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Wombats suffer from sarcoptic mange, a mite infection that ultimately leads to their death from secondary infections. In 2017, wildlife carers were granted legal approval to treat bare-nosed wombats (Vombatus ursinus) for sarcoptic mange in the field using 4 mL of topical Cydectin® per adult wombat. However, (limited) scientific field trials suggest approved protocols are inadequate which has been supported anecdotally by wildlife carers. Elucidating carer experience is key to holistically advancing understandings of sarcoptic mange treatment. We interviewed 18 wildlife carers regarding the use of Cydectin® to treat free-ranging adult wombats infected with sarcoptic mange which uncovered 43 detailed case studies for examination. Case studies revealed that wildlife carers have used 10–200-mL doses of topical Cydectin® to treat wombats to recovery. These results suggest there is no best-fit for treating wombats in the field, due to individual differences in observed levels of sarcoptic mange severity and differences in wombat behavior. Furthermore, wildlife carers suggested pour-on Cydectin® appeared non-toxic to wombats at rates as high as 200 mL per treatment. We recommend scientific trials should be undertaken to determine the impact and efficacy of the varying treatment regimens, including low and high doses of topical Cydectin® on bare-nosed wombats. This information is required for regulating authorities, and subsequently wildlife carers, and managers, to make fully informed decisions about wombat sarcoptic mange treatment.

Ticks (Arachnida: Acari) are vectors for pathogens and the biggest threat to animal health. Many Australian ticks are associated with pathogens that impact humans, domestic animals and livestock. However, little is known about the presence or impact of tick-borne pathogens in native Australian wildlife. Wombats are particularly susceptible to the effects of the ectoparasite Sarcoptes scabiei which causes sarcoptic mange, the reason for which is unknown. Factors such as other ectoparasites and their associated pathogens may play a role. A critical understanding of the species of ectoparasites that parasitise wombats and their pathogens, and particularly ticks, is therefore warranted. This review describes the ectoparasites of wombats, pathogens known to be associated with those ectoparasites, and related literature gaps. Pathogens have been isolated in most tick species that typically feed on wombats; however, there are minimal molecular studies to determine the presence of pathogens in any other wombat ectoparasites. The development of next-generation sequencing (NGS) technologies allows us to explore entire microbial communities in ectoparasite samples, allowing fast and accurate identification of potential pathogens in many samples at once. These new techniques have highlighted the diversity and uniqueness of native ticks and their microbiomes, including pathogens of potential medical and veterinary importance. An increased understanding of all ectoparasites that parasitise wombats, and their associated pathogens, requires further investigation.

Background Ticks are obligate haematophagous ectoparasites of vertebrate hosts and transmit the widest range of pathogenic organisms of any arthropod vector. Seven tick species are known to feed on bare-nosed wombats (Vombatus ursinus), in addition to the highly prevalent Sarcoptes scabiei mite which causes fatal sarcoptic mange in most bare-nosed wombat populations. Little is known about the pathogens carried by most wombat ticks or how they may impact wombats and wombat handlers. Methods Wombat ticks were sourced from wildlife hospitals and sanctuaries across Australia and identified to species level using taxonomic keys. Genomic DNA was extracted from a subsample, and following the amplification of the bacterial 16S rRNA gene V3–V4 hypervariable region, next-generation sequencing (NGS) on the Illumina MiSeq platform was used to assess the microbial composition. Results A total of 447 tick specimens were collected from 47 bare-nosed wombats between January 2019 and January 2020. Five species of ticks were identified comprising wombat tick Bothriocroton auruginans (n = 420), wallaby tick Haemaphysalis bancrofti (n = 8), bush tick Haemaphysalis longicornis (n = 3), common marsupial tick Ixodes tasmani (n = 12), and Australian paralysis tick Ixodes holocyclus (n = 4). Tick infestations ranged from one to 73 ticks per wombat. The wombat tick was the most prevalent tick species comprising 94% of the total number of samples and was present on 97.9% (46/47) of wombat hosts. NGS results revealed the 16S rRNA gene diversity profile was predominantly Proteobacteria (55.1%) followed by Firmicutes (21.9%) and Actinobacteria (18.4%). A species of Coxiella sharing closest sequence identity to Coxiella burnetii (99.07%), was detected in 72% of B. auruginans and a Rickettsiella endosymbiont dominated the bacterial profile for I. tasmani. Conclusions A new host record for H. longicornis is the bare-nosed wombat. One adult male and two engorged adult female specimens were found on an adult male wombat from Coolagolite in New South Wales, and more specimens should be collected to confirm this host record. The most prevalent tick found on bare-nosed wombats was B. auruginans, confirming previous records. Analysis of alpha-diversity showed high variability across both sample locations and instars, similar to previous studies. The detection of various Proteobacteria in this study highlights the high bacterial diversity in native Australian ticks. Graphical Abstract

There are many limitations when using traditional laboratory species. Limits on variation, may result in limited outcomes, at both the species and individual level, due to different individuals/species having diverse physiological processes, or differing molecular and genetic mechanisms. By using a variety of model species, we will be able to develop creative solutions to biological problems and identify differences of which we were not previously aware. The laboratory mouse has been a suitable model species for various mammalian studies, however most are bred specifically for laboratory research with limited variability due to selective breeding. Marsupial models offer unique research opportunities compared to eutherian models. We believe that there should be an expansion in marsupial model species, and the introduction of the red-tailed phascogale (Phascogale calura), a dasyurid marsupial, should be one of them. Phascogales are easily managed in captivity, and there are now multiple studies involving their development, reproduction, nutrition, behavior and immune system, which can serve as a baseline for future studies. The addition of the phascogale as a model species will improve future mammalian studies by introducing variability and offer alternate solutions to biological problems, particularly in the areas of genetics, nutrition, immunology, the neuro-endocrine system, and ageing, due to their semelparous reproductive strategy and hence, subsequent predictive physiology. In this review, we provide information based on existing research on red-tailed phascogales to support their inclusion as a model species.

