MICROBIOLOGY Overheads Unit 1

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MICROBIOLOGY Overheads Unit 1 Powered By Docstoc
					MICROBIOLOGY                 1

Isolation and Identification of
   Pathogens:
      What organisms?
      What antibiotic?
Study of:
   1. Bacteria
   2. Fungi
   (Molds and yeasts)
   3. Parasites
(Protozoans and worms)
   4. Miscellaneous
(Rickettsia, chlamydia)
   5. Viruses
Objectives                         2
Clinical Microbiology
1. Know which microbe are normal
flora in a certain area, and which are
pathogenic.
2. Know correct procedures for
isolating and identifying
microorganisms.
3. Know the characteristics of the
microorganisms so that positive ID
is made.
4. Know what procedures are
followed for assessing the drugs
which are best for treatment.
5. Understand how to maintain a
quality control program in a
Microbiology department.
3 Non Toxic Metabolites of
Staph aureus

1. Protease
2. Coagulase
3. Hyaluronidase
4. Deoxyribonuclease
5. Staphlokinase
6. Lipase
7. Gelatinase
8. Phosphatase
9. Protein A
Toxic Metabolite of Staph aureus
1 - 4 α β γ δ (alpha, beta, gamma,
delta) Lysins
   **act on host cell membranes to
   mediate cell destruction
2. Leukosidin
   Panton-Valentine Leukosidin
   (MRSA have to kill WBCs)
3. Dermonecrotic (affects skin)
4. Lethal toxin (rapidly fatal)
5. Enterotoxins (six): food
poisoning
6. TSST-1 (Toxin Shock Syndrome
Toxin-1)
7. Exfoliatin: scalded skin
syndrome in neonates
Dieases caused by Staph       4
***Staphylococcus aureus
      pimples, boils, carbuncles,
impetigo, acne, postoperative and
wound infections, pyelitis, cystitis,
scalded skin syndrome
      More serious:
Food poisoning(entertoxins),
septicemia, endocarditis,
meningitis, pneumonia,
osteomyelitis, TSS

***Staphylococcus epidermidis
  stitch abcesses, endocarditis,
bacteremia, prosthetic joints, CNS
shunts
                           4-2
***Staphylococcus saprophyticus
     UTIs especially in young,
sexually active women

***10 -12 other species rarely
isolated and implicated in infections
Classification of Streptococcus 5
Brown’s
  Classified by
  Hemolytic reaction when grown
on blood agar

4 types:
   1. Alpha α (partial hemolysis:
greenish)
   2. Beta β (complete: clear zone
around colony)
   3. Gamma γ (no hemolysis)
   4. Alpha prime α’ or wide zone
alpha
5--2
Factors affecting hemolysis:
   1. Media
   2. Production of Streptolysin O
   and S
      –Streptolysin O is O2 labile
      –Streptolysin S in O2 stable
   3. Various types of blood
   4. Aerobic or Anaerobic
   Incubation

What increases hemolytic activity?
 1. Anaerobic incubation
 2. Prolonged aerobic incubation
 3. Overnight refrigeration
Beta Steptococcus                6
   Group A - L, etc.
1. Group A Beta Strep (GAS)
   Strep pyogenes
      Streptococcal pharyngitis
      Scarlet fever
      Erysipelas
      Rheumatic fever
      Acute glomerulonephritis
   Test to ID Group A Beta Strep
      – Bacitracin disk (A disk)
      – PYR
2. Group B Beta Strep
   Strep agalactiae
   Normal flora in female vaginal
   Pathogen in newborns
                              6-2

Tests to ID Group B Beta Strep:
      **Camp test
      **Hippurate Hydrolysis

Alpha Strep:
1. Apha Strep viridans group
   (Strep salivarius, S. mitis,
    S. mutans, S. sanguis, etc)
   Normal flora throat
   Pathogenic in SBC (subacute
bacterial endocarditis)
2. Streptococcus pneumoniae
   (Pneumoncoccus)
   Gram positive diplococcus
                               6-3

