciparum malaria in and robust enough to use in the research environment. Normal humans. Type 1 diabetes is an autoimmune disease that destroys CT values have been established for NZA and DB rabbits and the insulin-producing beta cells, and non-obese diabetic (NOD) HOB guinea pigs. mice provide a good model of human type 1 diabetes. In this work, we examined the course of type 1 diabetes in NOD mice P142 Effect of Sub-tenon Injections on Intraocular Pressure in infected with P. c. chabaudi CR. SPF female NOD/Uni mice were Dutch Belted Rabbits (Oryctolagus cuniculus) inoculated intraperitoneally with red blood cells infected with P. c. chabaudi CR. Blood glucose was measured by using Roche DN Earnest*, S Yacoub, MA McLaughlin Accu-Chek Active kits in samples of tail blood collected every DIS-Pharmacology Screening, Alcon Research, Ltd, Fort Worth, other day for 11 d postinfection. The level of parasitemia was TX expressed as the percent of infected erythrocytes based on cell Progressive therapy for glaucoma will include routes of drug counts of tail blood smears stained with May-Grünwald-Giemsa. administration that allow sustained contact with the eye or The mice were classified according to the severity of their dia- sustained delivery of active drug from a device (such as a betes: moderate (300 to 500 mg/dl) or severe (>500 mg/dl). The suspension or tablet). No published accounts were found for control group consisted of BALB/c/Uni mice. The mortality sub-tenon drug delivery for intraocular pressure lowering in was recorded for healthy and infected NOD/Uni mice. Blood rabbits. Several parameters were investigated to define a model. glucose decreased at the height of parasitemia, around the 5th Intraocular pressures (IOP) in female Dutch Belted rabbits (n = d of infection. Infected diabetic mice survived longer after the 5 to 7) were measured with an applanation pneumatonometer onset of diabetes than the non-infected controls. Histologically, after light corneal anesthesia with topical 0.1% proparacaine. the main renal lesions included glomerular edema and necrosis, Animals were then anesthetized with isoflurane to effect. Using a cell infiltration, tubular atrophy and thickening of the glomerular 30-gauge needle on a sterile 1 ml syringe, various volumes (25, 50, Abstracts of scientific papers 2008 AALAS National Meeting 100, and 250 µl) and placements (2 mm from and perpendicular rabbit diets have consisted primarily of pellets with an alfalfa to the limbus, 2 mm from and parallel to the limbus, and 4 mm base. Exotic animal veterinarians now recommend low-protein, from and parallel to the limbus) of 3 vehicle depots (mannitol, high-fiber diets consisting primarily of timothy hay and timothy- saline, and BSS Balanced Salt Solution) under the tenon capsule based pellets, rather than alfalfa. An evaluation of the available were investigated. Following recovery from anesthesia, IOP commercial diets for laboratory rabbits found that all pellets, measurements were taken at 2, 4, 6, 8, and 24 h postinjection. even the high-fiber varieties, are alfalfa-based. Because alfalfa There was an unexpected pronounced volume effect on IOP; is higher in calcium than timothy, alfalfa-based pellets contain the greater the volume, the greater the IOP reduction (at 4 h 1.2% calcium, which is higher than even the published lactational postdose): 250 µl = –13.0%; 100 µl = –9.9%; 50 µl = –5.7%, 25 requirement of rabbits (0.75%). Timothy pellets, on the other µl = –5.9%. Contralateral, untreated eyes also responded with hand, contain about half the calcium of alfalfa-based diets. Our reduction in IOP with the maximum effect 2 h earlier than in hypothesis was that rabbits fed an alfalfa-based pelleted diet treated eyes: 250 µl = –11.8%; 100 µl = –3.0%; 50 µl = –11.1%; 25 would have increased serum calcium resulting in increased µl = –6.0%. IOP in treated eyes returned to baseline within 24 h; renal pathology compared to those rabbits fed a timothy-based in contralateral eyes, it returned by 4 h. The type of vehicle did pelleted diet. One group of 6-mo-old rabbits was fed a high- not influence outcome. Rabbits, whether through pseudofacility, fiber, alfalfa-based diet (control), while another group was fed, consensual neurological or barometric response, or some other upon arriving at our research facility, a commercially available cause, respond to sub-tenon injection with lowering of IOP both timothy-based diet that is not currently used for laboratory ipsilateral and contralateral in the absence of any pharmaco- species. Urine and blood was subsequently collected monthly logical agent. Care must be taken when using this model that for calcium evaluation. We found that both urine calcium and parameters are standardized. A separate control (vehicle-treated) serum calcium levels remained within the normal published group of animals is essential. ranges for both timothy and alfalfa groups. Comparisons of average serum calcium levels over time revealed no significant P143 Endpoints for Mouse Subcutaneous Tumor Models: Re- difference between the control and experimental groups. There finement of Current Criteria was also no significant difference in the overall urine calcium levels between the timothy and alfalfa groups. However, when E Paster1, K Villines2, DL Hickman*1 serum calcium levels were initially examined in the experimental 1Veterinary Medical Unit, VA Medical Ctr, Portland, OR; 2College group after switching from a alfalfa-based to a timothy-based of Veterinary Medicine, Oregon State University, Corvallis, OR diet, a difference was found that later normalized as time pro- gressed. Our data suggests that a timothy-based diet does not Currently accepted methods of humane endpoint determination lower serum or urine calcium over time; however, when a new, for subcutaneous tumor growth models in mice include tumor lower calcium diet is fed, there can be a change in the initial size (generally maximum size of 1 cm), presence or absence of serum calcium level. ulceration, and changes in body weight. Because tumor mass may mask pathologic weight loss, this study explored the use of body condition scoring (BCS) as an additional criteria to bet- P145 Infectivity of Giardia spp. in BALB/c Nude Mice In- ter assess morbidity in these animals. To complete this study, 2 oculated with Cysts of Effluent Wastewater after UV Light different tumor models were assessed: a skin papilloma model Disinfection with multiple tumors produced after exposure to an appropriate L Santos1, R Franco2, A Guaraldo2,3, R Cantusio Neto4, D Al- carcinogen, and a subcutaneous melanoma study where a single, ves*3 bulk-producing tumor was produced after subcutaneous injec- 1Laboratory of Advanced Oxidation Processes—Faculty of Civil tion of tumor cells. Animals were evaluated 3 times per wk for Engineering, 2Department Of Parasitology—Institute Of Biology, body weight, BCS, tumor number, size of the largest tumor (if 3CEMIB—Multidisciplinary Center for Biological Investiga- multiple), and general clinical presentation (such as grooming, posture, and responsiveness to stimuli). Data analysis revealed tion on Laboratory Animal Science, UNICAMP—University of Campinas, Campinas, Brazil; 4Laboratory of Microbiology, that BCS was significantly more reliable than body weight as an SANASA—Society for Water Supply and Sanitation, Campinas, indicator of anorexia when correlated with a moribund clinical Brazil condition. However, tumor growth in these models was found to be well tolerated. Tumors over 1 cm in diameter did not result Giardia spp. is a protozoa of public health interest that causes in a decrease in BCS. Instead, we found that BCS consistently gastroenteritis in humans and other animals, and is very common did not decline until the tumor became ulcerated or interfered in developing countries. In the last decades, outbreaks have been with mobility because of location on or near a limb. Based on recorded around the world, and the domestic sewer is suspected the conclusions of this study, our IACUC has updated its tumor to be the source of river contamination. In the city of Campinas in evaluation guidelines to allow tumors of up to 2 cm in diameter, southeast Brazil, giardiosis is endemic; the pathogen is detected as long as mobility is not decreased, there is no ulceration, and at high concentrations in effluent wastewater, which is a poten- the mouse maintains a BCS of 3. The results of this study will be tial reservoir for transmission. The Wastewater Treatment Plant of interest to investigators and animal care staff working with (WWTP) Samambaia in Campinas used a system of activated similar studies. sludge for sewage treatment and UV light for disinfection of ef- fluent wastewater. In order to evaluate this disinfection process P144 Does Feeding an Alfalfa-based Diet Predispose Labora- in inactivating cysts of Giardia spp., 2 samples types were selected tory Rabbits to Hypercalcemia and Renal Disease? for study: effluent wastewater without UV disinfection (EFL) and effluent wastewater with UV disinfection (EFL+UV). BALB/c E Paster, DL Hickman* nude immunodeficient mice were intragastrically inoculated Veterinary Medical Unit, VA Medical Center, Portland, OR with a mean dosage of 14 cysts of Giardia spp. collected from either EFL or EFL+UV. All animals inoculated with Giardia spp. Available literature on rabbit nutrition offers conflicting advice cysts developed the infection, but the animals inoculated with on feeding our laboratory lagomorphs. Historically, acceptable Abstracts of scientific papers 2008 AALAS National Meeting UV-exposed cysts released cysts in the feces with a lower average the animals were tested in the system under the same conditions. concentration than that animals inoculated with cysts without We were able to show significant differences (P < 0.05) between UV disinfection. Trophozoites were also observed in the animals the baseline and the 4-h samples post-carrageenan injection of both groups. These findings suggest that cysts exposed to UV in the following parameters: swing time, stance/swing ratio, were damaged, but were able to cause infection. stance time, % stance/stride, stride length, paw area, stride time, % swing/stride, % propulsion/stride, % braking/stride, P146 SCID Mice as Adjuncts to Rodent Sentinel Health Moni- propulsion time, % braking/stance and % propulsion/stance. toring Programs We validated that the system is a reliable, accurate and repeat- able system for detecting changes in gait related to acute pain. D Gettys*, CL Franklin, LK Riley, R Livingston We have also demonstrated the repeatability in both interday University of Missouri, Columbia, MO and intraday testing, as well as the lack of effect of anesthesia on gait dynamics. Pathogenic microorganisms can have many adverse affects on rodents used in biomedical research. As a result, the health status of contemporary rodent colonies is monitored regularly. P148 Use of m-CT to Assess Anatomical and Structural Features Researchers rely on sentinel programs as a measure of colony of Proximal Femurs in Commonly Used Laboratory Animals health, but there are published reports and abundant anecdotes ER Berryman*, C Bagi, MR Moalli of sentinel program failure. Our hypotheses were that certain World Wide Comparative Medicine, Pfizer Global Research and agents do not readily transmit to sentinel animals and that Development, Groton, CT incorporation of immmunodeficient (SCID) mice into sentinel programs would improve detection of these agents. For this The physical characteristics of bone include the morphology, di- study, soiled bedding from mice infected with a number of mensions, and distribution of apatite crystals that forms cortical pathogens was transferred to cages containing 3 sentinels (n = and cancellous bone structures. The skeleton is a mechanically 5 cages) or 2 sentinels and 1 SCID mouse (n = 5 cages). Every 2 optimized biological system whose composition and organization wk, sentinel mice were evaluated for exposure to multiple patho- reflects the functional demands made upon it differing between gens by serology, fecal PCR, oral bacterial culture, and perianal various species. Hydroxyl apatite accounts for approximately tape tests. The infectious pathogens tested for included: murine 65% of bone weight, provides most of the strength and stiffness norovirus, mouse parvovirus, minute virus of mice, Helicobacter of bone, and allows for use of radiological technologies to assess spp., Mycoplasma pulmonis, Pasteurella pneumotropica, and Theiler bone mass and structure. Development of imaging technologies murine encephalomyelitis virus. All viral pathogens readily such as microcomputed tomography (m-CT) has markedly im- transferred to most sentinel mouse cages. However, bacterial proved our ability to assess structural parameters of cortical and agents, including Helicobacter spp. and Pasteurella pneumotropica, cancellous bone. In the current study, we sought to determine were rarely transmitted to sentinels, and Mycoplasma pulmonis and describe differences in cortical and cancellous bone structure, did not transfer to any sentinel cages. The addition of SCID bone mineral density and morphology (geometry) at proximal mice did not result in increased detection of the latter patho- femur (hip) between most commonly used laboratory animals gens. This study supports published and anecdotal reports that including mice, rats, rabbits, dogs, and nonhuman primates. This Helicobacter spp. and Pasteurella pneumotropica are not reliably work enhances our understanding on bone physiology across lab detected by sentinel programs and demonstrates that inclusion animal species and should enable scientists to select appropriate of SCID mice does not improve their detection. Because of this, species for research involving skeleton including metabolic bone alternative methods of monitoring for these agents, such as test- diseases, bone fragility or skeletal toxicology. Preliminary data ing of colony animals, should be considered in design of health demonstrated cross-species differences in cancellous bone struc- monitoring programs. ture, including trabecular number, thickness, plate separation, and connectivity. Bone mineral density was greatest in rodents, P147 Digital Gait Analysis Validation in CD Rats Using the followed by dog, nonhuman primate, and rabbit. Differences in DigiGait System geometry and structure of the femoral head, neck, and midshaft were demonstrated in 2- and 3-dimensional analysis that may ER Berryman*, C Bagi, R Harris, C Andresen indicate adaptation to the mechanical loads that each species World Wide Comparative Medicine, Pfizer Global Research and experiences during normal cage activity. Development, Groton, CT The gait analysis system we tested combines a motorized tread- P149 Development of an Ambulatory Continuous Intravenous mill, high-speed digital imaging, and broadband video storage, Infusion Method in the Nonhuman Primate and provides visual assessment of gait dynamics in rodents and EA Lambert*, JR Hynick, JJ Setser, TD Morris, WR Miller, CP quantitative analyses of gait that may be related to alterations Chengelis in genes, disease, and drugs that affect coordinated motion. The Toxicology, WIL Research Laboratories, LLC, Ashland, OH advantage of the system is that we may conduct noninvasive, longitudinal studies and obtain quantitative assessments of As recent industry demands have broadened, a need for the de- response to therapy and correlate the data with traditional post- velopment of an effective ambulatory continuous infusion model mortem endpoints. The main goal of this study was to validate in nonhuman primates has increased. Most current continuous the system for use in the evaluation of movement/gait dynamics infusion models involve jacket and tether systems connected in the published carrageenan rat model of pain and inflamma- to syringe pumps that are externally attached to the cage. Our tion. Following IACUC review and approval, test subjects were objective was to develop a viable ambulatory infusion model in run on the treadmill system for 5 to 10 s to obtain at least 6 step nonhuman primates that resulted in an accurate administration cycles, and video images were obtained for analysis. The animals of a test article continuously while allowing freedom of move- were then anesthetized, and the right stifle was injected with ment and socialization within the cage, as well as manipulation either saline or carrageenan. Two hours and 4 h after injection, outside of the cage. A study was conducted in 4 phases using a Abstracts of scientific papers 2008 AALAS National Meeting total of 24 cynomolgus macaques, 2 animals of each sex during in contrast to conditioned cats, the Class B cats exhibited wide the first 3 phases and 6 animals of each sex during the last phase. variations in their coagulation parameters and indigenous inhi- All animals were catheterized in a femoral vein with a 3 to 6 Fr bitions of coagulations. Furthermore, Class B vendor cats may Cath-in-Cath system and fitted with infusion jackets. Infusion not be optimal for vascular or cardiovascular research due to the pumps and syringes were used to continually infuse saline over variations in the haemostatic parameters. a 28-d period. The syringes were weighed and changed 3 times weekly during the first 2 phases, 5 d a wk during the third phase P151 Helicobacter Detection in Murine Fecal Samples in Nine and 7 d a wk during the fourth phase. Specific endpoints, such Diagnostic Laboratories Using PCR as clinical pathology parameters, equipment-related issues, dose accuracy, and macroscopic examination of the infusion site, were E Mahabir*1, C Mörth1, M Brielmeier1, K Markoullis1, W Beisker2, evaluated to determine the effectiveness of this model. Twenty- J Needham3, H Balzer4, LE Bater5, A Bleich6, A Deeny7, J Dix5, two of the 24 animals placed on study successfully completed the K Jacobsen1, A Lorenz8, M Mähler6, W Nicklas9, JD Phipps5, KE 28-d infusion period. There were 2 incidences of localized inflam- Seidel10, M Seidelin11, MF Toft11, A Tomlinson7, P Wilhelm1, J matory reactions resulting in removal of these 2 animals from Schmidt1 study, but no significant alterations in clinical pathology. Dose 1Department of Comparative Medicine, 2Institute of Toxicology, volume accuracy for phases 1, 2, 3 and 4 were 93.5%, 96.6%, 96.0% Helmholtz Center Munich—German Research Center for Envi- and 97.8%, respectively, with an overall accuracy of 96.5%. Over ronmental Health, Neuherberg, Germany; 3 The Microbiology the course of the study, the number of incidences of equipment- Laboratories, North Harrow, United Kingdom; 4Department of related issues decreased, from 9 events for 4 animals in the first Molecular Biology, Vet Med Labor GmbH, Division of