Dissertationen zum Thema „Tammar wallaby“

Marsupials and eutherians are the two principal groups of modern mammals. Mammalian immunological studies, to date, have focused on eutherian systems with little or no comprehensive work having been carried out on marsupials. This project investigates the functional and developmental aspects of T-cell responses in the marsupial, Macropus eugenii (Tammar wallaby) in both adults and pouch young at various stages of development. Determination of the age at which the Tammar wallaby immune system becomes competent has been examined through the use of cellular and molecular studies carried out on developing pouch young tissue. The capacity for generating an immunological response in adult and pouch young marsupials has been studied by following cellular proliferation in response to mitogens or mixed lymphocyte culture (MLC). After examining adult responses to mitogens and allogenic lymphocytes, optimised conditions were then used to examine the development of responsiveness in pouch young. Several further tests were conducted and findings shown. The study has shown that the earliest age at which Macropus eugenii is capable of mounting a T-cell mediated immune response is between 5 to 13 days post-partum
Doctor of Philosophy (PhD)

Marsupials and eutherians are the two principal groups of modern mammals. Mammalian immunological studies, to date, have focused on eutherian systems with little or no comprehensive work having been carried out on marsupials. This project investigates the functional and developmental aspects of T-cell responses in the marsupial, Macropus eugenii (Tammar wallaby) in both adults and pouch young at various stages of development. Determination of the age at which the Tammar wallaby immune system becomes competent has been examined through the use of cellular and molecular studies carried out on developing pouch young tissue. The capacity for generating an immunological response in adult and pouch young marsupials has been studied by following cellular proliferation in response to mitogens or mixed lymphocyte culture (MLC). After examining adult responses to mitogens and allogenic lymphocytes, optimised conditions were then used to examine the development of responsiveness in pouch young. Several further tests were conducted and findings shown. The study has shown that the earliest age at which Macropus eugenii is capable of mounting a T-cell mediated immune response is between 5 to 13 days post-partum

[Truncated abstract] Understanding the effect that the disturbance of habitat by humans has on the population dynamics and ecology of wild animals is critical for the management of these populations. By understanding the demographic effects of disturbance the ways in which a population can be managed to increase or decrease its rate of change in size also become apparent. This thesis describes the effect that human disturbance, through the establishment of a large naval base, has had on the population dynamics and ecology of tammar wallabies (Macropus eugenii) on Garden Island, Western Australia. The disturbance of the environment on the HMAS Stirling Naval Base included the establishment of large areas of irrigated and fertilised couch grass (Cynodon dactylon) that increased and made virtually constant the amount of food available to the tammars in that area. In addition, traffic associated with the naval base resulted in large numbers of tammar wallabies being killed by vehicles. The effects of these disturbances were determined by comparing population dynamics, through vital rates of survival and fecundity and population growth rates, and spatial ecology, through the size of the animals' home ranges, in three areas of Garden Island. The three areas were the naval base (highly disturbed), southern bushland (adjacent to the naval base) and the northern bushland (undisturbed). The tammars on the naval base were in better body condition than those living in the two bushland areas of the island. ... When the impact of road-kills was removed, increased to 1.150.101 per year on the naval base and 0.960.076 per year in the southern bushland. Fecundity transitions, defined as the product of the rates of birth and pouch-young survival, and adult survival rates were lower in the bushland areas compared with the naval base in two of the three years, which were the main reasons for the lower estimates. There were no significant differences in the size of the tammars' home ranges between areas with modified or unmodified habitats or between the sexes (P>0.05). In summer the mean size of the home ranges was 3.90.66 ha, which was larger than winter when home ranges were 3.20.54 ha, but this difference failed to reach significance (P=0.058). These results indicate that the modification of the tammars' habitat has probably not caused significant changes in the size of the animals' home ranges. The size of the home ranges of tammar wallabies is likely to be determined by a complex interaction of many factors, and habitat modification alone has not been sufficient to cause substantial changes. The results presented in this thesis demonstrate that the disturbance caused by the establishment of the naval base on Garden Island has altered the population dynamics of the tammars wallabies, through increasing in the amount of food available to the tammars and through high numbers of road-kills. These results also demonstrate how gaining detailed knowledge of population dynamics can have direct application to managing the impact of disturbance on populations of wild animals.