Tests to ID:
   1. Bile solubility
   2. Optochin (P disk)
(ethylhydrocupreine HCL)
   3. Neufeld Reaction (Quelling)
   4. Inulin fermentation
**if tests are neg. for S. pneumonia,
   check for Enterococcus

Enterococcus (E. faecalis, E.
faecium)
   Hemolysis: usually gamma γ or
 alpha α , (rarely beta β)
                             6-4

Tests to ID Enterococcus:
  1. Bile Esculin Media (BEA)
  2. 6.5 % NaCl
  3. SF broth (Enterococcus broth)
  4. PYR test positive
 7
 Enterobacteriaceae: gnrs
 Citrobacter:
    **UTIs
    **motile; lactose +; H2S +

 Edwardsiella:
  **GI tract of reptiles
  **motile;lactose =; maybe H2S+

 Enterobacter:
  **motile; lactose +

  Escherichia coli:
   (Most common aerobe in GI)
**Most common cause of UTIs
 **Meningitis in infants
 7-1

**EIEC (Enteroinvasive E.coli)
**ETEC (Enterotoxic E.col
**EPEC (Enteropathogenic E.coli)
**EHEC (Enterohemorrhgic E.coli)
    E.coli O157:H7
       Test: Sorbital MacConkeys
             Sorbital negative

Hafnia: NLF; formerly Enterobacter

Klebsiella:
  Kleb. pneumoniae (Friedlandars
pneumonia– lobar pneumonia)
**nonmotile; lactose +
7-2
 K. granulomatis (formerly
Calymmatobacterium)
   **not grow on culture media
   **Granuloma inguinale: STD
   nodules (Donovan bodies)
   **safety pin appearance on gram
stain; encapsulated (blue rod
surrounded by pink capsule)

Morganella (formerly Proteus)
 **motile; NLF

Proteus:
  **swarming; NLF; rapid urease +
  P. vulgaris and P. mirabilis
 7-3
Providencia:
  **motile; NLF

Salmonella species:

  **Polyvalent O antisera
  **O, Vi, and H antigens
  **~1,800 serotypes and biotypes
  **NLF; H2S +; motile
    S. typhi (enteric fever; typhoid)
  Others:
    S. parayphi, S. typhimurium,
    S. cholerae-suis, S. enteritidis,
    S. arizonae
  **septicemia, gastroenteritis
  **Carriors: convalescent/chronic
 7-4
Serratia:
   **15 somatic groups
   **Lactose variable
   **S. marcescans– brick red
pigment

Shigella species:
   **4 serotypes (serovars)
     A , B, C, D
   **Cause of bacillary dysentary
   **shiga toxin
     Apoptosis and release of IL-1
and IL-8; inflammatory response to
cytokines damages colon mucosa
   **NLF, H2S neg, nonmotile
7-5

Yersinia:
   **nonmotile; NLF; grows 4⁰C
   2 species of medical importance:
   Y. pestis: plague
   (Bubonic, pneumonic,
septicemia)
   Y. enterocolitica:
   **CIN – bull’s eye

Newest Member:
  Plesiomonas shigellosis:
     **only one oxidase pos
     **GI disease
8
IMViC Reactions
A. Indole Production
   **Media contains tryptophane
   **If bacteria produces
tryptophase–> Indole produced
   **reagents:Kovac’s
(p-dimethylaminobenzaldehyde)
         Ehrlich’s
(p-dimethylaminocinnamaldehyde)
  Red ring = positive

B. Methyl Red Test
   **MR-VP Broth (Clark & Lubs
      Dextrose Broth)
   **If bacteria uses mixed acid
pathway to ferment glucose–> much
acid produced–>MR pos (red color)
8-2
C. Voges-Proskauer
   **MR-VP Broth
   **If bacteria uses butyleneglycol
(acetoin) pathway to ferment
glucose–> acetoin
[acetylmethycarbinol,(acetoin),
butyleneglycol, diacetyl]–>
VP positive = pink color
   **Reagents: alpha-naphthol and
40% KOH (potassium hydroxide)

