Search

Cryptococcosis is a fungal disease in humans and animals, caused by the Cryptococcus neoformans and Cryptococcus gattii species complexes. Clinical cryptococcosis primarily manifests as upper respiratory tract disease; however, dissemination to other organs, particularly the brain, can occur. Nasal colonisation and subclinical cryptococcosis are common in koalas (Phascolarctos cinereus) due to their shared environmental niche with Cryptococcus: Eucalyptus trees. However, for reasons that remain unclear, the prevalence of clinical disease is low in koalas. Interactions between respiratory pathogens and the nasal mycobiome are thought to play a role in the development and progression of numerous respiratory diseases. As such, this study aimed to characterise the mycobiome of the nasal vestibule in koalas with and without evidence of cryptococcal colonisation and subclinical disease via the next-generation sequencing (NGS) of the ITS1 region of the fungal internal transcribed spacer (ITS) gene. Samples were collected from 47 koalas from a population of free-ranging koalas in the Liverpool Plains, NSW, Australia, with a known history of Cryptococcus exposure and nasal colonisation. Of the 47 animals tested, 6.4% were culture-positive only, 4.3% were seropositive only, and 2.1% were culture- and seropositive. C. gattii was detected in four samples via NGS. C. neoformans was not detected via NGS. There were no significant differences in the nasal mycobiomes of Cryptococcus-positive and -negative animals; thus, we could not establish a definitive association between the mycobiome and infection outcomes. We identified a number of fungal genera that were significantly more abundant in samples from Cryptococcus-positive animals, but there was no apparent relationship between these genera and the development of cryptococcosis. This study represents the first investigation of the nasal mycobiota of wild koalas. Further studies involving koalas with clinical disease are necessary to determine the role of the nasal mycobiota in the development of cryptococcosis. [ABSTRACT FROM AUTHOR]

Infection by retroviruses and the mobilization of transposable elements cause DNA damage that can be catastrophic for a cell. If the cell survives, the mutations generated by retrotransposition may confer a selective advantage, although, more commonly, the effect of new integrants is neutral or detrimental. If retrotransposition occurs in gametes or in the early embryo, it introduces genetic modifications that can be transmitted to the progeny and may become fixed in the germline of that species. PIWI-interacting RNAs (piRNAs) are single-stranded, 21–35 nucleotide RNAs generated by the PIWI clade of Argonaute proteins that maintain the integrity of the animal germline by silencing transposons. The sequence specific manner by which piRNAs and germline-encoded PIWI proteins repress transposons is reminiscent of CRISPR, which retains memory for invading pathogen sequences. piRNAs are processed preferentially from the unspliced transcripts of piRNA clusters. Via complementary base pairing, mature antisense piRNAs guide the PIWI clade of Argonaute proteins to transposon RNAs for degradation. Moreover, these piRNA-loaded PIWI proteins are imported into the nucleus to modulate the co-transcriptional repression of transposons by initiating histone and DNA methylation. How retroviruses that invade germ cells are first recognized as foreign by the piRNA machinery, as well as how endogenous piRNA clusters targeting the sequences of invasive genetic elements are acquired, is not known. Currently, koalas (Phascolarctos cinereus) are going through an epidemic due to the horizontal and vertical transmission of the KoRV-A gammaretrovirus. This provides an unprecedented opportunity to study how an exogenous retrovirus becomes fixed in the genome of its host, and how piRNAs targeting this retrovirus are generated in germ cells of the infected animal. Initial experiments have shown that the unspliced transcript from KoRV-A proviruses in koala testes, but not the spliced KoRV-A transcript, is directly processed into sense-strand piRNAs. The cleavage of unspliced sense-strand transcripts is thought to serve as an initial innate defense until antisense piRNAs are generated and an adaptive KoRV-A-specific genome immune response is established. Further research is expected to determine how the piRNA machinery recognizes a new foreign genetic invader, how it distinguishes between spliced and unspliced transcripts, and how a mature genome immune response is established, with both sense and antisense piRNAs and the methylation of histones and DNA at the provirus promoter. [ABSTRACT FROM AUTHOR]

Effective detection techniques are important for wildlife monitoring and conservation applications and are especially helpful for species that live in complex environments, such as arboreal animals like koalas (Phascolarctos cinereus). The implementation of infrared cameras and drones has demonstrated encouraging outcomes, regardless of whether the detection was performed by human observers or automated algorithms. In the case of koala detection in eucalyptus plantations, there is a risk to spotters during forestry operations. In addition, fatigue and tedium associated with the difficult and repetitive task of checking every tree means automated detection options are particularly desirable. However, obtaining high detection rates with minimal false alarms remains a challenging task, particularly when there is low contrast between the animals and their surroundings. Koalas are also small and often partially or fully occluded by canopy, tree stems, or branches, or the background is highly complex. Biologically inspired vision systems are known for their superior ability in suppressing clutter and enhancing the contrast of dim objects of interest against their surroundings. This paper introduces a biologically inspired detection algorithm to locate koalas in eucalyptus plantations and evaluates its performance against ten other detection techniques, including both image processing and neural-network-based approaches. The nature of koala occlusion by canopy cover in these plantations was also examined using a combination of simulated and real data. The results show that the biologically inspired approach significantly outperformed the competing neural-network- and computer-vision-based approaches by over 27%. The analysis of simulated and real data shows that koala occlusion by tree stems and canopy can have a significant impact on the potential detection of koalas, with koalas being fully occluded in up to 40% of images in which koalas were known to be present. Our analysis shows the koala's heat signature is more likely to be occluded when it is close to the centre of the image (i.e., it is directly under a drone) and less likely to be occluded off the zenith. This has implications for flight considerations. This paper also describes a new accurate ground-truth dataset of aerial high-dynamic-range infrared imagery containing instances of koala heat signatures. This dataset is made publicly available to support the research community. [ABSTRACT FROM AUTHOR]

