Herpesviruses and macropods Fact sheet

Introductory statement

Despite the widespread distribution of herpesviruses across a large range of macropod there is a lack of detailed knowledge about these viruses and the effects they have on their hosts. While they have been associated with significant mortality events infections are usually benign, producing no or minimal clinical effects in their adapted hosts. With increasing emphasis being placed on captive breeding, reintroduction and translocation programs there is a greater likelihood that these viruses will be introduced into naïve macropod populations. The effects and implications of this type of viral movement are unclear.

Aetiology

Herpesviruses are enveloped DNA viruses that range in size from 120 to 250nm. The family Herpesviridae is divided into three subfamilies. Alphaherpesviruses have a moderately wide host range, rapid growth, lyse infected cells and have the capacity to establish latent infections primarily, but not exclusively, in nerve ganglia. Betaherpesviruses have a more restricted host range, a long replicative cycle, the capacity to cause infected cells to enlarge and the ability to form latent infections in secretory glands, lymphoreticular tissue, kidneys and other tissues. Gammaherpesviruses have a narrow host range, replicate in lymphoid cells, may induce neoplasia in infected cells and form latent infections in lymphoid tissue (Lachlan and Dubovi 2011, Roizman and Pellet 2001). There have been five herpesvirus species isolated from macropods, three alphaherpesviruses termed Macropodid Herpesvirus 1 (MaHV1), Macropodid Herpesvirus 2 (MaHV2), and Macropodid Herpesvirus 4 (MaHV4) and two gammaherpesviruses including Macropodid Herpesvirus 3 (MaHV3), and a currently unclassified novel gammaherpesvirus detected in swamp (Wallabia bicolor) (Callinan and Kefford 1981, Finnie et al. 1976, Johnson et al. 1985, Smith et al. 2008, Stalder unpublished, Vaz, et al. 2013, Wilcox et al. 2011).

Natural hosts

MaHV1 has caused mortalities in parma wallabies ( parma), while MaHV2 has caused mortalities in wallabies (Dorcopsis muelleri luctuosa) and (Setonix brachyurus). MaHV3 has been associated with cloacal ulceration, respiratory disease and possibly with mammary neoplasia in eastern grey (Macropus giganteus). MaHV4 infection was associated with rhinitis and conjunctivitis in an eastern grey . Unidentified herpesviruses have caused mortality in red kangaroos (Macropus rufus), brush-tailed (Bettongia penicillata), rufous bettongs (Aepyprymnus rufescens), long-nosed ( tridactylus), black tree kangaroos (Dendrolagus ursinus), dusky (Thylogale brunii), and brush-tailed rock wallabies (Petrogale penicillata). They have also been associated with cloacal ulceration in tammar wallabies (Macropus eugenii). A possible herpesvirus infection may have been responsible for fatal disease in a (Macropus fuliginosus) and a black (Macropus bernardus) (Acland 1981, Britt et al. 1994, Callinan and Kefford 1981, Canfield and Hartley 1992, Dickson et al. 1980, Finnie et al. 1976, Finnie 1980, Jerrett 2007, Smith et al. 2008, Vaz et al. 2013, Wilcox et al. 2011, Wilks et al. 1981). More recently a novel gammaherpesvirus species has been detected in free-living, healthy swamp wallabies (Stalder unpublished).

World distribution

MaHV3 has been found in a group of captive eastern grey kangaroos with cloacal ulcers in the United States (Smith et al. 2008). An unidentified herpesvirus was detected in a with hepatitis, also in the United States (Britt et al. 1994). Herpesvirus antibodies have been found in macropods in New Zealand (Webber and Whalley 1978). Given the high prevalence of macropod herpesviruses in captive Australian macropods it seems likely that prevalence is similarly high in outside .

Occurrences in Australia

Macropod herpesviruses occur Australia-wide with all macropod species assumed to be susceptible to infection (Vogelnest and Portas 2008).