Microsatellites are simple repetitive DNA sequences that are used as genetic markers throughout the biological sciences. The high levels of variation observed at microsatellite loci contribute to their utility in studies at the population and individual levels. This variation is a consequence of mutations that change the length of microsatellite repeat tracts. Current understanding suggests that most mutations are caused by polymerase slippage during DNA replication and lead to changes of a single repeat unit in length, but some changes involving multiple repeats can also occur. Despite this simplistic overview, there is evidence for considerable heterogeneity in mutation processes between species, loci and alleles. Such complex patterns suggest that other mechanisms, including those associated with DNA recombination, are also involved in the generation of microsatellite mutations. Understanding which mutational mechanisms are responsible for variation at microsatellite markers is essential to enable accurate data interpretation in genotyping projects, as many commonly used statistics assume specific mutation models. I developed microsatellite markers specific to the X and Y chromosomes and an autosome in the tammar wallaby, Macropus eugenii, and investigated their evolutionary properties using two approaches: indirectly, as inferred from population data, and directly, from observation of mutation events. First, I found that allelic richness increased with repeat length and that two popular mutation models, the stepwise mutation model and the infinite allele model, were poor at predicting the number of alleles per locus, particularly when gene diversity was high. These results suggest that neither model can account for all mutations at tammar wallaby microsatellites and hint at the involvement of more complex mechanisms than replication slippage. I also determined levels of variation at each locus in two tammar wallaby populations. I found that allelic richness was highest for chromosome 2, intermediate for the X chromosome and lowest for the Y chromosome in both populations. Thus, allelic richness varied between chromosomes in the manner predicted by their relative exposure to recombination, although these results may also be explained by the relative effective population sizes of the chromosomes studied. Second, I used small-pool PCR from sperm DNA to observe de novo mutation events at three of the most polymorphic autosomal markers. To determine the reliability of my observations I developed and applied strict criteria for scoring alleles and mutations at microsatellite loci. I observed mutations at all three markers, with rate variation between loci. Single step mutations could not be distinguished because of the limitations of the approach, but 24 multi-step mutations, involving changes of up to 35 repeat units, were recorded. Many of these mutations involved changes that could not be explained by the gain or loss of whole repeat units. These results imply that a large number of mutations at tammar wallaby microsatellites are caused by mechanisms other than replication slippage and are consistent with a role for recombination in the mutation process. Taken as a whole, my results provide evidence for complex mutation processes at tammar wallaby microsatellites. I conclude that careful characterisation of microsatellite mutation properties should be conducted on a case-by-case basis to determine the most appropriate mutation models and analysis tools for each locus. In addition, my work has provided a set of chromosome-specific markers for use in macropod genetic studies, which includes the first marsupial Y chromosome microsatellites. Sex chromosome microsatellites open a new range of possibilities for population studies, as they provide opportunities to investigate gene flow in a male context, to complement data from autosomal and maternally-inherited mitochondrial markers.

"January 2003"
Addendum on back page.
Bibliography: p. 175-184.
184 p. : ill. (some col.) ; 30 cm.
Title page, contents and abstract only. The complete thesis in print form is available from the University Library.
"In the study reported in this thesis, a PCR-based approach was used to isolate the b-like globin genes that are present in the tammar wallaby, Macropus eugenii, including the gene that encodes the w-globin chain. Three -like globin genes (b-, e-, w-) that had previously been described at the protein level in the tammar wallaby were characterised. w-globin orthologues were also identified in a wide range of marsupial species, and in one of these species, the dunnart (Sminthopsis crassicaudata), the complete DNA sequence of the w-globin gene was determined. Southern analysis in the dunnart and in situ hybridisation in the tammar wallaby, provided evidence for the unexpected conclusion that w-globin is not part of the -globin gene cluster in these species. RT-PCR studies using RNA isolated from a new-born dunnarts confirmed that w-globin is expressed in this species. Therefore, this is the first report of an "orphaned" b-like globin gene that is expressed in a vertebrate." --p. 6.