IDEXX phase, to 8 events for 12 animals in the fourth phase. All of these Laboratories, Ludwigsburg, Germany; 5LAS Diagnostics, Glaxo- issues were correctable, allowing animals to be maintained on SmithKline, Herts, United Kingdom; 6 Institute for Laboratory study without being compromised. As a result, a refined con- Animal Science and Central Animal Facility, Hanover, Germany; tinuous infusion model in nonhuman primates was successfully 7Molecular Biology Laboratory, Harlan UK, Hillcrest, United developed, in which saline was accurately administered up to 28 Kingdom; 8 GIMmbH, Wildenbruch, Germany; 9Microbiologi- d with a low rate of equipment issues, but still allowing freedom cal Diagnostics, German Cancer Research Center, Heidelberg, of movement and socialization among animals. Germany; 10Institute of Microbiology and Hygiene, Technical University Munich, Munich, Germany; 11Taconic Europe, Silke- P150 Coagulation Profiling of Class B Vendor Cats with Refer- borg, Denmark ences to Serine Protease Inhibitors Since Helicobacter infections in mice may influence the results of in E McGeehan*1, K Wielgus1, F Azarafrooz1, LM Cera1, R Bues- vivo experiments the Federation of European Laboratory Animal ing1, J Fareed2 Science Associations (FELASA) recommended investigations 1Comparative Medicine Department, 2Department of Pathology, for Helicobacter spp. during regular annual health monitoring. Helicobacter diagnostic is highly dependent on methods that are Loyola University Medical Ctr, Maywood, IL specific, sensitive and rapid. Currently, the polymerase chain Class B Vendor cats (Felis catus) are commonly used in biomedi- reaction (PCR) is the preferred diagnostic tool, but results ob- cal research; however, the laboratory parameters, including the tained in different laboratories from aliquots of the same fecal hemacologic and clinical chemistry, widely vary between each samples may vary considerably. To determine the PCR methods cat due to initial health status. A systemic study on the plasmatic used for detection of murine Helicobacter spp. in fecal pellets and coagulation profile and serine protease inhibition is not reported. to compare their sensitivity, their specificity, and their repro- It was hypothesized that due to the health status of these cats ducibility, a ring study involving 9 commercial and academic the hemostatic system is also compromised. To address this European laboratories was designed. The study was conducted hypothesis, 4 groups of Class B vendor cats (n = 42) were ini- over a 3-y period and consisted of 4 rounds during which fecal tially profiled for prothrombin time (PT), partial thromboplastin samples were analyzed. The initial round revealed up to 3 log time (aPTT), Heptest, and thrombin time (TT). The antiprotease differences in detection of Helicobacter. The sensitivity, specific- activities were also measured in terms of the anti-Xa, anti-IIa, ity and reproducibility of testing improved considerably during heparin cofactor II (HCII), and antithrombin III (ATIII) using the 4 rounds of analyses. After completion of the 4 rounds of specific chromagenic substrate methods. Along with the Class analysis, determination of the number of Helicobacter genomes B vendor groups, 1 additional group, comprising 10 clinically in fecal pellets from naturally infected mice by the organizing healthy cats which were habituated at a veterinary research laboratory revealed that each method used by the participating facility for 6 mo, were used as a control group. Citrated blood laboratories was sufficiently sensitive to reliably detect Helico- samples were drawn from all groups and centrifuged to obtain bacter in naturally infected mice. plasma samples. The samples were analyzed in groups using standard clotting assays (PT, aPTT, Heptest, TT and antiprotease P152 Increased Hemorrhagic and Antithrombotic Activities of activity). The ATIII and HCII activities were measured using Contaminated Heparin as Studied in an Animal Model commercially available chromagenic substrate assays. All results were compiled as groups, with applicable statistical methods E Litinas*1, E McGeehan2, W Jeske1, D Hoppensteadt1, J Fa- used to analyze the data. The Class B vendor cats did not show reed1 any variation in the PT and TT assays, and only weak anti-Xa 1Department of Pathology, 2Comparative Medicine Department, activity (>10%); however, there was a wide variation in the aPTT Loyola University Medical Center, Maywood, IL (12.2 to 49.9 s) and Heptest (20.8 to 51.7 s) assays between the The USFDA recently recalled heparin preparations contaminated 2 groups. The HCII activities measured by Dermatin Cofactor with hypersulfated chondroitin sulfate (HCS) due to reported also showed wide variations which were largely dependent on adverse events and deaths. Although the adverse events were the individual groups (range = 50% to 150%). In the ATIII as- reported to be associated with the activation of the contact sys- say, similar trends were also noted. These studies suggest that tem by HCS, no data on the anticoagulant and antithrombotic Abstracts of scientific papers 2008 AALAS National Meeting bleeding effects of these contaminated heparins is available. P154 Antitumor Efficacy and Safety of Pteris semipinnata Linn To address the in vitro anticoagulant profile and the bleeding Extracts in Chemical-induced Liver Cancer in Mice and antithrombotic actions of heparin, a contaminated heparin GG Chen*, J Leung, JH Yip, NM Choi, PS Lai (CH) and a noncontaminated heparin (CFH) were compared in standard laboratory methods. Both heparin preparations The Chinese University of Hong Kong, Hong Kong, China were compared in equigravimetric concentration and dosages; Pteris semipinnata Linn (PsL), is a traditional Chinese herbal coagulation profiles of both agents were carried out in normal medicine which has been used to subside lumps and to improve pooled rat plasma. The hemorrhagic effects of both groups were a variety of inflammatory conditions. We have shown that the tested in a rat tail transaction model in individual groups (n = 6). extracts of PsL, such as 5F and 6F, are able to inhibit the growth In the same groups, the antithrombotic effects were measured of a number of tumors in cell culture experiment. However, its using a jugular vein clamping model. Ex vivo analysis of blood efficacy and safety in in vivo tumor models are unknown. In samples were carried out to determine the pharmacodynamics this study, we have established a mouse model of liver cancer differences between the 2 agents. In the in vitro profiles using the induced by diethylnitrosamine (DEN), tested the efficacy of 5F partial thromboplastin time and antiprotease assays at potency on the growth of tumor and monitored the side effects. DEN was adjusted levels, the 2 preparations were comparable; however, administered by intraperitoneal injection to 12-d-old juvenile the contaminated product showed slightly stronger activity. In C3H/HeJ mice to induce HCC. At wk 17, 5F was given after the hemorrhagic model, both agents produced a dose-dependant DEN administration; mice were sacrificed 12 wk later. We found (2 to 5 mg/kg) prolongation of bleeding time compared to sa- that 5F (12 to 48 mg/kg) significantly reduced the number of line. At the higher dose (5 mg/kg) the contaminated heparin the tumors in the liver in a dose-dependent manner. It also ap- produced much stronger bleeding times (67.8 ± 1.8 min CH peared to markedly decrease the total volume of liver tumors. compared with 57.1 ± 1.9 min CFH). Similar trends were noted These findings are in agreement with results obtained in the cell in the antithrombotic model where a higher number of jugular culture study of 5F. Thus, the data have indicated that 5F is able clampings were needed in rats treated with CH (7.6 ± 0.8 clamps to kill liver tumor cells in vitro and in vivo. We did not observe CH compared with 6.0 ± 0.2 clamps CFH). Ex vivo analysis of convulsion, sedation, lack of grooming, loss of righting reflex, plasma also revealed a stronger anticoagulant/antiprotease increased or decreased respiration, gross ascites, signs of dehy- affect of the contaminated product. This data suggests that the dration, lethargy, emaciation, or weight loss during the period presence of HCS as a contaminant enhances the in vivo bleeding of treatments. Serum markers also showed no significant change and antithrombotic effects of the recalled product. Therefore, in the markers related to toxicity in liver and kidney. However, besides the nonanticoagulant effects, the CH may also modulate hyperactivity occurred in some treated mice. In conclusion, 5F the bleeding and antithrombotic efficacy of heparin, thus having can effectively inhibit the development or growth of liver cancer significant drug/drug interactions. in mice and does not generate significant side effects. Therefore, 5F is a promising anti-liver cancer agent. P153 Further Analysis of LCMV M1 Isolate F Ike*1, K Sakai3, T Mizutani3, K Takimoto4, F Bourgade2, J P 1 5 5 Ta c o n i c C 5 7 B L / 6 J B o m Ta c , C 5 7 B L / 6 N Ta c , a n d Jaubert2, X Montagutelli2, A Kajita1, H Nakata1, A Yoshiki1, S C57BL/10SgSnAiTac Mice Do Not Carry the Nnt Mutation Morikawa3 JL Seymour1, MA Young1, GW Bothe*1, AV Perez2 1BioResource Center Experimental Animal Division, RIKEN, 1R&D, 2Veterinary Services, Taconic, Hudson, NY Tsukuba, Japan; 2The Central Animal Facilities, The Institut Pasteur, Paris, France; 3Department of Virology 1, 4Division of Nicotinamide nucleotide transhydrogenase (NNT) is a trans- Experimental Animal Research, National Institute of Infectious membrane protein that catalyzes the interconversion of NADH Diseases, Tokyo, Japan and NADPH in the mitochondria. Under conditions of oxidative stress NADH will be converted to NADPH, which in turn re- We isolated lymphocytic choriomeningitis virus (LCMV M1 generates reduced glutathione. Studies suggest that the presence isolate) from wild-derived mice originally caught in Austria. of NNT is essential for mitochondrial defense against oxida- M1-infected cells were recognized by mouse anti-LCMV Arm- tive stress and for normal cellular metabolism. Furthermore, strong serum. But homology analyses on nucleoprotein (NP) mitochondrial dysfunction has been implicated in a number of nucleotide and amino acid sequences indicated that there was a degenerative diseases, including Parkinson disease, Alzheimer genetic distance between M1 and already-known LCMV strains, disease, and diabetes. Recently, several reports have documented which makes it difficult to create PCR primers for these types that C57BL/6J (B6J) mice carry a 17.8 kbp deletion in the Nnt of viruses. On the contrary, the recently identified LCMV-like gene including exons 7 to 11. We decided to screen the “Black” Dandenong virus showed a high homology with M1 on NP substrains, C57BL/6JBom (B6JBom), C57BL/6NTac (B6NTac), amino acid sequence (97%). To know the genetic location of M1 and C57BL/10SgSnAiTac (B10Tac) for this mutation. Two sets among LCMV, we determined full nucleotide sequences of S- of primers were used. The first set amplifies 312 bp in intron segment and L-segment of M1 and compared these with those 6, a region that is deleted in mutant mice. B6JBom, B6/NTac, of already-reported LCMV. RNA of M1-infected Vero E6 cells B10Tac, DBA/2NTac (DBA/2), and B6D2F1/Tac (B6D2F1) show was extracted and transcribed to cDNA using random hexamer the presence of the 312 bp band. The control C57BL/6J strain primer. Nucleotide sequences were determined and homology received from The Jackson Laboratory did not amplify the 312 was analyzed by Mega4 program. Amino acid homology between bp band, indicating the absence of the intron 6 DNA sequence. M1 and Dandenong virus was very high on NP, GPC protein, A second set of primers was used to amplify across the dele- Z protein, and L protein. LCMV M1 isolate seemes to belong to tion breakpoint, giving a 547 bp product if the DNA carried the the new group of LCMV as indicated on LCMV-like Dandenong deletion. The only amplified band is the one found in C57BL/6J virus. Dandenong virus was isolated from fatal transplant re- mice. Interestingly enough, B6JBom does not present an ampli- cipients. Adequate, thorough LCMV surveillance is necessary fied band. The amplification of intron 6 and the absence of the for mice and other animals, including hamsters. breakpoint amplified band indicate that B6JBom does not present Abstracts of scientific papers 2008 AALAS National Meeting the deletion of exons 7 to 11 of the Nnt gene. The B6JBom strain and intestines from mice contaminated with S. muris were PCR was obtained through Taconic Europe in 1988. The original stock positive, whereas feces and intestines from mice contaminated was received from The Jackson Laboratory in 1971. Therefore, we with other common protozoa but not S. muris were PCR negative. can assume that the Nnt mutation arose at The Jackson Labora- The assay was also found to have high sensitivity, detecting as tory after 1971. We can confidently say that C57BL/6JBomTac, few as 6 template copies per microliter of diluent. Importantly, C57BL/6NTac and C57BL/10SgSnAiTac do not carry the 17.8 all mice positive for S. muris on postmortem exams were also Kbp deletion in Nnt. positive by fecal PCR. Moreover, S. muris was detected by PCR in mice negative by postmortem examination but from colonies P156 Injection Variability Observed with Syringe Pumps known to be infected. Because S. muris loads have been reported to decrease with age, we adapted the PCR to a quantitative format GM Herrera*1, BD Hare1, SH Kojtari1, KR Zurn1, SI Dworkin2 and assessed protozoal loads in feces of naturally infected mice of 1Med Associates, Inc., Saint Albans, VT; 2Psychology, Jacksonville several ages. Fecal loads of S. muris ranged from 1.4×105 template State University, Jacksonville, AL copies per fecal pellet in a 4-wk-old mouse to 1.4×102 template cop- ies per fecal pellet in an 18-wk-old mouse, confirming that fecal The use of motor-driven syringe pumps in biomedical research is loads decreased with age. In conclusion, the PCR assay developed wide ranging, from studies of the direct intracranial infusion of promises to be a highly specific antemortem diagnostic assay with drugs to rapid fixation of tissues for neurobiological assessments. higher sensitivity than existing postmortem tests, especially in One of the most common uses of motor-driven syringe pumps animals harboring low levels of S. muris. is for evaluating the effects of intravenous drug administration. Specifically, drug self-administration studies allow nonhuman subjects to control the intravenous delivery of drugs typically P158 Footpad Sweating Is Related to Cardiovascular Auto- abused by humans. The results of these studies are used to nomic Responses in the Dog schedule drugs and evaluate behavioral and neurobiological H Yamamoto*1, Y Arasaki1, M Can2, E Hamade3, H Yokoi3, C Tsut- effects of these drugs as well as assess potential treatments for sumi1, S Ohtsuka1, E Hori2, K Umeno2, A Fujiki2, H Nishijo1,2 drug abuse. These studies involve training a subject to engage in 1Life Science Research Center, Toyama, Japan; 2Graduate School a lever press or other response that results in the activation of a of Medicine and Pharmaceutical Science for Research, The motor-driven syringe pump. The dosage of the drug infused is University of Toyama, Toyama, Japan; 3 Life Care Giken Inc, a function of the concentration of the drug placed in the syringe (mg/ml) and the time the pump is activated (approximately Toyama, Japan 1 to 5 s). While the average amount of drug infused during a To investigate the relationship between stress and sweating from session tends to be very accurate, we have noticed that there is the dog footpads, cardiovascular autonomic functions (heart considerable variability in the dosage delivered for individual rates, heart rate variability) and sweating volume in unstressed injections. We investigated the involvement of syringe con- and stressed conditions were analyzed in dogs. Sweating vol- struction, syringe size, and pump activation duration on the ume was measured as transepidermal water loss (TEWL), which variability of consecutive infusions. To conduct this study, we was semi-quantified using skin patches including color-reacting used polypropylene and polycarbonate syringes in 20 ml and papers impregnated with cobalt chloride. The skin patches were 10 ml sizes and a single-speed syringe pump (3.3 rpm). Each stuck to the footpad; the color of the skin patches changed in syringe was tested using 10 injections with 1-, 3-, 6-, and 10-s response to water in sweat from the footpads. This simplified injection times. No significant differences were found in injec- system enabled us to assay TEWL from conscious dogs. In the tion variability based on syringe type (P > 0.05 polypropylene first experiment, electrocardiograms (ECGs) and TEWLs were compared with polycarbonate). Injection variability does, how- recorded from laboratory beagles. The same experiment was re- ever, vary significantly as a function of injection time. With 20 peated again after the dogs were allowed to freely move around ml syringes, injection volumes vary by 17%, 9%, 6%, and 4% for for about 20 to 30 min. Finally, ECGs and TEWL were also mea- 1-, 3-, 6-, and 10-s injections, respectively. Using 10ml syringes sured from the anesthetized beagle dogs. Both heart rates and the pattern is similar, with injection volumes varying by 28%, TEWL were lower after the laboratory beagles were allowed to 11%, 3%, and 4% for 1-, 3-, 6-, and 10-s injections, respectively. In relax for 20 to 30 min. These parameters further decreased when order to minimize the impact of injection-to-injection variability they were anesthetized. Furthermore, spectral analysis of heart when using motor-driven syringe pumps, longer injection times rate variability indicated that TEWL was positively correlated (>3 s) are advisable whenever possible. to heart rates and sympathetic nervous activity, while TEWL was negatively correlated to parasympathetic nervous activity P157 Development of a Fecal PCR Assay for Spironucleus in the laboratory beagles. In the second experiment, the TEWL muris was indexed by visual inspection according to the color table, while arousal or stressed levels were indexed according to the G Jackson*, R Livingston, LK Riley, CL Franklin behavioral criteria using domestic dogs under various stressed Research Animal Diagnostic Laboratory, University of Missouri, and non-stressed conditions. These 2 indices were significantly Columbia, MO correlated. The results suggest that sweating from the footpads in dogs might corresponds to human sweating from the palms, Spironucleus muris is a protozoan of the intestinal tract in mice, and that