Habitat loss due to wildfire is an increasing problem internationally for threatened animal species, particularly tree-dependent and arboreal animals. The koala (Phascolartos cinereus) is endangered in most of its range, and large areas of forest were burnt by widespread wildfires in Australia in 2019/2020, mostly areas dominated by eucalypts, which provide koala habitats. We studied the impact of fire and three subsequent years of recovery on a property in South-East Queensland, Australia. A classified Differenced Normalised Burn Ratio (dNBR) calculated from pre- and post-burn Sentinel-2 scenes encompassing the local study area was used to assess regional impact of fire on koala-habitat forest types. The geometrically structured composite burn index (GeoCBI), a field-based assessment, was used to classify fire severity impact. To detect lower levels of forest recovery, a manual classification of the multitemporal dNBR was used, enabling the direct comparison of images between recovery years. In our regional study area, the most suitable koala habitat occupied only about 2%, and about 10% of that was burnt by wildfire. From the five koala habitat forest types studied, one upland type was burnt more severely and extensively than the others but recovered vigorously after the first year, reaching the same extent of recovery as the other forest types. The two alluvial forest types showed a negligible fire impact, likely due to their sheltered locations. In the second year, all the impacted forest types studied showed further, almost equal, recovery. In the third year of recovery, there was almost no detectable change and therefore no more notable vegetative growth. Our field data revealed that the dNBR can probably only measure the general vegetation present and not tree recovery via epicormic shooting and coppicing. Eucalypt foliage growth is a critical resource for the koala, so field verification seems necessary unless more-accurate remote sensing methods such as hyperspectral imagery can be implemented. [ABSTRACT FROM AUTHOR]

Simple Summary: After losing their habitat or having it broken up by infrastructure developments, one of the major threats to wild koalas is being hit by cars. Researchers analysed data for 345 koalas that were hit by cars on a 51 km section of the Peak Downs Highway in Queensland, Australia, from October 2014 to November 2023. We found that the spatial distribution of these accidents fit a random pattern along this stretch of road. Two factors seemed to predict where koalas might get hit: the amount of high-quality koala habitat nearby as defined by the local koalas' tree species preference, and driver visibility along the road. Because the landscape is relatively uniform in terms of landuse and vegetation, koalas can, and do, cross anywhere. More research, including detailed habitat mapping using geographic information systems and ground-truthing, is needed to help protect this important koala population and reduce the number of koala vehicle collisions and associated injury and mortality. After the loss and fragmentation of habitat, vehicle collisions are one of the main threats to the long-term survival of wild koalas. Koala road strike data were analysed for a section of the Peak Downs Highway between Nebo and Spencer's Gap, west of Mackay, Queensland, Australia. The analysis was carried out on 345 records (October 2014 to November 2023), and results suggested the spatial distribution of koala road strike followed a random pattern along this section of the highway, assuming a Poisson point pattern on a linear network. An analysis of the candidate predictors of koala vehicle collisions, including habitat and road variables, found that the amount of high-quality koala habitat (as defined by the local koalas' tree species preference) present and the driver visibility were the only significant predictors. The relative homogeneity of landuse and vegetation across this landscape may mean that koalas do not concentrate at specific crossing points. More research, including detailed habitat mapping, is needed into this population, which currently lacks government and conservation attention, to inform mitigation efforts and reduce mortality rates for this potentially nationally significant population. [ABSTRACT FROM AUTHOR]

Q fever is a zoonosis caused by Coxiella burnetii, primarily affecting those in close contact with domestic ruminants, the main source of human infection. Coxiella burnetii has also been detected in various wildlife species globally. In Australia, serological and molecular studies have shown exposure to and infection by C. burnetii in macropods, bandicoots, and koalas. However, the extent to which these species contribute to human infection remains unclear. An unpublished public health investigation into a Q fever case in a person involved in koala care could not conclusively link the infection to koalas due to the patient's broad animal exposure. This study aimed to explore the potential role of koalas in transmitting C. burnetii to humans by investigating the presence of C. burnetii DNA in urogenital tract (UGT) swabs from koalas. DNA was extracted from UGT swabs from koalas in three regions in New South Wales, Australia. An optimised multiplex qPCR assay detected C. burnetii DNA in 2 out of 225 samples (0.89%) at approximately 10 genome equivalents per reaction. Both positive samples amplified all three gene targets. MLVA genotyping identified two distinct C. burnetii genotypes previously isolated from Australian Q fever cases. These findings highlight the need for vaccination against Q fever for those in close contact with koalas. [ABSTRACT FROM AUTHOR]

Simple Summary: Chlamydial infections are a major threat to the health of koalas (Phascolarctos cinereus), an iconic Australian marsupial. Among the different chlamydial species, Chlamydia pecorum (C. pecorum) is the major pathogen infecting koalas, affecting their health and long-term survival, both in the wild and in captivity. Therefore, a deeper understanding of chlamydial infections, including their epidemiology, transmission mode, pathogenesis, host immune response, control, and prevention, is critical for the management of chlamydial infections in koalas. Herein, we discuss the current literature on C. pecorum infection in koalas, including the epidemiology, transmission, pathogenesis, immune response, and control strategies for chlamydial infection, with the aim of improving koala health and achieving effective conservation strategies. Chlamydial infections pose a significant threat to koala populations. Chlamydia pecorum (C. pecorum) remains the major chlamydial species affecting koala health, both in the wild and in captivity, and chlamydial infections are considered important factors affecting the long-term survival of koalas. A clear understanding of chlamydial infections, including the epidemiology, transmission mode, pathogenesis, immune response, control, and prevention thereof, is essential for improving the management of chlamydial infections in koalas. In this study, we discuss the important advances made in our understanding of C. pecorum infection in koalas, focusing on the epidemiology of chlamydial infections, and the transmission, pathogenesis, immune response, and control strategies for chlamydial infection, with the aim of improving koala health and achieving effective conservation strategies. [ABSTRACT FROM AUTHOR]

