Experimental infections of brush-tailed possums, common wombats and water rats with Leptospira interrogans serovars balcanica and hardjo

Of 12 brush-tailed possums (Trichosurus vulpecula) inoculated with Leptospira interrogans serovar balcanica 11 developed migroagglutination (MA) antibody to jardjo antigen by 14 days postincubation (PI). Leptospiruria was observed in 2 possums 117 to 145 days PI. Of 6 possums inoculated with serovar hardjo 4 developed low short-lived titres by day 18 PI. Two of 3 wombats (Vombatus ursinus) inoculated with balcanica had high MA titres (greater than or equal to 1:128) by day 16 PI and leptospiruria occurred by day 16. One wombat inoculated with hardjo developed a low MA titre. Low transitory MA titres to hardjo were found in 1 of 3 water rats (Hydromys chrysogaster) after inoculation with balcanica and 1 of 2 given hardjo. Histopathological examination of kidneys revealed mild to moderately severe focal interstitial nephritis in 4 of 8 possums, in 2 wombats and in 2 water rats following experimental infection with balcanica. Similar lesions were observed in 2 of 4 possums, 1 wombat and 2 water rats following experimental infection with hardjo.     

A serological survey of Australian wildlife for antibodies to Leptospires of the Hebdomadis serogroup.

A serological survey for antibodies to Leptospira interrograns serovar hardjo was conducted on 574 serum samples from 10 native and 4 introduced wildlife species in south-eastern Australia. The microscopic agglutination (MA) test was used, and titres to hardjo antigen were detected in 33.5% of 352 brushtailed possums (Trichosurus vulpecula) sampled in several areas of Victoria. Prevalence of reactors ranged from 14 to 66% in 4 populations examined intensively. Serovar balcanica was isolated from possums with hardjo antibodies from two different areas. Of 20 wombats Vombatus ursinus) examined in Victoria, antibodies to hardjo were found in sera from 4 and titres to Pyrogenes and Pomona serogroups were detected in another. Hardjo antibodies were demonstrated in sera from 13 of 19 rusa deer (Cervus timorensis). Negative MA test results to hardjo antigens were recorded in 55 mountain possums (T. caninus), 63 macropods (Macropus spp.), 17 water rats (Hydrmys chrysogaster), 39 fallow deer (Dama dama), 2 hog deer (Axis porcinus) and 2 water buffalo (Bubalus bubalus). No MA antibodies to any of 16 leptospiral serogroups were detected in 17 water rats tested. Kidneys were examined from 330 of these animals and focal interstitial nephritis suggestive of leptospirosis was found in kidneys of 63 of 169 T. vulpecula, 3 of 55 T. caninus, 12 of 18 V. ursinus, 6 of 22 Macropus spp., 9 of 16 H. chrysogaster, 5 of 11 C. timorensis and 3 of 39 D. dama. A statistical association between focal interstitial nephritis and MA antibodies to hardjo was found in T. vulpecula.     

Experimental infection of calves and sheep with Leptospira interrogans serovar balcanica

Two of four calves inoculated with Leptospira interrogans serovar balcanica developed low microscopic agglutinating (MA) titres to serovar hardjo. A third calf had an MA titre of 1:1024 by day 19 post-inoculation (PI). Transient leptospiruria was recorded in one calf on days 12 and 13 PI. An in-contact calf did not seroconvert. None of the calves had fever or other clinical signs of disease. Four ewes inoculated with balcanica developed MA titres to hardjo by day 13 PI, and a transient leptospiruria between days 14 and 25 PI. None of the ewes showed any evidence of clinical disease and three of them delivered healthy lambs 22 to 64 days PI. One ewe had mild lesions of focal interstitial nephritis.     

