Infection of naïve, free-living brushtail possums (Trichosurus vulpecula) with the nematode parasite Parastrongyloides trichosuri and its subsequent spread.

Despite the importance of spatial processes in host-parasite interactions, parasite dispersal has been the subject of few experimental studies. Introduced marsupial common brushtail possums (Trichosurus vulpecula) are a major environmental and agricultural pest in New Zealand. Parastrongyloides trichosuri, an intestinal rhabdiasoid nematode parasite specific to possums, is being evaluated as a self-disseminating delivery system for engineered fertility control vaccines. This study addressed whether an artificial infection could be established in a naïve, free-living possum population, by measuring the post-release dynamics of possum-parasite interactions at the release site, and by following the spread of the parasite into surrounding possum populations. Infection was established efficiently by applying infective larvae to the skin of possums on a single occasion. All experimentally infected possums recaptured 3 weeks after infection had parasite eggs in their faeces. Over the subsequent 2.5 years, infection spread steadily over an area of about 6000 ha. Infection persisted at the original release site for the 3.5 years of the study and at a nearby site infected by natural spread for more than 3 years. Seasonal changes in faecal egg counts were similar at the two sites. The rapid establishment of the parasite and its spread provide additional support for its ongoing development as a vaccine delivery system.

High microsatellite diversity and differential structuring among populations of the introduced common brushtail possum, Trichosurus vulpecula, in New Zealand.

An understanding of genetic variation and structure of pest populations has the potential to improve the efficiency of measures to control them. Genetic analysis was undertaken at five microsatellite loci in four native Australian and 14 introduced New Zealand populations of the common brushtail possum Trichosurus vulpecula in order to document these parameters. Genetic variation in New Zealand populations, and phylogenetic relationships among Australian and New Zealand populations, were largely predicted by the recorded introduction history. Populations on the two main islands of New Zealand had only slightly lower genetic diversity than did Australian populations, except that allelic richness on the South Is. was significantly lower. Diversity was higher in North Is. than in South Is. populations (although not significantly so) and mainland New Zealand populations as a group were significantly more diverse than offshore islands that represented secondary population size bottlenecks. In phylogenetic analyses South Is. and offshore island populations grouped with Tasmania, while North Is. populations grouped either with mainland Australia or were intermediate between the two Australian sources. This scheme was supported by admixture coefficients showing that North and South Is./offshore island populations were largely mainland Australian and Tasmanian in origin, respectively. Population structure differed markedly between the North and South Islands: populations were typically more genetically differentiated on the former than the latter, which also showed significant isolation-by-distance. Substantial linkage disequilibrium in most sampled New Zealand but no Australian population between microsatellite loci Tv16 and Tv27 suggests they may be physically linked.

Detection and isolation of Helicobacter mustelae from stoats in New Zealand.

AIM: To determine the incidence of Helicobacter mustelae in stoats (Mustela erminea) in New Zealand. METHODS: Helicobacter-like organisms and total genomic DNA were isolated from gastric tissue of stoats and identified using a combination of bacterial culture, phenotypic testing and molecular techniques. RESULTS: A Helicobacter-specific 16S rRNA polymerase chain reaction product was detected in 16/32 gastric tissue biopsies tested. Nine of 13 partially sequenced 16S rRNA DNA identified H. mustelae 16S DNA. Bacteria, subsequently identified as H. mustelae, were successfully cultured from the stomachs of 4/32 stoats. Other Helicobacter species were also identified by DNA sequence analysis, but were not cultured. CONCLUSIONS: Helicobacter mustelae is present in stoats from both the North and South Islands of New Zealand.

The relationship between prevalence of Mycobacterium bovis infection in feral ferrets and possum abundance.

AIM: To determine the relationship between the prevalence of macroscopic Mycobacterium bovis infection in feral ferrets (Mustela furo) and the abundance of brushtail possums (Trichosurus vulpecula). METHODS: The predictive power of a previously reported positive association between the prevalence of macroscopic M. bovis infection in ferrets and possum abundance was examined by undertaking surveys of M. bovis infection in ferrets at sites of low and high possum abundance. The association was then tested by a manipulative experiment that measured changes in the prevalence of macroscopic M. bovis infection in feral ferrets after reducing possum abundance. RESULTS: The positive relationship between the prevalence of macroscopic M. bovis infection in ferrets and possum abundance remained valid for new survey data, although the goodness of fit of the relationship was reduced. Experimentally reducing possum abundance reduced the odds of macroscopic M. bovis infection in ferrets by 80% in the years immediately following possum control (Odds Ratio=0.23, p=0.003). CONCLUSIONS: There is a causal link between possum abundance and the prevalence of macroscopic M. bovis infection in feral ferrets in areas in which M. bovis infection is known to occur in ferret populations. This suggests that possum-to-ferret transmission of disease occurs and accounts for most of the disease evident in ferret populations, though does not determine whether ferrets are spillover or maintenance hosts of M. bovis. Management to reduce the prevalence of M. bovis infection in ferrets should consider reducing possum abundance as a control tactic.