Climate change has altered the frequency and severity of extreme weather, which can affect vegetation condition and habitat quality for wildlife. Declines in vegetation productivity during droughts and heatwaves can negatively impact animals that depend on vegetation for water and nutrition. We used the normalised difference vegetation index (NDVI) to look at relationships between vegetation productivity and the presence of koalas (Phascolarctos cinereus) in potential habitat throughout much of their range. Using a large, long-term koala presence dataset, we tested the hypothesis that locations where koalas had been observed would exhibit higher NDVI values than a random, representative sample from the same vegetation group. We also identified the minimum NDVI threshold at which koalas occurred across time for each vegetation group and compared these to the minimum NDVI values across potential koala habitat before and during the Millennium Drought, one of the worst recorded in Australia. Additionally, we investigated whether vegetation above the minimum NDVI thresholds was significantly closer to perennial water than unsuitable vegetation. We found that koalas tend to occur at locations with higher NDVI than average for all vegetation groups. Prior to the drought, 49% of potential koala habitat maintained a minimum NDVI above the koalas' threshold, equating to 190,227 km2, which declined to 166,746 km2 during drought (i.e., a 12% reduction). We also found that unsuitable vegetation tended to occur farther from perennial water than suitable vegetation for some vegetation groups. Areas that remained above the NDVI thresholds during the drought should be considered potential refugia for populations during an event of similar magnitude and could indicate future habitat extent. [ABSTRACT FROM AUTHOR]

Simple Summary: This review reports on the current and potential utility of the "koala scat" sample to provide a range of ecological and physiological assessments both at the population and individual animal level and do so in a non-invasive manner. DNA recovered from the scat sample provides useful information on koala distribution, diet, genetics and disease, whereas hormone metabolites can inform physiology. While there are still limitations with respect to the decay of quality DNA (host, microbiome, and pathogen) over time related to climate and sample handling, some of these issues can be overcome with timely sample collection. Other current limitations include an inability to detect and quantify particular hormone metabolites such as oestrogens and/or an appropriate biological interpretation of glucocorticoid metabolite secretion when measured in the faecal sample. The use of samples or scats to provide important ecological, genetic, disease and physiology details on free-range populations is gaining popularity as an alternative non-invasive methodology. Koala populations in SE Queensland and NSW have recently been listed as endangered and continue to face anthropomorphic and stochastic environmental impacts that could potentially lead to their extinction. This scoping review examines the current and potential utility of the koala scat to contribute data relevant to the assessment of koala conservation status and decision making. Although we demonstrate that there is great potential for this methodology in providing details for both individual wild animal and population biology (distribution, abundance, sex ratio, immigration/emigration, genetic diversity, evolutionary significant unit, disease epidemiology, nutrition, reproductive status and stress physiology), the calibre of this information is likely to be a function of the quality of the scat that is sampled. [ABSTRACT FROM AUTHOR]

Cryptococcus is a genus of fungal pathogens that can infect and cause disease in a range of host species and is particularly prominent in koalas (Phascolarctos cinerus). Like other host species, koalas display a range of outcomes upon exposure to environmental Cryptococcus, from external nasal colonization to asymptomatic invasive infection and, in rare cases, severe clinical disease resulting in death. Host factors contributing to these varied outcomes are poorly understood. Due to their close relationship with eucalypt trees (a key environmental niche for Cryptococcus gattii) and suspected continual exposure to the pathogen, koalas provide a unique opportunity to examine host susceptibility in natural infections. Caspase recruitment domain-containing protein 9 (CARD9) is a key intracellular signaling protein in the fungal innate immune response. Humans with mutations in CARD9 succumb to several different severe and chronic fungal infections. This study is the first to sequence and explore CARD9 variation in multiple koalas using Sanger sequencing. Four CARD9 exons were successfully sequenced in 22 koalas from a New South Wales, Australia population. We found minimal variation between koalas across all four exons, an observation that was also made when CARD9 sequences were compared between koalas and six other species, including humans and mice. Ten single-nucleotide polymorphisms (SNP) were identified in this study and explored in the context of cryptococcal exposure outcomes. While we did not find any significant association with variation in cryptococcal outcomes, we found a high degree of conservation between species at several SNP loci that requires further investigation. The findings from this study lay the groundwork for further investigations of CARD9 and Cryptococcus both in koalas and other species, and highlight several considerations for future studies. [ABSTRACT FROM AUTHOR]