D. Citrate (Simmons)
   **Prussian Blue color = positive
   **The organism can utilize
citrate as its only source of Carbon.
9
Nonfermentative Gram Neg Bacilli
   A. Oxidase + motile rods:
      Pseudomones
      Alcaligenes
      Burkholderia
   B. Oxidase - nonmotile
diplobacilli:
      Acinetobacter
   C. Oxidase + nonmotile,
diplobacilli (diplococci)
      Moraxella

Oxidase reagent:
  tetramethyl,p-
phenyldiaminedihydrochloride
9-2
O-F Media(Oxidative-Fermentative)
   oxidizer vs. fermentative vs,
   nonoxidizer (nonfermenter)

Pseudomonas and Burkholderia
**Aerobic, straight & slender gnr
**Motile (except B. mallei)
**Mesophilic(but can grow at 4C)
**Very resistant (can grow in
disinfectants)

Pseudomonas aeruginosa:
  Colony appearance:
     Bluish green pigmentation
     Maybe beta hemolytic
9-3
Identification:
   1. Oxidase positive
   2. TSI: K/NC (nonfermenter of
glucose, lactose, and sucrose)
   3. Good growth at 4⁰C
   4. Pyocyanin production
      (Bluish-green pigment)
   5. Odor: grape-like (fruity)
      or corn tortilla odor
Burkholderia cepacia (formerly
Pseudomonas)
   **Cystic fibrosis
Burkholderia pseudomallei
  **Melioidosis (survives in
macrophages)
9-4
Burkholderia mallei
  **Glanders in horses
  (Equines–>humans rarely)

Oxidase negative gnr:
Acinetobacter:
  **environmental bacteria
  **colonizer of hospital patients
  **gram neg plump coccobacilli

Stenotrophomonas maltophilia
(formerly Pseudomonas and
Xantrhomonas)
   **nosocomial infections
10
Gram Negative Coccobacilli

Pasteurella
Former species:
     Yersinia pestis
     Fancisella tularensis

Pasteurella multocida:
  **common in animals
  **humans from infected bites or
     scratches
  **Fails to grow on MacConkeys
  **Oxidase positive
  **Indole positive
10–2
Actinobacillus
   *1 species NF human mouth
   A.actinomycetemcomitans
   (Most common infection)
   *puff balls in blood culture
   *3 species normal domestic
   animals
   *may become pathogens in
   humans (bites and scratches)
   *gram neg coccibacillus
Kingella: short plump coccobacillus
catalase negative (differ Neiserria)
Capnocytophaga: fulminant, life
threatening infect after dog bites
10-3
Haemophilus:
   Differentiated by:
     **Requirement for
           A. X Factor = Hemin
           B. V Factor = NAD
   (Nicotine Adenine Dinucleotide;
     Coenzyme I)
     **Hemolytic characterisitics
Sources of V Factor:
     Yeast
     Potato extract
     Certain Bacteria (especially
Staph, Neisseria, Strp pneumoniae)
Haemophilus influenzae
   **Satellitism (Requires X & V)
10–4
  Diseases:
     Pink Eye (subsp: aegypticus)
     Acute epiglottitis
     Acute Bacterial meningitis
  Vaccination for H.influenzae b
Haemophilus ducreyi:
  STD: soft chancroid
  “school of fish” on gram stain

 Brucellosis:
   **Facultative intracellular
      parasites
   **Parasites of domestic animals
(cattle: contagious abortion)
   **Can infect humans
10–5
Culture:
   **Blood cultures: keep 30 days
(~4 weeks). Do weekly “blind”
subcultures since may appear neg
in broths.
   **Plate to Brucella agar and
incubate in CO2. Save 3 weeks.
Pasteurization:
   63⁰C for 30 mins, or
   72⁰C for 30 sec.
Identification of Brucella:
**Isolates are Level III pathogens
   **catalase and oxidase pos
**gram neg coccobacilli( poor stain)
**nonmotile
**rapid urease positive
10–6
Serodiagnosis
   **serum agglutination test (SAT)
   **titer > 160 is diagnostic
4 Species:
   Br. abortus: cattle
   Br. suis: swine
   Br. canis: dogs
   Br. melitensis: sheep/goats (AKA
malta or undulent fever in humans)