Genetic analysis of the mating system of the common brushtail possum (Trichosurus vulpecula) in New Zealand farmland.

We examined male reproductive success in a common brushtail possum population in New Zealand farmland. Paternity was assigned to 66 of 91 pouch young (maternity known), using a likelihood approach applied to genotypes at six microsatellite loci having an overall average exclusion probability of around 99%. The distribution of number of offspring per male was L-shaped with a standardized variance of 1.52. At least 46% of the 76 sampled reproductively mature males, bred, siring between one and four offspring each. Although breeding males were on average older and larger than nonbreeding males, the small differences did not result in a significant overall difference between the two groups in a multivariate permutation test analagous to a t-test. Paternity analysis of 22 sibling pairs (resulting from experimental removal of pouch young early in the breeding season, inducing a second oestrous) suggested that sequential mating of females with the same male was uncommon ( approximately 16-27%). Whilst there was a tendency for female possums to mate with nearby males, consistent with previous observations of territorial mating behaviour in Australian populations, some interhabitat matings were also inferred. The study population displayed only a low degree of polygyny, which may in part reflect population and habitat characteristics of the study site. A comprehensive understanding of the mating system of Trichosurus vulpecula awaits genetic paternity analysis in additional populations from both Australia and New Zealand, using quantitative approaches undertaken in this study.

Effects of sustained control of brushtail possums on levels of Mycobacterium bovis infection in cattle and brushtail possum populations from Hohotaka, New Zealand.

AIMS: To examine the effect of reducing the abundance of brushtail possums (Trichosurus vulpecula) on the distribution and prevalence of bovine tuberculosis (Mycobacterium bovis, Tb) in possums and the incidence of Tb in domestic cattle on a group of farms in the central North Island, New Zealand. METHODS: The cumulative yearly incidence of Tb infection from 12 cattle herds was estimated from annual tuberculin testing and abattoir inspection data over the period 1983-98. Intensive control of possum populations began for six of the herds in 1988, five herds in 1994 and the remaining herd in 1996. The prevalence and distribution of macroscopic M. bovis infection in possums and an index of possum abundance was estimated during yearly cross-sectional surveys from 1988 to 1998. This enabled formal testing of the link between the abundance of tuberculous possums and the incidence of Tb in cattle. RESULTS: Before possum control, infected possums were clustered in foci on or adjacent to the farms with the highest annual incidence of tuberculosis in cattle, and had an overall prevalence of macroscopic M. bovis infection of 2.3%. The prevalence of disease declined to zero with ongoing possum control, although infected possums continued to be found during the first 5 years of control. Maintaining the possum population at an average of 22.1% of its pre-control density significantly reduced the odds of the cumulative yearly incidence of Tb in cattle by 77% during the first 5 years of possum control and a further 65% in the second 5-year period. Nine of 11 tuberculous possums identified since the start of possum control were found within the areas where infected possums were clustered during the pre-control survey, suggesting that the persistence of infection within these clusters rather than infected immigrants was the source of ongoing disease. Annual estimates of the prevalence of tuberculous possums broadly followed the predictions of Barlow's possum-Tb model for a controlled possum population. CONCLUSION: The results support the hypothesis that tuberculous possums transmit bovine tuberculosis to domestic cattle, and therefore that reducing the abundance of tuberculous possums reduces the incidence of Tb in cattle. If the level of possum culling is sufficient, it appears that M. bovis infection may be eradicated from possum populations. Better information on population density, rate of increase and annual culling rates would have been needed for a truly independent examination of the Barlow possum-Tb model.

Australian and New Zealand mammal species considered to be pests or problems.

In New Zealand and Australia, 25 and 16 introduced mammals are viewed as pests, respectively, as well as a further 17 native mammals in Australia. Most introductions were deliberate and the deleterious effects became apparent later. These pests affect primary production, act as a sylvatic reservoir of disease, cause degradation of natural ecosystems, or threaten rare or endangered native animals and plants. Many species have multiple impacts. In Australia, some native mammals, particularly kangaroos and wallabies, are also controlled because of their adverse impacts on primary production. In both countries, current control depends largely on the use of poisons, shooting, the spread of disease (in the case of rabbits), trapping, habitat alteration, and commercial or recreational hunting. Methods of control by interfering with fertility (immunocontraception) are currently being investigated for rabbits, house mice, foxes, and kangaroos in Australia, and for the brushtail possum in New Zealand. If these methods prove effective, they may be applied to other mammal pests, but the need to tailor the particular approach to the ecology and behaviour of the species means that there will be a necessarily long lead time.