Simple Summary: The 2019–2020 Australian bushfire season had a devastating impact on native fauna. It was estimated that 3 billion animals were affected by the fires, but there are few accounts of the species or numbers of animals rescued and rehabilitated post-fire. We reviewed rescue, triage, rehabilitation and release reports for marsupials from two regions: the state of New South Wales (NSW) and Kangaroo Island, South Australia. In NSW, only 889 marsupial rescues were reported, despite estimates of 46.8 million marsupials inhabiting the fire zones. Three marsupial groups dominated the rescue statistics: kangaroos and wallabies (n = 458), koalas (n = 204), and possums (n = 162), with smaller numbers of other marsupial species. The probability of survival and release was lowest for kangaroos and wallabies (15% ± 4%) compared with koalas (47% ± 4%) and possums (55% ± 10%). Injury type was a significant predictor of survival for all groups, with malnourished/moribund animals or those with traumatic injuries less likely to survive. In both jurisdictions, koalas were over-represented, and possums under-represented, in rescue statistics relative to baseline population densities and pre-fire wildlife rescue trends. Triage decisions also varied by species, with koalas more likely to enter care, whereas other species were more likely to be euthanised at triage. Koalas were more likely to die during rehabilitation, with 73% dying or being euthanised between day 1 and 30 post-rescue, representing a potential welfare concern. These species differences in presentation post-fire warrant further investigation, as do the differences in triage, survival and release outcomes. These data highlight the need for detailed record keeping and data sharing, and the development of consistent and evidence-based triage, treatment and euthanasia guidelines for all species. The 2019–2020 Australian bushfire season had a devastating impact on native wildlife. It was estimated that 3 billion native animals were impacted by the fires, yet there are few estimates of the number of animals that were rescued and rehabilitated post-fire. Focusing on the state of New South Wales (NSW) and Kangaroo Island, South Australia, we used a case study approach to determine the number of marsupials that were reported rescued due to the 2019–2020 bushfires in these areas and analysed species-specific trends in rescue and release success. In NSW, we found 889 reports of fire-affected marsupials in 2019–2020, mostly comprising kangaroos and wallabies (macropods; n = 458), koalas (n = 204), and possums (n = 162), with a smaller number of wombats (n = 43) and other marsupial species. Most reports of fire-affected marsupials occurred 6–8 weeks after fire ignition, and there was no difference in temporal frequency of rescues between marsupial groups. For the three main groups, the probability of survival and subsequent release differed, with macropods having the lowest probability of release after rescue (0.15 ± 0.04) compared to koalas (0.47 ± 0.04) and possums (0.55 ± 0.10). The type of injury was the main predictor of survival during rehabilitation for all three marsupial groups, with those malnourished/moribund or with traumatic injuries less likely to survive rehabilitation. Death or euthanasia occurred on the day of rescue for 77% of macropods, 48% of possums and 15% of koalas. Koalas most often died during rehabilitation rather than on the day of rescue, with 73% either dying or being euthanised between day 1 and 30 post-rescue, representing a potential welfare concern. On Kangaroo Island, koalas were the most frequently rescued marsupial species; most euthanasia cases and deaths occurred in a hospital, whereas other marsupials were mostly euthanised at triage. In both jurisdictions, koalas were over-represented while possums were under-represented relative to baseline population densities and wildlife rescue trends in the years before the 2019–2020 bushfires. These species differences in presentation post-fire warrant further investigation, as do the differences in triage, survival and release outcomes. It is hypothesised that the high intensity and large scale of the 2019–2020 fires impeded marsupial fire evasion tactics, as evidenced by the small number of animals found for rescue, and the differing rates of presentation relative to underlying population densities for the main marsupial groups. Based on our findings, there is a need for detailed record keeping and data sharing, development of consistent and evidence-based triage, treatment and euthanasia guidelines and deployment of trained wildlife emergency rescue teams with advanced search techniques to minimise animal suffering where safe to do so. [ABSTRACT FROM AUTHOR]

Simple Summary: The koala has recently been listed as an endangered species in the northern part of its range where it faces significant challenges associated with habitat fragmentation, climate change, and disease. Fundamental to its conservation is a detailed knowledge of its reproductive system, particularly with respect to what represents normal anatomy and/or pathology. In order to conserve we must first understand. This article brings together findings from two PhD theses that provide a detailed description of the female koala reproductive anatomy and histology through different presumptive stages of the reproductive cycle. The koala (Phascolarctos cinereus), while being an iconic Australian marsupial, has recently been listed as endangered. To establish an improved understanding of normal reproductive anatomy, this paper brings together unpublished research which has approached the topic from two perspectives: (1) the establishment of an artificial insemination program, and (2) the definition of Chlamydia spp.-derived histopathological changes of the female koala urogenital system. Based on the presentation and histological processing of over 70 opportunistic specimens, recovered from wildlife hospitals in Southeast Queensland (Australia), we describe the gross and microanatomy of the koala ovary, oviduct, uteri, vaginal complex, and urogenital sinus during the interestrous, proliferative, and luteal phases of the reproductive cycle. [ABSTRACT FROM AUTHOR]

Simple Summary: In the summer of 2019–2020, multiple states of Australia experienced catastrophic bushfires, with koalas being the main wildlife species rescued. Many suffered burns which affected all parts of their body, but particularly their footpads. It can be more difficult to assess the severity of burns in animals than in humans due to the variation in their foot morphology (footpads, hooves, paws), and because fur covers the majority of their body. Here, we describe the clinical and microscopic findings of burns in koalas, following biopsies obtained from animals that were euthanised on welfare grounds after rescue from bushfire. We also describe microscopic findings in the lungs of koalas due to smoke inhalation. These findings will assist veterinarians involved with triaging koalas affected by burns and help predict if they are candidates for rehabilitation or if their prognosis is too poor. In the wake of increasingly frequent bushfires emerging as a threat to wildlife worldwide, koalas have notably been the most rescued species in Australia. However, our understanding of burns and their severity in koalas is limited; hence, this study investigated the histopathological features and depth of burns in koala skin, as well as the presence of smoke-induced respiratory tract damage. In four bushfire-affected koalas that had been euthanised on welfare grounds, skin burns in various body regions were scored based on clinical appearance as superficial, partial thickness, or full thickness. Histological sections of affected regions of skin were assessed as Grades I–IV and showed that furred regions on the ear margins and dorsum were histologically more severe, at Grade III, compared with the clinical score. There was a similar finding for footpad burns, which were the most common body region affected. In the respiratory tract, pulmonary oedema and congestion were evident in all koalas. Overall, the results highlight that cutaneous burn lesions on furred and palmar/plantar surfaces can have higher severity based on the burn depth than is clinically apparent. Therefore, there is a need to consider this when developing treatment plans and establishing prognosis for burnt koalas at triage, as well as that a high likelihood of pulmonary oedema exists. [ABSTRACT FROM AUTHOR]