Multiple studies have described the anatomy and function of the external ear (pinna) of bats, and other placental mammals, however, studies of marsupial pinna are largely absent. In bats, the tragus appears to be especially important for locating and capturing insect prey. In this study, we aimed to investigate the pinnae of Australian marsupials, with a focus on the presence/absence of tragi and how they may relate to diet. We investigated 23 Australian marsupial species with varying diets. The pinnae measurements (scapha width, scapha length) and tragi (where present) were measured. The interaural distance and body length were also recorded for each individual. Results indicated that all nectarivorous, carnivorous, and insectivorous species had tragi with the exception of the insectivorous striped possum (Dactylopsila trivirgata), numbat (Myrmecobius fasciatus), and nectarivorous sugar glider (Petaurus breviceps). No herbivorous or omnivorous species had tragi. Based on the findings in this study, and those conducted on placental mammals, we suggest marsupials use tragi in a similar way to placentals to locate and target insectivorous prey. The Tasmanian devil (Sarcophilus harrisii) displayed the largest interaural distance that likely aids in better localization and origin of noise associated with prey detection. In contrast, the smallest interaural distance was exhibited by a macropod. Previous studies have suggested the hearing of macropods is especially adapted to detect warnings of predators made by conspecifics. While the data in this study demonstrate a diversity in pinnae among marsupials, including presence and absence of tragi, it suggests that there is a correlation between pinna structure and diet choice among marsupials. A future study should investigate a larger number of individuals and species and include marsupials from Papua New Guinea, and Central and South America as a comparison., We investigated the pinnae of marsupials, with a focus on the presence/absence of tragi and how it may relate to diet. Of the 23 Australian marsupial species studied, only nectarivorous, carnivorous, and insectivorous species had tragi. The results suggest a correlation between pinnae structure and diet choice among marsupials.

Hearing is essential for communication, to locate prey and to avoid predators. We addressed the paucity of information regarding hearing in Australian native mammals by specifically assessing the hearing range and sensitivity of the red-tailed phascogale (Phascogale calura), the kultarr (Antechinomys laniger) and the spinifex hopping-mouse (Notomys alexis). Auditory brainstem response (ABR) audiograms were used to estimate hearing thresholds within the range of 1–84 kHz, over a dynamic range of 0–80 dB sound pressure level (SPL). Phascogales had a hearing range of 1–40 kHz, kultarrs 1–35 kHz and hopping-mice 1–35 kHz, with a dynamic range of 17–59 dB SPL, 20–80 dB SPL and 30–73 dB SPL, respectively. Hearing for all species was most sensitive at 8 kHz. Age showed no influence on optimal hearing, but younger animals had more diverse optimal hearing frequencies. There was a relationship between males and their optimal hearing frequency, and greater interaural distances of individual males may be related to optimal hearing frequency. Because nocturnal animals use high-range hearing for prey or predator detection, our study suggests this may also be the case for the species examined in this study. Future studies should investigate their vocalizations and behaviour in their natural environments, and by exposing them to different auditory stimuli.

People's behaviour towards the conservation of Australian wildlife is important, because people's actions are the main causative factor associated with the demise and future recovery of wildlife conservation. We investigated the type and prevalence of behaviour performed by NSW residents towards wildlife conservation, investigate participants’ willingness to perform conservation behaviours in the future, and identify the barriers preventing people from engaging in conservation action. New South Wales (NSW) residents (n=312) participated in an online questionnaire in a cross-sectional study. Overall, participants’ intention to engage, and rates of actual participation, in conservation behaviours was low. The latter was mostly due to lack of time, money, knowledge, and the impact of health and fitness levels. The knowledge gained from this study can be harnessed to inform future policy and management decisions, design interventions to change conservation behaviours, and provides a foundation for further enquiry into the ‘human dimensions of wildlife’. There remains a wildlife conservation ‘attitude-intention-action gap’, where we are limited with our knowledge on how to transform pro-conservation attitudes into conservation action, which warrants further research.

Marsupials and eutherians are mammals that differ in their physiological traits, predominately their reproductive and developmental strategies; eutherians give birth to well-developed young, while marsupials are born highly altricial after a much shorter gestation. These developmental traits also result in differences in the development of the immune system of eutherian and marsupial species. In eutherians, B-cells are the key to humoral immunity as they are found in multiple lymphoid organs and have the unique ability to mediate the production of antigen-specific antibodies in the presence of extracellular pathogens. The development of B-cells in marsupials has been reported and hypothesised to be similar to that of eutherians, except that haematopoiesis occurs in the liver, postpartum, until the bone marrow fully matures. In eutherians, specific genes are linked to specific stages in B-cell development, maturation, and differentiation processes, and have been identified including immunoglobulins (heavy and light chains), cluster of differentiation markers (CD10, 19, 34 and CD79α/β), signal transduction molecules (BTK, Lyn and Syk) and transcriptional regulators (EBF1, E2A, and Pax5). This review aims to discuss the known similarities and differences between marsupial and eutherian B-cells, in regards to their genetic presence, homology, and developmental stages, as well as to highlight the areas requiring further investigation. By enhancing our understanding of the genes that are involved with B-cells in the marsupial lineage, it will, in turn, aid our understanding of the marsupial immune system and support the development of specific immunological reagents for research and wildlife conservation purposes.