Mucosal immunity in the brushtail possum (Trichosurus vulpecula): detection of antibody in serum and at female reproductive sites after intranasal immunization

Vaccination strategies for the brushtail possum, which rely upon stimulation of mucosal immunity, are being developed for biocontrol purposes. As little is known about how to stimulate possum immune responses via a mucosal site, groups of possums were immunized intranasally with keyhole limpet haemocyanin (KLH) alone or in combination with known or novel mucosal adjuvants. Antigen-specific antibody titres in female reproductive secretions were measured by ELISA and compared with antibody titres in the serum. Antigen-induced lymphocyte proliferative responses were measured as an indicator of cell-mediated responses. Intranasal immunization with KLH alone stimulated a weak serum antibody response that was significantly increased when KLH was given with cholera toxin subunit B (CTB), recombinant possum tumour necrosis factor alpha (TNF alpha) or live Mycobacterium bovis bacillus Calmette Guerin (BCG). Antibody titres in secretions from ovarian follicles and the uterus were very low in animals administered KLH alone. Significantly higher antibody titres to KLH were present in the reproductive secretions of possums immunized with KLH plus CTB, BCG or heat-killed Mycobacterium vaccae. Antibody titres were lower in mucosal secretions than in the serum, but there was a significant correlation between the two. In addition, coadministration of live BCG with KLH produced a strong antigen-specific cell-mediated response to KLH. This study has shown that an immune response to a protein antigen can be stimulated in possums by intranasal immunization and that antigen-specific antibodies can be detected in secretions from the female reproductive tract.     

Absence of Ross River virus amongst Common brushtail possums (<Emphasis Type="Italic">Trichosurus vulpecula</Emphasis>) from metropolitan Sydney,Australia

Ross River virus (RRV) is a mosquito-transmitted Alphavirus emerging in urban centres throughout Australia. The Common brushtail possum (Trichosurus vulpecula), a native marsupial that has successfully adapted to human settlement, has been implicated as a maintenance reservoir for RRV. In the present study, RRV exposure was assessed amongst 72 urban-adapted possums from Northern Sydney and ten possums from a woodland area, remote from urbanisation. Serological screening was performed using an enzyme-linked immunosorbent assay to detect RRV antibodies in possum sera. Findings indicated that both possum populations from urban and woodland habitats were negative for the presence of RRV antibodies. Lack of exposure to RRV highlights that the host status of possums is contingent upon factors other than their abundance and proximity to human settlement. In view of the potential for climate change to favour transmission of mosquito-borne disease in Australia, identification of wildlife populations entirely absent of RRV may prove useful for monitoring the predicted spread of the virus.     

Spatial variation in impacts of brushtail possums on two Loranthaceous mistletoe species

Browsing by introduced brushtail possums is linked to major declines in mistletoe abundance in New Zealand, yet in some areas mistletoes persist, apparently unaffected by the presence of possums. To determine the cause of this spatial variation in impact I investigated the abundance and condition (crown dieback and extent of possum browse) of two mistletoes (Alepis flavida, Peraxilla tetrapetala) and abundance and diet of possums in two mountain beech (Nothofagus solandri var. cliffortioides) forests in the central-eastern South Island of New Zealand. Mistletoe is common and there are long-established uncontrolled possum populations in both forests. Mistletoes were abundant (216?1359 per hectare) and important in possum diet (41?59% of total diet), but possum density was low (c. 2 per hectare) in both areas. Possum impacts were slight with low browse frequencies and intensities over much of the study sites. However, impacts were significantly greater at a forest margin, where possum abundance was highest, and at a high-altitude site where mistletoe density was lowest. Mistletoe crown dieback was inversely proportional to intensity of possum browsing. These results suggest that the persistence of abundant mistletoe populations at these sites is due to mistletoe productivity matching or exceeding consumption by possums in these forests of low possum-carrying capacity, rather than low possum preference for the local mistletoe populations.     

Leptospirosis serology in the common brushtail possum (Trichosurus vulpecula) from urban Sydney, Australia

The common brushtail possum (Trichosurus vulpecula) is indeed a common marsupial in major cities of Australia. This species is known to be susceptible to leptospirosis and often lives in close contact with humans, raising concerns about the potential for transmission of this disease in urban areas. A total of 192 brushtail possum blood samples were collected from 136 individuals in suburban areas of metropolitan Sydney from November 2002 to November 2004. Sera were screened against a reference panel of 21 Leptospira spp. using the microscopic agglutination test. Leptospiral antibodies were detected in 9.6% (13/136) of tested brushtail possums and represented two serovars; antibodies to Leptospira interrogans serovar Hardjo were most frequently identified (11/136). A representative of the exotic sero-group Ballum, most likely serovar Arborea, was found in two of 136 brushtail possums. Exposure to leptospirosis seemed to be associated with age, as older animals had a higher incidence, but there was no distinction in relation to gender. Antibody prevalence varied between the different sampling sites and seropositive animals were clustered and restricted to a few sites. These data support the possible role of brushtail possums as a maintenance host for Leptospira spp. in urban environments and also identified them as a previously unknown and potential source of serovar Arborea.     