Simple Summary: Bushfires are a regular occurrence in the Australian landscape. The 2019/20 megafires were unprecedented in intensity and scale, impacting many native species. Koalas were particularly impacted by the fires, with many of those coming into care requiring extensive rehabilitation and treatment. Very little is known about the health parameters of rehabilitated koalas following their release and how these parameters may differ between individuals living in a burnt landscape compared to those in an unburnt landscape. This study tested haematological and serum biochemical parameters, chlamydial shedding and body condition scores of rehabilitated koalas and non-rescued residents living in burnt and unburnt habitats. All koalas received a health check 5–9 months post-fire and one 12–16 months post-fire. Rehabilitated koalas also received a health check when coming into care initially. While some variation in parameters was recorded, the majority of measurements in each koala group fell within the normal reference ranges, suggesting that resident and rehabilitated koalas were in good health at the time of release and when recaptured. These results show that koalas can be supported in burnt landscapes (provided there is adequate nutrition) and that the health of rehabilitated koalas is not compromised by returning them to burnt habitats. Many koalas (Phascolarctos cinereus) required rehabilitation after the 2019/20 Australian megafires. Little is known about how the post-release health of rehabilitated koalas compares to non-rescued resident koalas. We evaluated health parameters in rehabilitated koalas and resident koalas in burnt and unburnt habitat in southern New South Wales, Australia. Health checks were undertaken within six weeks of fire (rehabilitated group), 5–9 months post-fire and 12–16 months post-fire. Body condition improved significantly over time in rehabilitated koalas, with similar condition between all groups at 12–16 months. Rehabilitated koalas therefore gained body condition at similar rates to koalas who remained and survived in the wild. The prevalence of Chlamydia pecorum was also similar between groups and timepoints, suggesting wildfire and rehabilitation did not exacerbate disease in this population. While there was some variation in measured serum biochemistry and haematology parameters between groups and timepoints, most were within normal reference ranges. Our findings show that koalas were generally healthy at the time of release and when recaptured nine months later. Landscapes in the Monaro region exhibiting a mosaic of burn severity can support koalas, and rehabilitated koala health is not compromised by returning them to burnt habitats 4–6 months post-fire. [ABSTRACT FROM AUTHOR]

Simple Summary: Carnivores play an important part in maintaining ecosystem health by limiting the population sizes of their prey. However, their feeding habits can also place populations of threatened species at risk of extinction or decline. We examined the diet of Australia's largest terrestrial carnivore, the dingo, to determine whether it threatens at-risk small macropods in two subtropical forests. We found that although dingoes may prey upon some threatened macropods, they do not appear to do so at rates that will affect population persistence. We show that dingoes in some subtropical Australian forests generally target prey according to availability but also according to accessibility. Our study suggests that at present, dingoes do not appear to pose a threat to threatened macropods or some other threatened mammals in either of the forests surveyed. Carnivores fulfil important ecological roles in natural systems yet can also jeopardise the persistence of threatened species. Understanding their diet is, therefore, essential for managing populations of carnivores, as well as those of their prey. This study was designed to better understand the diet of an Australian apex predator, the dingo, and determine whether it poses a threat to at-risk small macropods in two floristically different yet geographically close reserves in subtropical Australia. Based on an analysis of 512 scats, dingo diets comprised 34 different prey taxa, of which 50% were common between reserves. Our findings add support to the paradigm that dingoes are opportunistic and generalist predators that prey primarily on abundant mammalian fauna. Their diets in the Border Ranges were dominated by possum species (frequency of occurrence (FOC) = 92.5%), while their diets in Richmond Range were characterised by a high prevalence of pademelon species (FOC = 46.9%). Medium-sized mammals were the most important dietary items in both reserves and across all seasons. The dietary frequency of medium-sized mammals was generally related to their availability (indexed by camera trapping); however, the avoidance of some species with high availability indicates that prey accessibility may also be important in dictating their dietary choices. Other prey categories were supplementary to diets and varied in importance according to seasonal changes in their availability. The diets included two threatened macropods, the red-legged pademelon and black-striped wallaby. Our availability estimates, together with earlier dietary studies spanning 30 years, suggest that the red-legged pademelon is resilient to the observed predation. The black-striped wallaby occurred in only two dingo scats collected from Richmond Range and was not detected by cameras so the threat to this species could not be determined. Two locally abundant but highly threatened species (the koala and long-nosed potoroo) were not detected in the dingoes' diets, suggesting dingoes do not at present pose a threat to these populations. Our study highlights the importance of site-based assessments, population monitoring and including data on prey availability in dietary investigations. [ABSTRACT FROM AUTHOR]

Genome sequencing is a powerful tool that can inform the management of threatened species. Koalas (Phascolarctos cinereus) are a globally recognized species that captured the hearts and minds of the world during the 2019/2020 Australian megafires. In 2022, koalas were listed as 'Endangered' in Queensland, New South Wales, and the Australian Capital Territory. Populations have declined because of various threats such as land clearing, habitat fragmentation, and disease, all of which are exacerbated by climate change. Here, we present the Koala Genome Survey, an open data resource that was developed after the Australian megafires. A systematic review conducted in 2020 demonstrated that our understanding of genomic diversity within koala populations was scant, with only a handful of SNP studies conducted. Interrogating data showed that only 6 of 49 New South Wales areas of regional koala significance had meaningful genome-wide data, with only 7 locations in Queensland with SNP data and 4 locations in Victoria. In 2021, we launched the Koala Genome Survey to generate resequenced genomes across the Australian east coast. We have publicly released 430 koala genomes (average coverage: 32.25X, range: 11.3–66.8X) on the Amazon Web Services Open Data platform to accelerate research that can inform current and future conservation planning. [ABSTRACT FROM AUTHOR]