Thesis (Ph.D.)--University of Adelaide, Dept. of Molecular Biosciences, 2003

As a representative of Australian marsupials, the recently sequenced genome of a model kangaroo, the tammar wallaby (Macropus eugenii) provides unique opportunities to understand the organisation and evolution of the genome in marsupials, and in mammals in general. Comparisons with the fully sequenced genome of the Brazilian short-tailed opossum Monodelphis domestica allow comparing the genomes of American and Australian marsupials. The general aims of this thesis were to examine the extent to which part of the genome has been conserved in marsupials and in therian mammals, as well as to explore the organisation and evolution of the largest gene family in mammals, whose members code for olfactory receptors. As part of the KanGo’s task of establishing a map of the tammar genome, the comparative map of the long arm of chromosome 6 in the tammar wallaby was prepared. Several syntenic blocks of genes were mapped to tammar wallaby 6q and it was found that there are only few rearrangements between the tammar wallaby and the opossum in this part of the genome. However, the genomic parts orthologous to tammar wallaby 6q reside on several chromosomes in human, dog, and chicken, suggesting that the fusion occurred in the marsupial ancestors and remained conserved during marsupial evolution. I then developed a strategy to explore the OR gene (ORG) family in the tammar wallaby. Sequences corresponding to ORGs were extracted from the first assembly of the tammar wallaby genome and sequences classified into families and subfamilies. BACs bearing conserved mammalian ORG clusters were isolated and physically mapped in tammar wallaby. Comparison with the opossum OR repertoire revealed that these two distantly related marsupials share a very similar ORG superfamily. Conserved features include the total numbers of genes, families, and subfamilies, gene distribution across the families and subfamilies, patterns of expansions and contractions in families and subfamilies and genomic location of major ORG clusters. I then examined in detail the genomic organisation of a highly conserved ORG cluster that lies near the MHC locus in several mammals. By making a BAC contig over the entire chromosome region I found that this cluster is conserved in tammar wallaby and carries almost the same genes as in the opossum. Preliminary analysis of platypus ORGs dates the origin of this cluster back to the common ancestor of therian and monotreme mammals more than 166 million years ago, and provides examples of both conservation and adaptation of some genes in this cluster. My general conclusion is that the two distantly related marsupial species have retained very similar genomes since their divergence 70 million years ago. This conservation is reflected both at the level of genome arrangement, and at the organisation and evolution of gene families. This conservation is in marked contrast to the variability observed between eutherian groups, both in gross gene arrangement and in the constitution of the ORG family, suggesting that marsupial genomes have been evolving more slowly than other mammals, possibly due to some unique features of their physiology and way of life.

This thesis describes a comprehensive study of the cellular responses of a number of endangered marsupial species with a principal focus on the tammar wallaby (Macropus eugenii) as a model macropod species. The development of in vitro experimental assays for the assessment of immune responses in this model species are described, which provided a set of benchmarks for comparisons with other members of the Macropodidae and with eutherian mammals. Once this data was collected and protocols were established, the study was extended to include investigations of the immune responses in opportunistic samples obtained from the Rufous Hare-wallaby (Lagorchestes hirsutus), the Long-footed potoroo ( Potorous longipes) and the more common, but nonetheless still vulnerable, Long-nosed potoroo (Potorous tridactylus) with a view to investigating their apparent susceptibility to infection with intracellular pathogens, particularly mycobacterial species. The findings from the application of these assays suggest that the cellular immune responses of these species are relatively complex and involve a level of sophistication that rivals their eutherian counterparts. Specifically peripheral blood and tissue leukocytes were morphologically similar to those of other mammals, with the exception of tammar wallaby monocytes that appeared to contain few lysosomal granules, and the basophils of the Rufous Hare-wallaby that contained very large atypical granules. The overall findings of this study suggest that the immune systems of macropod species possess most of the sophistication associated with that of eutherian mammals. Whilst some differences were apparent in cells and their products in the test species, no single factor common to all macropods was identified as a cause for immune dysfunction. It appears likely that as yet undefined factors related to their confinement rather than an inherent defect in their immunocapacity is responsible for the apparent disease susceptibility of these animals.
