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Salmonella in Australian Macropods FACT SHEET

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Salmonella in Australian Macropods FACT SHEET Powered By Docstoc
					                     Salmonella in Australian Macropods
                                FACT SHEET

Introductory statement
Human salmonellosis is one of the most common and economically important zoonotic diseases in the
world (OIE, 2008). It is therefore important to recognise that macropods can harbour salmonellae
without displaying clinical signs of disease (Samuel, 1982). It is not uncommon for people to come into
direct contact with macropods or their faeces in the urban environment, as kangaroos are commonly
found grazing at local parks and golf courses and wildlife carers often hand-rear orphaned joeys. The
risk posed by Salmonella to both people and their pets through carcass contamination during
harvesting and processing of kangaroos is not known.

Aetiology
Salmonella is a gram-negative, rod-shaped bacilli belonging to the family Enterobacteriaceae, genus
Salmonella (CDC, 2008). Salmonella nomenclature is complex and evolving (Brenner et al., 2000).
Currently, the genus Salmonella consists of only two species, S. enterica and S. bongori. S. enterica is
further divided into six subspecies, which are referred to by a Roman numeral and a name (I, S.
enterica subsp. enterica; II, S. enterica subsp. salamae; IIIa, S. enterica subsp. arizonae; IIIb, S.
enterica subsp. diarizonae; IV, S. enterica subsp. houtenae; VI, S. enterica subsp. indica (OIE, 2008;
Brenner et al., 2000). The individual names of each serotype in subspecies I continue to be used for
identification, whilst antigenic formulae are cited for unnamed serotypes described after 1966 within the
remaining subspecies and in S.bongori. S. bongori, formally classed as subsp. V, still retains this
symbol in classification despite being recognised as a separate species. Antigenic formulae include
subspecies designation, followed by O (somatic) antigens and flagellar protein antigens (H) in
accordance with the Kauffmann–White scheme (OIE, 2008; Brenner et al., 2000). At the first citation of
a serotype in subsp. I, the genus name is given followed by the word “serotype” or the abbreviation
“ser.”, then the serotype name. Subsequently, the name may be written with the genus followed directly
by the serotype name. The serotype name is capitalised and not italised (Brenner et al., 2000),
although variations of this exist in the literature.


Natural hosts
Salmonellae are known to infect a wide range of natural hosts, including macropods. Within Australia,
quokkas (Setonix brachyurus) on Rottnest Island, pet kangaroos, kangaroos held in captivity and hand-
reared joeys are particularly susceptible to infection (Speare and Thomas, 1988; Hart et al., 1985;
Iveson and Bradshaw, 1973; Arundel, 1981). Records collated by the National Enteric Pathogens
Surveillance System (NEPSS) in Australia up until 2006 indicate that Salmonella spp. are most
commonly isolated from red (Macropus rufus), eastern grey (Macropus giganteus) and western grey
(Macropus fuliginosus) kangaroos. Salmonella has also been reported in the boodie or Lesueur’s
bettong/rat kangaroo (Bettongia lesueur). A significant number of isolates were obtained from
macropods whose species was recorded only as kangaroo, rat kangaroo or in one instance, as wallaby
(National Enteric Pathogens Surveillance Scheme, 2007). Consequently, it is possible that the list of
natural macropod host species is more extensive.

World distribution
Salmonella has a world-wide distribution, but appears to be most prevalent in areas of intensive animal
husbandry, particularly where pigs, calves and poultry are housed in confinement (OIE, 2008).

Occurrences in Australia
Salmonella is present throughout Australia. Incidence of foodborne salmonellosis is highest among
young children and their carers, together with residents of the Northern Territory (OzFoodNet Working
Group, 2007).