Simple Summary: Habitat loss, urbanisation and climate change may cause stress in koalas. Non-invasive monitoring of faecal cortisol metabolites (FCMs) can be utilised to evaluate the impact of stress. The effectiveness of two enzyme immunoassays (EIAs), 50c and cortisol, in measuring FCM values in wild, stressed koalas was tested. Faecal samples of 234 diseased, injured and control koalas in Queensland, Australia were analysed. Diseased and injured koalas had significantly higher FCM values than clinically healthy control animals as measured by the 50c EIA. Only the 50c EIA detected higher absolute values in males, and also found that females showed a more elevated response to stress manifested by injury and disease. The cortisol EIA was also found unreliable in detecting stress in rehabilitated koalas treated with synthetic glucocorticoids as it cross-reacts with these chemicals. Loss of habitat, urbanisation, climate change and its consequences are anthropogenic pressures that may cause stress in koalas. Non-invasive monitoring of faecal cortisol metabolites (FCMs) can be utilised to evaluate the impact of stressors. The aim was to determine if the tetrahydrocorticosterone (50c) and cortisol enzyme immunoassays (EIAs) could be effective in measuring FCM values in wild, stressed koalas. This research included 146 koalas from the Australia Zoo Wildlife Hospital (AZWH) and 88 from a study conducted by Endeavour Veterinary Ecology (EVE), Queensland, Australia. Faecal samples of diseased, injured and control koalas were analysed. The effect of hospitalisation on FCM values was also investigated. Diseased and injured koalas had significantly higher FCM values than clinically healthy control animals as measured by the 50c EIA. FCM values with the cortisol EIA differed significantly between control and diseased koalas, but not between control and injured ones. Moreover, only the 50c EIA detected higher absolute values in males compared to females, and also found that females showed a more elevated response to stress manifested by injury and disease compared to males. The 50c EIA detected stress during hospitalisation better than the cortisol EIA. The cortisol EIA was also found unreliable in detecting stress in rehabilitated koalas treated with synthetic glucocorticoids as it cross-reacts with these steroids providing artificially high values. [ABSTRACT FROM AUTHOR]

Koala retrovirus (KoRV) exists in both endogenous and exogenous forms and has appeared as a major threat to koala health and conservation. Currently, there are twelve identified KoRV subtypes: an endogenous subtype (KoRV-A) and eleven exogenous subtypes (KoRV-B to -I, KoRV-K, -L, and -M). However, information about subtype-related immune responses in koalas against multiple KoRV infections is limited. In this study, we investigated KoRV-subtype (A, B, C, D, and F)-related immunophenotypic changes, including CD4, CD8b, IFN-γ, IL-6, and IL-10 mRNA expression, in peripheral blood mononuclear cells (PBMCs) obtained from captive koalas (n = 37) infected with multiple KoRV subtypes (KoRV-A to F) reared in seven Japanese zoos. Based on KoRV subtype infection profiles, no significant difference in CD4 and CD8b mRNA expression was observed in the study populations. Based on the different KoRV subtype infections, we found that the IFN-γ mRNA expression in koala PMBCs differs insignificantly (p = 0.0534). In addition, IL-6 and IL-10 mRNA expression also did not vary significantly in koala PBMCs based on KoRV subtype differences. We also investigated the Toll-like receptors (TLRs) response, including TLR2–10, and TLR13 mRNA in koala PBMCs infected with multiple KoRV subtypes. Significant differential expression of TLR5, 7, 9, 10, and 13 mRNA was observed in the PBMCs from koalas infected with different KoRV subtypes. Therefore, based on the findings of this study, it is assumed that co-infection of multiple KoRV subtypes might modify the host innate immune response, including IFN-γ and TLRs responses. However, to have a more clear understanding regarding the effect of multiple KoRV subtypes on host cytokines and TLR response and pathogenesis, further large-scale studies including the koalas negative for KoRV and koalas infected with other KoRV subtypes (KoRV-A to -I, KoRV-K, -L and -M) are required. [ABSTRACT FROM AUTHOR]

Simple Summary: This study examined brain/body ontogenetic growth in marsupials and compared it with placental mammals. While marsupials display morphology and cerebral organization diverse from placentals, their neocortical arrangement and cellular composition is unclear. Unfortunately, knowledge of marsupial ontogenetic brain/body size allometry is limited. Since marsupial brain structure and volume differ when compared with those of placentals, marsupials are considered to possess simple behavioural patterns. This is misleading, since even at a basic observation, Australian marsupials display many of the same mental capacities as other mammals. Consequently, the study findings support further investigation into the intellectual abilities of marsupials. There exists a negative allometry between vertebrate brain size and body size. It has been well studied among placental mammals but less is known regarding marsupials. Consequently, this study explores brain/body ontogenetic growth in marsupials and compares it with placental mammals. Pouch young samples of 43 koalas (Phascolarctos cinereus), 28 possums (Trichosurus vulpecula), and 36 tammar wallabies (Macropus eugenii) preserved in a solution of 10% buffered formalin, as well as fresh juveniles and adults of 43 koalas and 40 possums, were studied. Their brain size/body size allometry was compared to that among humans, rhesus monkeys, dogs, cats, rats, guinea pigs, rabbits, wild pigs, and mice. Two patterns of allometric curves were found: a logarithmic one (marsupials, rabbits, wild pigs, and guinea pigs) and a logistic one (the rest of mammals). [ABSTRACT FROM AUTHOR]