Background Roads have major ecological impacts on wildlife. Vehicle collisions most frequently impact large herbivores due to their larger home range compared to smaller animals, and higher population density compared to carnivores. Kangaroos (Macropus spp.) account for a large proportion of reported wildlife vehicle collisions that occur in N S W, Australia. We aimed to evaluate what influenced road mortality of eastern grey kangaroos (Macropus giganteus) in a temperate rural/suburban region. The location of roadkilled kangaroos found on or near two 1 km stretches of road in Richmond NSW was recorded throughout 2014 and 2015. Weather and moon phase data were recorded for the date of each roadkilled kangaroo. Transects were setup on both roads, and multiple road and landscape features, including the width of roadside, fence construction, habitat type, and distance from street lights measured at 50 m intervals. Data were analyzed to explore which landscape features and temporal factors influenced the occurrence of a roadkill hotspot. Results More kangaroo road mortalities occurred during periods of low temperature and low rainfall, and these factors are likely to affect forage quality. Fewer mortalities occurred when rain was falling. A greater number of mortalities occurred during the waning gibbous phase of the lunar cycle. Significantly more road mortalities occurred a short distance from the end of a section of street lights. Conclusions The findings suggest that illumination influences the likelihood of kangaroo road mortalities. Large herbivores are particularly sensitive to habitat fragmentation because they need unrestricted access to large continuous habitat. Knowledge of factors that influence where and when kangaroos are most likely to cross roads can be used to inform more targeted management strategies and improve future road design and habitat connectivity to reduce the incidence of wildlife vehicle collisions.

Moxidectin (MOX) is a macrocyclic lactone approved worldwide for the treatment of both endo- and ecto-parasites in many mammalian species. The aim of this study was to assess the efficacy of MOX as a treatment against parasites in a range of mammalian species. An electronic literature search was performed for publications to the 1st September 2020. A total of 205 papers were retrieved and screened against all required criteria; hence, 35 were papers were reviewed in this study. The level of evidence and methodological quality was analysed, where a total of 13 publications were categorised as a ‘randomised control trial’, seven were categorised as a ‘non-randomised control trial’ and 15 as an ‘experimental control trial’. The overall methodological quality of the publications was considered low, low to moderate, moderate, moderate to high and high in ten, four, twelve, five and a further four, respectively. We assessed the treatment and possible toxicity of MOX in 13 mammalian species, six investigations reported adverse effects to MOX in a small percentage of individuals. The authors reported observed reactions that were typically mild symptoms that did not require additional therapies, and/or resolved themselves. Further studies are needed to assess the efficacy of MOX treatment in a larger number of species, particularly in wildlife.

B-cells are key to humoral immunity, are found in multiple lymphoid organs, and have the unique ability to mediate the production of antigen-specific antibodies in the presence of pathogens. The marsupial immunoglobulin (Ig) heavy (H) chain locus encodes four constant region isotypes, IgA, IgG, IgM and IgE, but no IgD, and there are two light (L) chain isotypes, lambda (Igλ) and kappa (Igκ). To gain an understanding of the marsupial humoral immune system, B-cell transcriptomes generated by single-cell RNA sequencing from gray short-tailed opossum (Monodelphis domestica) splenocytes, and peripheral blood mononuclear cells were analysed. The cells used were from a single unimmunized animal and the majority of B-cells were transcribing IgM heavy chains. The ratio of Ig light chain use was roughly 2:1, Igλ:Igκ in this individual. This was not predicted due to Igκ being the more complex of the two L chain loci. The variable (V) gene segment pairs used in individual B-cells confirm greater diversity provided by the L chain V. This study is the first to report on using single cell analysis to investigate Ig repertoires in a marsupial and confirms a number of prior hypothesis, as well as revealing some surprises.

Ticks (Arachnida: Acari) are vectors for pathogens and the biggest threat to animal health. Many Australian ticks are associated with pathogens that impact humans, domestic animals and livestock. However, little is known about the presence or impact of tick-borne pathogens in native Australian wildlife. Wombats are particularly susceptible to the effects of the ectoparasite Sarcoptes scabiei which causes sarcoptic mange, the reason for which is unknown. Factors such as other ectoparasites and their associated pathogens may play a role. A critical understanding of the species of ectoparasites that parasitise wombats and their pathogens, and particularly ticks, is therefore warranted. This review describes the ectoparasites of wombats, pathogens known to be associated with those ectoparasites, and related literature gaps. Pathogens have been isolated in most tick species that typically feed on wombats; however, there are minimal molecular studies to determine the presence of pathogens in any other wombat ectoparasites. The development of next-generation sequencing (NGS) technologies allows us to explore entire microbial communities in ectoparasite samples, allowing fast and accurate identification of potential pathogens in many samples at once. These new techniques have highlighted the diversity and uniqueness of native ticks and their microbiomes, including pathogens of potential medical and veterinary importance. An increased understanding of all ectoparasites that parasitise wombats, and their associated pathogens, requires further investigation.