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Epidemiology
The morbidity and mortality rate of Salmonella infection in wild macropods is believed to be negligible.
Although theoretically possible under conditions of environmental stress, there have been no confirmed
reports of primary clinical disease in these animals in their natural habitat (Speare et al., 1989).
Contamination of kangaroo meat, however, does suggest that wild macropods are capable of
harbouring the organism. Studies have also demonstrated the presence of infection in a large number
of healthy, captive macropods. In captivity, these animals may become infected as a result of oral
consumption of contaminated feed or water (Samuel, 1982). Shedding of Salmonella is variable and
cannot always be related to the level of stress the animal is exposed too. Healthy macropods (species
unspecified) housed in hygienic, large, grassed enclosures were found to have higher rates of infection
than macropods held at a research institute that were regularly exposed to stress associated with
handling, fighting and travel. Despite the introduction of an infected kangaroo to this latter group, all
animals remained negative. There were no differences in the prevalence which could be attributed to
the sex, age or species of macropod (Samuel, 1982). Three agile wallabies (Macropus agilis) orally fed
a dose of S. ser. Infantis began shedding the organism within twenty four hours, despite the absence of
clinical signs. Prevalence of Salmonella infection in orphaned joeys is considered to be significant. A
severely ill joey is likely to excrete more Salmonella than a healthy joey, posing a greater risk to its
carer (Speare and Thomas, 1988).

Clinical signs
Salmonellosis is the clinical manifestation of infection with Salmonella (Blood and Studdert, 1999). In
hand-reared joeys and kangaroos held in captivity, salmonellosis can cause varying levels of
gastroenteritis and septicaemia, manifesting most commonly as diarrhoea. Infection has also been
reported to cause death as a primary pathogen and as an opportunistic invader of the intestinal tract in
animals in captivity (Samuel, 1982). Joeys often present as dull and lethargic with a reduced appetite.
Diarrhoea becomes haemorrhagic and foul smelling with time (Staker, 2006). Clinical disease is
possible in free-ranging macropods subjected to environmental stress, but has not been reported
(Speare et al., 1989). Many infected kangaroos are asymptomatic, yet are still capable of shedding
large numbers of Salmonella.


Diagnosis
Diagnosis in kangaroos is based on isolation of the organism from aseptically collected tissue at
necropsy or from faeces or rectal swabs. As many animals are asymptomatic carriers, other evidence
of disease compatible with salmonellosis must be present. Prior or current infection of animals by some
serovars may also be diagnosed serologically (OIE, 2008), although this has not been reported in
macropods.


Clinical Pathology
Clinical pathology in macropods has not been described.


Pathology
Due to the large number of healthy macropods found to carry Salmonella yet exhibit no clinical signs, it
is often difficult to attribute illness, death and consequently, pathology, to this organism. The natural
history of Salmonella in wild macropods has largely been studied in the quokka (S. brachyurus) on
Rottnest Island, located off the coast of Western Australia (Hart et al., 1985; Iveson and Bradshaw,
1973). Despite virtually all animals being positive throughout the summer months, no evidence of
disease has been found in infected quokkas (Hart et al., 1985). Consequently, there are no reports of
pathology. Agile wallabies (M. agilis) infected with S. infantis were found to shed the bacteria within a
day of being inoculated however no signs of illness were displayed. Whilst no gross or histological
changes were noted at post mortem, Salmonella was recovered from the walls of the ileum, caecum,
stomach, stomach contents and mesenteric lymph nodes draining the intestine (Samuel, 1983).
Salmonella has been isolated less commonly from the urine, liver, lung, kidney, heart, abdomen and
spleen of macropods. Carriers often harbour the organism in their mesenteric and ileocaecal lymph
nodes (Thomas et al., 2001).




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Differential diagnoses
In captive macropods, differential diagnoses of Salmonella include other causes of gastroenteritis and
diarrhoea. Coccidiosis is a common differential. In the event of acute death, toxoplasmosis should also
be considered. Strongyloides has been found to cause diarrhoea and death in eastern grey
(M.giganteus), western grey (M. fuliginosus) and red kangaroos (M. rufus) as well as a red-necked
wallaby (Macropus rufogriseus) (Arundel, 1981; Winter, 1957). In hand-reared joeys, diet and
environmental stress, coccidiosis, yeast/candidiasis, roundworms, antibiotic administration and
cryptosporidia are all differential diagnoses of diarrhoea, listed from most through to least likely (Mason,
2007). Other causes of bacterial diarrhoea, including E. coli and Klebsiella, should also be considered
(Gage, 2002).