Effective management of threatened and invasive species requires regular and reliable population estimates. Drones are increasingly utilised by ecologists for this purpose as they are relatively inexpensive. They enable larger areas to be surveyed than traditional methods for many species, particularly cryptic species such as koalas, with less disturbance. The development of robust and accurate methods for species detection is required to effectively use the large volumes of data generated by this survey method. The enhanced predictive and computational power of deep learning ensembles represents a considerable opportunity to the ecological community. In this study, we investigate the potential of deep learning ensembles built from multiple convolutional neural networks (CNNs) to detect koalas from low-altitude, drone-derived thermal data. The approach uses ensembles of detectors built from combinations of YOLOv5 and models from Detectron2. The ensembles achieved a strong balance between probability of detection and precision when tested on ground-truth data from radio-collared koalas. Our results also showed that greater diversity in ensemble composition can enhance overall performance. We found the main impediment to higher precision was false positives but expect these will continue to reduce as tools for geolocating detections are improved. The ability to construct ensembles of different sizes will allow for improved alignment between the algorithms used and the characteristics of different ecological problems. Ensembles are efficient and accurate and can be scaled to suit different settings, platforms and hardware availability, making them capable of adaption for novel applications. [ABSTRACT FROM AUTHOR]

A Central Queensland University (CQU) partnership with the Queensland Government National Park management agency has developed a koala (Phascolarctos cinereus) habitat managers' toolkit for vegetation health assessment. Private and public landholders use the field-based toolkit to assess habitat suitability or monitor conservation outcomes for the koala—an iconic Australian arboreal herbivorous marsupial. The toolkit was upgraded recently with instructions to process European Space Agency (ESA) Sentinel-2 multispectral satellite-derived selected vegetation maps for areal vegetation health trend monitoring. A field campaign sought to validate the relatively coarse spatial resolution derived indices (photosynthetic health, leaf area index and leaf water content) to verify their suitability for the habitat management decision-support toolkit. Other user requirement-driven criteria for including remote sensing in the toolkit were imagery and associated processing software costs and ease of map production for habitat managers without cost-effective access to spatial science skills. Despite moderate-to-low field and image vegetation proxy correlations, discussing the results with stakeholders indicates that, at a landscape scale, the use of cost-free, suitable temporal resolution, 10-m spatial resolution imagery is satisfactory when aligned with the design outcomes of a habitat health toolkit. [ABSTRACT FROM AUTHOR]

Keywords: assisted reproductive technologies; artificial reproductive technologies; biobanking; captive breeding; genetic diversity; genome resource banking; heterozygosity; inbreeding; wildlife hospitals EN assisted reproductive technologies artificial reproductive technologies biobanking captive breeding genetic diversity genome resource banking heterozygosity inbreeding wildlife hospitals N.PAG N.PAG 18 04/26/22 20220415 NES 220415 1. In parallel with the development of improved koala sperm cryopreservation technology, it is comforting to note koala spermatozoa also have the capacity of survival for up to 40 days in a chilled (4 °C) state; given the estrous cycle of the koala is approximately 33 days, such physiology lends itself for the immediate application of artificial insemination with chilled semen in wild koalas. However, without legislative support to protect koala habitat, investment in any conservation action to support koala recovery, including our proposed additions to the koala conservation toolbox, cannot function effectively for their intended purpose - to guarantee koala survival in the wild. Based on the significantly increased costs for long-term conventional captive programs aimed at ambitious and optimal genetic diversity retention modelled in the present study (Table 1 and Table 2) and previous studies [[3], [18]], it is unlikely that conservation practitioners will be able to: (1) provide long-term captive insurance in perpetuity against mass mortality or rapid, unforeseen declines of wild koala populations due to stochastic events (e.g., bushfire, climate change [[32], [35]]); (2) continue to bolster wild koala populations and match the current rate of I in situ i decline of these populations and their unique genetic material; and (3) cost-effectively hold viable colony sizes long enough to meet globally accepted genetic diversity targets (e.g., 90% heterozygosity retention for 100 years [[3], [18]]). [Extracted from the article]

Cryptococcosis is typically a sporadic disease that affects a broad range of animal species globally. Disease is a consequence of infection with members of the Cryptococcus neoformans or Cryptococcus gattii species complexes. Although cryptococcosis in many domestic animals has been relatively well-characterized, free-living wildlife animal species are often neglected in the literature outside of occasional case reports. This review summarizes the clinical presentation, pathological findings and potential underlying causes of cryptococcosis in various other animals, including terrestrial wildlife species and marine mammals. The evaluation of the available literature supports the hypothesis that anatomy (particularly of the respiratory tract), behavior and environmental exposures of animals play vital roles in the outcome of host–pathogen–environment interactions resulting in different clinical scenarios. Key examples range from koalas, which exhibit primarily C. gattii species complex disease presumably due to their behavior and environmental exposure to eucalypts, to cetaceans, which show predominantly pulmonary lesions due to their unique respiratory anatomy. Understanding the factors at play in each clinical scenario is a powerful investigative tool, as wildlife species may act as disease sentinels. [ABSTRACT FROM AUTHOR]

Transspecies transmission of retroviruses is a frequent event, and the human immunodeficiency virus-1 (HIV-1) is a well-known example. The gibbon ape leukaemia virus (GaLV) and koala retrovirus (KoRV), two gammaretroviruses, are also the result of a transspecies transmission, however from a still unknown host. Related retroviruses have been found in Southeast Asian mice although the sequence similarity was limited. Viruses with a higher sequence homology were isolated from Melomys burtoni, the Australian and Indonesian grassland melomys. However, only the habitats of the koalas and the grassland melomys in Australia are overlapping, indicating that the melomys virus may not be the precursor of the GaLV. Viruses closely related to GaLV/KoRV were also detected in bats. Therefore, given the fact that the habitats of the gibbons in Thailand and the koalas in Australia are far away, and that bats are able to fly over long distances, the hypothesis that retroviruses of bats are the origin of GaLV and KoRV deserves consideration. Analysis of previous transspecies transmissions of retroviruses may help to evaluate the potential of transmission of related retroviruses in the future, e.g., that of porcine endogenous retroviruses (PERVs) during xenotransplantation using pig cells, tissues or organs. [ABSTRACT FROM AUTHOR]