the present simplified method using color-reactive skin rats, and hamsters. While its effect on the health of animals and patches is useful to assess degrees of stress in dogs. its capacity to confound research are debated, it is often included in rodent health monitoring programs. Diagnosis usually relies on postmortem examination of the intestinal tract for trophozoites. P159 Migration of Ascaris suum Larvae in Mice In this study, we sought to develop a PCR-based antemortem HZ Naghashyan*, OV Shcherbakov diagnostic test with improved sensitivity and specificity. A PCR assay targeting the 16S-like rRNA gene of S. muris was designed Veterinary Medicine, Armenian State Agrarian University, Ye- and optimized. The assay was found to be highly specific as feces revan, Armenia Abstracts of scientific papers 2008 AALAS National Meeting Research of Ascaris suum larvae migration in mice has a great air or ground transportation has negligible impact on the tumor importance, because mice serve as an experimental model for growth kinetics in the A431 xenograft mouse model. Therefore, study of early migration in pigs, as well as for syndrome of the importation of tumor-bearing mice is a viable alternative to visceral larva migrans in humans. Many authors deal with the creating xenograft models in house. problem of ascarides larvae migration in uncommon hosts, but some aspects of migration remain controversial, and not well P161 Withdrawn studied. According to literature data, A. suum larvae penetrate the mouse cecum and colon and reach the liver by 4 h postinocu- P162 Comparison of Diagnostic Methods for Surveillance of lation. As for other authors, larvae appear in liver at days 2 to 7 Helicobacter Species Infection in Mouse Colonies postinoculation. Taking into account this information, we aimed to study migration of A. suum larvae in laboratory mice. For this JL Lofgren*, A Potter, M Fortin, NS Taylor, MT Whary, JG Fox purpose, 10 adult males (weight, 20 g) were orally infested with Division of Comparative Medicine, Massachusetts Institute of 1000 to 5000 eggs. Mice were euthanized by ether every other Technology, Cambridge, MA day postinfection and dissected with larvae detection by one of the following methods: compression with dyeing by 1% solution Helicobacter species naturally infect research mice and may con- of methyl-violet, compression with clearance by 1% solution of found experimental studies; thus, surveillance for helicobacter lactic acid, or digestion in artificial stomach juice. Results show infection status should be a component of murine health moni- that the most informative and efficacious method for larvae toring. Although choice of diagnostic method should be based detection appears to be compression with clearance by 1% so- on available sample type, technical requirements and cost, assay lution of lactic acid. It allows detect maximal number of larvae limitations of sensitivity and specificity are critical to obtaining in the internal organs of mice. The first free larvae are detected reliable results. We compared genus-specific PCR based on a 1200 in the gastrointestinal tract, mainly in stomach and cecum, on base-pair 16s RNA product, culture, and Helico-Stix Test strips d 2 after infestation. On d 4, larvae are detected in mesenteric for detection of Helicobacter spp. infection using feces collected at lymph nodes, liver, and lungs, and quantity of migrating larvae necropsy from 34 sentinel CD-1 mice. Sentinels were housed for in gastrointestinal tract is maximal. Larvae were detected in the 6 mo in 31 rooms, representing 5 facilities, and were exposed to internal organs up to 10th d postinoculation. Therefore, our re- weekly dirty bedding transfer from colony mice of varying ge- search shows that A. suum larvae migration in uncommon hosts, netic and SPF health status. Fresh fecal pellets from each sentinel namely mice, is rather longer than described in literature, and (n = 8 to 10) were solubilized in 1 ml of laboratory-grade distilled does not depend on inoculating eggs dosages. This circumstance water for the Helico-Stix Test assay. Several additional feces were may be considered in clinical and laboratory practice when early frozen at –20 °C for subsequent DNA extraction and at –70 °C in migration phase of ascariasis is studied. 30% glycerol broth for subsequent microaerobic culture at 37 °C on blood agar and antibiotic-selective media with and without filtration through a 0.45 μ filter. Nine of 34 fecal samples were P160 Impact of Transportation on Murine Xenograft Tumor PCR-positive for helicobacter, and helicobacter isolates were re- Models covered by culture from 7 of these 9 PCR-positive samples. Only H Liang*, R Driscoll, K Harbert, LA Conour 4 of the 9 PCR-positive samples were positive when screened Discovery Services, Charles River, Wilmington, MA for helicobacter infection using the Helico-Stix Test assay. The Helico-Stix Test had a preponderance of false positive outcomes; Xenotransplantation of human tumors into immunodeficient 20 PCR-negative samples were strip-positive. In comparison to rodents is a frequently used animal model for assessing the PCR, these results indicate that the sensitivity and specificity of efficacy of antineoplastic therapies. Among them, the subcu- culture were 78% and 100%, and for Helico-Stix Test 44% and taneous xenograft model is the most common due to technical 20%, respectively. Thus, genus-specific PCR was the most sensi- ease of tumor cell implantation and tumor growth monitoring, tive diagnostic method for testing fecal samples from sentinel permitting high-throughput assessment of compound efficacy. mice and can be confirmed by culture when indicated. Depending on the particular cell line, tumor growth rate can vary in the live animal model. Time required for tumor growth P163 Laser-assisted IVF and Modified ICSI Markedly Improve prior to therapeutic testing results in associated losses in valuable the Efficiency of IVF in C57BL/6J Mice animal housing space, personnel time, and laboratory resources. To address these resource constraints, many pharmaceutical and J Mkandawire*1,2, T Chavarria1, P Qi1, A Discua1, A Viana1, SE biotechnology companies outsource the creation of xenograft Erdman1, JG Fox1, KJ Maurer1 tumor models with import of the animals in house for compound 1Division of Comparative Medicine, 2Center for Cancer Research, screening. To date, however, the impact of shipping stress on Massachusetts Institute of Technology, Cambridge, MA tumor-bearing mice and the tumor kinetics in vivo has not been reported in the literature. To determine whether air or ground Assisted reproductive technologies (ART) are an essential com- transportation influences tumor growth, we have evaluated ponent of a modern mouse husbandry program. Of all ARTs, in the NU/NU (Crl:NU-Foxn1nu) mouse model subcutaneously vitro fertilization (IVF) has perhaps the most potential because implanted with A431 human epidermal carcinoma cells. At 4 or only a single male is necessary to provide a large number of pups. 8 d post–tumor cell injection, the mice were randomly divided Unfortunately, in some inbred strains of mice IVF is not highly ef- into 3 groups and either shipped by air or by ground or remained ficient. Recently, we tried two modifications of IVF to see if we could in-house. Travel time for air and ground transportation was 20 h increase the effectiveness. Specifically, we performed laser-assisted and 30 h, respectively. In addition to daily clinical observations, IVF and a simplified version of intracytoplsamic sperm injection. animal body weight and tumor mass were measured 2 times All studies described herein used wild-type C57BL6/J oocytes per wk for 5 wk. Our data showed no statistically significant and sperm. Laser-assisted IVF was performed using a zyclone to differences in animal body weight and tumor volume between make 2 to 3 holes in the zona pellucida of eggs. Ablated oocytes groups. Our study suggests that shipping stress sustained during were co-cultured with sperm in human tubal fluid (HTF) for 5 h, washed, and cultured overnight in KSOM at 37 oC in 5% CO2. Modi- Abstracts of scientific papers 2008 AALAS National Meeting fied intracytoplasmic sperm injection (ICSI) was conducted using 1Comparative Medicine, 2Pharmacology, Pfizer, Inc, Saint Louis, simple manual microinjection of sonicated sperm heads deposited MO within the oolema with gentle, positive, micropipette air pressure. Intranasal (IN) dosing has long been the prominent technique A sperm microinjection micropipette was calibrated in-house for used when targeting the lungs. Different dosing methods can easy aspiration of sperm cells and entry past the zona and oolema affect systemic delivery of compounds, inhibiting the lung in- barriers while averting oocyte lysis. After injection of sperm in fluid flammatory process and costing valuable time. An innovative holding medium (FHM), eggs were transferred to KSOM and incu- method of delivery is modified intratracheal (ModIT) dosing bated overnight at 37 oC in 5% CO2. Standard IVF was conducted which historically has been used with much less frequency. as described for laser assisted IVF without ablating the zona prior Specifically, mice are anesthetized with isoflurane and verti- to incubation with sperm. Fertilization rates for all procedures cally suspended on a bar by the upper incisors. The tongue is were scored by analyzing the amount of eggs that proceeded to extended by lined forceps, a finger is placed over the nostrils, the 2-cell stage following overnight incubation. Both laser-assisted and 50 µl of compound is pipetted onto the back of the tongue. IVF and the simplified ICSI method produced significantly greater Air is blown into the face of the animal prior to the release of the fertilization rates (55% and 46%, respectively) compared to standard tongue, causing the animal to inhale; this reaction is unique, as IVF (2%; P < 0.0001). In conclusion, both laser-assisted IVF and the there is no discovered documentation explaining this occurrence. modified ICSI technique are simple, rapid methods to increase the We examined 2 in vivo dosing techniques—intranasal (IN) and fertilization rate of IVF in inbred mice. ModIT—to evaluate and compare methods for time and resource management, dosing variability measured by lung dispersion P164 Withdrawn and deposition, and efficacy using compounds. Compound efficacy was determined by dosing BALB/C mice (ModIT and P165 Commercially Available Corn-cob Bedding Naturally IN for fluticasone, PF-xxx, and ± LPS controls) 1 h prior to IN Contaminated with Zearalenone Significantly Advances the LPS challenge with a takedown 4 h post-LPS. Inflammatory cell Time of Vaginal Opening in Immature CD-1 Mice counts (total cells and PMNs) in BAL fluids were analyzed by flow cytometry. Separately, compound dispersion was examined JE Thigpen*, E Padilla-Banks, G Kissling, GF Caviness, TE Whi- using injection of coummasie blue dye (50 µl) IN and ModIT with teside, J Locklear, MG Grant, DB Forsythe endpoints of 30 min and 4 h. After euthanasia, the lungs were National Institutes of Health, Research Triangle Park, NC harvested and observed for dispersion of dye in the interstitial We reported that corn-cob bedding contains the mycoestrogen lung tissue. Data points (in min) were also taken to determine zearalenone (ZEN). This report determines the concentration the time elapsed/dosing group. Animals dosed with IN and of ZEN in twelve different types of corn-cob bedding, and its ModIT compounds showed equivalent efficacy in the number of effects on time of vaginal opening (VO) in CD-1 mice. Liquid lung infiltrates for each compound. ModIT coumassie blue dye chromatography tandem mass spectrometry was used to mea- groups displayed a higher dispersion in the lungs with less dye sure the concentration of ZEN and alpha-Zearalenol in samples in the stomach or intestines. Variability within the ModIT group of plasma, urine, feed, and bedding spiked at 0, 5, or 10 ppb/g. was minimal in contrast to the IN groups, and was supported by Upon arrival, mice were given a phytoestrogen-free diet and necropsy images. Plotted dose time per animal varied, revealing reverse osmosis/deionized water ad libitum. Pups were weaned that ModIT has a 40% to 50% quicker dose time per animal in at PND 15, and 15 mice per group were randomly assigned to comparison to ModIT. Ultimately, ModIT dosing was proven to control or test bedding. Body weights at weaning were similar be equally efficacious, have a quicker dosing time, and provided among the groups. Mice were weighed weekly and on the day of less variability in lung dispersion. VO and observed daily for VO. Plasma and urine samples were collected on PND 22 and PND 34. Low, medium, or high levels P167 Characterization of the Charles River GK Rat (GK/To- of ZEN were detected in corn-cob bedding from all vendors. The hiCskCrljCrl) time of VO was significantly (P < 0.05) advanced in mice exposed K Harbert*1, ST Chou2, R Driscoll1, L Barbeau1, D Mendonca1, H to high levels of ZEN (4202 to 5284 ng/g) as compared to mice Liang1, C Clifford3, LA Conour3 exposed to low levels of ZEN (<500 ng/g). The time of VO was 1 DiscoveryServices, 2Discovery Services/GEMS, 3Research advanced, but not significantly in mice exposed to medium levels of ZEN (1900 to 2200 ng/g). The time of VO was significantly Models and Services, Charles River, Wilmington, MA (P < 0.05) advanced in mice exposed to bedding spiked with The Goto-Kakisaki Rat (GK Rat) is a well-established model of 10,000 ng/g of ZEN as compared to controls. Concentrations of non–insulin-dependent, nonobese Type II diabetes mellitus (T2DM). ZEN and alpha-zearalenol in the plasma and urine of the mice Distinct substrain colonies have been maintained around the world were correlated with the concentration of ZEN in bedding. We since the model’s introduction in the 1970s and have demonstrated concluded that ZEN contaminated corn-cob bedding can expose similar levels of glucose intolerance and impaired insulin response. mice to ZEN by contact, ingestion, or aerosol exposure, and Some of these source colonies are known to differ from each other in can significantly impact time of VO, an estrogen-sensitive bio- pancreatic morphology and insulin level. Currently, we use a sub- marker. This is the first report to determine that corn-cob bedding strain of GK Rat (GK/TohiCskCrljCrl) for internal and sponsored naturally contaminated with ZEN can significantly advance the studies. We have performed an in-Departmenth characterization of time of VO in CD-1 mice and has the potential to impact other this substrain, comparing it to data reported in the literature. Both hormonal endpoints. male and female GK rats demonstrate a similarly reduced growth rate and average body weights compared to Wistar (Crl:WI) con- P166 Comparison of Intranasal Dosing and Modified Intra- trols. As expected, the GK/TohiCskCrljCrl rats have significantly tracheal Dosing in BALB/c Mice: Compound Efficacy, Time elevated fasting blood glucose levels (P < 0.0001) and an altered Management, and Compound Dispersion response to oral glucose tolerance test at as early as 8 wk of age. In addition, the baseline blood glucose is significantly higher in J Daller*1, JD Blom2 male GK/TohiCskCrljCrl rats than female cohorts at 16 wk of age Abstracts of scientific papers 2008 AALAS National Meeting (P < 0.02). Indirect blood pressure measurements and pathological Oregon National Primate Research Center, Beaverton, OR assessment at 11 and 22 wk of age do not indicate signs of cardiac Positive reinforcement training (PRT) is a type of operant condi- disease. As previously reported, the male GK/TohiCskCrljCrl rats tioning in which the trainer reinforces desired behaviors (such also maintained a significantly lower pulse rates (323 ± 68 bpm, P as coming to the front of the cage) by rewarding the animal < 0.05 at 22 wk). Unexpectedly, the male GK/TohiCskCrljCrl rats when it performs the target behavior. PRT increases the animal’s demonstrate proteinuria (>200 mg/dl) compared to both Wistar control over its environment and reduces the stress associated controls and female cohorts at as early as 9 wk of age (P < 0.001). with procedures (such as blood draw or injection) and as such is Histopathological examination of the pancreas showed chronic beneficial to the well-being of the animal. While there are many diffuse pancreatic islet degeneration and regeneration, hemosi- benefits associated with training, it often involves a large initial derosis, and islet cell anisocytosis. However, examination of the input of time resources, which may preclude some facilities from kidney did not reveal any significant changes in relation to diabetic using PRT. Finding alternate training methods that can make the nephropathy at up to 26 wk of age. Our findings maintain that the process less time consuming would be of great value. Because GK/TohiCskCrljCrl rats have similar diabetic characteristics as some primates are known to imitate the actions of others, one those found in established substrains published in the literature. potential alternate technique is to have subjects watch conspecif- The source of sex-specific proteinuria in young male rats suggest ics being trained to perform various tasks. In the present study, early glomerular injury and filtration deficits, and is under further we examined whether observing others would reduce training investigation for serving as an indicator for monitoring the develop- time in adult, male rhesus macaques (Macaca mulatta). Monkeys ment of nephropathy associated with this diabetic model. (n = 12) were housed in individual cages located in a single room. We used PRT techniques to train the monkeys to present a thigh P168 Effects of Feed and Water Deprivation on Japanese Quail at the front of the cage and remain stationary for injection. We (Coturnix japonia) first trained 6 “demonstrator” monkeys, all housed on the same KL Arora*, B Biswas side of the room, for approximately 4 mo. We then trained the other 6 monkeys (“observers”) using the same techniques. The Department of Veterinary Science, Fort Valley State University, observers were located directly across from the demonstrators. Fort Valley, GA For both groups, training sessions were 5 to 10 min, 2 to 3 times This study resulted from an accidental deprivation of feed and per wk. Training was considered successful when the task was water in Japanese quails over 1 weekend. Apparently, the birds performed on command during 3 consecutive training sessions. didn’t exhibit any adverse effect, and accepted feed and water While we were able to train all 12 monkeys to present their readily when offered, and appeared satisfied in a couple of min- thigh at the front of the cage, it took the observers about 25% utes. This experiment was planned to address adverse effect of fewer training sessions than the demonstrators to complete the deprivation on physiological parameters. Six adult males and six task (P < 0.01). Additionally, significantly more observers than females (layers) of uniform size and weight were deprived of both demonstrators allowed needle insertion (P < 0.05). These results feed and water for 20 h or 40 h and the effects on body weight, suggest that observing others being trained may help to reduce hematocrit, plasma proteins, egg production, and shell quality overall training time in macaques. were studied. At the start of experiment, the birds were weighed and blood collected from the wing vein for evaluation for sugar, P170 Locomotor Activity in Rhesus Macaques (Macaca mulatta) PCV, and plasma proteins. Blood sugar was measured using a on Long-term Calorie Restriction glucometer, PCV using hematocrit tubes and plasma proteins using a refractometer. After fasting, the birds were weighed daily K Hagelin*1, J Young1, J Mattison2, D Ingram3 for 9 consecutive d and the blood was evaluated on d 1 and 3. 