Doctor of Philosophy (PhD)

The newborn marsupial is born at a highly altricial stage of development when compared to eutherian newborn mammals. Interestingly, the marsupial young are born into a non-sterile environment and lack a developed immune system which prevents them from mounting an adaptive immune response. Two reviews are presented in this thesis. The first review examined the complementary mechanisms which protect the developing marsupial young. Immune protection through forms of innate immunity and maternal contributions has been examined in marsupials. However, as there are many facets to immune protection, our understanding of the protection of the developing marsupial young is far from complete and much of the developing marsupial's innate immune system remains unexplored. Importantly, the review also recognised that the availability of marsupial genomes should be exploited to help direct further research on the protection of marsupial young. The second review examined the role of the pouch in marsupial reproduction. While it is clear that the pouch provides immune protection to those species that have a pouch, the pouch cannot provide immune protection to the young of those species without a pouch. Thus, instead of using the pouch to explain immune protection of marsupial young, the review examined the role of the pouch in supporting highly altricial young and promoting reproductive success in various environments where a pouch may confer significant advantages over free hanging young. The reviews set the direction for the experimental research presented in this thesis. The model species tammar wallaby (Macropus eugenii) was used to further examine innate immune genes. First, innate immune genes, mucin and lysozyme were identified or predicted in the tammar wallaby. The mucin and lysozyme genes were then physically mapped to conserved regions of the tammar wallaby genome which facilitated the confirmation of the identification of the genes. Second, the expression of immune genes was examined in the skin and lung of developing tammar wallabies considered immune-incompetent and immune-competent using transcriptome sequencing. There were 390 and 429 immune genes expressed in developing tammar wallaby skin and lung, respectively, suggesting that both tissues play a role in immune defense. Unexpectedly, mucin and lysozyme were not expressed in the skin and lung of wallabies considered to be immune-incompetent. Further, enrichment analysis of genes identified in Gene Ontology terms showed that specific immune genes, complement component 3, complement factor B and apolipoprotein A1, have increased expression in the skin and lung of wallabies considered immune-incompetent. The increased expression of the immune genes suggests that there may be specific innate immune system processes in play to compensate for the delayed development of the adaptive immune system in marsupial. The research presented in this thesis identified gaps in the research of immune protection of marsupial young and identified, mapped and examined the expression of innate immune genes in the developing tammar wallaby. Thus, we are closer to understanding the complexity of the developing marsupial immune system. The identification of immune genes and their expression, lays the foundation for further functional research in marsupial developmental immunology.

This thesis describes a comprehensive study of the cellular responses of a number of endangered marsupial species with a principal focus on the tammar wallaby (Macropus eugenii) as a model macropod species. The development of in vitro experimental assays for the assessment of immune responses in this model species are described, which provided a set of benchmarks for comparisons with other members of the Macropodidae and with eutherian mammals. Once this data was collected and protocols were established, the study was extended to include investigations of the immune responses in opportunistic samples obtained from the Rufous Hare-wallaby (Lagorchestes hirsutus), the Long-footed potoroo ( Potorous longipes) and the more common, but nonetheless still vulnerable, Long-nosed potoroo (Potorous tridactylus) with a view to investigating their apparent susceptibility to infection with intracellular pathogens, particularly mycobacterial species. The findings from the application of these assays suggest that the cellular immune responses of these species are relatively complex and involve a level of sophistication that rivals their eutherian counterparts. Specifically peripheral blood and tissue leukocytes were morphologically similar to those of other mammals, with the exception of tammar wallaby monocytes that appeared to contain few lysosomal granules, and the basophils of the Rufous Hare-wallaby that contained very large atypical granules. The overall findings of this study suggest that the immune systems of macropod species possess most of the sophistication associated with that of eutherian mammals. Whilst some differences were apparent in cells and their products in the test species, no single factor common to all macropods was identified as a cause for immune dysfunction. It appears likely that as yet undefined factors related to their confinement rather than an inherent defect in their immunocapacity is responsible for the apparent disease susceptibility of these animals.