Laboratory diagnostic specimens
There are numerous methods for isolation of Salmonella in use world-wide. In macropods, culture is the
most common method of diagnosis and is used to isolate Salmonella from tissues collected aseptically
at post mortem or from faecal and rectal swabs in infected animals. Studies have shown that the
relative yield of Salmonella in faecal material is greater than from rectal swabs (Hart et al., 1982).
Lymph nodes, primarily the mesenteric, commonly harbour Salmonella in carrier animals, even when
they are not shedding. The detection rate of Salmonella was found to be 2.4 times greater in joeys that
had died than in live joeys as a result of the isolation of Salmonella spp. from lymph nodes (Speare and
Thomas, 1988). Consequently, these are particularly useful tissue samples to collect at post mortem.
Salmonella has also been isolated from ileocaecal lymph nodes, liver, lung, urine kidney, spleen and
gall bladder (Samuel, 1982; Thomas et al., 2001). Samples should be collected as aseptically as
possible during the acute phase of disease or as soon after death as possible. It is also preferable to
collect samples prior to the commencement of any antibiotic treatment (OIE, 2008).


Laboratory procedures
Salmonellae may be isolated using a variety of culture techniques, involving pre-enrichment to
resuscitate sublethally damaged salmonellae, enrichment media containing inhibitory substances to
suppress competing organisms and selective plating agars to differentiate salmonellae from other
enterobacteria. Care must be taken when selecting an enrichment media as some can be toxic to
certain Salmonella serovars. More detailed information on the culture of Salmonella can be obtained
from the ‘Manual of diagnostic tests and vaccines for terrestrial animals 2008’ (OIE, 2008).

Other forms of Salmonella detection in use include immunomagnetic separation (IMS), electrical
conductance/impedence and PCR. Serological tests to detect infected herds/flocks have also been
developed and include the serum agglutination test (SAT) and ELISA (OIE, 2008). Their use has not
been reported in macropods.


Treatment
There are no published reports of the efficacy of antibiotics in the treatment of Salmonella infection in
macropods. Culture and sensitivity is likely to provide a useful means of determining the most effective
antibiotic to use, given it is not contraindicated in the species. Gage (2002) recommends the use of a
broad spectrum sulfa/sulfa-trimethoprim antibiotic at a dose of 5mg/kg of the trimethoprim component
P/O OID for 5-7 days (Gage, 2002). Gentamycin and amoxicillin have also been suggested (Staker,
2006). Additional oral or Intravenous fluids coupled with glucose/electrolytes, probiotics, a multivitamin
and a reduction in diet and environmental related stress may assist in recovery (Staker, 2006). There is
no requirement to treat asymptomatic infection in macropods.


Prevention and control
Minimising diet and environmental related stress is paramount in preventing and controlling Salmonella
in macropods in captivity and joeys in care. Dogs should be prevented from cleaning the cloaca of hand
reared joeys and carers should ensure their hands are washed immediately after contact with the
animal. Artificial pouches should be washed/disinfected if soiled, avoiding contamination of the joey’s
fur. Regular cleaning and disinfection of enclosures, including the removal of faecal matter will assist in
reducing environmental contamination. Salmonellae are susceptible to a range of disinfectants,
including 1% sodium hypochlorite, 70% ethanol, 2% glutaraldehyde, iodine-based disinfectants,
                                                                         C
phenolics and formaldehyde. They can also be killed by moist heat (121° for a minimum of 15 min) or
                     C
dry heat (160-170° for at least 1 hour) (The Center for Food Security and Public Health, 2005). In




                                                  Page 3                                        27/06/2009
captivity, feed contamination can be a source of infection and should be investigated for the presence
of salmonellae.

Professional shooters working in the commercial harvesting industry need to ensure the gut is not
perforated during the evisceration process, resulting in contamination of the carcass at harvesting.
Cross contamination at the processor can be minimised by thoroughly washing knives between
carcasses and implementing adequate decontamination procedures. All individuals coming into contact
with kangaroos need to maintain adequate levels of hygiene. Care should be taken when handling and
storing kangaroo meat to avoid cross contamination of other foodstuffs. Thorough cooking of meat
intended both for human and pet consumption will also reduce the risk of salmonellosis.


Surveillance and management
Notifiable Salmonella serovars within Australia include Salmonella ser. Enteriditis in poultry, Salmonella
ser. Abortus equi, Salmonella ser. Abortus ovis and Salmonella. ser. Pullorum. A number of these are
also notifiable to the OIE. These serotypes are largely species specific and are unlikely to cause
infection in macropods.