1Division of Veterinary Resources, NIHAC SoBran, Inc., Pool- After 20-h deprivation, the males lost 7% (118.3 g compared with esville, MD; 2Laboratory of Experimental Gerontology, NIA, 109.5 g) and females 13% (141.1 g compared with 122.4 g) of their Poolesville, MD; 3Nutritional Neuroscience and Aging Laborato- body weight (P < 0.05). After restoring feed and water, the males ry, Pennington Biomedical Research Center, Baton Rouge, LA recovered 96% and females 93% of their prefasting weights by d Decline in locomotor activity is a robust feature of aging that 3. The blood glucose decreased (P > 0.05), and PCV and plasma has been documented in both humans and nonhuman primates. proteins increased (P < 0.05) during fasting, then returned to The effects of long-term calorie restriction (CR) on aging and its normal levels by d 3. Females resumed laying after skipping 1 impact on behavioral processes are still being investigated. Age or 2 d. After 40-h deprivation, males lost 12% (123.2 g compared and diet effects on locomotor activity were evaluated in a group with 107.9 g) and females 25% (144.6 g compared with 107.6 g) of of healthy young and aged male and female rhesus monkeys their body weight (P < 0.05). After restoring feed and water, the involved in a long-term longitudinal study of CR. All monkeys males recovered 94% and females 90% of their prefasting weight are provided a highly nutritious diet, with the CR group receiv- by d 3, with full recovery by d 9. Eggs were smaller, and egg shells ing 30% less than that provided to comparable age, gender, and lacked proper pigmentation. The PCV increased during fasting body size controls (CON). CR was initiated when the monkeys (48.9% compared with 42.9%; P < 0.05) and returned to prefasting were juveniles, adults, or aged. At the time of data collection, level after d 3. Results indicated that water and feed deprivation monkeys were grouped as either Young (CON: n = 28, age = 15 could seriously alter the physiochemical parameters, with the ef- to 21 y; CR: n = 25, age = 15 to 27 y) or Old (CON: n = 7, age = 28 fects increasing with prolonged fasting; these, in turn, could affect to 35 y; CR: n = 6, age = 32 to 37 y); monkeys had been on CR for experiments in progress. Furthermore, the data could also be used 16 to 18 y. Locomotor activity was analyzed using noninvasive as a resource for animal care committees if faced with protocols activity monitors that were attached to a primate collar. These that include bird feed and water deprivation. monitors directly record activity counts by analyzing the occur- rence and intensity of motion through omnidirectional sensors. P169 Watching Conspecifics Being Trained Helps Rhesus Ma- All activity was collected in a monkey’s home cage over a period caques (Macaca mulatta) Learn Faster of 24 h for 7 consecutive d. Because the activity levels of rhesus K Mueller, K Moore*, A Maier, K Coleman monkeys are highly dependent upon the circadian cycle, activ- Abstracts of scientific papers 2008 AALAS National Meeting ity data was analyzed for 24-h, day, and night periods. Young The GE Lunar PIXImus II (PIXImus) is used for dual energy CON monkeys were significantly more active than Old CON x-ray absorptiometry (DEXA) in laboratory mice. However, the monkeys for the 24-h (R2 = 0.12, P = 0.04) and Night (R2 = 0.12, image acquisition area is only 80 by 65 mm, limiting the animal P = 0.04) periods, while Young CR monkeys were significantly size and weight (<50 gm) for which it can accommodate. The GE more active than Old CR monkeys for the Day (R2 = 0.15, P = 0.03) Lunar Prodigy (Prodigy) is an FDA-approved imaging modality and Night (R2 = 0.37, P = 0.003) periods. In terms of diet effect, for human use and validated for animals weighing over 200 gm. Young CR monkeys were significantly more active than Young When using these 2 densitometers, a diagnostic gap therefore CON monkeys, regardless of time of day. Conversely, Old CON exists for evaluation of obese mice and young rats for which no monkeys were significantly more active than Old CR monkeys. validated guidelines exist. A study was conducted to evaluate To gain a better understanding of the effect of CR on aging and animals weighing less than 200 gm using the Prodigy. Twenty- its associated health benefits, locomotor activity will be examined three adult, mixed-sex C57BL/6NCrl (n = 9) and Crl:CD-1(ICR) every 3 y. In addition, we hope this longitudinal assessment will (n = 14) mice with body weight ranging from 19 g to 41 g were provide additional information regarding whether or not the age used. Each animal was scanned using both densitometry models, in which CR is first introduced negatively or positively impacts and values for bone mineral density (BMD), bone mineral con- various behavioral functions. tent (BMC), fat, lean, and total tissue recorded. For the Prodigy, an average value was calculated from a minimum of 3 scans on P171 Evaluation of Three Catheter Maintenance Schedules on each mouse. Each data point was plotted against the animal’s Patency of Jugular Vein Catheters in CD Rats body weight to generate best-fit linear regression lines. Despite good correlation between body weight and DEXA data, 2-tailed KL Walter*1,2, Y Luo3, Y Luo3, T Fisher4, GB Mulder4 Wilcoxon matched pairs test showed a statistically significant dif- 1HarleysvilleVeterinary Hospital, Harlerysville, PA; 2Virginia- ference (P ≤ 0.03) for all 5 parameters measured. Except for total Maryland Regional College of Veterinary Medicine, Virginia tissue, the slopes of best-fit lines do not differ significantly for Tech, Blacksburg, VA; 3Charles River, Raleigh, NC; 4Charles the parameters, suggesting that the inaccuracy may be attributed River, Wilmington, MA to software differences or inappropriate calibration standards. It is possible to calculate for a variation factor for data correction, Surgical placement of jugular vein catheters (JVCs) in rats is a but validation will require comparison of body composition by common procedure to allow central venous access in conscious chemical carcass analysis. Despite inaccurate data, repeat mea- animals. Recommendations for catheter maintenance in rats, such surements taken on individual animals using the Prodigy proved as frequency of catheter flushing and types of lock solution used, to be consistent. In addition, there is no significant difference in are variable between institutions and generally based on limited the values calculated for mice scanned either as an individual or scientific evidence. We conducted a study investigating the opti- as a group, with individual bodies analyzed as a region of interest mum maintenance schedule for rats with JVCs. Sixty male Crl:CD (P > 0.05). We do not recommend the Prodigy for quantitative (SD) rats weighing 225 to 250 g were used; 30 with polyurethane analysis on rodents weighing less than 200 g without calibration (PU) JVC catheters and 30 with PU-silastic (Blended) JVC catheters adjustment. However, the consistency of data suggests its poten- originating from 2 different Charles River commercial barrier sur- tial use as a high-throughput tool for detecting general bone and gical facilities. Animals were randomly assigned to 1 of 6 groups body composition differences between animal groups. (n = 10) based on surgical facility of origin, catheter material, and maintenance frequency (catheter flushing every 3 (Q3d), 5 (Q5d) or 7 (Q7d) d after surgery). The day of surgery was considered d 0. P173 Effect of Buprenorphine Analgesia on Growth in Neo- Catheters were considered patent if blood was successfully with- natal Mice drawn. Treatment consisted of weighing each animal, aspiration of K Vail*1, A Echeverry2, R Adkins2, J Zaias1 the lock solution (50% heparinized dextrose) and blood, followed 1Division by flushing with physiologic saline and replacement of lock solu- of Veterinary Resources, 2Immunology, University of tion. Catheter patency rates for treatment groups were 100% on the Miami, Miami, FL first assessment for Q3d and Q5d, and 80% for Q7d. At d 14, 70% Experimentation and handling of neonates can often incite ma- of Q7d catheters remained patent and at d 15, 65% of Q5d catheters ternal cannibalism or neonatal mortality from lack of maternal remained patent. Thirty percent of Q3d catheters remained patent care. Therefore, studies involving neonates must be carefully at d 15. At the conclusion of the study, d 21 (Q7d) and d 20 (Q5d), done in order to obtain accurate results and ensure that the 35% and 55% of the catheters remained patent, respectively. Fifteen results obtained are from the experimental intervention rather percent of Q3d catheters remained patent on d 21. No significant than handling or anesthetic effects. Further complicating experi- difference in patency rates between catheter types (blended com- mentation with neonatal rodents is the issue of addressing pain pared with PU) were found for the Q5d and Q7d treatment groups or distress in neonates. Anesthesia and analgesia for surgery or (P > 0.966). For the Q3d treatment group, PU catheter performance procedures involving neonatal rodents are challenging. Scant was significantly improved (P = 0.007) compared to the blended information exists on the metabolism and biotransformation of catheters. In summary, patency rates were greater than 80% for each analgesics in neonatal rodents, and the routes of administration treatment group from d 3, 5, or 7 through d 10. Longer-term patency of analgesics to suckling neonates are more limited as compared rates were greatest for catheters flushed every 5 or 7 d. to adults. The purpose of this study is to determine whether there are untoward effects of administering subcutaneous analgesia P172 Evaluation of the GE Lunar Prodigy Densitometer for to young pups. The study assessed the effects on weight gain Use in Mice of subcutaneously injected buprenorphine to neonatal BALB/c mice both uninfected and infected with a pathogen (Yersinia en- KL Walter*2, ST Chou1, E Ahearn1, D Mendonca1, H Liang1, LA terocolitica) over a 2-wk period. We weighed neonates beginning Conour1 on postnatal d 7 to post natal d 23 and administered buprenor- 1Charles River, Wilmington, MA; 2 Harleysville Veterinary Hos- phine (0.05 mg/kg) twice daily for 14 d beginning on postnatal pital, Harleysville, PA d 10. There were 3 treatment groups each for uninfected and Abstracts of scientific papers 2008 AALAS National Meeting infected animals: 1 group was handled only (weighed twice nonsurgical approach to fight this disease, our group developed daily); the second group was injected twice daily with PBS; and a canine model to mimic human gastric emptying delay. The the third group was injected twice daily with buprenorphine at model increased the sensation of stomach fullness, decreasing 12-h intervals. No mortality was encountered. Weight gain did caloric consumption, which caused eventual weight loss in not differ between the noninfected groups as analyzed by 1-way the dog. An endoscopic procedure was developed for pylo- ANOVA. However, buprenorphine had an inhibitory effect on ric restriction (PR) using sutures to narrowing the pylorus. To growth in the neonates infected with Yersinia enterocolitica rela- conduct these experiments, 16 male dogs weighing between 13 tive to the handled-only and control mice. In addition, we found and 18 kg were randomized into 3 groups. Group 1 underwent that injecting mice subcutaneously affected the spleen weights the actual PR technique (8 dogs), group 2 were shams (4 dogs), compared to handled-only or infected-only mice. These findings and group 3 were controls (4 dogs). Gastric emptying was mea- suggest there might be synergistic effects between pathogens sured in all dogs using the octanoic acid gastric motility breath and analgesics that may alter development of neonatal mice. test (GMBT). Dogs in all groups were monitored daily for food This information should permit investigators to make educated intake and weight gain or loss.The PR dogs decreased their decisions regarding the effects of analgesics alone or in combina- food consumption by a mean of 48% (P < 0.002). At the same tion with infectious agents, and of potential handling or injection time, the sham and control dogs showed an 8% increase in food stress in neonatal mice. consumed, resulting in a 56% difference between the PR-sutured dogs and the sham and control groups of dogs (P < 0.0002). P174 Percutaneous Intrasalivary Gland Dosing Technique in There was a delay in gastric emptying between the presuturing Rats baseline and first postsuturing gastric emptying tests by 14% (P < 0.02). In the sham and control dogs, there was an increase in JG Tontini1, LM Serfilippi*1, DR Pallman1, CV Mujer2, CB Spain- gastric emptying between the presuturing baseline and suturing hour2 gastric emptying test by 18% (P = 0.08).We found a significant 1Department of Laboratory Animal Medicine, 2Department of delay in the PR dogs’ gastric emptying compared with the sham Toxicology, Calvert Laboratories, Inc., Olyphant, PA and control dogs’ gastric emptying. PR dogs lost 13% of their initial body weight (P < 0.03). Sham and control dogs gained Intrasalivary gland injections have been an area of increased 14% of their initial body weight. There was a 26% difference interest recently. This route has become important in develop- in the change in weight (P < 0.0002) between the PR dogs and ment of treatments for sialorrhea as well as delivery of local the sham and control dogs. Significant statistical difference was radiation and potential gene therapy. Difficulties with this dos- demonstrated in all parameters (weight loss, gastric emptying, ing technique are the ability and necessity to isolate the gland and food consumption) in PR dogs compared with sham and effectively to ensure that the dose is placed into the gland and control dogs. This canine model proved to be a reproducible not the contiguous tissues. Different modalities have been used model for an effective cure of human disease. The dog model to assist in the attainment of this goal, such as ultrasound or was suitable enough to achieve our goals of using a permanent surgical exposure of the gland. However, ultrasound requires (but reversible) nonsurgical procedure to induce weight loss by specialized equipment, is costly, and requires extensive train- restricting the pylorus. ing, while surgical exposure is associated with risk and is not conducive to repeat dosing since it induces scar tissue formation at the site. Our goal was to develop a noninvasive percutane- P176 Assessment of Chemotherapy-related Fatigue in Mice ous technique in the rat model which is easy to use and would MA Ray, R Trammell, S Ran, LA Toth* eliminate the need to use these other methods. Sprague-Dawley rats were anesthetized using isoflurane and placed in dorsal SIU School of Medicine, Springfield, IL recumbency with necks in dorsoflexion. The area over the sub- The frequent occurrence of fatigue in cancer patients and survi- mandibular salivary glands was shaved and swabbed with 70% vors and the negative impact of this problem on quality of life isopropyl alcohol. Using atraumatic tissue forceps, each gland and clinical outcome underscore the need to identify mechanisms was elevated, then isolated and a 0.1-ml injection of Evan blue that underlie fatigue, with a view toward developing treatments. was made through the skin into the glands’ parenchyma. The rats To further that goal, we developed a model for quantifying fa- were then euthanized by carbon dioxide asphyxiation and the tigue and determining its mechanisms during chemotherapy in glands were harvested. Glands were cross-sectioned to determine mice. Female BALB/cJ mice were implanted with a telemetry accuracy of the technique by noting the location of the Evan blue device that transmitted information on both core temperature dye, intraglandular compared with periglandular. With practice, and horizontal locomotor activity. Mice were also given access the success rate of delivery of dye to the submandibular gland to a running wheel in the home cage. Activity, wheel running, using this technique appears to be over 90%, which supports temperature, food intake, and body weight were monitored be- the claim that percutaneous salivary gland dosing may be an fore, during, and after administration of paclitaxel (10 mg/kg IV effective noninvasive alternative to deliver test article to the daily for 5 sequential d; n = 9 per group) either as the standard submandibular salivary glands. formulation prepared in cremophor EL vehicle (Taxol) or as the albumin-embedded nanoparticle formulation nab-paclitaxel P175 Canine Model of Obesity Using a Minimally Invasive prepared in saline (Abraxane). Taxol and Abraxane both reduced Pyloric Restriction horizontal locomotor activity and wheel running in mice. With either drug, mice showed essentially normal amounts of activity L Bright*1, K McIntyre1, M Salvatore1, A Agelan1, L Miller2, A during the first 2 h of the dark phase, indicating the ability and Vegesna2, M April1 initial desire to run during the active phase. However, during 1UniversityLabratory Animal Resources, 2Gastroenterology, the 5 d of drug administration, activity fell below normal for Temple University, Philadelphia, PA both measures during the remainder of the dark phase, with further reductions during the first week after chemotherapy. Obesity is a worldwide pandemic disease and a major concern Mice treated with Abraxane resumed normal amounts of dark- in the US healthcare system. In search of a noninvasive, quasi- Abstracts of scientific papers 2008 AALAS National Meeting phase activity during wk 2 after treatment, whereas mice