Heterogeneity in vertebrate and invertebrate herbivory and its consequences for New Zealand mistletoes

Vertebrate herbivores generally have greater effects than invertebrates on plants. However, few studies have investigated the effects of both invertebrate and vertebrate herbivores on a single plant species. In New Zealand, nationwide declines in mistletoe populations have often been attributed to possum herbivory, but never to insect herbivory. The main goal of the present study was to document levels of vertebrate and invertebrate herbivory on endemic New Zealand mistletoe plants to suggest whether herbivory is leading to mistletoe decline. In the present study, the annual amount of leaf loss from herbivory by the brushtail possum (Trichosurus vulpecula), insect herbivory and leaf abscission were measured in two populations each of three mistletoe species (Alepis flavida, Peraxilla colensoi, and Peraxilla tetrapetala, Loranthaceae). In two populations of each species from February 1997 to February 1998, abscission accounted for the most leaf loss (range 10–84% of total mean leaf area, mean 33%), whereas insects and possums usually removed small and similar amounts (less than 3%). Possum browse caused large amounts of abscission in only one population (A. flavida at Eglinton). Observed possum browse was more heterogeneous than insect browse among branches within a plant (possum coefficient of variation = 2.63, insect CV = 1.98, P < 0.001), among plants in a population (possum CV = 2.15, insect CV = 0.69, P < 0.001), and between populations (possum CV = 1.36, insect CV = 1.09). Moreover, insects damaged 100% of the study plants but never removed more than 16% of leaf area on a single plant, whereas possums only browsed 32% of the study plants but severely defoliated some plants. Thus, while the mean amount of biomass removed across a population may have important consequences for mistletoe survival, the effect of possums on mistletoe populations may also depend on the heterogeneity of browse among individuals in the population.     

Serologic survey for Toxoplasma gondii and Neospora caninum in the common brushtail possum (Trichosurus vulpecula) from urban Sydney, Australia

The common brushtail possum (Trichosurus vulpecula) has well adapted to increasing urbanization, resulting in greater interaction with humans and their domestic pets. Wildlife species in urban areas face a higher risk of exposure to zoonotic pathogens and may be affected by parasites hosted by cats (Toxoplasma gondii) or dogs (Neospora caninum), yet it is unknown to what extent urban T. vulpecula are exposed to these parasites. Antibodies to T. gondii and N. caninum were assayed in sera of 142 adult possums from the city of Sydney, Australia. Using the modified agglutination test, antibodies to T. gondii were found in 9 (6.3%) of the 142 animals in titers of 1:25 (4), 1:50 (1), 1:100 (1), 1:800 (1), 1:3,200 (1), 1:6,400 (1), and 1:12,800 (1). Of some T. vulpecula multiple sera samples within a 2-yr frame could be collected, but seropositive animals in general were not recaptured after initial seroconversion. One possum had a high T. gondii titer on 2 consecutive bleedings, 14 mo apart, and seropositive possums appeared normal when captured. Sex seemed not to have an affect on antibody prevalence, but age and location may play a role. Antibodies to N. caninum were not detected in 1:25 dilution of sera in the N. caninum agglutination test, indicating that T. vulpecula may not have been exposed to this parasite. This is the first serological survey for T. gondii and N. caninum infections in urban T. vulpecula.     