Bordetella:
   All species cause a whooping
cough-like infection
   B. pertussis: whooping cough
(pertussis)
10–7
Media:
   A. Bordet-Gengou agar (potato-
blood-glycerol-methicillin agar)
   Short shelf life
   “Cough plate”
   Colonies resemble mercury
droplets (grow within 7 days)
   B. Regan-Lowe agar (charcoal-
blood-cephalexin)
      Longer shelf life (4 -8 weeks)

Francisella:
   F.novicidan: animal pathogen (not
involved in human disease)
10–8
Francisella tularensis: animal and
human tularemia
(wild rodents, rabbits, beavers,
muskrats)
Human infection by:
   1. Handling infected animal skins
or carcasses
   2. Insect vectors (deerflies, ticks)
   3. Being biten by carnivores who
have eaten infected animals
**Mostly disease of adult men
**Most labs do not attempt to work
with bacteria–>special state labs
**Commonly lab acquired
      Level II clinical material
(Gloves and biolog. Safety cabinet)
10–9
  Level III if isolated
  Media:
**Blood-cystine-dextrose of Francis
     **cystine-heart agar

Gardnerella vaginalis:
   **formerly Haemophilus
   ** “clue cells”
   **NSV: bacterial vaginosis in
females; asymptomatic in males
or BV (bacterial vaginitis)
   *now known to be polymicrobial
   *10% KOH (Whiff test)–>
fishy odor
10-10

**no WBCs (vaginosis not
vaginitis)
**catalase and oxidase negative
**Beta-hemolysis on selective
Tween (HBT) agar

HACEK Group of organisms
causing slowly progressive bacterial
endocarditis
Haemophilus aphrophilus
Actinobacillus
Cardiobacterium
Eikenella
Kingella
11
Anaerobic, gram negative, non-
spore forming (NSF) bacilli:

The most commonly encountered
anaerobes in infection:

Bacteroides and Fusobacter

B.fragilis:
*Anaerobe most frequently isolated
 from infections;
*predominant organism of normal
GI tract

Fusobacterium: tapered ends
11-2
Anaerobic, gram positive, spore
forming (SF) rods:

Clostridium species:
  *obligate anaerobes
  *sporangia (drumstick/tennis
  racquet)
  *exotoxins (cause intoxications)
Culture methods:
  *immediately
  *egg yolk media
  *Nagler Reaction
  *Reverse CAMP test
  *Lecithinase test
11-3
Anaerobic gram positive NSF
bacilli:
Bifodobacterium
Propionibacterium
Eubacterium
Lactobacillus
Actinomyces
Mobiluncus

Lactobacillus: Doderleins bacillus
                L. acidophilus
Actinomyces sp:
   *human actinomycosis
   *sulfur granules
   * “molar tooth” colonies
A. israelii, A. naeslundii, A. bovis
12
Aerobic Gram Positive Rods:
Spore Forming:
Bacillus species:B.subtilis
                  B.anthracis
                  B.cereus
three types of Anthrax:
   1. Cutaneous
   2. Pulmonary (woolsorter’s)
   3. GI
                                 rd
Two potent Exotoxins:depend on 3
protein for activity=
Protective Antigen
   *Lethal factor + PA
   *Edema factor + PA
-Medusa head colonies
-Ascoli test
12-2
Aerobic,gram positive, non-spore
forming Rods:
   Listeria: major pathogen
      L.monocytogenes
         *Neonates
         *Pregnant women
         *Immunocompromised
   usually a meningitis
Identification:
   *Beta hemolytic
   *Camp +
   *Grows well on most media
   *Can grow at 4⁰ C
   *Hydrolyses Na Hippurate
   *Gram pos bacillus with some
coccibacillary forms
12-3
How to differ from Grp B. Strep
  *gram stain
  *Esculin hydrolysis–>Black
  *Motility on “wet mount”
     end over end tumbling
     at Room Temp (<motility at
     37⁰C)
     Umbrella shaped motility in
        motility media