treated data to support or refute these claims. In this study, the effects of with Taxol remained depressed until wk 4 after treatment, con- perioperative administration of buprenorphine, an opiate anal- sistent with the greater toxicity of Taxol and/or its cremophor gesic, and carprofen, an NSAID, were evaluated for effect on the vehicle. Assessment of potential causes of fatigue revealed that development of the CCI model of neuropathic pain. Thirty male during periods of fatigue, mice did not show anemia, elevated Sprague Dawley rats underwent chronic constriction of the left serum concentrations of proinflammatory sleep-modulatory sciatic nerve or sham surgery. Test groups received perioperative cytokines, or disturbed sleep. However, they did show mild treatment with buprenorphine (0.05 mg/kg SC twice daily for general debilitation (weight loss, anorexia, and hypothermia) 2 d), carprofen (5 mg/kg SC once daily for 2 d), or subcutane- and mild neuromuscular impairment. Thus, the combined as- ous saline twice daily for 2 d. Animals were tested at days –1, sessment of running wheel activity and horizontal locomotor 3, 7, 14, and 21 postsurgery for mechanical allodynia, thermal activity suggests that mice treated with taxane chemotherapy hyperalgesia, and incapacitance. The percentage of body weight develop fatigue. change in each group was measured for 3 d following surgery and weekly thereafter. Statistical analysis by 2-way repeated P177 Acclimation to Changes in the Light:Dark Cycle in Mice measures ANOVA demonstrated significant difference (P < 0.05) between the carprofen-treated and control group, as evidenced by CA Bosgraaf, LA Toth* higher withdrawal thresholds, higher latency times, and increased Southern Illinois University School of Medicine, Springfield, IL weight-bearing. There was no difference at any time point between the buprenorphine-treated and control group in any of the behav- Accomplishing research goals can sometimes be facilitated by ioral tests. No difference in the percentage of body weight change changing the light:dark (L:D) cycles of the experimental animals. was demonstrated between the carprofen and control groups, For example, performing sample or tissue collection during while percentage reduction in body weight was significantly the dark phase may be logistically easier if the animals are ac- greater in buprenorphine-treated rats compared to other groups climated to a reverse light cycle that allows their dark phase to on days 3 and 7. These results provide evidence that perioperative occur during normal work hours. However, prior to experimental treatment with buprenorphine can be used without significantly use, animals subjected to such changes must be permitted suf- impacting behavioral tests in the rat CCI model. ficient time to adapt physiologically and behaviorally to the new light:dark rhythm. To determine the minimal time likely to be required for such adaptation, we subjected individually P179 Interference of the Clomiphene Citrate in BALB/c/Uni housed male BALB/cByJ mice (n = 13) to a 12 h delay in dark Females Estrous Cycle onset, followed by resumption of a reversed L:D pattern. For A Salgado, M Corat*, VL Dias, A Gimenes, L Passos 14 consecutive d, mice were evaluated continuously for core CEMIB/UNICAMP, Campinas, Brazil body temperature and horizontal locomotor activity by using a telemetry device implanted in the abdomen. In addition, mice Most animal facilities that work with mouse embryos usually use had continuous access to a running wheel in the home cage. After superovulation to maximize the number of embryos for use in 7 d during which light and dark onset occurred at 0900 h and embryo banks, strain decontamination, transgenesis, and other 2100 h, respectively, the light phase was extended by 12 h, such techniques. This protocol is important into foundation colonies, that dark onset did not occur until 0900 h, with light and dark where the number of breeding pairs are limited and expensive. onset thereafter maintained at 2100 h and 0900 h, respectively. Usually pregnant mare serum gonadotropin (PMSG) is used for Mice showed reduced locomotor and running wheel activity for follicular stimulation and for synchronization of estrus. Due to approximately 24 h after the L:D change occurred. However, both the expense and delivery time of this hormone to South America, activities then returned to basal levels, with the normal circadian we decided to explore alternative methods of follicular stimula- pattern of maximal activity during the dark phase. During the tion. Our objective was evaluate the use of clomiphene citrate baseline period, peaks and nadirs of core temperature occurred to influence the ovulation of BALB/c/UNI mice females. All during the dark and light phases, respectively. After the time the animals used in this work were specific pathogen-free and change, temperature peaks were reduced for approximately 2 were supplied and housed in a sterile environment in ventilated to 3 d. Thereafter, normal rhythms resumed, although the circa- racks. Female mice were dosed by gavage with clomiphene cit- dian temperature range appeared greater than during the basal rate (0.250, 0.375, 0.500, or 0.750 mg). On d 3, females received period. Taken together, these data indicate that the behavioral 10 UI of human chorionic gonadotropin (HCG) and were bred (activity) and physiological (temperature) stabilization of mice with males. On d 1.5 of gestation, we collected the embryos (2- to drastic (12 h) changes in light cycles occurs within 72 to 96 h. cell stage) and checked their integrity. This methodology was However, because other parameters (such as hormone secretion) compared with superovulation by PMSG and natural ovulation. may nonetheless adapt more slowly, longer acclimation periods Clomiphene citrate outcomes were promising. The synchroni- may be warranted. zation of estrus with PMSG was highest, but with clomiphene citrate synchronization was significantly higher than control, P178 Evaluation of Perioperative Analgesia on Behavioral Tests which did not receive any hormone stimulus. Our results show in the Chronic Constriction Injury Neuropathic Pain Model that the number of viable eggs was the same between groups, although the number of collected embryos was higher in the LE Clarey*, C Huang, L Jackson group stimulated with PMSG. We conclude that clomiphene Biogen Idec, Cambridge, MA citrate is an acceptable alternative to PMSG for reproductive studies, including those related with embryo banks. Surgical models of neuropathic pain are used to evaluate candi- date therapeutic agents for clinical use. Investigators have been reluctant to provide perioperative analgesia in surgical models P180 Evaluation of Different Ovariectomy Techniques on the of neuropathic pain, citing concerns that the anti-inflammatory Induction of Experimental Menopause in Rats properties of analgesics would affect model development or con- M Guzman-Silva*1, HN Henriques2, AB Carvalho3, C Pollastri3, found interpretation of behavioral tests. There is little published JS Pantaleão3 Abstracts of scientific papers 2008 AALAS National Meeting 1Pathology, 2Biomedical School, 3Postgraduation Program in P182 Multiphase Surgical Approach for the Evaluation of Pathology, UFF, Niteroi, Brazil Active and Passive Fixation Pacing Leads for Telemetered Electrocardiograms in the Dog Many researchers use ovariectomy in rats for induction of meno- pause and evaluation of varied aspects related to the lack of sex MG Taschwer* hormones or the effect of hormonal replacement. Details of the Department of Laboratory Animal Medicine, Covance Labora- technique and the success of ovariectomy is not always reported. tories Inc, Madison, WI Two different ovariectomy techniques were compared and evalu- ated through vaginal cytology at the time of surgery and after The use of pacing leads in conjunction with telemetery trans- 30 d. Bilateral oophorectomy was performed in 35 adult female mitters has become a popular method for the acquisition of Wistar rats (222 ± 19 g) under xylazine and ketamin anesthesia. relatively artifact free electrocardiograms (ECG). This study In one group (OVX, n = 20), ovaries identified at laparotomy employed a multiphase surgical approach to compare the per- were excised; in the other group (OVX+PF), ovaries plus about formance of active and passive fixation intracardiac (IC) pacing 1 cm of surrounding periovarian fat were removed. Vaginal leads. Four purebred beagles were used per IC fixation type. In smears were fixed in 95% alcohol, stained (Papanicolaou), and Phase I pacing leads were introduced into the right ventricle classified by microscopy as trophic (estrus, metestrus, diestrus, through the jugular vein and guided into position by fluorogra- proestrus) or atrophic (anestrus). At the time of surgery, all rats phy and ECG signal. The distal ends of the leads were tunneled showed trophic smears. One month later, estrus vaginal citology into the subcutaneous tissue layer over the right lateral thorax. was found in 2 OVX rats; all OVX+PF rats showed anestrus vagi- Surgical phase II was performed 1 wk later. In this phase, a small nal citology. Effective menopause status was achieved in 100% incision was made to expose the distal IC connector and con- OVX+PF and in 90% OVX. No statistical difference was found nect it to a sterile transmitter for evaluation of the ECG signal. (Z statistic), but biological implications are evident since animals Candidates for transmitter implantation were selected based still showing sex hormonal influence can not be used in any kind on this evaluation. The final surgical phase required 6 IC-lead– of forward research where the menopause condition is required. implanted animals to receive telemetry transmitters. Results The refinement introduced in the ovariectomy technique led to indicated 2 of the 4 passive fixation pacing leads shifted position improved effectiveness and increased the number of research within 1 wk post implantation, creating less desirable ECG mor- subjects available under menopause status. phologies. One of the 4 passive leads dislodged completely 4 wk post implantation. In contrast, all 4 active fixation pacing leads remained in their implanted positions allowing these animals to P181 Species Variation in the Heparin Contaminant-mediated be chosen for transmitter implantation and further testing. The Generation of Kallikrein multiphase surgical approach employed two minor procedures M Clark*1, E McGeehan2, O Iqbal1, C Adiguzel1, J Cunanan1, D prior to a major surgery. This approach provided an intermit- Hoppensteadt1, J Fareed1 tent step to verify good IC lead placement and signal quality 1Department of Pathology, 2Comparative Medicine Department, after 1 wk of healing. Although this intermittent step required Loyola University Medical Center, Maywood, IL additional anesthesia and time, doing so confirmed that the ani- mal selected for implantation with the telemetry device would One of the major pathopysiologic responses related to the adverse be a viable candidate for test and would not require corrective reactions due to recalled contaminated heparins is attributed to surgery to remove and reimplant the IC lead. In addition, study the generation of kallikrein, leading to the formation of brady- time schedules would not be disrupted for corrective surgery. kinin. The generation of kallikrein is reported to be due to the We conclude that the performance of the active fixation lead, deliberate adulteration of heparin by hypersulfated chondroitin with respect to maintaining its implanted position, was superior sulfate. The contaminated heparins were found to contain up to to the passive fixation lead. The multiphase surgical approach 30% of this contaminant on a weight basis in heparin. The re- provides the advantage of evaluating the IC lead position and ported hypotensive effects of the contaminated heparin and the ECG quality prior to implantation of the telemetry unit. contaminant were found to be species dependent. To further in- vestigate the variations in the contaminant-mediated generation of kallikrein in different species, citrated plasma samples from P183 Establishment of DMN-induced Liver Cirrhosis Model human, dog, rat, cow, rabbit, and monkey were supplemented in NOD/SCID Mouse for Evaluation of the Efficacy and Safety with contaminated heparin (CH) and contaminant-free heparin, of Human Mesenchymal Stem Cells the isolated heparin contaminant (HC), and hemisynthetic hy- M Hyon *1, E Kwon 1, H Choi 1, M Jang 1, K Cho 1, J Che 1, B persulfated chondroitin sulfate (HCS-1 and HCS-2). In human Kang1,2 plasma, all agents produced varying degrees of concentration- 1Department of Experimental Animal Research, Clinical Research dependent activation of kallikrein formation (range, 0.3 to 0.7/15 Institute, Seoul National University Hospital, Seoul, South min). In the monkey plasma, the activity was somewhat weaker Korea; 2 Collage of Medicine, Seoul National University, Seoul, (0.13 to 0.36/15 min). The generation of kallikrein was found to South Korea be higher in dog plasma (0.27 to 0.65/15 min). However, in rat plasma relatively weaker activity was observed (0.12 to 0.18/15 Recently, many studies have suggested the potential of human min). Interestingly, no activation of prekallikrein was noted in the mesenchymal stem cells (hMSC) to differentiate into a hepatocyte- cow plasma. Various agents produced different degrees of activa- like lineage. In this study, we tried to establish an animal model tion with the rank order of HC > HCS preparations > CH. CFH of dimethylnitrosamine (DMN)-induced liver cirrhosis in NOD/ showed the least activity. These variations in the activation of SCID mouse for evaluation of the efficacy and safety of cell therapy the contact system in different species may be responsible for the such as human mesenchymal stem cells. NOD/SCID mice were differences in the observed hemostatic and vascular responses. treated with DMN (1, 5, or 10 mg/kg IP) for 3 consecutive d per These observations should be considered while screening the wk for 5 wk and sacrificed at week 3, 4, 5, and 6 after first DMN effects of drugs and agents with vasoactive actions. injection. In the 10 mg/kg group, most mice were critically ill or died. In the 5 mg/kg group, there were obvious clinical signs such Abstracts of scientific papers 2008 AALAS National Meeting as abdominal distention and piloerection. However, there was no mechanisms and interventions of diabetes and “diabesity” (dia- difference between the control group and the 1 mg/kg group in betes with obesity). Automated clinical chemistry analyzers have clinical findings. Based on serum biochemistry for evaluation of been used for clinical chemistry and glucose monitoring of rats the liver function, ALT, AST, ALP, and total bilirubin in the 5 mg/ and larger species, but blood volume requirements often in excess kg group were significantly increased and albumin was decreased of 100 µl, and the time to process serum and plasma for typical compared with the 1 mg/kg or control group. In histopathological diagnostic clinical chemistry instruments preclude practical use findings in the liver, there were central venous necrosis, hemor- with mice. Hand-held glucometers are used for diabetic patients. rhage, and fibrillar septa formation in the 5 mg/kg group at 4 wk. They are gaining popularity for mouse studies, but comparisons The extracellalar matrix (collagen), iron, and glycogen deposition of different instruments and validation against clinical chemistry were confirmed by Masson trichrome (MT), Prussian blue (PB) and analyzers for mouse specimens are difficult to find. We compared periodic acid Schiff (PAS) staining, respectively. We also found 2 glucometers that require < 10 µl of whole blood, Accu-Chek degenerative changes of renal structure and splenomegaly in the Compact Plus and OneTouch Ultra2, with each other and against 5 mg/kg group. In conclusion, treatment of NOD/SCID mice our VetACE clinical chemistry system for glucose measurement. with 5 mg/kg of DMN for 4 wk can be used to create a pertinent We collected 33 blood samples between 1100 and 1130 h, over a liver cirrhosis model and can be useful to evaluate the efficacy and 2-wk period by cardiac puncture of adult mice of various age, safety of cell therapy such as human mesenchymal stem cells. sex, and strain. Fresh blood specimens were tested immediately on each glucometer. The remaining blood was separated into P184 Sleep Deprivation Impairs Acquisition, Consolidation, EDTA tubes (to be tested 15 min later) and serum separator and Retrieval of an Ethical Discriminative Avoidance Task tubes. Sera were tested within 2 h of collection. Data were ana- in Rats lyzed to compare the instruments and condition effects (whole fresh blood, whole blood EDTA, and serum) on each specimen TA Alvarenga1, ML Andersen*1, CL Patti2, RH Silva3, MB Calza- using 2 tailed paired t-test. Significant and consistent differences vara2, GB Lopez2, R Frussa-Filho2, S Tufik1 (SD < 0.05) were found between glucometers both immediately 1Psychobiology, 2Pharmacology, Univ Fed Sao Paulo, Sao Paulo, after blood extraction and after 15 min at room temperature Brazil; 3Physiology, Univ Fed Rio Grande do Norte, Rio Grande in EDTA. Whole fresh blood results from ACCU-CHEK were do Norte, Brazil similar to serum results of our analyzer. Results from OneTouch were slightly but significantly higher than the analyzer and The plus-maze discriminative avoidance (PM-DAT) is a new ani- ACCU-CHEK. EDTA blood results from both glucometers were mal model of learning and memory that provides simultaneous significantly lower than the analyzer. Both glucometers yielded information about anxiety. This behavioral paradigm complies consistent results that are comparable to the chemistry analyzer. with ethical protocols, since there is no foot shock as imposed The low cost, cage-side operation, small volume requirement, by classical avoidance tests. As sleep deprivation promotes and rapid results of these glucometers make them a practical tool. important behavioral alterations in both humans and rodents, However, results from different instruments, or from different the aim of the present study was to investigate the effects of specimen types may be difficult to compare. Protocols should paradoxical sleep deprivation (PSD) for 96 h on the learning and be developed to standardize specimen handling and testing to memory processes in rats submitted to the plus-maze discrimina- optimize utility. tive avoidance task (PM-DAT), which simultaneously evaluates learning, memory, anxiety, and motor function. Four experiments were performed in which rats were submitted to: I) posttraining P186 Effects of Blood EDTA Saturation on Selected Mouse