Since 1980, the National Enteric Pathogens Surveillance System (NEPSS) has collated, analysed and
disseminated data on human enteric bacterial infections diagnosed in Australia. Reports of isolates are
received from the five laboratories capable of serotyping and phage typing Salmonella within Australia.
Salmonella isolates are submitted to these laboratories by other diagnostic laboratories throughout the
country. The NEPSS publish their findings via Communicable Diseases Intelligence (CDI) (Department
of Health and Ageing, 2008).

In 2000, the OzFoodNet network was established by the Australian Government Department of Health
and Ageing to undertake national investigations into gastroenteritis outbreaks and clusters of disease
potentially related to food products. Results are announced quarterly and annual reports have been
published in CDI since 2002 (Department of Health and Ageing, 2008).


Statistics
According to the National Wildlife health Surveillance Database (Event No. 556), a mass mortality event
suspected to be associated with drought in Meekatharra, Western Australia, resulted in the death of
approximately 3000 red kangaroos (M. rufus). Salmonella was cultured from three animals, one of
which had bacteria associated with the intestinal lining. Whilst it is possible Salmonella played a role in
the eventual death of this animal, it is likely that dehydration and malnourishment were the primary
problems. AWHN also provides reports on wildlife isolates to the NEPSS, which maintains a summary
database of Salmonella isolates from wildlife in Australia. This is a valuable resource for Australia.


Research
Macropods were first suspected of carrying high levels of Salmonella in the 1960’s when it became
evident that contamination of kangaroo carcasses in pet meat preparation was a definite risk factor for
infection in pets (Anderson et al., 1964). Following a case of salmonellosis in a young infant on Rottnest
Island, off the coast of Western Australia (Iveson and Bradshaw, 1973), a significant amount of
research was undertaken to investigate whether quokkas (S. brachyurus) were carriers of the organism.
It was soon discovered that infection in these small macropods peaked as high as 70-100% in the
summer months as a result of disruption to the digestive physiology associated with a decline in feed.
No signs of clinical disease were noted (Hart et al., 1985). Whilst studies investigating the natural
history of Salmonella in macropods has been largely limited to the quokka (S. brachyurus), it is
important to note that these animals are unlikely to be representative of large-bodied macropods on the
mainland. Consequently, care should be taken in extrapolating this data.

In a prevalence study of Salmonella in macropods (varying species) from Queensland, 51% of animals
tested were found to be positive. Thomas et. al. (2001) undertook a similar study, culturing faeces and
organs of macropods over a period of twenty years. From the 57 animals sampled, 62 isolates were
cultured, belonging to 24 different Salmonella serotypes. Whilst these studies report high levels of
infection, it is important to realise that environmental and diet related stress associated with living in
captivity is likely to result in a potentially higher prevalence than in macropods in their natural
environment (Samuel, 1982; Thomas et al., 2001). For similar reasons, orphaned joeys are considered
an occupational risk for their wildlife carers. Under care, joeys often experience stress related to
sudden withdrawal from their mother, a change in nutritional diet and unaccustomed exposure to
humans and other animals. Between 1981 and 1985, Speare and Thomas (1988) examined orphaned
macropod young. Samples were taken from 65 live joeys and 38 dead joeys. A total of 26.8% were



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found to be infected with Salmonellae spp. whilst 21.7% were actively excreting the bacteria in faeces
(Speare and Thomas, 1988).

Kangaroo meat intended for human and pet food consumption has been associated with high rates of
Salmonella contamination for some time now. Whilst early studies reported contamination rates as high
as 44.9% (Suzuki et al., 1967), enforced regulatory standards within the industry have seen this figure
decrease significantly. More recent studies have reported contamination rates between 0.84% and 18%
(Holds et al., 2008; Eglezos et al., 2007; Bensink et al., 1991). Presence of Salmonella on kangaroo
carcasses suggests that natural infection does occur in wild macropods. However, there are no
published studies reporting the true prevalence of infection in these animals.

Research is currently being undertaken at Murdoch University in Western Australia to determine the
prevalence of Salmonella in kangaroos (predominantly M.fuliginosus) in their natural habitat. It is
important to draw comparison between the level of natural infection and the rate of carcass
contamination. Any significant differences are likely to highlight the need to improve hygiene practices
at the processor and evisceration techniques in the field. More extensive studies in all states,
comparing the rate of carcass contamination between processors for human consumption and those for
pet meat need to be investigated, as different standards exist. Contaminated pet meat is a direct source
of infection to household animals, who are then likely to contaminate the household environment.
Contaminated pet meat is also a source of infection to household members who handle the food and
store it in their own refrigerators.