Corynebacteria
   Pleomorphic, aerobic or
facultative, gram pos rods
   Gram stain: clubbing of bacillus;
   “Koryne” “Chinese Letters”
   “Picket fence” formations
 12-4
 Diphtheroids: Resembles pathogen
 but are normal flora
 (AKA “coryneforms”)
    **most are harmless
    **C.jeikeium & C.urealyticum
   **can cause infection in
      compromised hosts
Corynebacterium diphtheriae
      (Klebs-Loeffler bacillus)
   *Media: grows on most
      Loeffler (serum) slants
      Cystine tellurite blood agar:
      Black/gray colonies; garlic odor
   *Elek test: demonstrate toxin prod
   *Metachromatic granules on
      methylene blue stain
13
Mycobacteria
(Acid-Fast Bacillus;AFB)
   *Coccibacillary to long rod
   *aerobic
   *Waxy cell wall (mycolic acid)
      Difficult to stain; once stained
hard to decolorize even with acid-
alcohol decolorizer
Gram stain: poorly stains, “beaded”
“gram-ghosts”
Mycobacterium tuberculosis
complex:
   M.tuberculosis, M.bovis,
M.bovis BCG), M.africanum
   *All capable of causing TB
   *most common M.tuberculosis
 13-2
   *inhale single organism to infect
   *15-20% infected–>disease
   *M.bovis from milk from
infected cows
   *M.bovis bacilleCalmette-Guerin
      (BCG) used as vaccine
ID of TB:1. Stained smear for AFB
          2. Isolation on culture
Types of cultures:sputum, gastric,
urine
Staining: Presumptive ID of AFB
examine a minimum of 300 fields
1. Ziehl-Neelsen Stain (“hot stain”)
   Carbolfuchsin Stain
   Acid-Alcohol
(3% Hcl & 95% ethanol)
13-3
Red = AFB pos Blue = AFB neg
2. Kinyoun Stain (“cold stain”)
      **same reactions
      **without heating
      **20 - 80% sensitivity
3. Fluorochrome Auramine-
Rhodamine
   *>sensitivity than carbolfuchsin
   **fluorescent stain: bacilli stand
out brightly against background
Other Acid Fast organisms:
      -Norcardia
      -Legionella (Pittsburgh pneu)
      -Cryptosporidium**
      -Isospora**
**coccidian protozoan parasites
13-4
Culturing of Mycobacterium:
1. Digestion/Decontamination
   *NaOH
   *Zephiran-trisodium
   *N-acetyl-L-cysteine
     (NALC-NaOH)
2. Media:
   *L-J slants (Lowenstein-Jensen)
   *Middlebrook 7HIO
   *Middlebrook Cohn 7H9 broth
Inoculate two tubes: Incubate 1 in
dark and 1 in light @ 37⁰C
**Examine weekly
**Report neg@ 8 wks.
Mycobacterium tuberculosis:
   *Luxuriant growth (eugonic)
13-5
   *Buff and rough
   *Cauliflower-like (2-3 wks)
Niacin Test: pos
Catalase: neg/neg for two types:
(1. semiquantitative >45mm of
bubbles; 2. At 68⁰C)
Tellurite Reduction: neg
Nitrate Reduction: pos
Tween Hydrolysis: neg
Molecular Methods: DNA

Nontuberculous Mycobacteria:
MOTT(Mycobacterium other than
tuberculosis)
13-6
All others including
  M.aviumComplex (MAC)
      M.avium-intracellulare
      *important pathogen in HIV
  M.ulcerans (Buruli ulcer/skin)
  M.kansasii (more common in
white males; in tap water)
  M.paratuberculosis: isolated from
bowel mucosal of Crohn’s disease