Epidemiology

Herpesviruses tend to form latent infections that are periodically reactivated during times of stress or immune-compromise. This capacity has been demonstrated in macropods through a study of MaHV1 seropositive eastern grey kangaroos that began excreting an alphaherpesvirus similar to MaHV1/2 during treatment with corticosteroids. Virus was found in nasal swabs indicating a probable respiratory route of transmission. Herpesvirus DNA was also detected in the trigeminal nerve ganglia of two kangaroos with either reactivated or latent infection. No herpesvirus DNA was detected in cloacal swabs or blood from these animals (Guliani et al. 1999). Herpesviruses have been identified in cloacal swabs of tammar wallabies suffering ulcerative cloacitis, indicating that virus transmission may also occur via the venereal route (Holz, unpublished.).

Herpesviruses are fragile and do not survive well outside the body. They are killed by all common disinfectants including bleach and F10 (benzalkonium chloride/polyhexamethylene biguanide hydrochloride).

Transmission of other herpesviruses generally requires close contact such as mating, licking, nuzzling or sneezing resulting in aerosol spread over short distances.

Clinical signs

Clinical disease has predominantly been described in captive animals. Signs include sudden death, depression, fever, incoordination, conjunctivitis, increased respiratory sounds, and vesicles and ulcers on the oral mucosa, cloaca and penis (Ladds 2009). Tammar wallabies have developed cloacal ulcers which heal but then recur

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(Holz unpublished.). An association between MaHV3 infection and mammary neoplasia has also been postulated (Smith et al. 2008). In a mob of free ranging eastern grey kangaroos, MaHV3 was implicated in an outbreak of respiratory disease, lethargy, inappetence and ataxia, leading to mortality in some cases (Wilcox et al. 2011). MaHV4 was isolated from a free ranging showing signs of rhinitis and conjunctivitis (Vaz et al. 2013).

Diagnosis

Diagnosis in the live is largely dependent on demonstrating a rising serum antibody titre via the serum neutralisation test. The collection of swabs from conjunctivae, nostrils, oropharynx or cloaca may also demonstrate the presence of herpesvirus DNA in actively shedding infections (Stalder unpublished). This test may be more sensitive in the presence of cloacal lesions or mucocutaneous ulceration. At necropsy multifocal hepatic necrosis together with intranuclear inclusion bodies is suggestive of herpesvirus infection. This can be confirmed through electron microscopy, viral culture or PCR (Ladds 2009, Vogelnest and Portas 2008).

Pathology

Detailed information on the pathology of macropod herpesviruses is available at the Australian Registry of Wildlife Health (ARWH: http://www.arwh.org/). Post mortem lesions can include a mucoid tracheitis, pulmonary congestion and pneumonia and pale foci in the liver corresponding to multifocal areas of necrosis. Affected liver cells may contain intranuclear inclusion bodies. Diphtheritic plaques can be found on the oesophageal and gastric mucosa and focal ulceration and necrosis of the genitalia may also occur. Inclusion bodies are also found on occasion in affected areas of skin and mucosa (Vogelnest and Portas 2008).

Differential diagnoses

Differential diagnoses include macropod diseases associated with sudden death, such as and encephalomyocarditis virus infection, diseases causing hepatic necrosis, such as yersiniosis and salmonellosis and diseases causing mucosal ulceration, such as Treponema spp. infection (Vaughan-Higgins et al. 2011).

Laboratory diagnostic specimens

Detailed information on laboratory diagnostic specimens required for diagnosis of macropod herpesvirus infections is available at the ARWH (http://www.arwh.org/). For live animals, submit serum and plain swabs from oculonasal, oropharyngeal and cloacal mucosae. For dead animals a complete necropsy should be performed. Collect a range of tissues, including liver and any obvious lesions, and submit them in formalin for histopathology. Fresh tissues should also be submitted for viral culture and PCR (Vogelnest and Portas 2008).