Human health implications
In Australia every year, there are approximately 17.2 million cases of gastroenteritis, of which 5.4
million are estimated to have originated from contaminated food. Salmonella is one of the three most
common pathogens responsible for foodborne gastroenteritis (Hall and Kirk, 2005). Food products of
animal origin are considered to be the major source of human Salmonella infections (OIE, 2008). A
number of serovars commonly responsible for salmonellosis have been isolated from kangaroos,
including S. ser. Typhimurium, S. ser. Saintpaul, S. ser. Chester and S. ser. Muenchen (National
Enteric Pathogens Surveillance Scheme, 2007). At this stage, no confirmed outbreaks of foodborne
gastroenteritis have been linked to the consumption of kangaroo meat. There is also an occupational
risk associated with salmonellosis. Meat workers may acquire infection from carcasses contaminated
during the evisceration process as can individuals working in close association with joeys and
kangaroos in captivity.


Conclusions
Salmonella infections in animals intended for human consumption are considered to be a major source
of salmonellosis. Whilst kangaroos have not yet been linked with an outbreak of disease, these animals
do present a potential risk of infection both to people and their pets. Despite the prevalence of infection
in wild macropods, other than the quokka (S. brachyurus), remaining unknown, the contamination of
kangaroo meat suggests that macropods in the wild are capable of harbouring Salmonella.
Furthermore, studies have demonstrated that high rates of infection occur in orphaned joeys and
captive macropods. Consequently, it is important for all individuals coming into close contact with these
animals and their meat products to practice adequate hygiene measures at all times.


References and other information
Anderson K, Crowder EF and Woodruff P. 1964. The isolation of samonellae from kangaroo meat sold
as pet food. Med. J. Aust. 2. 668 - 669.

Bensink JC, Ekaputra I and Taliotis C. 1991. The isolation of Salmonella from kangaroos and feral pigs
processed for human consumption. Aust. Vet. J. 68. (3). 106-107.

Blood DC and Studdert VP. 1999. Saunders Comprehensive Veterinary Dictionary. WB Saunders;
London.

Brenner FW, Villar RG, Angulo FJ, Tauxe R and Swaminathan B. 2000. Salmonella nomenclature. J.
Clin. Microbiol. 38. (7). 2465-2467.
Speare R and Thomas AD. 1988. Orphaned kangaroo and wallaby joeys as a potential zoonotic source
of Salmonella spp. Med. J. Aust. 148. (12). 619, 622-623.

CDC. 2008. Salmonellosis; (Accessed: 12th May, 2009).
http://www.cdc.gov/nczved/dfbmd/disease_listing/salmonellosis_ti.html




                                                 Page 5                                        27/06/2009
Department of Health and Ageing. 2008. Surveillance systems: NEPSS; (Accessed: April 29th, 2009).
http://www.health.gov.au/internet/main/publishing.nsf/Content/cda-surveil-surv_sys.htm

Department of Health and Ageing. 2008. Surveillance systems reported in CDI, 2008; (Accessed: 25th
June, 2007, 2007). http://www.health.gov.au/internet/wcms/publishing.nsf/Content/cda-surveil-
surv_sys.htm

Eglezos, Sofroni, Huang, Bixing and Stuttard. 2007. A survey of the microbiological quality of kangaroo
carcasses processed for human consumption in two processing plants in Queensland, Australia. J.
Food Prot. . 70. (5). 1249 -1251.

Gage L. 2002. Hand-rearing wild and domestic mammals Wiley-Blackwell; Australia.

Hall G and Kirk M. 2005. Foodborne illness in Australia; Annual incidence circa 2000. Department of
Health and Ageing.

Hart RP, Bradshaw SD and Iveson JB. 1985. Salmonella infections in a marsupial, the quokka (Setonix
brachyurus), in relation to seasonal changes in condition and environmental stress. Appl. Environ.
Microbiol. 49. (5). 1276-1281.

Hart RP, Iveson JB, Bradshaw SD and Speed TP. 1982. A study of isolation procedures for multiple
infections of Salmonella and Arizona in a wild marsupial, the quokka (Setonix brachurus). J. Appl.
Bacteriol. 53. 395-406.