Factors influencing brushtail possum (

Damage by introduced brushtail possums (Trichosurus vulpecula) to Pinus radiata trees was assessed in 41 compartments of a commercial forestry plantation on the Coromandel Peninsula, New Zealand. All the trees assessed were less than 3 years old. Possum damage in the compartments was low (median prevalence 3.3%) but highly variable (range 0?30%). Eight of 37 measured habitat factors differed significantly (P < 0.05) between the sites with damaged and undamaged trees. The best predictor of mean damage was stand age, but this explained only 21% of the variation in damage among compartments. Including both stand age and New Zealand bracken (Pteridium esculentum) cover improved the model significantly and explained 36% of variation in damage. Damage was apparently unrelated to compartment size, distance from the compartment boundary, and possum den-site availability. Surprisingly, the relationship between browse damage and a trap-catch index of possum abundance was weakly negative (r_S = −0.53, P = 0.05). The dense understorey associated with young pine stands tends to increase possum damage to associated P. radiata trees, but the possums in such stands may be less mobile at ground level and thus less easily trapped. Assessment of stand age and understorey characteristics, together with visual inspection for early signs of damage, is likely to be more cost-effective than possum surveys for identifying forest compartments at risk from possum browse.     

Mapping of B cell epitopes on the zona pellucida 2 protein of a marsupial, the brushtail possum (Trichosurus vulpecula)

Immunocontraceptive vaccines against zona pellucida (ZP) proteins are being developed for brushtail possum (Trichosurus vulpecula) management in New Zealand. Mapping of B cell epitopes on the ZP2 protein of possums was undertaken in this study to define the antigenic regions that may be crucial to sperm-egg binding. The amino acid sequence of the full-length possum ZP2 protein (712 amino acids) was used to synthesize a complete set of 71 (15-mer) biotinylated peptides with an offset of five amino acids. The peptides were used in a modified enzyme-linked immunosorbent assay (ELISA) to identify continuous epitopes recognized by antibodies in the sera of possums immunized with recombinant possum ZP2 (rZP2) constructs. Seventeen continuous epitopes were located on possum ZP2 protein. Comparisons of the peptide binding pattern of antibodies in individual sera with the fertility status of the same immunized possums revealed three significant infertility-relevant peptide epitopes (amino acids 111-125, 301-315, and 431-445). One of these (amino acids 431-445) bound to possum spermatozoa from the caudal epididymis. The implications of these findings for developing immunocontraceptive vaccines for possum control are discussed.     

Diet and impacts of brushtail possum populations across an invasion front in South Westland, New Zealand

Impact of the irruptive fluctuation in abundance of brushtail possum populations since their initial colonisation was investigated in the forests of South Westland, New Zealand. Possum abundance, fecundity, and diet, the condition of common possum-palatable tree species, and the abundance of common forest birds were measured at three sites occupied by possums for c. 10, 20, and 30 years. Possum densities were highest at the site where possums had been present for c. 20 years. Possum fecundity was higher at the site they had recently colonised than at the two sites where they had been present for c. 20–30 years. Diet of possums where they had recently colonised was dominated by highly preferred foods (72% of total diet), whereas these same foods contributed just 36% of total diet at the site where possums had been present for more than two decades. Canopy condition of common possum-preferred trees was scored progressively lower in areas with increasing length of possum occupation, especially at the site where the possum population had apparently declined from its maximum density. Native forest bird abundance also declined with increasing length of possum occupation. The implications of these results for management are discussed.     

Assessing Movements of Brushtail Possums (Trichosurus vulpecula) in Relation to Depopulated Buffer Zones for the Management of Wildlife Tuberculosis in New Zealand

In New Zealand, managing the threat of bovine tuberculosis (TB) to livestock includes population reduction of potentially infectious wildlife, primarily the brushtail possum (Trichosurus vulpecula). Population control is often targeted on forested buffer zones adjacent to farmland, in order to limit movements of possums across the buffer and reduce the risk of disease transmission to livestock. To assess the effectiveness of buffers in protecting livestock we analysed GPS telemetry data from possums located in untreated forest adjacent to buffers, and used these data to characterise patterns of movement that could lead to possums reaching farmland during the season when most dispersal occurs. Analyses of movement data showed that the direction of dispersal by sub-adult and adult possums and the extent of long exploratory movements were not biased toward forest buffers, even though these provided vacant habitat as suitable for possums as untreated forest. Instead, dispersal and exploratory movements were uncommon even for sub-adult possums and such events typically lasted <10 days. Dispersing possums settled predominantly in river valleys. A simulation model was developed for the 3-6-month dispersal season; it demonstrated a probability of <0.001 that an infected possum, originating from a low-density population with low disease prevalence in untreated forest, would move across 3 km of recently controlled forest buffer to reach farmland. Our results indicate short-term reduction in the risk of TB transmission from possums to livestock in New Zealand by the use of depopulated buffer zones, while acknowledging that the threat of disease spread from untreated forest is likely to increase over time as possum population density and, potentially, TB prevalence among those possums, increase in the buffer zone.     