Laboratory procedures

Detailed information on laboratory procedures required for diagnosis of macropod herpesvirus infections is available at the ARWH (http://www.arwh.org/). Serum from live animals can be tested for antibodies using the serum neutralisation test. As many macropods have antibodies to herpesviruses paired samples and a rising titre are required to demonstrate active infection. Both viral culture and PCR can be performed on tissues to demonstrate the presence of herpesvirus spp. PCR performed on oculonasal, oropharyngeal or cloacal swabs can confirm infection in the live animal.

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Treatment

Currently there is no treatment. An in vitro study evaluated the ability of the anti-herpetic compounds (E)-5- (2’-bromovinyl)-2’-deoxyuridine (BVDU), acyloguanosine (ACV), 5-iodo-2’-deoxyuridine (IdU), 5-iodo-2’- deoxycytidine (IdC), triflurothymine deoxyribose (TFR), adenine 9-β-D-arabino-furanoside (Ara-A), cytosine β- D-arabino-furanoside (Ara-C), and thymine 1-β-D-arabino-furanoside (Ara-T) to inhibit the ability of MaHV2 to form plaques in kidney cells. Ara-A was the best performing compound, followed by BVDU, but there are no reports of in vivo use in macropods (Smith 1996).

Prevention and control

Given the high prevalence of antibodies in captive macropods it is difficult to maintain a herpesvirus free population. As clinical disease appears to occur infrequently the difficulty and expense associated with maintaining negative populations will likely outweigh the benefits. If a negative population is required newly arrived animals should be quarantined and serviced separately from collection macropods until their herpesvirus status can be ascertained. Macropods with cloacal ulcers should not be used in breeding programs as joeys are presumably susceptible to infection during the birth process. Significant clinical disease and mortality outbreaks have generally been associated with periods of stress, primary exposure of immunologically naïve individuals or in novel host species, which should be considered in the management of captive or free-living macropods.

Surveillance and management

Wildlife disease surveillance in Australia is coordinated by the Wildlife Health Australia. The National Wildlife Health Information System (eWHIS) captures information from a variety of sources including Australian government agencies, zoo and wildlife parks, wildlife carers, universities and members of the public. Coordinators in each of Australia's States and Territories report monthly on significant wildlife cases identified in their jurisdictions. NOTE: access to information contained within the National Wildlife Health Information System dataset is by application. Please contact [email protected].

There is no targeted surveillance program or AUSVETPLAN for macropod herpesviruses in Australia. Cases identified as part of Australia’s general wildlife surveillance system are recorded in the national wildlife health information system.

Statistics

Neutralising antibodies are widespread in both captive and wild populations. Results from one survey are presented in Tables 1 and 2 (Webber and Whalley 1978). A second survey found 98 out of 129 eastern grey kangaroos and 52 out of 89 western grey kangaroos with antibodies (Kerr et al. 1981). Unpublished surveys in 2007 identified seven out of 11 seropositive tammar wallabies and six out of 14 brush-tailed rock wallabies housed in captive institutions in South Australia, and two out of two seropositive yellow-footed rock wallabies (Petrogale xanthopus), four out of four brush-tailed rock wallabies, and two out of eight tammar wallabies housed in Victoria. An unpublished survey conducted in 2011 at a fauna park in the ACT found no actively shedding herpesvirus infections on the basis of PCR performed on swabs from eight brush-tailed rock wallabies, three yellow-footed rock wallabies, four eastern bettongs (Bettongia gaimardi), nine long-nosed potoroos, and eight tammar wallabies despite two of the tammar wallabies having cloacal ulcers at the time of sampling. The serological status of these animals is yet to be determined.

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Unfortunately, these serological reports do not differentiate between viruses. As cross-neutralisation is common between herpesviruses, and virus isolation was not performed, it is not possible to determine which viruses these animals were exposed to. A recent study found that antibodies against MaHV4 cross-neutralised MaHV2 (Vaz et al. 2013).

Table 1. Serum neutralising antibodies to macropod herpesvirus in wild macropods (Webber and Whalley 1978).