Wombats suffer from sarcoptic mange, a mite infection that ultimately leads to their death from secondary infections. In 2017, wildlife carers were granted legal approval to treat bare-nosed wombats (Vombatus ursinus) for sarcoptic mange in the field using 4 mL of topical Cydectin® per adult wombat. However, (limited) scientific field trials suggest approved protocols are inadequate which has been supported anecdotally by wildlife carers. Elucidating carer experience is key to holistically advancing understandings of sarcoptic mange treatment. We interviewed 18 wildlife carers regarding the use of Cydectin® to treat free-ranging adult wombats infected with sarcoptic mange which uncovered 43 detailed case studies for examination. Case studies revealed that wildlife carers have used 10-200-mL doses of topical Cydectin® to treat wombats to recovery. These results suggest there is no best-fit for treating wombats in the field, due to individual differences in observed levels of sarcoptic mange severity and differences in wombat behavior. Furthermore, wildlife carers suggested pour-on Cydectin® appeared non-toxic to wombats at rates as high as 200 mL per treatment. We recommend scientific trials should be undertaken to determine the impact and efficacy of the varying treatment regimens, including low and high doses of topical Cydectin® on bare-nosed wombats. This information is required for regulating authorities, and subsequently wildlife carers, and managers, to make fully informed decisions about wombat sarcoptic mange treatment.

Infrared cameras are now used universally to study wildlife. In this study camera traps were used to report the use of bare-nosed wombat Vombatus ursinus burrows by other species. Eleven s...

Many species have adapted their behaviour to survive in anthropogenically developed environments (hereafter referred to as developed). Eastern grey kangaroos Macropus giganteus are common ...

Roadkill is a worldwide issue that can cause local population extinctions. In Australia numerous species are affected, however the bare-nosed wombat (Vombatus ursinus) is particularly vulnerable due to its preference for disturbed habitats. Collisions with motor vehicles causes significant damage to the vehicle and sometimes the driver, as well as wounding or killing the wombat. Virtual fences are light and sound-based devices, originally developed in Austria that can be used to reduce roadkill through mitigation. They have had mixed results. In this study a virtual fence was installed along a 1.5 km stretch of road in semi-rural NSW, Australia, with the aim of reducing wombat roadkill through the use of this form of mitigation strategy. The number of roadkilled wombats was counted before and after the fence was installed in March 2020. Prior to the fence being installed 23 wombats were killed and after the fence was installed six wombats were killed. Along Old Bega Road, outside of the fenced area 64 wombats were killed pre-fence installation and 17 post fence installation. Bare-nosed wombats are semi-fossorial ecosystem engineers with an important role in the ecosystem and despite being listed as Least Concern are readily impacted by roadkill. Virtual fencing implemented in regions that have high wombat roadkill rates may aid in reducing road deaths and species conservation. However, we recommend that more research is required to assess virtual fencing, as a roadkill mitigation strategy, including an investigation into a larger number of species in a range of different habitats.

B-cells are key to humoral immunity, are found in multiple lymphoid organs, and have the unique ability to mediate the production of antigen-specific antibodies in the presence of pathogens. The marsupial immunoglobulin (Ig) heavy (H) chain locus encodes four constant region isotypes, IgA, IgG, IgM and IgE, but no IgD, and there are two light (L) chain isotypes, lambda (Igλ) and kappa (Igκ). To gain an understanding of the marsupial humoral immune system, B-cell transcriptomes generated by single-cell RNA sequencing from gray short-tailed opossum (Monodelphis domestica) splenocytes, and peripheral blood mononuclear cells were analysed. The cells used were from a single unimmunized animal and the majority of B-cells were transcribing IgM heavy chains. The ratio of Ig light chain use was roughly 2:1, Igλ:Igκ in this individual. This was not predicted due to Igκ being the more complex of the two L chain loci. The variable (V) gene segment pairs used in individual B-cells confirm greater diversity provided by the L chain V. This study is the first to report on using single cell analysis to investigate Ig repertoires in a marsupial and confirms a number of prior hypothesis, as well as revealing some surprises.

Sarcoptic mange is caused by the mite Sarcoptes scabiei and has recently been recognized as an emerging infectious disease of wildlife worldwide. The mite is one of the main causes of population decline in southern hairy-nosed (Lasiorhinus latifrons) and bare-nosed wombats (Vombatus ursinus). This review focuses on Sarcoptes scabiei infestations in wombats and provides insights into why the disease may be so prevalent in wombats. Current treatment practices and trials conducted in the field to reduce the incidence of sarcoptic mange in wombats are described and critically reviewed. Current and potential future avenues of research are discussed.

Short-beaked echidnas (Tachyglossus aculeatus) are myrmecophages, or ant and termite insectivore specialists, and replicating their exact diet in captivity is problematic. Diets for captive animals often incorporate raw meat, eggs and cat food mixed together with water, and vitamin and mineral supplements. These diets have promoted a number of health problems in captive echidnas, such as gastritis, cystitis, gut impaction, obesity, and diarrhea. A manufactured diet was designed and three echidnas from two zoos were transitioned onto this diet to assess the acceptability and digestibility of this diet for echidnas. The new "test" diet was readily accepted by the echidnas with a 1 week transition period. Daily digestible energy intake was 280 kJ kg-0.75 d-1 , similar to another myrmecophagous species. Digestibility values were above 74% for all macronutrients. It was determined that this diet was an acceptable replacement for the previous diets and it was decided that the remaining echidnas at both institutions would be transitioned to the new diet. The diet will also be used for wild echidnas being rehabilitated in the zoo hospitals prior to release and commercially available within Australia. Further data are being collected to assess the use of this diet for seasonal weight management, transitioning hand-reared puggles and effects on gastrointestinal tract health. Zoo Biol. 36:56-61, 2017. © 2017 Wiley Periodicals, Inc.