The South Australian mainland tammar wallaby (Macropus eugenii eugenii) was presumed extinct in the wild from the early 1930's, until a feral population was re-discovered in New Zealand. Eighty-five animals were returned to Australia as part of a repatriation program into their former range. The establishment phase after a reintroduction is a critical time as animals may fail to survive if they cannot find resources and avoid predators in an unfamiliar habitat. To maximise reintroduction and establishment success, reintroductions need to be planned with a good understanding of the animals’ ecology and anti-predator strategies. To improve this understanding requires experimental reintroductions and detailed monitoring. This thesis investigates the experimental reintroduction of 46 wallabies into Innes National Park in South Australia and examines the influence of release group familiarity on establishment. Part of this was an investigation of home range, habitat requirements and social behaviours during establishment and seasonally post-establishment. The thesis includes three data chapters which focus on (1) home range and core area, home range stability and degree of overlap with conspecifics; (2) habitat selection at the landscape scale and for day and night use within home range; and (3) factors influencing fine scale habitat use and social grouping behaviours in light of predation risk. In this study, the animals’ perceived risk of predation is assumed to be reflected by surrogate measures of risk, such as distance to cover, likelihood of using cover, group size, and distance to nearest neighbour. Release group familiarity was established by housing animals together in captivity for at least one month prior to release (“familiar” groups), whereas “unfamiliar” release groups comprised animals housed separately but released together. After each reintroduction the first month was considered to be a time of “establishment” in the environment. During the establishment month, home ranges were not randomly located within the landscape, as indicated by the biased occupancy of particular habitat types. Habitat types selected at the landscape scale were similar for familiar and unfamiliar release groups. However, animals released in unfamiliar groups showed a stronger preference for denser high cover habitat during their nocturnal activities within their home range. As tammars use cover to conceal themselves from predators, this result suggests that animals released in unfamiliar groups were more cautious than those released in familiar groups. Indeed at the fine scale, it was also found that animals released in unfamiliar groups were more likely to be found in high cover habitat, and forage closer to cover at night than did those released in pre-established familiar groups. Using habitat with more caution and capitalising on communal vigilance in an unfamiliar habitat may ultimately improve the likelihood of survivorship and overall reintroduction success. Comparing habitat decisions and social behaviours during the establishment period to similar times of year post-establishment suggested that animals' naivety about their new environment influenced some decisions they made. While habitat selection at the landscape scale was similar during establishment and at an equivalent time of year post establishment, analyses showed that they preferred to use melaleuca (a high cover habitat) during their nocturnal activities during establishment much more strongly than they did once they had established. Home range and core areas were also significantly smaller during the establishment month than at an equivalent time of year post establishment. This result supports the idea that animals will restrict their movements when they are unfamiliar with the habitat and predation risks. It also suggests that some habitat choices improved once they were familiar with their new environment and presumably predation risks. Their habitat choices reflected better anti-predator behaviour than those made during the establishment period: they were more likely to use high cover habitat, they remained significantly closer to cover while foraging, and group sizes were larger than during establishment. Seasonal habitat selection at the landscape and home range scales suggested that the five habitat types within the study area provided different fundamental resources for the animals, as they were preferred at different times of year. Some differences in habitat selection were observed between the sexes, and the females were more selective in their diurnal and nocturnal activities. These differences most likely reflected, in part, differences in predator avoidance and reproductive strategies of the sexes, where females' preference at the landscape scale shifted towards high cover habitat during spring, the time of year when pouch young vacate the pouch and start to become independent. While no such selection at the landscape scale was observed for the males, it was also observed that within their daily activities both sexes were more likely to be found in high cover habitat during spring than any other time of the year, perhaps suggesting that as males were following the females. Overall, females generally selected Eucalyptus diversifolia, Acacia anceps and grassland at the landscape scale, and used E.diversifolia for refuge during the day and the other two habitats for foraging at night, whereas males generally preferred Melaleuca halmaturorum instead of E.diversifolia for diurnal refuge. Eucalyptus rugosa was mostly avoided by both sexes. From month to month, both sexes expanded rather than shifted their home ranges to incorporate new areas, and these new areas were explored with conspecifics (when the amount of new area increased the amount of sharing also increased), highlighting their reliance on communal vigilance when in unfamiliar habitat. More new areas were incorporated into home ranges from July to December than from January to May. Time since release was not influential, which also supported the conclusion that perhaps home ranges moved to follow resources or overlap conspecifics more. Indeed, in one circumstance when neighbouring animals had died, a male wallaby was observed to move four kilometres through unfamiliar habitat and completely shift his home range in search of other residence. Compositional analysis of habitat use versus availability indicated that monthly home ranges were selectively positioned