Species No. tested No. positive % positive

Petrogale (rock-) species 102 20 19.6 Thylogale thetis and stigmata (red-necked and 7 2 28.6 red-legged ) conspicillatus (spectacled hare- 2 2 100 wallaby) Macropus parma () (Aus) 13 0 0

Macropus parma (parma wallaby) (NZ) 23 12 52.2

M. robustus robustus (eastern wallaroo) 5 2 40

M. robustus erubescens (euro) 12 1 8.3

M. giganteus (eastern grey kangaroo) 7 5 71.4

M. rufogriseus (red-necked wallaby) 6 0 0

M. rufus (red kangaroo) 2 0 0

Potorous tridactylus (long-nosed potoroo) 13 3 23.1

Total 192 47 24.5

Table 2. Serum neutralising antibodies to macropod herpesvirus in captive macropods (Webber and Whalley 1978).

Species No. tested No. positive % positive

Macropus eugenii () (healthy) 17 5 29.4

M. eugenii (sick) 10 10 100

M. eugenii (Garden Island) 16 0 0

Setonix brachyurus () (Garden Island) 2 0 0

M. parma (parma wallaby) 13 4 30.8

M. parma x M. eugenii 2 1 50

M. robustus erubescens (euro) 4 3 75

M. robustus robustus (eastern wallaroo) 9 9 100

M. giganteus (eastern grey kangaroo) 1 0 0

M. rufus (red kangaroo) 6 5 83.3

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(M. r. erubescens x M. r. robustus) 1 1 100 x M. r. erubescens M. r. robustus x M. r. erubescens 2 1 50 M. r. erubescens x (M.r. erubescens 1 1 100 backcross hybrid) M. r. robustus x M. rufus 1 1 100

Petrogale (rock-wallaby) species 4 4 100

Total 89 45 50.6

Research

As most of the available prevalence data is based on information that was acquired over 30 years ago prior to the molecular identification and classification of macropod herpesvirus species that is currently accepted, further surveys are required, and are currently in progress, to determine the present prevalence of macropod herpesviruses in wild macropod populations. Research is also necessary to determine how infection is acquired, the relationship between infection and clinical disease and the impact of infection at a population level. Thought needs to be given to the implications and management of herpesviruses in re-introduction programs. Possible treatment options for captive macropods could also be explored. As cloacal ulcers can occur in the absence of herpesvirus infections further research is required to determine the aetiology of non- herpesvirus-associated ulcers.

Human health implications

None.

Conclusions

Herpesvirus infections appear to be extremely common in both captive and wild macropods. However, clinical disease is rare indicating that the virus is likely well adapted to its host. As with other herpesvirus infections disease generally occurs if the virus finds its way into a naïve, novel or immunocompromised host. With ongoing research, it seems likely that further macropod herpesvirus species will be discovered.

References and other information

Acland HM. Parma wallaby herpesvirus infection. Journal of Wildlife Diseases 1981:17:471-477.

Britt JO, Frost DF and Cockrill JM. Fatal herpesviral hepatitis in a red kangaroo (Macropus rufus). Journal of Zoo and Wildlife Medicine 1994:25:580-584.

Callinan RB and Kefford B. Mortalities associated with herpesvirus infection in captive macropods. Journal of Wildlife Diseases 1981:17:311-317.

Canfield PJ and Hartley WJ. A survey and review of hepatobiliary lesions in Australian macropods. Journal of Comparative Pathology 1992:107:147-167.

Dickson J, Hopkinson WI, Coackley W, Spence T and Fairfax R. Herpesvirus hepatitis in rat kangaroos. Australian Veterinary Journal 1980:56:463-464.

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Finnie EP, Littlejohns IR and Acland HM. Mortalities in parma wallabies (Macropus parma) associated with probable herpesvirus. Australian Veterinary Journal 1976:52:294.

Finnie EP. A herpesvirus. In: Montali RJ and Migaki G, editors. The Comparative Pathology of Zoo Animals. Smithsonian Institution Press, Washington, DC. 1980:179-182

Guliani S, Smith GA, Young PL, Mattick JS and Mahony TJ. Reactivation of a macropodid herpesvirus from the eastern grey kangaroo (Macropus giganteus) following corticosteroid treatment. Veterinary Microbiology 1999:68:59-69.