Differences in steroid metabolism by testis, prostate and epididymis in the immature and adult possum (Trichosurus vulpecula)

Immature possums 126-195 days old and adults over 1 year old were used. Testicular homogenates from immature possums converted [3H]progesterone to nine different products, of which greater than 63% were 5 alpha-reduced androstane metabolites. The major product from adult testis was testosterone in yields greater than 60%. While metabolism of [3H]testosterone by the epididymis of immature possum was minimal, in adults 5 alpha-reduced products constituted greater than 80% of the yield. In contrast, prostatic tissue from adults converted less than 4% of [3H]testosterone to 5 alpha-reduced products, while the yields were greater than 80% from prostates of immature animals. The results showed that like in rodents, the testis of immature possum has a high 5 alpha-reductase and low 17 beta-hydroxysteroid dehydrogenase activity, which reverses in the adult state. The implications of the findings are discussed.     

Combining Aspirin with Cholecalciferol (Vitamin D3) – A Potential New Tool for Controlling Possum Populations

The introduced Australian brushtail possum is a major vertebrate pest in New Zealand, with impacts on conservation and agriculture being managed largely through poisoning operations. Cholecalciferol (vitamin D₃) is registered for use in controlling possums and despite its many advantages it is expensive and relatively inhumane. Combination of a high proportion of aspirin with a low proportion of cholecalciferol was effective in killing high proportions of groups of acclimatised, caged possums: this is attributed to both an unexpectedly high toxicity of the type of cholecalciferol used, and a proposed synergistic mechanism between the two compounds. Death was caused by localised damage to heart ventricles by aspirin, and inhibition of tissue repair by both aspirin and cholecalciferol. The observed toxicosis had lower impact on the welfare of possums than either compound administered alone, particularly aspirin alone. Residue analyses of bait remains in the GI tract suggested a low risk of secondary poisoning by either compound. The combination of cholecalciferol and aspirin has the potential to meet key requirements of cost-effectiveness and humaneness in controlling possum populations, but the effect of the combination in non-target species has yet to be tested.     

Comparison of antibodies to Leptospira in white-tailed deer (Odocoileus virginianus) and cattle in Ohio

A survey was conducted to determine the prevalence of leptospiral antibodies in sera from 248 white-tailed deer (Odocoileus virginianus) in Ohio. The sera were collected at check stations during the hunting season in 1983. The microscopic agglutination microtiter test was used to determine the presence of antibodies to Leptospira interrogans serovars pomona, icterohemorrhagiae, canicola, hardjo, and grippotyphosa. Eighteen of 248 (7.3%) serum samples had antibody titers (greater than or equal to 1:100) to at least one of the five serovars tested, with three of these samples reacting to more than one serovar. Prevalence did not differ significantly between sex or age groups. The serovar antigens reacting most frequently with serum antibodies were grippotyphosa (10 of 22, 45.5%) and pomona (eight of 22, 36.4%). Sera agglutinating with pomona antigen had higher titers (ranging from 1:200 to 1:6,400) than did sera agglutinating with the other serovars. These results were compared to results obtained from cattle tested at the Ohio Department of Agriculture Laboratories during 1983. There was a significant relationship between pomona infections detected in deer and cattle (P less than 0.05), but not with grippotyphosa.     

Serologic evidence of leptospirosis in woodchucks (Marmota monax) in central New York State.