combined with pretest PSD; II) posttraining or pretest PSD; III) Hematology Variables pretraining PSD or paradoxical sleep (PS) rebound (24 h); and NA Forbes*, CF Brayton IV) pretest PS rebound. Posttraining PSD combined with pretest PSD induced memory deficits, an anxiolytic-like behavior, and an Molecular and Comparative Pathobiology, Johns Hopkins School increase in locomotor activity. Both posttraining PS-deprived and of Medicine, Baltimore, MD pretest PS-deprived groups showed memory deficits; however, Ethylenediaminetetraacetic acid (EDTA) is a widely used antico- only the pretest PS-deprived animals presented anxiolytic-like agulant for blood submission for complete blood count (CBC). behavior and increased locomotor activity. In experiment III, Blood tubes contain the appropriate amount of dry (powder) pretraining PS-deprived rats showed learning and memory defi- or liquid anticoagulant for the recommended blood volume as cits, anxiolytic-like behavior, and increased locomotor activity. A indicated on each tube. Overfilling blood collection tubes results 24-h sleep recovery period after the PSD abolished the learning in insufficient chelating of calcium and blood clots. Underfilling and memory deficits, but not anxiety and locomotor alterations. tubes is reported to result in cell shrinkage and potentially al- Finally, sleep rebound did not modify acquisition (experiment tered blood cell parameters and morphology. Many mouse blood III) and retrieval (experiment IV). This study strengthened the submissions to our laboratory are less than 20% of the recom- critical role of sleep (but not sleep rebound) in all the steps of mended volume for the anticoagulant tubes (40 to 70 µl of blood learning and memory formation. In addition, it suggests that PSD in a 500-µl tube). The purpose of this study is to determine the effects on acquisition and consolidation do not seem to be related optimal blood volume for EDTA K2 to have confidence in RBC to other behavioral alterations induced by this procedure. parameters and CBC results. Cardiac blood samples obtained at euthanasia from 40 adult mice of various ages, sexes, and strains P185 Glucometer Validity in Mouse Research were used to investigate the effect of underfilling anticoagulant blood tubes on CBC parameters obtained with our Hemavet NA Forbes*, CF Brayton 950 impedance-based blood analyzer. Becton Dickinson 500 µl Molecular and Comparative Pathobiology, Johns Hopkins School Microtainer EDTA K2 tubes filled with 50, 100, 175, 250, and 500 of Medicine, Baltimore, MD µl of blood (10%, 20%, 35%, 50% and 100% of the recommended tube volumes, respectively) were analyzed by our Hemavet 950 Blood glucose monitoring is essential to various metabolic instrument, and results were examined by paired t-test. At 10% studies, especially those of genetic, dietary and therapeutic Abstracts of scientific papers 2008 AALAS National Meeting and 20% of recommended tube volume, hematocrit, WBC, RBC, separate from the video recording devices. New technology in and Hb were found to be significantly lower than those with telemetry which uses time-synchronized video cameras seems 100% recommended volume of 500 µl (P < 0.05). At 35% recom- to be the solution. Here we describe the use of the synchronized mended volume, WBC and Hb were significantly lower than video function of our validated telemetry acquisition and analy- at 100% recommended volume. At 50% recommended volume, sis system to identify behavior and positional-associated artifacts. there were no significant differences between values (P > 0.05). ECG, blood pressure, and body temperature were acquired from In conclusion, within the parameters of this study relevant to 5 telemetered cynomolgus monkeys (Macaca fascicularis). ECG submissions to our laboratory, multiple CBC parameters may be and blood pressure waveforms and body temperature were ex- impacted by underfilling of EDTA K2 anticoagulant tubes. We amined in correlation with the time-synchronized videos. Four currently use and recommend 100-µl anticoagulant tubes, filled animals exhibited T wave morphological changes associated with to at least 50% and not overfilled, for mouse blood submissions positional changes. Noisy blood pressure waveforms were noted to our laboratory. There are several options for anticoagulants, in all animals when they scratched with their leg containing the blood-collecting tubes, and the analyzers, and it is the responsi- pressure catheter (motion-related artifact). Wet-dog–shaking bility of investigators to use appropriate protocol and techniques behavior observed in all animals caused characteristic fluctua- for optimal and valid results. tions in both ECG and blood pressure waveforms (myogenic and motion-related artifacts). Body temperature was not affected by P187 New Method for Determining Urine Frequency in Rats the animals’ behavior/positioning. Based on our observations, animal behavior and position are important factors to consider NN Nikolaidis*, R Guinn in the analysis of telemetry data. A time-synchronized video R&D, Bioanalytical Systems, Inc, West Lafayette, IN system allows for accurate identification of physiological artifact in telemetry data. Urine is often collected in pharmacological studies in order to analyze metabolic compounds. These analyses often require that urine remain chilled during collection. Measurements of P189 Mouth Cell Collection Device for Newborn Rodents urine volume and urinary frequency can also provide useful NK MacLennan*1, Y Zhang1, B Huang1, K Eastman1, M Bedernik1, data during these experiments, especially in animal models with LL McCabe1, E McCabe1,2 altered urination, such as diabetes or overactive bladder. While 1 Pediatrics/Human Genetics, 2 Bioengineering, UCLA, Los measuring urine volume over time is relatively simple, tracking Angeles, CA urinary frequency can be more difficult. We developed a simple method to automatically track urine frequency in rats during For efficient and accurate genotyping of transgenic and knockout urine collection, while keeping the urine chilled for further me- mice, the ability to reduce pain and suffering and to obtain DNA tabolite analysis. Using metabolic cages and funnels, urine was early in life are critical. We have developed a novel method to collected into scintillation vials placed into a chilling chamber, sample buccal cells from neonatal mice to obtain DNA. Our which chilled the urine to 0 °C. A temperature probe was placed mouse mouth cell collection process includes an oral speculum into the vial, and the temperature of the interior was recorded by and collection device which enables rapid extraction of enough a computer once every 10 s. Each urinary event caused a transient DNA for up to 50 PCRs from each buccal sampling. This cell increase in vial temperature. Eight male Sprague Dawley rats collection device fills a clear need for buccal sampling from were placed in metabolism cages. Urine was collected for 24 h to neonatal mice, greatly facilitating research in mouse models of provide baseline frequency and volume data. The rats were then human disease. We have data to support a decreased incidence injected intraperitoneally with 1 mg/kg oxybutynin (a bladder of cannibalism of pups by mothers with use of this device. smooth muscle inhibitor), and urine was collected for another 24 Eliminating the pain and distress caused by traditional cell collec- h. After a 24-h washout period, the rats were injected intraperi- tion procedures lessens the potential for confounding variables toneally with 20 mg/kg furosemide (a diuretic), and urine was between control and experimental animals. In conclusion, our again collected for 24 h. Oxybutynin did not significantly affect mouse mouth cell collection process can be applied to very small urinary frequency. However, furosemide caused an immediate animals for which there exists no current device. increase in urinary output. Both urinary frequency and overall urine volume increased in furosemide-treated animals. The P190 Prevalence of Bordetella hinzii in Laboratory Mouse temperature tracking method clearly indicated the time of each Facilities in Japan urination event, and distinguished urinary events as close as 90 s apart. This method allows urinary frequency to be tracked easily N Hayashimoto*, M Yasuda, T Ishida, S Kameda, K Goto, A during a urine collection study, and can provide additional data Takakura, T Itoh during metabolic studies. Central Institute for Experimental Animals, Kawasaki, Japan Bordetella hinzii is commensal in the respiratory tracts of fowl, and P188 Identification of Behavior- and Position-related Artifacts has been previously shown to have some pathogenic potential in Nonhuman Primate Telemetry Data Using a Time-synchro- in immunocompromised humans. Isolation of B. hinzii from nized Video Telemetry System nonhuman mammalian species remains extremely rare. Recently, N Crosby*, Y Mori, K Maulding, J Montalbano, D Moskowitz, E we isolated B. hinzii from the trachea and lungs of a C57 BL/6 Bou-Abboud, S Meyer mouse showing sneezing with a chattering noise. The agent also induced respiratory disease in the same way as in the initial case Scientific Services, SNBL USA, Ltd., Everett, WA in ICR and NOD-SCID mice in our experimental study. Although In nonhuman primate telemetry studies, abnormal data can be the etiological role of B. hinzii in mice was suggested by our observed due to physiological artifacts. Animal behavior and study, the prevalence of B. hinzii in laboratory mouse facilities position may cause physiological artifacts ,and should be consid- has never been reported, and the actual figure is still unknown. ered in telemetry data analysis. It can be difficult to identify these The objective of this study is to reveal the prevalence of B. hinzii artifacts using the traditional telemetry configuration, which is in laboratory mouse facilities in Japan. A survey was performed Abstracts of scientific papers 2008 AALAS National Meeting on a total of 6,040 mice from 1,020 facilities (5,254 mice from 890 Central pain has been well documented in humans and occurs facilities in university research institutes and 786 mice from 130 with lesions all along the neuraxis involving neural structures of facilities in pharmaceutical companies) in Japan for microbiologic the spinothalamocortical pathway. However, most animal models monitoring at the Central Institute for Experimental Animals of central pain are based on spinal cord injury. The objective of this from April 2007 to January 2008. The detection of B. hinzii was study was to evaluate pain perception in rats following an intrac- performed by culture of tracheal swabs using horse blood agar, erebral hemorrhage in basal ganglia. Twelve male Sprague-Dawley and suspected colonies were identified by phenotypic tests and rats weighing between 175 and 250 g were used. Two groups of 6 PCR using B. hinzii specific primers designed for this study. In animals were used to evaluate thermal sensitivity (Hargreave test) university research institutes, 88 out of 5,254 mice (1.6%) derived to a progressive intensity of light set at 35 or 50% (groups 1 and 2, from 24 out of 890 facilities (2.7%) were positive. On the other respectively) of the maximal output. Following 3 d of behavioral hand, no B. hinzii-infected mice were found in 130 pharmaceuti- testing (baseline values), rats were injected intracerebrally with cal companies surveyed. The overall prevalence of B. hinzii in 0.5 U collagenase Type VII in the right caudoputamen nucleus, mouse facilities in this survey was 2.4%. No clinical symptoms inducing a hematoma. They were then tested for the next 7 to and no gross lesions were observed in any B. hinzii-infected 9 consecutive d. With the light intensity set at 35%, no changes mice, but moderate sinusitis and mild rhinitis were observed were observed following surgery when compared to presurgical in all selected 14 B. hinzii-infected mice free from other known baseline results. However, at 50% of maximal output, reaction pathogens by histopathology. Our survey provided evidence for times were significantly faster on days 3, 4, 5, 6 (P < 0.0001) and 7 the prevalence of B. hinzii in laboratory mouse facilities in Japan, (P < 0.006). Rats returned to presurgical baseline reaction times on and indicated the pathogenicity of B. hinzii in naturally infected days 8 and 9 (P = ns). Histopathology showed that the lesion was mice. Based on these findings, B. hinzii should be eliminated from located mainly in the caudoputamen nuclei; however, the internal laboratory mouse facilities. capsule and the thalamus were also affected. In conclusion, a tran- sient central pain was induced in rats with a collagenase-induced P191 Quantitative Comparison of Ovarian FSH receptors intracerebral hemorrhage of the caudoputamen nuclei most among Six Mouse Strains probably associated with lesions of the internal capsule as well as the thalamus. These results suggest that animal models used O Suzuki* for studies other than pain, and with lesions affecting structures Laboratory of Experimental Animal Models, National Institute along the spinothalamocortical pathway, should be evaluated for of Biomedical Innovation, Ibaraki, Japan pain perception, as this may have an impact on the interpretation of the results. Superovulation induction by gonadotropin treatment is widely used in reproductive experiments. Previously, I showed that the differential response among various mouse strains to gonadotro- P193 Spontaneous Tumors in VEGFP-GFP/FVB and Tie2P-GFP/ pins is still a serious problem for practical applications. In order FVB Transgenic Mice to elucidate the mechanism underlying the strain difference in P Huang*, M Dawson, R Lanning, RK Jain, D Fukumura superovulation induction, I planned to compare the FSH-receptor Radiation Oncology, Massachusetts General Hospital, Boston, in various mouse strains qualitatively and quantitatively. I have MA already reported that no difference was found in the FSH-receptor amino acid sequences among 12 mouse strains. In this study, I com- The phenomena in our previous observations indicated that the pared ovarian FSH-receptor mRNAs and proteins among mouse presence of green fluorescence protein (GFP) reporter trans-genes, strains quantitatively. Total RNA (5 mice per strain) and proteins such as VEGFP-GFP and Tie2P-GFP, may affect the incidence of (4 mice per strain) were extracted from the ovaries of 4-wk-old fe- spontaneous tumors as well as their histopathological and biological males of 6 strains: C57BL/6Cr, A/J, DBA/1, DBA/2, C3H/HeJ, and characteristics in transgenic mice with FVB/N background. In this BDF1. The FSH-receptor mRNA was quantified using quantitative study, a total of 94 retired transgenic mice breeders (14 males and real-time RT-PCR, with normalization by the amount of GAPDH 80 females; 37 of VEGFP-GFP/FVB and 57 of Tie2P-GFP/FVB mice) mRNA. The amounts of FSH-receptor (approximately 70 kDa) from our defined-flora mice facility were observed during their and β-tubulin (approximately 55 kDa) proteins were measured natural lifespan. Our preliminary results showed the incidence of using Western blot analysis, visualizing band fluorescence with spontaneous tumors was 71.4% (10/14) in males and 57.5% (46/80) two rabbit antibodies against mouse FSH-receptor and β-tubulin, in female transgenic mice. Primary tumors found in the males were respectively, as primary antibodies, and a fluorescent-labeled sec- 3 liver hepatocellular carcinomas, 2 lung alveolar-bronchiolar (AB) ondary antibody against rabbit IgG. The amounts of FSH-receptor tumors, 2 lymphomas, and 3 others. In females, pathologic examina- proteins were normalized with those of β-tubulin proteins. Neither tion revealed 27 mammary tumors, 3 liver hepatocellular tumors, 3 transcripts (P = 0.54) nor proteins (P = 0.29) differed significantly lung AB tumors, 3 fibrosacomas, 3 lymphomas, and 7 others. Com- among strains judged using 1-way ANOVA (P > 0.05). Therefore, it pared to our wild-type FVB/N retired breeds, these transgenic mice is unlikely that the strain difference in the response to gonadotropin demonstrated a significantly lower incidence of spontaneous lung treatment is due to differences in the structure or abundance of AB tumors (P < 0.01) and a higher rate of liver tumors (P < 0.01). ovarian FSH-receptors. As the next target, LH-receptors should be Examination by confocal microscopy found that GFP was expressed compared among mouse strains to elucidate the mechanism of the strongly in fibrosarcoma cells (arose in VEGFP-GFP/FVB), or in strain difference in superovulation induction. adenocarcinoma stromal cells (arose in VEGFP-GFP/FVB and Tie2P- GFP/FVB mice). Several of these tumors have been transplanted P192 Transient Central Pain following Collagenase-induced into the 6- to 8-wk-old transgenic and wild-type FVB/N mice. When Intracerebral Hematoma in Rats these isografts grew in secondary FVB/N mice, the fibrosarcomas continuously expressed GFP in their tumor cells; the adenocarcino- H Orlando, C Savard, P Vachon* mas brought their own stromal cells (such as endothelial cells) to the Veterinary Biomedicine, University of Montreal, St-Hyacinthe, new hosts, and these stromal cells could express GFP and survive QC, Canada for more than 2-wk postimplantation. These histopathological and Abstracts of scientific papers 2008 AALAS National Meeting biological characteristics of spontaneous tumors arising from the mutations in Lep/Lepr are rare in the human population, the transgenic mice are clearly different than those of spontaneous tu- ZDSD/Pco diabetic (type II) rat model has been designed and mors in wild-type FVB/N mice. In particular, a significantly lower developed without these mutations. This rat model was initiated incidence of spontaneous lung AB tumors in the transgenic mice by crossing a diet-induced obese (DIO) rat that was derived was observed. Further studies to better understand the differences, from the Crl:CD(SD) strain (exhibiting insulin resistance and such as lung AB tumor incidence, within these spontaneous tumors polygenic obesity) with a homozygous lean ZDF/Crl rat (which in transgenic and wild-type FVB/N mice are warranted. will express beta cell failure with the Leprfa/Leprfa genotype). Selective inbreeding for obesity and diabetes (19 generations) has P194 Acute Oxidative Injury Promotes Venous Endothelial resulted in a strain that exhibits these characteristics without the Dysfunction without Thrombogenesis in a Rodent Model Leprfa mutation. At 6 wk of age the ZDSD males average 217 g, compared to 187 g for the obese ZDF/Crl-Leprfa male rats. Only RM Moore*1,2, A Hawley2, D Farris2, S Wrobleski2, R Sigler3, D males develop diabetes, with glucose levels averaging >500 mg/ Myers2,1 dl. The strain also expresses