Mosquito-borne viruses are well recognized as a global public health burden amongst humans, but the effects on non-human vertebrates is rarely reported. Australia, houses a number of endemic mosquito-borne viruses, such as Ross River virus, Barmah Forest virus, and Murray Valley encephalitis virus. In this review, we synthesize the current state of mosquito-borne viruses impacting non-human vertebrates in Australia, including diseases that could be introduced due to local mosquito distribution. Given the unique island biogeography of Australia and the endemism of vertebrate species (including macropods and monotremes), Australia is highly susceptible to foreign mosquito species becoming established, and mosquito-borne viruses becoming endemic alongside novel reservoirs. For each virus, we summarize the known geographic distribution, mosquito vectors, vertebrate hosts, clinical signs and treatments, and highlight the importance of including non-human vertebrates in the assessment of future disease outbreaks. The mosquito-borne viruses discussed can impact wildlife, livestock, and companion animals, causing significant changes to Australian ecology and economy. The complex nature of mosquito-borne disease, and challenges in assessing the impacts to non-human vertebrate species, makes this an important topic to periodically review.

In this review we investigated the diet and nutrition of wombats and highlight areas for future research. The low energy requirements of wombats, conservative foraging behaviours and burrowing lifestyle allow them to subsist on low-quality food. Furthermore, their digestive anatomy and physiology is well adapted for digesting the resources available in their habitats. All species feed predominantly on native grasses; however, the nutritional requirements of juvenile and aged wombats are largely unknown. Changes to food availability and grass species in wombat habitats may alter their diet and nutritional intake. Also, despite evidence suggesting that wombats can satisfy their water requirements via the food they eat, the minimum threshold of water content at which wombats will cease feeding is unknown. Other areas for future research include investigating factors that affect digesta retention times (such as age or lactation), and the development of a method to determine age, because unlike most other mammals, wombat teeth grow continuously, hence teeth cannot be used as a reliable predictor of age in wombats.

Much of the literature on wombat diet is concerned only with the broad categories of consumed matter, and there is little about their specific nutritional requirements despite diet playing a major role in health. This study gathered information by surveying zoological parks and wildlife carers about current feeding regime, diet and the prevalence of nutritional or dental disorders of wombats held in captivity, with the aim of improving the diets of captive wombats, thus their health and welfare outcomes. Data on the composition, variation and basis of diets for 31 bare-nosed or common wombats (Vombatus ursinus) and 19 southern hairy-nosed wombats (Lasiorhinus latifrons) were gathered from 13 facilities. The nutritional values of diets fed to wombats in captivity were calculated and compared with known dietary requirements for bare-nosed wombats. Results demonstrate that the current diet of wombats held in captivity often consists of a high portion of fruits, vegetables and livestock feed, with the fruit and vegetable portions providing higher levels of simple carbohydrates and starch compared with natural grass diets. Generally, the facilities are providing diets that are exceeding reported maintenance energy and nitrogen requirements. Due to a lack of details provided regarding quantity and species of grasses fed, we could not provide an accurate ration calculation for the proportion of grasses in the diets. Some of the health concerns noted included insufficient weight gain, endoparasites, malocclusion and insufficient tooth wear. Further research is required to assess the requirements of reproductive, lactating and juvenile wombats; however, it is clear that wombats in captivity need to be fed larger volumes and varieties of grasses, forbs and sedges than they are currently being fed to maintain good health.

Quolls are carnivorous marsupials in the family Dasyuridae with characteristic white spots. They are distributed throughout Australia and New Guinea, but uncommonly seen due to their mostly nocturnal solitary nature, and large home ranges. All Australian quolls are listed as ‘near threatened’ or ‘endangered’ at state, national and international levels, largely due to human-induced threats. Threats include introduced predators, habitat loss through clearing and modifications including changed fire regimes, disease, human persecution, vehicle collisions and accidental or targeted poisoning by humans and cane toads (Rhinella marina). Conservation efforts that have focussed on reducing introduced predators, and minimising the impact of cane toads, have aided some translocations, hence species recovery in some local areas of Australia has occurred. Where species conservation has required captive breeding for translocation, successful captive management has been crucial. We summarise research conducted in captivity on aspects of birth and development, health and disease, and blood and nutrition parameters of quolls, and suggest future directions for research. Further research on captive and wild quoll populations will benefit future translocations, reintroductions and conservation through increased knowledge, improved maintenance and husbandry of captive colonies, and monitoring of wild populations.

Quolls are carnivorous marsupials in the family Dasyuridae with characteristic white spots. They are distributed throughout Australia and New Guinea, but uncommonly seen due to their mostly nocturnal solitary nature, and large home ranges. All Australian quolls are listed as ‘near threatened' or ‘endangered' at state, national and international levels, largely due to human-induced threats. Threats include introduced predators, habitat loss through clearing and modifications including changed fire regimes, disease, human persecution, vehicle collisions and accidental or targeted poisoning by humans and cane toads (Rhinella marina). Conservation efforts that have focussed on reducing introduced predators, and minimising the impact of cane toads, have aided some translocations, hence species recovery in some local areas of Australia has occurred. Where species conservation has required captive breeding for translocation, successful captive management has been crucial. We summarise research conducted in captivity on aspects of birth and development, health and disease, and blood and nutrition parameters of quolls, and suggest future directions for research. Further research on captive and wild quoll populations will benefit future translocations, reintroductions and conservation through increased knowledge, improved maintenance and husbandry of captive colonies, and monitoring of wild populations.