Internal parasites of possums (Trichosurus vulpecula) from Kawau Island, Chatham Island and Stewart Island.

As part of a search for pathogens that might be useful agents for biological control of possums, the three largest offshore islands of New Zealand that still have possums were surveyed to determine the pathogens present in these isolated populations. Brushtail possums from Kawau Island (n = 158), Chatham Island (n = 214) and Stewart Island (n = 194) were examined for internal parasites. Possums from Kawau Island were infected with Eimeria spp. (16.7%), Bertiella trichosuri (5.2%) and Purustrongyloides trichosuri (15.5%). No Paraustrostrongylus trichosuri or Trichostrongylus spp. were found. Possums from Chatham Island were infected with Eimeria spp. (10.9%). Bertiella trichosuri (3.6%), T colubriformis (6.6%), T retortaeformis (1%) and T. vitrinus (0.5%). No Parastrongyloides or Paraustrostrongylus spp. were found. Possums from Stewart Island were infected only with Eimeria spp. (4.6%). Because of their paucity of some parasites, the opportunity exists to use these offshore islands to study the introduction and spread of a parasite into a possum population, and what technology would be required to bring it to hyperendemicity.

Endoparasites of brushtail possums (Trichosurus vulpecula) from the South Island, New Zealand.

As part of a study to assess whether endoparasites could assist in the biological control of brushtail possums in New Zealand, we investigated the composition and distribution of possum endoparasites in the South Island. Possums were collected near five of the original release areas in the South Island : Banks Peninsula, Hokitika, Nelson, Dunedin and Invercargill. Among the nematodes, those most frequently encountered were Trichostrongylus spp., which were present in possums from all five study areas. Trichostrongylus species from possums in the Invercargill area comprised 4.5% T. colubriformis, 0.9% T. vitrinus and 11.3% T. retortaeformis. Paraustrostnmgylus trichosuri and Parastrongyloides trichosuri were found only in the Invercargill area, where they infected 1.4% and 14% of possums respectively. The cestode Bertiella trichosuri was present in possums from all locations except Dunedin. The protozoan Eimeria spp. occurred in all areas. These are the first records of Parastrongyloides trichosuri, Paraustrostrongylus trichosuri, T. vitrinus, T. retortaeformis and Eimeria spp. in South Island possums. The prevalence of endoparasites and the intensity of infection was very low compared to the lower North Island of New Zealand. Endoparasites at the existing levels in the South Island probably have very little effect on possum populations.

Possum biocontrol: prospects for fertility regulation.

Research has begun recently into biocontrol of brushtail possums as the only long-term, cost-effective solution to the possum problem in New Zealand, where possums cause significant damage to native forests, threaten populations of native plants and animals, and infect cattle and deer with bovine tuberculosis. Fertility regulation as a means of biocontrol has the support of major animal welfare and conservation groups in New Zealand. Systems are being investigated, mostly in reproduction and development, with the ultimate aim of developing immunologically-based fertility regulation (immunocontraception), but much basic information essential to such an approach for possums is lacking. The key components for the success of this approach--suitable vectors expressing possum-specific reproductive antigens sufficiently to block reproduction--are reviewed. The social and political issues of local, national and international risk and acceptability arising from such an approach are also discussed.

Foraging and denning patterns of brushtail possums, and their possible relationship to contact with cattle and the transmission of bovine tuberculosis.

Radio-tracking and direct observation were used over 18 months in 1990-92 to investigate both the use of sleeping dens and foraging activity by possums (Trichosurus vulpecula) on a 21 ha site in the Wairarapa used for a longitudinal study of bovine tuberculosis. Males had larger home ranges than females, and both sexes had larger activity areas during the autumn mating season than at other times of the year. Possums typically foraged in only a small area of their home ranges (termed an activity area) on any one night, and the areas used by individuals were commonly very similar over a series of nights. Activity areas overlapped extensively among possums. Possums used a limited number of dens, commonly in a small and in most cases a circumscribed part of their home range. No simultaneous den-sharing was found, with the exception of mother-joey pairs. The mortality of juveniles after independence was 36%. Only two of 25 juveniles under surveillance to detect dispersal dispersed more than 500 m off the study site, and both subsequently died. Grazing patterns of cattle meant that almost all accessible areas of the paddock were covered by at least some grazing cattle, and so all activity areas of possums within the paddock were covered by areas where cattle foraged. However, possums avoided contact with cattle, and when some cattle were excluded from access to the part of the paddock principally used by both tuberculous and healthy possums for denning, transmission of Mycobacterium bovis from possums to these cattle ceased, although there was subsequent transmission to deer. Cattle which grazed the area used principally for possum denning continued to become infected, and these denning areas appeared to be of importance in the transmission of tuberculosis.