Serum samples from 153 woodchucks (Marmota monax) from Tompkins County, New York, obtained in 1976 and 1977, were examined by plate agglutination tests for antibodies against five Leptospira antigens. Fourteen sera showed significant titers against either L. hardjo, L. icterohemorrhagiae and/or L. pomona. Reactions against L. hardjo were the most frequent. Woodchucks collected from two dairy farms with histories of bovine leptospirosis did not have a greater prevalence of antibodies than woodchucks collected from other locations. Each of two woodchucks experimentally-inoculated with L. hardjo developed titers to L. hardjo. Maximum titers occurred approximately 30 days post-inoculation. L. hardjo was not observed in urine specimens of these animals.     

Prevalence and Genetic Characterization of Cryptosporidium Isolates from Common Brushtail Possums (Trichosurus vulpecula) Adapted to Urban Settings

The common brushtail possum (Trichosurus vulpecula) is one of the most abundant native marsupials in urban Australia, having successfully adapted to utilize anthropogenic resources. The habituation of possums to food and shelter available in human settlements has facilitated interaction with people, pets, and zoo animals, increasing the potential for transmission of zoonotic Cryptosporidium pathogens. This study sought to examine the identity and prevalence of Cryptosporidium species occurring in possums adapted to urban settings compared to possums inhabiting remote woodlands far from urban areas and to characterize the health of the host in response to oocyst shedding. Findings indicated that both populations were shedding oocysts of the same genotype (brushtail possum 1 [BTP1]) that were genetically and morphologically distinct from zoonotic species and genotypes and most closely related to Cryptosporidium species from marsupials. The urban population was shedding an additional five Cryptosporidium isolates that were genetically distinct from BTP1 and formed a sister clade with Cryptosporidium parvum and Cryptosporidium hominis. Possums that were shedding oocysts showed no evidence of pathogenic changes, including elevated levels of white blood cells, diminished body condition (body mass divided by skeletal body length), or reduced nutritional state, suggesting a stable host-parasite relationship typical of Cryptosporidium species that are adapted to the host. Overall, Cryptosporidium occurred with a higher prevalence in possums from urban habitat (11.3%) than in possums from woodland habitat (5.6%); however, the host-specific nature of the genotypes may limit spillover infection in the urban setting. This study determined that the coexistence of possums with sympatric populations of humans, pets, and zoo animals in the urban Australian environment is unlikely to present a threat to public health safety.     

ELISA for the detection of specific IgM and IgG in human leptospirosis

ELISA was used to detect specific IgM and IgG in sera from humans with current or past leptospirosis. A serological pattern of a high IgM titre (greater than or equal to 1280), or moderately increased IgM (160-640) in conjunction with a low IgG titre (less than or equal to 20), with serovar copenhageni antigen was characteristic for approximately two-thirds of the sera from serovar icterohaemorrhagiae patients obtained in the first two months of the disease. The antigen was the supernatant of a heated and centrifuged culture of leptospires. Antigens were prepared from serovars copenhageni, grippotyphosa, hardjo and patoc. Sera from patients with icterohaemorrhagiae, grippotyphosa and hardjo infections showed cross-reactivity when different antigens were used. In past infections the IgG titres were clearly higher with the homologous antigen. ELISA for IgM and IgG allows the rapid diagnosis of acute leptospirosis.     

Bovine tuberculosis in brushtail possums: models, dogma and data

Three different models of bovine tuberculosis (Tb) in brushtail possums were evaluated against their stated purpose, and testable assumptions and predictions evaluated against available data where possible. Not surprisingly, two of the models may be falsified based on currently available data with respect to either important model assumptions or predictions, and the third may suffer from being right for the wrong reason. This does not mean that these models are not useful. To the contrary, I argue that all models, especially those published in the scientific literature have largely addressed their stated purpose, and have contributed to our understanding of and ability to manage bovine tuberculosis infection in brushtail possum populations. No model, however, satisfactorily explains the pronounced spatial clustering of possum Tb, and the models critiqued have provided little strong inference as to the routes of transmission of Tb among possums. This situation is not helped by the scarcity of datasets on Tb in uncontrolled possum populations that are readily available to confront competing possum/Tb models with. As time passes, there is a very real risk that these data sets will be lost. This is of particular concern, as the expansion in the area of New Zealand under active possum management means the future opportunity to collect further data on Tb in uncontrolled possum populations is severely limited.