Expressed coding sequences for interleukin-6 (IL-6) and interleukin-6 receptor α (IL-6R) were examined in five marsupial species. Full length expressed coding sequences for IL-6 and IL-6R were identified and characterized in the gray short-tailed opossum (Monodelphis domestica). For IL-6, ∼225 bp fragments of the mRNA sequence were identified in the red-tailed phascogale (Phascogale calura), kultarr (Antechinomys laniger), and stripe-faced dunnart (Sminthopsis macroura), while ∼563 bp fragments of mRNA encoding IL-6R were identified in the red-tailed phascogale, kultarr, stripe-face dunnart and fat-tailed dunnart (Sminthopsis crassicaudata). Relative expression levels of IL-6 and IL-6R were examined in the heart, muscle, lung, liver, spleen and kidney of adult red-tailed phascogales, and IL-6 gene expression was found to be significantly higher in the lung and spleen than the other tissues examined, while the expression of IL-6R was significantly higher in the liver, lung and spleen. These results now serve as a reference point for examining the role and levels of IL-6 and IL-6R in the health and disease of these marsupial species. The pro-tumorigenic nature of IL-6 is of particular interest, and the identification of these IL-6 and IL-6R coding sequences provides a platform for further work to evaluate the potential role of IL-6 in marsupial cancers.

Marsupials are born immunologically premature, relying on cells and molecules in maternal milk for immune protection. Both immunoglobulin and complement proteins have been identified in marsupial milk, but the expression of specific complement proteins remains largely unexplored. We report partial cDNA sequences for two complement-activating proteins, C3, C1r, CFP and MASP2, in liver tissues from red-tailed phascogale (Phascogale calura). Conservation of functionally relevant motifs were identified in the translated cDNA sequences from phascogale C3, CFP and MASP2 and their eutherian homologues. Gene expression of representative molecules from each of the major complement pathways was also investigated in whole body tissues from 1 to 18 day old animals and liver tissues from 31-day to 14-month old animals. Average complement expression in whole bodies and liver tissues of C1r, CFP, MASP2 and C3 increased significantly in juveniles compared to pouch young, presumably due to the maturation of the young's own complement system. Comparing expression in liver tissues only, we found that the average CFP expression were higher in pouch young compared to juveniles, while results were still statistically similar to the average expression of all tissues for C1r, MASP2 and C3. The average complement expression then significantly decreased as the animals aged into adulthood.

In mammals, interleukin-21 is a member of the common gamma chain cytokine family that also includes IL-2, IL-4, IL-7, IL-9 and IL-15. IL-21 has pleiotropic effects on both myeloid and lymphoid immune cells and as a consequence, the biological actions of IL-21 are broad: regulating both innate and adaptive immune responses and playing a pivotal role in antiviral, inflammatory and antitumour cellular responses. While IL-21 genes have been characterized in mammals, birds, fish and amphibians, there are no reports for any marsupial species to date. We characterized the expressed IL-21 gene from immune tissues of two macropod species, the tammar wallaby (Macropus eugenii), a model macropod, and the closely related endangered bridled nailtail wallaby (Onychogalea fraenata). The open reading frame of macropod IL-21 is 462 nucleotides in length and encodes a 153-mer putative protein that has 46% identity with human IL-21. Despite the somewhat low amino acid conservation with other mammals, structural elements and residues essential for IL-21 conformation and receptor association were conserved in the macropod IL-21 predicted peptides. The detection of IL-21 gene expression in T-cell-enriched tissues, combined with analysis of the promotor region of the tammar wallaby gene, suggests that macropod IL-21 is expressed in stimulated T cells but is not readily detected in other cells and tissues. The similarity of gene expression profile and functionally important amino acid residues to eutherian IL-21 makes it unlikely that the differences in B- and T-cell responses that are reported for some marsupial species are due to a lack of important functional residues or IL-21 gene expression in this group of mammals.

Embryonic development is strongly affected by incubation temperature in most animals, and the effects of incubation temperature continue after partition. However, relatively little is known about the effects of incubation temperature on the development of morphological features in reptiles. As a result, whether developmental rates of different traits change uniquely in response to incubation temperature remains unknown. In this study, we compared embryonic development of an Australian turtle incubated at different temperatures. We examined 43 internal and external morphological features of embryos at each week of development and compared presence and absence of each feature within and across weeks and temperature treatments, using a multivariate statistical approach. We identified morphological features that develop at different rates in turtle embryos incubated at different temperatures. Embryos incubated at 30°C hatched 3 weeks earlier, grew to hatchling size faster, and developed advanced morphological features earlier than those incubated at 26°C. Furthermore, embryonic features that appear late in development appeared progressively earlier in embryos incubated at 30°C. Embryos incubated at 26°C also retained some early embryonic features for longer periods of time during development, especially from weeks 2–4. Despite temperature effects, some traits appeared at the same week of development in embryos from both treatments, including a clearly distinguishable carapace and distinct gut villi at week 3. Variation in appearance and retention of features means that not all external features consistently appear in the same order, depending on incubation temperature. From a practical perspective, cool- and warm-incubated embryos may exhibit different sets of embryological features, which may not be captured by previously published embryological staging guides.