Jerrett I. Presumptive macropod herpesvirus infection and chronic renal disease in a . Veterinary Pathology Report 2007:73:6-7.

Johnson MA, Whalley JM, Littlejohns IR, Dickson J, Smith VW, Wilks CR and Reisner AH. Macropodid herpesvirus 1 and 2: Two herpesviruses from Australian differentiated by restrictions endonucleases, DNA composition and hybridization. Archives of Virology 1985:85:313-319.

Kerr A, Whalley JM and Poole WE. Herpesvirus neutralising antibody in grey kangaroos. Australian Veterinary Journal 1981:52:347-348.

Lachlan, NJM. and Dubovi EJ. Herpesvirales. In: Fenner's Veterinary Virology. 4th ed. Elsevier, London. 2011:179-201.

Ladds P. Viral diseases in terrestrial . In: Pathology of Australian Native Wildlife. CSIRO Publishing, Collingwood. 2009:9-24.

Roizman, B. and Pellett PE. The Family Herpesviridae: A Brief Introduction. In: Fields Veterinary Virology. Lippincott Williams & Wilkins, Philadelphia. 2001:2381-2397.

Smith G. In vitro sensitivity of macropodid herpesvirus 2 to selected anti-herpetic compounds. Journal of Wildlife Diseases 1996:31:117-120.

Smith JA, Wellehan JFX, Pogranichniy RM, Childress AL, Landolfi JA and Terio KA. Identification and isolation of a novel herpesvirus in a captive mob of eastern grey kangaroos (Macropus giganteus). Veterinary Microbiology 2008:129:236-245.

Vaughan-Higgins R, Buller N, Friend JA, Roberson I, Monaghan CL, Fenwick S and Warren K. Balanoposthitis dyspareunia, and Treponema in the critically endangered Gilbert’s potoroo (Potorous gilbertii). Journal of Wildlife Diseases 2011:47:1019-1025.

Vaz PK, Motha J, McCowan C, Ficorilli N, Whiteley PL, Wilks CR, Hartley CA, Gilkerson JR, Browning GF and Devlin JM. Isolation and characterization of a novel herpesvirus from a free-ranging eastern grey kangaroo (Macropus giganteus). Journal of Wildlife Diseases 2013:49:143-151.

Vogelnest L and Portas T. Macropods. In: Vogelnest L and Woods R, editors. Medicine of Australian Mammals. CSIRO Publishing, Collingwood. 2008:133-225.

Webber CE and Whalley JM. Widespread occurrence in Australian marsupials of neutralizing antibodies to a herpesvirus from a parma wallaby. Australian Journal of Experimental Biology and Medical Science 1978:56:351-357.

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Whalley JM and Webber CE. Characteristics of parma wallaby herpesvirus grown in marsupial cells. Journal of General Virology 1979:45:423-430.

Wilcox RS, Vaz P, Ficorilli NP, Whiteley PL, Wilks CR and Devlin JM. 2011. Gammaherpesvirus infection in a free-ranging eastern grey kangaroo (Macropus giganteus). Australian Veterinary Journal 2011:89:55-57.

Wilks CR, Kefford B and Callinan RB. Herpesvirus as a cause of fatal disease in Australian wallabies. Journal of Comparative Pathology 1981:91:461-465.

Acknowledgements

We are extremely grateful to Peter Holz and Kathryn Stalder who provided the initial draft of this fact sheet and to those individuals, agencies and organisations that provided comment and external review.

Updated: 12 February 2013

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Disclaimer

This fact sheet is managed by Wildlife Health Australia for information purposes only. Information contained in it is drawn from a variety of sources external to Wildlife Health Australia. Although reasonable care was taken in its preparation, Wildlife Health Australia does not guarantee or warrant the accuracy, reliability, completeness, or currency of the information or its usefulness in achieving any purpose. It should not be relied on in place of professional veterinary consultation. To the fullest extent permitted by law, Wildlife Health Australia 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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