Holds G, Pointon A, Lorimer M, Kiermeier A, Raven G and Sumner J. 2008. Microbial profiles of
carcasses and minced meat from kangaroos processed in South Australia. Int. J. Food Microbiol. 123.
88 - 92.

Iveson JB and Bradshaw SD. 1973. Salmonella javiana infection in an infant associated with a
marsupial, the quokka, Setonix brachyurus, in Western Australia. J. Hyg. 71. 423 - 432.
Arundel JH. 1981. Wildlife diseases of the Pacific Basin - Australia. (Accessed

Mason P. 2007. Diarrhoea in Joeys. Healesville Sanctuary; Australian Wildlife Health Centre.
(Accessed 15th May, 2009).
http://www.wildlifevictoria.org.au/cms/images/stories/docs/diarrhoea%20in%20joeys.pdf

National Enteric Pathogens Surveillance Scheme. 2007. National enteric pathogen surveillance system
data: Salmonella in macropods. Microbiological Diagnostic Unit; Department of Microbiology and
Immunology, The University of Melbourne.

OIE. 2008. Salmonellosis 2.9.9; Manual of diagnostic tests and vaccines for terrestrial animals 2008.
(Accessed: 2009, 12th May).
http://www.oie.int/eng/normes/mmanual/2008/pdf/2.09.09_SALMONELLOSIS.pdf

OzFoodNet Working Group. 2007. Monitoring the incidence and causes of diseases potentially
transmitted by food in Australia: Annual report of the OzFoodNet Network, 2006 CDI. 31. (4).

Samuel JL. 1982. Salmonella in macropods. In: Wildlife diseases of the Pacific Basin and other
Countries; Proceedings of the 4th International Conference of the Wildlife Disease Association.
Sydney. 61 - 63.

Speare R, Donovan JA, Thomas AD and Speare PJ. 1989. Diseases of free-ranging Macropodoidea.
In: Kangaroos, wallabies and rat-kangaroos. 705-734.

Staker L. 2006. The complete guide to the care of macropods; a comprehensive guide to the
handrearing, rehabilitation and captive management of kangaroo species. Lynda Staker;
Samuel JL. 1983. Jaw disease in macropod marsupials: bacterial flora isolated from lesions and from
the mouths of affected animals. Vet. Micro. 8. (4). 373 - 387.

Suzuki A, Kawanishi K, Suzuki N and Takayama S. 1967. A survey of Salmonella contamination of
imported meats during 1965 - 1966. Bull. Natn. Inst. Hyg. Sci. 85. 188 - 195.

The Center for Food Security and Public Health. 2005. Salmonellosis; Paratyphoid, non-typhoidal
salmonellosis. (Accessed: 19th May, 2009).
http://www.cfsph.iastate.edu/Factsheets/pdfs/Nontyphoidal_Salmonellosis.pdf




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Thomas AD, Forbes-Faulkner JC, Speare R and Murray C. 2001. Salmonelliasis in wildlife from
Queensland. J. Wildl. Dis. 37. (2). 229 - 238.
Winter H. 1957. Salmonella birkenhead from a kangaroo. Australia Veterinary Journal. 33. (3). 67.

Updated: 22/05/2009

To provide feedback on this fact sheet
The Australian Wildlife Health Network would be very grateful for any feedback on this fact
sheet. Please provide detailed comments or suggestions to rwoods@zoo.nsw.gov.au. We
would also like to hear from you if you have a particular area of expertise and would like to
produce a fact sheet (or sheets) for the network (or update current sheets). A small amount
of funding is available to facilitate this.

Disclaimer

This fact sheet is managed by the Australian Wildlife Health Network for information purposes
only. Information contained in it is drawn from a variety of sources external to the Australian
Wildlife Health Network. Although reasonable care was taken in its preparation, the Australian
Wildlife Health Network does not guarantee or warrant the accuracy, reliability, completeness,
or currency of the information or its usefulness in achieving any purpose. To the fullest extent
permitted by law, the Australian Wildlife Health Network will not be liable for any loss, damage,
cost or expense incurred in or arising by reason of any person relying on information in this
fact sheet. Persons should accordingly make and rely on their own assessments and
enquiries to verify the accuracy of the information provided.




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