Whey proteins of the common brushtail possum (Trichosurus vulpecula): isolation, characterization and changes in concentration in milk during lactation of transferrin, alpha-lactalbumin and serum albumin.

1. Transferrin and serum albumin were purified from both whey and serum and alpha-lactalbumin was purified from whey from the common brushtail possum (Trichosurus vulpecula). 2. The N-terminal amino acid sequences for transferrin and serum albumin were identical for the proteins from both whey and serum and showed homologies with transferrin or serum albumin from other species. 3. N- and C-terminal regions of possum alpha-lactalbumin were also sequenced and have been compared with wallaby alpha-lactalbumin and several eutherian alpha-lactalbumins. 4. Antisera raised to each of the three proteins were species specific and Western blots further confirmed the identity of the serum and whey transferrins and serum and whey serum albumins. 5. The concentration of transferrin increased ten-fold between days 110 and 130 of lactation, whereas no significant changes in the concentration of alpha-lactalbumin could be detected after day 60.

Changes in milk composition during lactation in the common brushtail possum, Trichosurus vulpecula (Marsupialia: Phalangeridae).

The milk constituents of Trichosurus vulpecula, a folivorous marsupial, showed marked quantitative and qualitative changes during the course of lactation. The milk produced in the early stages of lactation was dilute, with about 9-13% (w/w) solids during the first 3 weeks, comprising mostly carbohydrate and protein (35-40%). At 20 weeks, about three-quarters of the way through lactation, the milk was much more concentrated, about 28% solids, with lipid the predominant fraction (30-35%), after a marked decline in carbohydrate content (20-25%). Concentrations of the electrolytes sodium and potassium also underwent marked changes. The changes in milk composition of T. vulpecula during the first three-quarters of lactation were similar to those described for a range of herbivorous, insectivorous and carnivorous marsupials. In the last quarter of lactation, however, brushtail possum milk maintained a relatively stable composition, with higher levels of carbohydrate and lower levels of lipid than for other marsupials. There appears to be a uniform pattern of changes in milk composition throughout the Marsupialia over most of lactation, with family differences evident only in the latter stages.

Intestinal lactase (beta-galactosidase) and other disaccharidase activities of suckling and adult common brushtail possums, Trichosurus vulpecula (Marsupialia:Phalangeridae).

Small-intestinal disaccharidase activities of eight suckling T. vulpecula, aged from 34 to 150 days, and of two adult animals were investigated. Intestinal maltase, isomaltase and sucrase activities increased with age, whereas lactase activities decreased. Trehalase activities were relatively high in all animals and showed no obvious age-related changes. Three separate beta-galactosidase activities, one neutral and two acid, acted on lactose. The neutral beta-galactosidase activity appeared to be due to a brush border enzyme similar to that of eutherian mammals, whereas the acid beta-galactosidases were soluble and probably of lysosomal origin. One of these, acid beta-galactosidase-1, had similar properties to the sole intestinal beta-galactosidase of macropodid marsupials, whereas the other, acid beta-galactosidase-2, has not previously been described. Galactosyl oligosaccharides isolated from macropodid milk were readily hydrolysed by both acid beta-galactosidases but not by the neutral beta-galactosidase. The total intestinal lactase activity in animals aged up to 125 days was due mainly to acid beta-galactosidase-1, whereas in older animals it was due mostly to the neutral beta-galactosidase; this suggests that late in lactation the young T. vulpecula change from a macropodid mode of digestion of galactosyl oligosaccharides to a eutherian mechanism for the digestion of lactose. These findings may have implications for the hand-rearing of orphaned T. vulpecula.

Changes in milk carbohydrates during lactation in the common brushtail possum, Trichosurus vulpecula (Marsupialia:Phalangeridae).

Milk samples (186) were obtained at various stages of lactation from 27 common brushtail possums (Trichosurus vulpecula). Qualitative and quantitative changes in the milk carbohydrates during early and mid-lactation were similar to those previously seen in other marsupials; the principal carbohydrate was lactose early in lactation and higher oligosaccharides in mid-lactation, and the hexose concentration reached a peak during mid-lactation. However, the late-lactation milk was unusual in that the carbohydrate was mainly lactose and its concentration remained relatively high (3.5 to 5.5%). In contrast to earlier findings on the milk of the tammar wallaby (Macropus eugenii), little or no nucleotide pyrophosphatase, beta-galactosidase and alkaline phosphatase activities were detected late in lactation.