high insulin levels before the onset 1 Unit for Laboratory Animal Medicine, 2 Jobst Vascular Re- of diabetes (approximately 5 ng/ml), which decrease with the search Laboratories, Section of Vascular Surgery, University of progression to overt diabetes to <1 ng/ml. This rat also exhib- Michigan, Ann Arbor, MI; 3 Walker Downey & Associates, Inc., its hyperlipidemia with triglyceride levels of >600 mg/dl and Verona, WI cholesterol levels of >100 mg/dl. Micro-albumin excretion is >75 mg/d. The majority become diabetic between 15 and 21 wk of age Oxidative stress has been associated with many cardiovascular on Purina 5008. Dietary manipulation can reliably prompt and diseases, including endothelium dysfunction. Little is known synchronize the onset of diabetes in animals that are not already regarding the role of oxidative injury to venous endothelium, diabetic by dietary supplementation with corn syrup (30% solids and we sought to achieve this using a rodent model. We hy- in an extra water bottle) or a change to Purina 5015 or Research pothesized that venous endothelial dysfunction subsequent Diets D12468 to synchronize the onset of diabetes. Treatments to oxidative injury would activate adhesion molecules, thus with metformin (150 mg/kg twice daily); rosiglitazone (3 mg/ promoting venous thrombosis in a rat model. Sprague Dawley kg twice daily) and exenitide (1 µg twice daily) prevent the rats (n = 72) underwent photochemical injury to the inferior onset of diabetes in this model. In summary, the ZDSD/Pco rat vena cava (IVC) by injection of Rose Bengal dye (50 mg/kg, IV) is an important new model that expresses obesity and diabetes and green light laser (540 nm) exposure. Experimental groups without apparent alterations in the leptin pathway. This novel included controls, shams (Rose Bengal, but no laser), and ani- model appears to more closely mimic the human condition, than mals that underwent oxidative injury. Groups were evaluated current models, since it is obese and develops diabetes without at baseline, 1 h, and 1 d post-oxidative injury (POI). Vein well leptin pathway defects. It is thus likely to become more useful inflammatory cell counts per high power field, gene array (fold than other rat models in testing anti-diabetic and anti-obesity change relative to aldolase), real-time reverse transcriptase compounds for human obesity and diabetes. polymerase chain reaction (expression relative to β-actin), tis- sue inhibitor of metalloproteinase 1 (TIMP-1) levels (pg/mg total protein), and tissue factor activity (pM/mg total protein) P196 Alternative Method for Intrathymic Injections in Mice were evaluated. Polymorphonuclear cells significantly increased R Blair*2, SL Hoogstraten-Miller1, K Mueller2, P Schwartzberg2 in the 1-h group compared with controls (26 ± 13.5 compared 1Office with 6.6 ± 4.6 cells/5HPFs, P < 0.05). Gene array and real-time of Laboratory Animal Medicine, 2Genetic Disease Re- polymerase chain reaction (RT-PCR) identified angiotensinogen search Branch, NIH/NHGRI, Bethesda, MD (AGT) and L-selectin as genes upregulated due to injury, while The thymus is an organ located in the anterior portion of the metalloproteinase-14 (MMP-14) was downregulated. Plasma thoracic cavity, just behind the sternum. The thymus is the site of TIMP-1 levels significantly increased at 1 h (P < 0.05) and 1 d (P development of T cells, which are a critical part of the immune < 0.01) compared with controls POI. Vein wall TIMP-1 levels at system, required for fighting infection. The widespread use of 1 d were also elevated compared with controls (P < 0.01). Tissue transgenic and gene-targeted mice has allowed expansion into factor activity was significantly decreased in both the 1-h (1.95 many important areas in the study of T cell development. Because ± 0.5 compared with 4.0 ± 0.8 pM/mg total protein, P < 0.05) of its crucial role in immunity, the thymus is a frequent target of and 1-d (1.5 ± 0.3 compared with 4.0±0.8 pM/mg total protein, experimental manipulation in the quest for understanding various P < 0.01) POI groups compared with shams. Oxidative injury to immune processes in these animals. Our lab wanted to perform the rat vena cava produces moderate endothelial dysfunction, intrathymic injection with fluorescein isothiocyanate (FITC) to characterized by altered gene expression and inflammatory cell locally label thymocytes and follow the fate of developing T cells populations. This study is the first to demonstrate that focused as they exit the thymus and enter the peripheral circulation in oxidative injury to venous endothelium alters cellular transcrip- mouse models of primary immunodeficiencies. Intrathymic injec- tional properties without acute thrombogenesis in this model. tion has traditionally been performed using surgical procedures to open the chest cavity for direct visualization and injection into P195 ZDSD/Pco Rat: A Model for Diabetes, Metabolic the thymus. This technique subjects the mice to significant pain Syndrome and Obesity without Leptin or Leptin Receptor and distress, which can alter T cell development and potentially Mutations increase mortality. Intrathymic injections can also be performed by blind injection into the thoracic cavity. However, the success RG Peterson*, KL Coy, Y Wang, CR Stauss, KA McAndrews, W rate for this technique is usually low. In order to improve suc- Yeh cess rates with nonsurgical intrathymic injections, our lab has PreClinOmics, Inc, Indianapolis, IN performed intrathymic injections in isoflurane-anesthetized mice using ultrasound (US)-mediated guidance. The success rate us- The purpose of this study is to develop an obese-diabetic rat ing this technique has been high, as the needle and injectate can model that does not have monogenic mutations such as those both be visualized with the US probe as they enter the thymus. that produce leptin (Lep) or leptin receptor (Lepr) defects. Since Abstracts of scientific papers 2008 AALAS National Meeting Complications have been very rare and include the accidental University, New York, NY; Research Animal Resource Center, laceration of an intrathoracic vessel, which resulted in the eutha- Memorial Sloan-Kettering Cancer Center and the Weill Medical nasia of the mouse; this complication occurred in 1 mouse out College of Cornell University, New York, NY of approximately 85 injected to date. Moreover, it is important Myobia musculi (MB) and Myocoptes musculinus (MC) are 2 of to note that even though this mouse was euthanized, we were the most common fur mites affecting laboratory mice. Detection able to see the immediate hemorrhage on US and euthanize the can be difficult due to low levels of infestation or inadequate mouse while it was still anesthetized, thereby avoiding any pain monitoring programs. During a recent outbreak, we tested the or distress. Our results suggest that use of US-mediated guidance sensitivity of our sentinel monitoring program to detect mites. We is a refinement that can help reduce morbidity, distress and animal compared skin scrapes (SS) from 3 different sites (head, back, and usage in experiments involving intrathymic injection. ventrum) and found that mites were most prevalent on the head. Contact sentinels, Tac:(SW), tested positive for either MB or MC P197 Efficacy of Ad Libitum Antibiotics as Countermeasures within 2 d of exposure to infested mice (C57BL/6NTac). We then against Acute Radiation Syndrome in C57BL/7 Mice exposed naïve Crl:CFW(SW) mice to different concentrations of A Plett1, R Crisler-Roberts*2, H Chua1, N Nguyen1, ES West1, KJ mite contaminated/clean bedding (100/0%, 50/50%, 25/75%, Kohlbacher1, CH Sampson2, CA Johnson2, BE Juliar3, BM Katz3, 2.5/97.5% and 0/100%; groups A through E, respectively). LJ Brossia2, CM Orschell1 Each group consisted of 1 cage with 2 mice. SS were performed weekly on every animal until both mice per cage tested positive Department of Medicine, 1Division of Hematology/Oncology, on 2 consecutive wk. Groups A, B, and C tested positive on wk 2Laboratory Animal Resource Center, 3Division of Biostatistics, 1, 2, and 3, respectively. Group D first tested positive on wk 7. Indiana University School of Medicine, Indianapolis, IN All groups remained positive the following week after the first Our laboratory has developed a murine model of the hematopoi- positive test. Group E remained negative. As mite eggs are per- etic syndrome of the acute radiation syndrome (HS-ARS) for manently attached to the hair shafts, they persist on the animal efficacy testing of potential radiomitigators. Since first-line after successful treatment, potentially confounding posttreat- treatment after irradiation includes antibiotics and wetted feed, ment confirmatory testing. Four BALB/cAnNCrl, Crl:CFW(SW), feed and water consumption were evaluated in irradiated mice and Crl:CD1(ICR) mice were colored with hair dye and changes to gauge how to use these as vehicles for antibiotic delivery. To in coat color were monitored to determine the rate at which this end, consumption of water from sipper tubes was followed mice shed fur. After 5 mo, all mice had shed most, but not all of after exposure of C57BL/6 mice to the LD50/30 dose of 137Cs the dyed hair. In summary, soiled bedding sentinels are able to (67 cGy/min). Among our significant findings, we discovered detect fur mites within a 2-mo testing cycle; the head needs to that healthy 20 g mice consume approximately 4 ml of acidified be included as a sampling site, and egg-positive SS taken after water/d; that by d 1 postexposure, consumption is reduced by treatment need to be interpreted cautiously. 30% and continues to decrease to about 1.5 ml/d over the next 3 wk; that overall consumption of water from sipper tubes decreased P199 Toxicologic and Efficacy Testing of Ivermectin-impreg- by approximately 50% when wetted chow is available; and that nated Feed for Murine Fur Mite Eradication consumption of acidified water containing ciprofloxacin after irra- RJ Ricart Arbona*1,2, N Lipman1,2, FR Wolf1,2 diation was similar whether grape flavoring or sugar were added for increased palatability. With this background, we compared Tri-Institutional Training Program in Laboratory Animal Medi- the efficacy of 100 mg/kg/d of ciprofloxacin (Cipro), a broad- cine and Science, Memorial Sloan-Kettering Cancer Center, The spectrum fluoroquinolone antibiotic clinically used in neutropenic Rockefeller University, and the Weill Medical College of Cornell humans, to that of 125 mg/kg/d of the tetracycline doxycycline University, New York, NY; Research Animal Resource Center, plus 300 mg/kg/d of the non-absorbed aminoglycoside neomycin Memorial Sloan-Kettering Cancer Center and the Weill Medical (Doxy+Neo) in irradiated mice (n = 912 total mice). Mice were College of Cornell University, New York, NY provided 0.67 mg/ml Cipro in sipper tubes and wetted chow or Fur mite outbreaks have been a persistent problem in labora- 2 mg/ml Neo in drinking water plus wetted chow containing tory mouse colonies. Multiple treatment protocols have been 0.0625% Doxy. Thirty-day survival of mice exposed to a range of published. All are either labor intensive, expensive, or unreli- doses (775 to 900 cGy, n = 633) was increased in mice provided able. During a recent outbreak with Myobia musculi (MB) and Doxy+Neo (P = 0.005), but only marginally in mice provided Myocoptes musculinus (MC) in a large colony (approximately Cipro (P = 0.061) compared to untreated mice. Antibiotics tended 30,000 cages), we searched for a treatment modality that would to increase survival in females to a greater extent than in males. prove safe, efficacious, not labor intensive, and cost effective. We Mean survival time (MST) of decedents was increased by 4 to 5 d compounded and tested irradiated rodent feed impregnated with in both antibiotic groups (P < 0.0001), although Cipro increased 3 different concentrations (12, 24, 48 ppm) of ivermectin (IVM). MST more than Doxy+Neo (P = 0.031). These data demonstrate Postproduction analysis by ESI-MS revealed an approximately efficacy of antibiotics provided ad libitum in feed and water, and 40% loss of IVM during the manufacturing process. An 8-wk define important parameters required for the development of toxicity study was conducted to determine the highest dose that murine models of HS-ARS that can assess effectiveness of coun- resulted in the least adverse effects on a C57BL/6NTac mouse termeasures against radiation. breeding colony. No adverse clinical effects were noted, except 1 of 64 adult mice in the 48 ppm feed group developed tremors P198 Diagnostic Considerations in Murine Fur Mite Detec- and ataxia. The number of litters, pups born or pups/litter did tion not differ significantly between groups. Pup mortality did not RJ Ricart Arbona*1,2, N Lipman1,2, FR Wolf1,2 differ between the control and 12 ppm groups, but increased to 77% and 100% in the 24 ppm and 48 ppm groups, respectively. Tri-Institutional Training Program in Laboratory Animal Medi- No significant differences between groups were detected by cine and Science, Memorial Sloan-Kettering Cancer Center, The CBC, serum chemistry, body weight and gross necropsy. Plasma Rockefeller University, and the Weill Medical College of Cornell drug concentrations plateaued (concentration peak was dose Abstracts of scientific papers 2008 AALAS National Meeting dependent) at d 7 and decreased to undetectable levels 6 to 9 stress indicators. Metabolism excretion balance studies can in- d following discontinuation of treatment. We then determined volve the single housing of rats up to 168 h. The welfare of rats the efficacy in eradicating mites from a small colony of MB and in laboratory conditions is important to ensure physiological MC infested mice using the 12 ppm feed. Based on estimated and behavioural parameters fall within acceptable ranges. In mouse food consumption and production-induced drug loss, the order to assess the level of stress response (physiological and ingested dose was approximately 1 mg/kg. A 1-wk treatment behavioral parameters) of the singly housed rat an investigative schedule was sufficient to eradicate both MB and MC as evident study was conducted in which group-housed and singly housed from weekly testing for up to 21 wk. We have determined a (5 males and 5 females per group) rats were clinically observed relatively safe dose of IVM-medicated feed to effectively and daily. Feces was collected at 24-h time points, up to 168 h. Fecal efficiently eradicate fur mites from a large mouse colony. glucocorticoid hormone and metabolite analysis is being used as a tool by more animal researchers in the laboratory to provide P200 Homologous Collagen Implant to Repair a Tracheal Wall a noninvasive biomarker as an indication of the stress response. Segment By subjecting the collected feces samples to solid phase extrac- tion to remove impurities from the matrix, and then using liquid F Villegas-Álvarez, A González-Maciel, RE Soriano * , B chromatography-tandem mass spectrometry analysis, it can be Pérez-Guille, JF González-Zamora, L Padilla-Sánchez, R Reynoso- seen that the level of stress response in the singly housed rat Robles, A Ramos-Morales, E Zenteno Galindo, A Pérez-Torres is higher than the group-housed rat. Levels of glucocorticoid Experimental Surgery, Instituto Nacional de Pediatria, Mexico hormone in the singly housed rat is much higher than the levels City, Mexico seen in the group-housed rat. Tracheal stenosis might lead to tracheal malfunction, which is potentially life-threatening. Among other treatments, the P202 Transmission of a Bordetella bronchiseptica between bioprosthesis has been suggested as a tracheal substitute. This Rabbits and Guinea Pigs kind of prosthesis self-integrates to the respiratory epithelium S Pritt*1, RM Bunte2, D Sinha3, E Harvill3 without collapse, inflammatory response or secondary stenosis to 1 Covance Research Products, Denver, PA; 2University Laboratory maintain tracheal function. The goal of this study was to evaluate Animal Resources, University of Pennsylvania, Philadelphia, PA; a polyester prosthesis mesh covered with homologous collagen 3Department of Veterinary and Biomedical Science, Pennsylvania as tracheal wall segmental replacement (TWSR). We conducted State University, University Park, PA a single-blind study using randomly selected adult Wistar rats. The prosthesis was created from a homologus group that was Bordetella bronchiseptica is a Gram-negative coccobacillus consid- implanted with a subcutaneous polyester mesh. The control ered to be a veterinary pathogen infecting a number of species group had no surgical procedure. In the experimental group, with several strains of varying virulence. While rabbit-to-guinea we implanted the prosthesis to replace a segment of the anterior pig transmission is postulated through direct transmission, tracheal wall. Clinical, histomorphological, and immunochemi- fomites, or facility ventilation, it has not been demonstrated. cal postimplant evaluations were performed at 4 different times. This study examined the transmission of B. bronchiseptica (rabbit Rats in the experimental group presented with a few respiratory strain of low virulence) between rabbits and guinea pigs. Twelve malfunctions during the first week. The intraluminal area and B. bronchiseptica-free guinea pigs (6 males and 6 females) were thickness of the bioprosthesis wall in the experimental group was housed in a conventional facility also housing rabbits colonized higher (P = 0.001) than the control group at 4 mo; this increased with B. bronchiseptica. These guinea pigs were isolated in one thickness was due mainly to angiogenesis, without evidence of room, and personal protective equipment (hair cover, gloves, fibrosis or chronic inflammation. At the beginning of the study, fresh lab coat, shoe covers, surgical mask) was required for entry. undifferentiated cells were predominate on the implant surface; Personnel entered the guinea pig room before entering other however, in the last 2 mo, the epithelial and ciliated cells were animal rooms. During the study period, no animals became similar to the controls. At 4 mo postimplant, no histomorpho- clinically ill. At the end of 30 d, cultures of the nasal cavity, tra- logical or functional differences were found between groups. In chea, and lung were negative for B. bronchiseptica. Another 12 conclusion, collagen-polyester homologous prosthesis as TWSR B. bronchiseptica-free guinea pigs (6 males and 6 females) were in rats was able to produce proliferation of normal respiratory inoculated with 5,000 CFU per nostril of B. bronchiseptica upon epithelium and maintained tracheal function without collapse, arrival. During the study period, no animals became clinically inflammatory response, or secondary stenosis. ill. At the end of 30 d, 11/12 (92%) animals had positive nasal cavity cultures, while 8/12 (67%) had positive tracheal cultures P201 Comparison of Fecal Biochemical Stress Indicators in for B. bronchiseptica. No organisms were retrieved from lung Group-housed and Singly Housed Rats cultures. Histopathology revealed mild to moderate inflamma- tion in most animals, with a lymphoid infiltrate in nasal cavities SE Wall* and heterophilic inflammation in the trachea. This study dem- Rodent Toxicology, Covance Laboratories Ltd, Harrogate, United onstrates that guinea pigs may be maintained up to 30 d, with Kingdom special precautions, in conventional facilities also housing rabbits colonized with a B. bronchiseptica strain of low virulence and not It is widely recognized that commonly used laboratory rats in become colonized. Direct inoculation with the strain will result in general display fewer signs of stress and experience enhanced colonization and inflammation of the nasal cavities and tracheas welfare when housed in social groupings, compared to in isola- of guinea pigs, but no clinical disease or lung lesions. tion. Historic stress assessments at our facility have included quantification of glucocorticoid hormone levels in the blood, obtained by venipuncture. Developments in analytical tech- niques have enabled the monitoring of glucocorticoid hormone in the feces of experimental animals, providing a less invasive and more readily available means for monitoring biological Abstracts of scientific papers 2008 AALAS National Meeting P203 Optimization of Locking Catheter Ports for Rats lated and sentinel mice at 6 wk. The cage lid with the partitioning barrier we developed is very useful when sentinel animals are S Pawl*, J Agate, CM Dugan used in the quarantine of laboratory mice, such as genetically Toxicology Services, Covance Laboratories, Madison, WI modified or immunodeficient mice, because sacrifice of valuable The use of a heparin-dextrose solution to lock catheterized genetically modified mice should be avoided, and because im- ports (hep-lock) in rats is common practice in a nonclinical set- munodeficient mice cannot produce serum antibodies. ting. Due to the large volumes of animals requiring hep-lock, it became necessary to evaluate and streamline this process. The P205 Assessment of RNA Polymerase B Gene Compared with goal was to evaluate and maximize the amount of time between 16S Gene as a Target for PCR-based Detection of Pasteurella hep-lock procedures while minimizing the risk of failing to es- pneumotropica tablish patency. Different lengths of time between flushing and VS Dole*, LA Banu, RD Fister, KS Henderson replacing the hep-lock were evaluated. Replacing the hep-lock daily, every other day, and every 3, 4, and 6 d resulted in 100% Research Animal Diagnostic Services, Charles River, Wilming- patency. The remaining groups tested every 2 and 5 d, which ton, MA resulted in 97.6% and 90.9% patency, respectively. The ease of Currently, DNA-based differentiation of Pasteurella pneumotro- establishing patency was also evaluated. Harness removal and pica from other Pasteurellaceae involves PCR amplification of time required to complete the hep-lock were the determining the 16S gene region, although some attention has been given to factors for the optimal procedure. Replacing the hep-lock daily the RNA polymerase B (rpoB) gene. To assess whether the rpoB and every other day resulted in no instances of harness removal gene sequence may serve as an additional diagnostic target, and took approximately 1 min per animal. All other intervals partial rpoB gene sequences of various P. pneumotropica culture resulted in harness removal for less than 20% of animals; how- isolates were determined. P. pneumotropica culture isolates were ever, the time required varied. Replacing the hep-lock every 2, initially identified by microbiological and biochemical techniques 3, and 4 d took approximately 1.40 min per animal; replacing and further characterized as Heyl or Jawetz biotypes using it every 5 d took approximately 2.3 min per animal. Although a 16S-based fluorogenic nuclease PCR (fnPCR) assay. Partial results showed that performing the lock every 6 d was possible, sequencing of the rpoB gene (540bp) and the 16S gene (680bp) this took longer per animal, and it was determined that 2 to 4 d was done following PCR amplification. The rpoB sequences of was optimal for maintaining patency and lowering time needed 4 out of 5 isolates identified as Heyl by 16S-based fnPCR assay per procedure. were 100% identical to each other. Similarly, rpoB sequences of the 5 isolates identified as Jawetz by our 16S based fnPCR P204 Lid with Barrier for Partitioning Cage for Laboratory assay were 99% to 100% identical to the rpoB Jawetz sequence Animals in the GenBank (AY314034). However, alignment of rpoB gene sequences indicated that it will be challenging to develop an T Kokubo*, T Nishikawa, S Matsushita rpoB-based conventional two primer PCR (gel-based) or fnPCR National Institute of Radiological Sciences, Chiba, Japan assay to distinguish between the Heyl and Jawetz biotypes, Genetically modified laboratory mice are frequently transferred since the sequence variability between these two biotypes is not among research institutions. When these mice are quarantined, clustered in any particular region of the rpoB sequence. Also, sentinel animals are used to indirectly monitor the health of the the current sequence information available for the rpoB gene mice in order to avoid having to sacrifice the valuable geneti- (our sequencing data and the 324 Pasteurellaceae sequences in cally modified mice. Recent data on the health monitoring of the GenBank) does not indicate any conserved regions to dis- laboratory mice indicate that opportunistic infections are more tinguish P. pneumotropica from other members of Pasteurellaceae common than severe contagious infections. During quarantine, family. In contrast, a total of 1427 16S Pasteurellaceae sequences test animals and sentinel animals should be placed in the same are available in GenBank, and the sequence information indicates cage because opportunistic infections often exhibit low levels that amongst Pasteurellaceae, the 16S gene is more conserved of transmissibility. However, it is important to prevent fighting than rpoB. However, unique island clusters of differentiating among the animals. Thus, we developed a new cage lid made of nucleotides in the 16S gene can be exploited for specific PCR a stainless steel wire screen, with a screen barrier for partitioning detection of P. pneumotropica. Complete sequencing of the rpoB a cage into compartments. To evaluate the effectiveness of the gene from a larger number of isolates from various members of cage lid with the partitioning barrier, the transmission of cilia- Pasteurellaceae family may eventually be useful to further char- associated respiratory (CAR) bacillus was tested. A total of 40 acterize the Pasteurellaceae family. specific pathogen-free (SPF) mice were used. Sixteen mice were inoculated intranasally with the SMR strain of the CAR bacillus P206 Oral Transmission of Experimental Chagas Disease by under light ether anesthesia. Two inoculated mice were placed in Açaì Pulp Contaminated with T. cruzi Y Strain one compartment, and 3 uninfected sentinel mice were placed in VL Dias*1, RL Barbosa2, AR Salgado1, A Gimenes1, M Corat1, AM the other compartment. Each cage was covered with the lid we Guaraldo2, LA Passos1 developed. At weekly intervals, the 5 mice were sacrificed by ex- Laboratory—CEMIB, 2Parasitology—IB, UNICAMP, 1Genetic sanguination under anesthesia and were examined. Using a PCR assay, the 16S rRNA gene from the CAR bacillus was detected Campinas, Brazil from swab samples of the larynx and the respiratory organs of Because of its socio-economic impact, Chagas disease (T. cruzi) is both inoculated and sentinel mice at 4 wk. Histopathologically, the most important parasitic infection in South America. Despite CAR bacilli were shown on the surface of the epithelium of the the progresses in vector transmission control, new specific cases respiratory tracts of both inoculated and sentinel mice at 4 wk. showed the oral route as an important factor for disease dis- Typical bronchopneumonia and peribronchitis were observed semination. Recent outbreaks that arose in north Brazil indicate in both inoculated and sentinel mice at 5 wk. Serologically, IFA likely contamination of açaí pulp, a highly nutritional fruit that antibodies against the CAR bacillus were detected in both inocu- serves as a major food source for the poor people of this region. Abstracts of scientific papers 2008 AALAS National Meeting However, no data has been reported regarding outbreaks of T. compared with baseline). During fluid resuscitation, MAP cruzi in people through consumption of açaí pulp. Thus, in this increased to 95 ± 5 mmHg (P = NS compared with baseline) work, we evaluated the survival of T. cruzi in the açaí pulp and and PETCO2 increased to 33 ± 3 mmHg (P = NS compared with the potential for oral contamination through açaí pulp consump- baseline). Landrace/ Large White swine are suitable models for tion. Tripomastigotes (1×105) of T. cruzi Y strain were extracted hemorrhagic shock. Various parameters of acute blood loss can from the blood plasma of infected mice. The plasma was mixed be recorded in this model. at a 1:4 proportion in the açaí pulp and kept at room temperature for 28 h. The vitality of the parasite was checked at 6, 8, 12, 24, P208 Assessment of QTc Formulas in Conscious Dogs with and 28 h; it was observed directly in 5 μl of mix with trypan blue Pacing-induced Heart Failure on optical microscopy and in the eluate obtained from a forced straining. Parasite movement was classified as greatly active A Kijtawornrat1,2, Y Panyasing*1,2, JL Lolly2, JJ Schmidt2, Y Nishi- (GA), active (AT), and slow (S). The results demonstrated that jima2, BM Roche2, RL Hamlin1,2 at 6 h, 100% of parasites were GA; at 28 h, 67% were GA and 1The Ohio State University, Columbus, OH; 2 QTest Labs, LLC, 33% were S. To confirm the hypothesis of oral transmission, 100 Columbus, OH μl of T. cruzi mix was administered either intraperitoneally, per Drugs given to people with naturally occurring diseases have been os, or by gavage to 10- to 12-wk-old immunodeficient female shown to be more toxic than when given to those without disease; mice (CB17scid/Uni); plasma from infected animals was used therefore, it appears reasonable to search for potential toxicity in a as a positive control and negative control animals received only model possessing a pathophysiological substrate known to sensitize fruit pulp. Animals were observed for 40 d, and infection was humans to toxicity. Since a search for potential effects of drugs on confirmed by the Brener method. Death was noted at d 14 for ventricular repolarization is so critical, models possessing heart positive control animals dosed intraperitoneally and by gavage failure (HF) have become increasingly popular to investigate poten- and at d 18 for those dosed per os. For the test group we ob- tial toxicity. Prolongation of heart rate (HR) corrected QT interval served death at d 24 in animals dosed per os; no negative control (QTc) is one of the surrogate markers for potential torsadogenicity animals died. These results showed the pulp does not interfere of a test article. This study was conducted to determine which cor- in the parasite’s capacity to cause disease; rather, it seemed only rection method best accounts for the effects of changes in the RR to delay infection for 6 to 8 d when orally administered. interval on the QT interval of conscious, sling-trained dog with HF. Bipolar, transthoracic ECGs were obtained from 18 conscious P207 Developing a Hemorrhagic Shock Model in Landrace/ dogs at baseline and after 12 wk of rapid ventricular pacing. RR Large White Swine and QT intervals were measured and QTc were calculated using 3 T Xanthos, X Balkamou*, S Konstantinos, L Papadimitriou, I common algorithms (Bazette, Fridericia, and Van de Water). After Dontas 12 wk of pacing dogs with HF confirmed by reduced LV shortening fraction had significantly slower HR (P < 0.01) and significantly Experimental Surgery and Surgical Research, University of longer QTc (P < 0.01) when compared to baseline. The following Athens, Athens, Greece regression equations were related QTc to RR: QTc(V) = 239.4–2.68 Shock following acute blood loss is a daunting medical emer- RR, r2 = 0.00 for the healthy dog; QTc(F) = 258.4–0.01 RR, r2 = 0.01 gency. Various organs are affected due to the compromised for the dogs in HF. The Van de Water method demonstrated least oxygen delivery. The aim of the present study was to describe dependency of QTc on RR interval in conscious, healthy dogs, the model of hemorrhagic shock developed in our department. but the Fridericia method was best for correcting dogs in HF. In Twenty Landrace/ Large White swine with an average weight conclusion, the rate-corrected QT formula of Fridericia appears of 19 ± 2 kg were included in the present study. The animals to be useful as a biomarker for assessing effects of test articles on were premedicated with ketamine hydrochloride (10 mg/kg), ventricular repolarization in dogs with HF. midazolam (0.5 mg/kg ) and atropine (0.05 mg/kg). Anesthesia was induced with propofol (2 mg/kg) cis-atracurium (0.15 mg/ P209 Food Restriction Changes Physiological and Pathological kg) and fentanyl (2 mγ/kg). Intubation followed and the swine Parameters in Zucker Diabetic Fatty Rats were ventilated with FiO2 21% and with a tidal volume 12 ml/ kg. The respiratory frequency was adjusted in order to maintain Y Kirihara*1, M Takechi1, A Ago1, S Kaidzu2, Y Notsu3, Y Saito4, PETCO2 = 35 to 40 mmHg. The left carotid artery was surgically M Ito5, T Yamada1 prepared, and an arterial catheter was advanced in the thoracic 1Experimental Animals, Center for Integrated Research in Sci- aorta. The right internal jugular vein was also surgically pre- ence, 2Ophthalmology, 3Medical Technology attached to School pared, and a Swan-Ganz catheter was inserted for measurements of Medicine, 4Anesthesiology, Shimane University, Izumo, Japan; of the right atrial pressures. The animals were left to stabilize for 5Technical Center, Charles River Japan, Aikougun, Japan 30 min following surgical manipulations. Afterward, 50% of the ZDF rats (Charles River, Japan) are a disease model of human total blood volume was drained over a period of 20 min. Animals type 2 diabetes. It is well known that dietary restriction is the were left in shock for 90 min. Lactated Ringer solution (20 ml/kg) first therapy for human diabetes. However, there is no data about was then infused over 20 min. Animals were monitored continu- effects of food restriction in ZDF rats. Therefore, we examined the ously for a period of 4 h. The animals were then euthanatized effects of 80% food restriction on physiological and pathological with 2 g of pentobarbital. All animals were routinely examined parameters in ZDF rats. We also examined the relationship of postmortem to record any underlying pathology and for organ ZDF rats to diabetic retinopathy and neuropathy. We used ZDF/ harvesting. Mean arterial pressure (MAP) was 100 ± 12 mmHg CrlCrlj-Leprfa/Leprfa (Fatty) rats for a diabetic model and ZDF/ following surgical manipulations. PETCO2 was 35 ± 4 mmHg. CrlCrlj-?/+ (Lean) rats for control. We divided the rats into four Hemoglobin was 10.3±0.8 mg/dl. Following the hemorrhagic groups: group A (n = 7), Fatty rats given free-feeding; group B (n phase MAP declined to 45 ± 12 mmHg (P < 0.01 compared with = 7), Fatty rats given 80% food restriction; group C (n = 8), Lean baseline), PETCO2 declined to 26 ± 3 mmHg (P < 0.05 compared rats given free-feeding; group D (n = 8), Lean rats given 80% with baseline) and hemoglobin was 6.6 ± 0.9 mg/dl (P < 0.001 food restriction. When rats were 6 wk old, food restriction was Abstracts of scientific papers 2008 AALAS National Meeting started. Every day, we measured food consumption of groups P210 Monitoring of Infectious Bacteria in Poultry A and C, then gave 80% of that amount of food to groups B D Sullivan*, A Byers, W Willis, J Allen and D the next day. The experiment was conducted for 30 wk. Body weights were measured once a week. We analyzed GOT, Animal Science, North Carolina A&T State University, Greens- GPT, T-Cho, HDL-Cho, LDL-Cho, TG, BUN, glucose (Glu), and boro, NC glycated albumin (GA). Groups A and B were given a glucose tolerance test (GTT). To evaluate pain thresholds, a paw pressure Determination of the health status of the flock and offspring has (PP) test was used. We also measured electroretinogram (ERG). traditionally been accomplished by analysis of serum antibod- Statistical analysis was performed using ANOVA with repeated ies. This study compared the antibacterial activities of chicken measures, followed by a Scheffe test. Body weights of the groups antibody in egg yolk compared with serum in the detection of given food restriction were significantly lower than those of the Salmonella typhimurium, Campylobacter, and Salmonella enteritidis groups fed ad libitum. There were no significant differences for monitoring the health of flock. Salmune, a live avirulent strain between groups C and D in laboratory data. Group A showed of Salmonella typhimurium, is typically given to birds at d 1 and significantly higher To-Cho, LDL-Cho, BUN, Glu, and GA than wk 6, 10, and 18 to vaccinate the flock against S. typhimurium the other groups. Group A also showed significantly higher Glu and S. enteritidis. Samples of egg yolk and sera were collected for in the GTT than that of Group B. Group A indicated significant 5 mo from 15 broiler breeder hens previously vaccinated with lower pain thresholds compared to the other groups on the PP Salmune. Antibody specificity to these 3 bacteria was determined test. There were significant differences between Fatty rats (groups by western blot assay. The nitrocellulose membranes were probed A and B) and Group D in the ERG data. However, all rats in with purified IgY antibody isolated from the serum and egg yolks Group A developed cataracts by wk 30. The results indicated of each hen. At wk 27 and 36, all 15 hens had IgY titers in their that food restriction changed physiological and pathological serum and egg yolks specific for S. typhimurium, S. enteritidis and parameters in ZDF rats. ZDF rats might be a useful model for Campylobacter. These studies show that antibodies to specific bac- diabetic retinopathy and neuropathy. teria present in both serum and egg yolk samples from vaccinated hens can be detected by western blot assay. The analysis of egg yolk can be a rapid and effective alternative method for screening flocks for the presence of antibodies to specific bacteria.