in the landscape and were always larger than 4ha. Males' home ranges were larger than females', and males shared more of their home ranges than females did, supporting the usual sex bias observed for polygamous species. Core areas were proportional to the size of the home range, with similar sizes held by males and females and throughout the year. Core size was not influenced by the degree of overlap with conspecifics, with similar amount of core area shared by both sexes year round. The time of year influenced home range size, the smallest were held in winter when food resources were likely to be most abundant, but also when inclement weather was likely to restrict movements, as the animals' ability to detect predators may be hindered due to wet and windy conditions. Living with conspecifics is known to assist predator detection by group vigilance. Indeed, this study found the amount of home range overlap and the time two individuals spend together was positively correlated, and the size of home ranges decreased when more of it was shared with conspecifics, which suggested that sharing of home range was important. The degree which home ranges were shared was observed to be a fairly stable requirement for both sexes and did not change with season or time since release. Despite previous isolation from predators, the wallabies displayed anti-predator behaviours which incorporated interrelated benefits obtained from group vigilance and using protective cover. Additionally, these behaviours were adjusted according to their familiarity with the habitat. Post-establishment, animals were observed to go further from cover when they were a greater distance from their nearest neighbour but surrounded by larger numbers of conspecifics. Whereas during the establishment period, animals ventured further from cover when they were closer to a nearest neighbour, but group size was not influential. It is known that larger groups of animals have more false alarms to predators, and false alarms result in the animals‟ wasting energy in fleeing. If false alarms are more prevalent while occupying unfamiliar habitat with unfamiliar risks, then relying on large numbers of conspecifics while establishing may have been more of a liability than a benefit during the establishment period. However, some anti-predator strategies were commonly used, regardless of familiarity with their habitat. During the establishment month and post-establishment, animals were always more likely to be found in high cover habitat when they were further from their nearest neighbour, or were surrounded by fewer conspecifics. Some strategies and habitat decisions may have reflected differences in reproductive needs. While females, with and without pouch young did not differ in how far they would forage from protective cover, females with pouch young remained closer to their nearest neighbour than those without. This finding perhaps reflected the importance of relying on communal vigilance when their flight time from a predator may be hindered due to increased weight and bulk of a pouch young. These findings supported the theory that group vigilance anti-predator strategies are somewhat innate in the tammars, as having previously been completely isolated from predators their responses could have been lost, and once released there was no opportunity to socially learn the appropriate responses off an established population. Some habitat and social grouping behaviours were occasionally unexpected but may have been balanced out by other behaviours. This study observed that animals foraged further into the open during winter than at any other time of year, which contradicts findings by other authors where tammars foraged further into the open when the weather was fine. However, I also found that animals remained closer to their nearest neighbour during autumn and winter than at any other time of year. Perhaps foraging further into the open in inclement weather is actually safer if it provides a greater chance to detect and react to an approaching predator. For example, a fox approaching from the scrub edge would not give the wallaby enough time to respond. An additional benefit of foraging further away from cover in larger group sizes, is that animals can flee in different directions confusing the fox, as it would have to make a quick decision and chose one animal to pursue. Animals were furthest from their nearest neighbours in spring which was also somewhat surprising as this is when pouch young vacate the pouch and it was observed that females with pouch young remain closer to their nearest neighbour suggesting they gain some anti-predator benefit from doing so. However, during spring the animals were also more likely to be found in high cover habitat than at any other time of year, so this may have somewhat balanced out the need for a close neighbour. Overall, this study confirmed that tammars retain anti-predator behaviours despite previous isolation from predators. However, their habitat and social decisions improved with time since release. In this experimental study, animals released in groups with unfamiliar conspecifics appeared to be at an advantage as they displayed habitat use and social groupings which suggested they were using their new habitat with more caution. Therefore, this study recommends releasing groups of unfamiliar conspecifics. Releasing animals at different times of year also had an influence on how they used their habitat. Animals released in spring displayed behaviours suggesting that they were more cautious in avoiding predators: they were more likely to be found in cover, foraged closer to cover, and were in larger group sizes than those animals released in winter. Therefore, it is recommended that animals are released at a time of year where conditions are fine and resources are abundant. Releasing females with pouch young did not appear to hinder the animals after their release (compared to females without pouch young) and could be recommended as young permanently evacuating the pouch in the wild are at a greater advantage than juveniles released from captivity. Results from this experimental study were used to assist ongoing management decisions and were imperative in the planning of subsequent reintroduction events for this species, and can be applied more generally to other species with similar anti-predator strategies.
Thesis (Ph.D.) -- University of Adelaide, School of Earth and Environmental Sciences, 2011