Studies of gene diversity are used to investigate population dynamics, including immunological fitness. Aside from the selection of an appropriate gene target, an important factor that underpins these studies is the ability to recover viable DNA samples from native animals that are protected, threatened or difficult to sample or locate such as the bare-nosed wombat (Vombatus ursinus). In this study, we used genomic DNA extracted from muscle tissue samples and also identified the optimal method to extract DNA from fresh wombat scat samples to enable further analyses to be performed using non-invasive techniques. The DNA was probed via the polymerase chain reaction using previously targeted marsupial Major Histocompatibility Complex (MHC) gene primers. These genes are highly variable and associated with binding and presentation of pathogens in the immune system. Twenty-three unique MHC Class II DAB V. ursinus gene sequences were translated to 21 unique predicted peptide sequences from 34 individual tissue or scat samples. Vombatus ursinus MHC Class II DAB gene and peptide sequences were most similar to other marsupial DNA and peptide sequences. Further analysis also indicated the likelihood of MHC Class II DAB family membership through motif identification. Additional sampling is required to assess the full level of diversity of MHC Class II DAB genes among V. ursinus populations; however, this study is the first to identify MHC genes in a wombat and will advance immunological and disease studies of the species.

Wombats are large marsupial grazers native to Australia. They are ‘ecosystem engineers’ due to their burrowing, impacting rates of soil turnover, assisting with nutrient cycling and water infiltration, and creating habitat for other species. The aim of this review is to provide an overview of available information on the habitat and distribution, abundance and threats to bare-nosed wombats (Vombatus ursinus), and to provide a critical review of the current gaps in our knowledge to aid future management of the species. Bare-nosed wombats were previously widespread. Since European settlement their range has reduced. While habitat loss and contraction impact population numbers, bare-nosed wombat populations are also threatened by roadkill, introduced species, legal and illegal human persecution, and disease. Further research is required to understand the historic range of bare-nosed wombats, as well as current population numbers, so that we can determine the threats that most significantly impact this species. Increasing our understanding of habitat requirements, distribution and population numbers, as well as how to mitigate threats, will aid future management and conservation strategies for the species.

Objectives The genetic diversity of an eastern grey kangaroo (Macropus giganteus) population surrounded by landscape barriers was examined. DNA was extracted from tissue samples from 22 road-killed kangaroos, and blood samples from four live captured kangaroos. Amplified loci were used to determine relatedness between individual kangaroos. The level of relatedness and location of road-killed kangaroos were compared to evaluate spatial autocorrelation. Results The expected and observed heterozygosity confirmed the loci were polymorphic and highly informative for use in this population. One pair of kangaroos were identified to be full siblings, and a high proportion were identified as half siblings. Six positive parentage assignments were detected. No correlation between relatedness and crossing site was detected.

Background The red-tailed phascogale is a ‘Near Threatened’ dasyurid marsupial. Males are semelparous and die off shortly after the breeding season in the wild due to a stress-related syndrome, which has many physiological and immunological repercussions. In captivity, males survive for more than 2 years but become infertile after their first breeding season. Meanwhile, females can breed for many years. This suggests that captive males develop similar endocrine changes as their wild counterparts and undergo accelerated aging. However, this remains to be confirmed. The health status and immune function of this species in captivity have also yet to be characterized. Results Through an integrative approach combining post-mortem examinations, blood biochemical and hematological analyses, we investigated the physiological and health status of captive phascogales before, during, and after the breeding season. Adult males showed only mild lesions compatible with an endocrine disorder. Both sexes globally maintained a good body condition throughout their lives, most likely due to a high quality diet. However, biochemistry changes potentially compatible with an early onset of renal or hepatic insufficiency were detected in older individuals. Masses and possible hypocalcemia were observed anecdotally in old females. With this increased knowledge of the physiological status of captive phascogales, interpretation of their immune profile at different age stages was then attempted. During the breeding season, males developed a stress leukogram characterized by a marked lymphopenia, further aggravated by a severe leukopenia after the breeding season. To determine whether these changes were limited to the peripheral blood or had more profound implications, histopathology of the spleen was performed opportunistically. Adult males showed white pulp atrophy, at various degrees. The atrophy was mainly lymphoid and more severe in 1.5-year-old males than in 3.5-year-old females. These results suggest that captive males undergo accelerated immunosenescence. Conclusions Functional studies are now needed to characterize the underlying mechanisms leading to immunosenescence in marsupials. Semelparous dasyurids present great potential for studying the effects of sex and stress on immunity in marsupials. Characterization of these immune-endocrine interactions may help refine veterinary treatment plans, husbandry protocols and conservation programs to maintain the health of captive and wild populations.

Cells expressing the surface markers CD3, CD4, CD79b, IgM, MHC class II, and ModoUG (nonclassical MHC class I) were detected in red-tailed phascogale tissues using immunohistochemistry, and the appearance and localization of cells observed here was consistent with previous observations in other marsupial species. CD3+ cells were first detected at one day postpartum (dpp) in the thymus, followed by ModoUG+ cells at 5-7 dpp in the thymus and lymph nodes. CD79b+ cells were first detected at 12-14 dpp in bone marrow, spleen, and lymph nodes. IgM+ cells were first detected at 12-14 dpp in thymus, bone marrow, spleen, and lymph nodes. MHC class II+ cells were first detected at 12-14 dpp in thymus, bone marrow, and lymph nodes. CD4+ cells were detected in adult thymus and spleen only. The presence of the mature immune cell populations and their localization to characteristic T and B cell zones in mature lymphoid tissues with normal histological structure indicates that red-tailed phascogales develop immunocompetence by the end of pouch life. Anat Rec, 302:1985-2002, 2019. © 2019 American Association for Anatomy.