Non-cultivatable
  M.leprae
    Hansen’s bacilli
    Leprosy
    Humans only known host
13-7
Runyon Classes of NTM:
   I. Photochromogens: develop
pigment following exposure to light
after growth in dark
   II. Scotochromogens: develop
pigment in the dark
   III. Nonphotochromogens:
nonpigmented in light or dark
   IV. Rapid growers: < 7 days
14
Spirochetes and Curved Rods:
Treponema              Spirillum
Leptospira
Borrelia
1. Spirillum minus (minor)
   *curved rod (with 2-3 coils)
   *Rat bite fever (Sodoku)
   *Never been grown in culture
2.Treponema
Disease             Organism
*Syphilis           T.pallidum,
                    biotype
                    pallidum
*Yaws               T.pallidum
                    biotype
                    pertenue
14-2
*Pinta               T carateum
*Vincent’s
gingivitis        various treponemes
(Trench mouth) with anaerobes
Tests for syphilis:
   A. Treponemal:
      Darkfield Microscopy
      Immunofluorecent staining
      FTA-ABS (most sensitive)
  B. Non-treponemal
     VDRL (Venereal Disease
        Research Lab)
     RPR (Rapid Plasma Reagin)
     Complement Fixation
14-3
3. Leptospira interrogans
      (hooked ends)
   Leptospirosis:
      1. Anicteric: septicemia and
         aseptic meningitis
      2. Icteric: Weil’s disease
         Most severe: death in 10%
   Infectious Leptospiral Jaundice

4. Borrelia species:
   **Relapsing fever: >15 species
        Vector: tick or louse
     Diagnosis: microscopic exam
     of blood during febrile phase
        Darkfield microscopy
        Wright or Giemsa stain
14-4
  **Lyme Disease
     Borrelia burgdorferi: isolated
1982 by Dr. Willy Burgdorfer
  in Lyme, Connecticut
     Vector: Ixodes dammini
              Ixodes pacificus (Ca)
     Natural hosts for ticks:
              Deer and rodents
     3 Stages of disease:
        1. Erythema migrans (EM)
           Ring shaped skin lesion
        2. Arthritis and neurologic
        3. Chronic arthritis
15
Miscellaneous Pathogens:
1. Streptobacillus moniliformis
      Haverhill fever: non-spirillar
      rat bite fever
2. Mycoplasma sp. PPLO
(Pleuro Pneumonia-Like Orgs)
   *Primary Atypical Pneumonia
      cold agglutinins
      walking pneumonia
   *M.hominis
      non-gonococcal urethritis
3. Legionella pneumophilia
   Legionaire’s disease
   BCYE agar (Buffered Charcoal-
Yeast-Extract agar) 2 plates
   Faint gram neg stain
  15-2
  Silver or Giemsa stain of tissue
  Direct immunofluorescence Ab
(DFA) of respiratory secretions
  Molecular techniques (DNA)
16
Obligate Intracellular Parasites:
Chlamydia:
   C. trachomatis
     Oculogenital Infections
        most common STD
        acute conjunctivitis
        Neonatal pneumonia
     Trachoma:
        Chronic inflammation of
        Conjunctiva blindness
     LGV (Lymphogranuloma
   Venereum) STD (not in US)
16-2
C.psittaci
  Common in birds
  Aerosol inhalationhumans
C.pneumoniae (TWAR)
  Mild respiratory disease
Life Cycle
  *Reticulate Body (RB) intra-
  Cellular replication form
  *Elementary Body (EB) inert
  Extracellular infective form
Diagnosis:
  *DFA slides of infected cells
16-3
*Wright’s stain: granular
cytoplasmic inclusions
     **contain glycogen (stain
          with Iodine)
*Transport mediacultivate
     in cell culture(McCoy or
     Green Monkey Lines): stain
     monolayer
*Chlamydia Antigen tests
*DNA probe
(Chemiluminescent)
16-4
Rickettsia:
  * small, gram- coccobacillus
  *no LPS or peptidoglycan
  *Live in wild animals
  *Vectors: arthropod (insect)
  *multiply in vascular
  endothial cells
  *Serological test: Weil-Felix
     **serum agglutinates with
       Proteus vulgaris
  *Rocky Mountain spotted
  fever/vector tick
17
Viruses:
   *nucleic acid (RNA or DNA)
   *capsid: protein coat
        Capsomeres:subunits
   *may have envelope/spikes
Terms:
   Syncytia: aggregates of cells
fused together1 large cell
with multiple nuclei
   TORCH testing:Toxoplasma,
Rubella, Cytomegalovirus,
Herpes Simplex (Cause neonatal
infections)
17-2
Immunizations available:
  Poliomyelitis (Sabin/Salk)
  Measles
  Mumps
  Rubella (German measles)
  Hep B & Hep A
  Influenzae
  Yellow fever
  Small pox
  Chickenpox
     Varicella/Shingles
  Rotavirus
  HPV
17-3
Antiviral Therapy:
  Acyclovoir (HSV & Varicella)
  Rimantadine (Influenzae A)
  Amantadine (Influenzae A)
  Famciclovir (HSV
&Varicella)
  Azidothymidine (HIV)
  Protease Inhibitors (HIV)
  Ribavirin (RSV)*
  Ganciclovir (CMV)
  Fascarnet (CMV)
*Respiratory Syncytial Virus
17-4
Viral Classifications
  DNA or RNA (single or
  double strand)
Latent Infections:
  Most DNA and Retroviruses
can integrate into host genome
and replicate with host cell
  Can reactivate into viral
shedding