Objective Marsupials suffer from an increasing number of stressors in this changing world. Functional studies are thus needed to broaden our understanding of the marsupial immune system. The red-tailed phascogale (Phascogale calura) is a small Australian marsupial previously used in descriptive immunological studies. Here, we aimed to develop functional assays by isolating and stimulating blood and spleen mononuclear cells in vitro. Results While peripheral blood mononuclear cell (PBMC) were relatively easy to isolate, only 105 mononuclear cells (> 90% purity and > 75% viability) could be recovered from the spleen, independently of the sex and age of the animal or the centrifugation time and speed tested. The pores of the mesh sieve used for tissue homogenization might have been too big to yield a single cell suspension. Nevertheless, in spite of the overall low number of cells recovered, PBMC and splenic mononuclear cells were successfully activated in preliminary trials with phytohemaglutinin. This activation state was evidenced by a change in shape and the presence of small cell aggregations in the mitogen-stimulated cultures. A non-radioactive colorimetric assay was also performed to confirm cell proliferation in these wells. This work highlights the importance of developing and reporting detailed methodological protocols in non-traditional research species.

Expressed coding sequences were identified for the T cell receptors TCRα, TCRβ, TCRγ, TCRδ, TCRμ, and co-receptors CD3e, CD4, and CD8α in the red-tailed phascogale (Phascogale calura). Expression of these genes was examined in the developing thymus and spleen of pouch young and juvenile individuals to detect any changes in expressionrelative to developmental stages. Transcripts of CD3e, CD4, TCRδ and TCRμ were first detected in the thorax at 1 day postpartum (dpp), and transcripts of all target genes were detected in the thymus from 3 dpp, and the spleen from 17 dpp. The relative expression of all target genes changed significantly over the course of pouch life and was associated with histological milestones of both the thymus and the spleen. Peak gene expression was observed early in pouch life in the thymus, with a decrease in expression associated with timing of thymic involution, while expression peaked later in the spleen and was associated with the initial differentiation of the red and white pulp and a population of T cells forming the periarterial lymphatic sheath (PALS). Active expression of these target genes within the thymus and spleen indicates that the young marsupials are actively developing immunocompetence during pouch life.

Red-tailed phascogale (Phascogale calura) pouch young at birth were relatively underdeveloped in comparison with their eutherian counterparts, and the lymphoid tissues of the immune system were found to be histologically immature. The phascogale thymus rapidly developed in the first few days of pouch life and was quickly populated with lymphocytes. By the end of pouch life, involution of the thymus was underway. The bone marrow started to develop in the early stage of pouch life, although adipocytes and megakaryocytes were not observed until slightly later. The liver was hematopoietic from birth and reached histological maturity toward the end of pouch life. The lymph nodes were difficult to detect macroscopically because of their small size, but were easily identified microscopically later in pouch life, particularly in the mesentery, and these lymph nodes exhibited germinal centers by the end of pouch life. The early spleen was predominately mesenchymal, but exhibited some erythropoiesis. Follicles with well-developed germinal centers were not observed until the latest stage of pouch life. Although intraepithelial lymphocytes were detected in the intestines early in pouch life, the discrete lymphoid aggregates and Peyer's patches characteristic of the gut-associated lymphoid tissue (GALT) were not detected until later in pouch life. This is the first report of histological development in phascogale pouch young, as well as the first report of the thymus, bone marrow, and lymph nodes in this dasyurid species at any age.

Torpor is an important physiological process used to reduce body temperature and thus energy needs in mammals and birds. Rewarming from torpor can be costly. In some small mammals basking is used to reduce the cost of rewarming from torpor. The use and duration of basking was measured in two native Australian marsupials, the red-tailed phascogale (Phascogale calura) and kultarr (Antechinomys laniger), using a heat rock. Animals were observed via video cameras to determine whether they used basking behaviours and whether this behaviour was affected by food availability. Both species became accustomed to the heat rock as time progressed. Absence of food increased basking duration and frequency in some individuals but not all animals. Generally kultarrs basked for longer periods than phascogales, and kultarrs would bask at all hours of the day while phascogales showed a strict nocturnal regime (1900-0700h). The results demonstrate that basking may be an important behavioural adaptation of the kultarr; while the phascogale requires further research.

Very few assays that are used to assess the status of mammalian immunity have proved useful for assessment of marsupial health and/or diagnosis of disease. This is largely due to the lack of species cross-reactive reagents that underpin such experiments. To begin to address this deficit, we describe the activation of classical and alternative complement pathways of red-tailed phascogales (RTP; Phascogale calura). Using standard haemolytic assays, the existence of both complement pathways were established in RTP serum based on its ability to lyse unsensitised rabbit erythrocytes (RbE) and sensitised sheep erythrocytes (SE), respectively. The alternative complement pathway assays were conducted using pooled serum of male and female RTPs, and the remaining RTP sera were opportunistically used to test the presence of a functional classical complement system in individual animals, a first in non-eutherian animals. Observations from this study suggest that the activation of these two complement pathways in RTPs are comparable to that seen in other mammals. Since this assay was able to be used on very small samples of blood, it could serve as a useful tool to gather data for comparative immunological studies and to further our knowledge of the mechanisms of immunity available to marsupial young.