Viral Highlights:
 A.Oncogenic viruses
   a. Human Papilloma
17-5
    b. HBV: Liver cancer
    c. HHV: Kaposis
     Epstein-Barr: Burkitt
     Lymphoma
    d. HTLV: T-cell leukemia
 B. Retroviruses
     Reverse transcriptase
       RNADNA
     HIV
     HTLV
 C. Diseases:
     Adenovirus: most colds
17-6
   Herpes Viruses:encephalitis
      Epstein-Barr: IM
      Herpes Simplex
      Varicella: chickenpox
      Herpes Zoster: shingles
      CMV: slow grower (NBs)
   RSV: infect upper respiratory
ciliated cells (nasalpharynx
specimen)
   Flavivirus: Yellow fever
   Spread: Aedes sp. mosquito
   Rhabdovirus: rabies
        (Negri bodies)
17-7
Viral Specimens:
*Place on ice or refrigerate
     DO NOT FREEZE
*Swabs(cotton,rayon,dacron)
NOT Ca alginate:inactivate HSV
*Process quickly
*Transport Media:
   (Stuarts, Amie’s, Hanks
balanced salt solution, Eagles)
*use biological safety cabinet
Tissue smears/biopsy: look for
intracelluar inclusions
17-8
Detection:
 A.Tzanck test (VZV or HSV)
 B. Negri bodies (Rabies)
 C. Immunofluorescent stains
 D.ELISA
 E. ELVIS (Enzyme Linked
    Virus Inductible System)
 F. Nuclei Acid Probes
 G.PCR
17-9
Cell culture:
    a. PMK (primary monkey
     kidney)
    b. Hep-2 (human
     epidermoid carcinoma)
    c. HDF (human diploid
     fibroblast)
In cell cultures produce CPEs
Cytopathic Effect on cells
   Areas of dead cells
Shell Viral Culture
   More rapid way to see CPEs
Viral Serology: detect Antibody
in patient’s serum

Paired sera
   1. Acute (drawn ASAP)
   2. Convalescent 2-3 weeks
Positive Test: fourfold Ab titer
increase between

IgM detected first 1-4 months
Then IgG detectable for life
IgM:indicates a recent infection
17-11
Routine Testing for Antibodies:

Rubella: pregnancy
Varicella/Measles: heath care
workers
CMV: organ transplant donors
  Premature infants: only CMV
neg blood (transmitted by
WBCs)

				
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posted:12/1/2011
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