Koala populations in southeast Queensland are under threat from many factors, particularly habitat loss, dog attack, vehicle trauma and disease. Animals not killed from these impacts are often rescued and taken into care for rehabilitation, and eventual release back to the wild if deemed to be healthy. This study investigated current rescue, rehabilitation and release data for koalas admitted to the four major wildlife hospitals in southeast Queensland (Australia Zoo Wildlife Hospital (AZWH), Currumbin Wildlife Sanctuary Hospital (CWH), Moggill Koala Hospital (MKH) and the Royal Society for the Prevention Against Cruelty to Animals Wildlife Hospital atWacol (RSPCA)), and suggests aspects of the practice that may be changed to improve its contribution to the preservation of the species. It concluded that: (a) the main threats to koalas across southeast Queensland were related to urbanization (vehicle collisions, domestic animal attacks and the disease chlamydiosis); (b) case outcomes varied amongst hospitals, including time spent in care, euthanasia and release rates; and (c) the majority (66.5%) of rescued koalas were either euthanized or died in care with only 27% released back to the wild. The results from this study have important implications for further research into koala rescue and rehabilitation to gain a better understanding of its effectiveness as a conservation strategy. [ABSTRACT FROM AUTHOR]

The acceptance and application of citizen science has risen over the last 10 years, with this rise likely attributed to an increase in public awareness surrounding anthropogenic impacts affecting urban ecosystems. Citizen science projects have the potential to expand upon data collected by specialist researchers as they are able to gain access to previously unattainable information, consequently increasing the likelihood of an effective management program. The primary objective of this research was to develop guidelines for a successful regional-scale citizen science project following a critical analysis of 12 existing citizen science case studies. Secondly, the effectiveness of these guidelines was measured through the implementation of a citizen science project, Koala Quest, for the purpose of estimating the presence of koalas in a fragmented landscape. Consequently, this research aimed to determine whether citizen-collected data can augment traditional science research methods, by comparing and contrasting the abundance of koala sightings gathered by citizen scientists and professional researchers. Based upon the guidelines developed, Koala Quest methodologies were designed, the study conducted, and the efficacy of the project assessed. To combat the high variability of estimated koala populations due to differences in counting techniques, a national monitoring and evaluation program is required, in addition to a standardised method for conducting koala population estimates. Citizen science is a useful method for monitoring animals such as the koala, which are sparsely distributed throughout a vast geographical area, as the large numbers of volunteers recruited by a citizen science project are capable of monitoring a similarly broad spatial range. [ABSTRACT FROM AUTHOR]

10.3390/ani11123550 6 Kayesh M.E.H., Hashem M.A., Tsukiyama-Kohara K. Toll-like receptor expression profiles in koala (Phascolarctos cinereus) peripheral blood mononuclear cells infected with multiple KoRV subtypes. The successful treatment of individual koalas with chlamydial disease is also important, and it has recently been recognised that many antibiotics [[4]] - as well as analgesics used for pain relief [[5]] - can have altered pharmacokinetics in koalas, which may change their efficacy. The koala ( I Phascolarctos cinereus i ) is an arboreal marsupial that is found throughout much of eastern and southeastern Australia, and it relies primarily on eucalypt trees for food, water and shelter. [Extracted from the article]

Surveying threatened and invasive species to obtain accurate population estimates is an important but challenging task that requires a considerable investment in time and resources. Estimates using existing ground-based monitoring techniques, such as camera traps and surveys performed on foot, are known to be resource intensive, potentially inaccurate and imprecise, and difficult to validate. Recent developments in unmanned aerial vehicles (UAV), artificial intelligence and miniaturized thermal imaging systems represent a new opportunity for wildlife experts to inexpensively survey relatively large areas. The system presented in this paper includes thermal image acquisition as well as a video processing pipeline to perform object detection, classification and tracking of wildlife in forest or open areas. The system is tested on thermal video data from ground based and test flight footage, and is found to be able to detect all the target wildlife located in the surveyed area. The system is flexible in that the user can readily define the types of objects to classify and the object characteristics that should be considered during classification. [ABSTRACT FROM AUTHOR]

Simple Summary: Conservation management is critical for threatened wildlife species and an effective conservation approach relies on good understanding of animal behaviour in natural habitat. Currently, koalas are listed as vulnerable to extinction in Australia with declining populations, but their social and breeding system remains unclear. Male koala vocalisations, known as bellows, are believed to be closely related to their breeding behaviour. Bellows incorporate callers' body size information which can be perceived by other koalas. In this study, we tested the behavioural responses of 20 GPS-collared free ranging koalas to bellow recordings collected from small (<6 kg) and large (>8.5 kg) adult male koalas. We report evidence of intra-male competition, with adult males approaching bellow playbacks, particularly those from small-sized males. In contrast, juvenile males under three years of age showed avoidance of the playbacks. No patterns in the response of females were detected. Our results provide the strongest evidence yet that bellows are primarily a means by which males occupy and control habitat space during the breeding season. Future studies are required to see if female response to bellows depends on their reproductive status. Effective conservation strategies rely on knowledge of seasonal and social drivers of animal behaviour. Koalas are generally solitary and their social arrangement appears to rely on vocal and chemical signalling. Male koala vocalisations, known as bellows, are believed to be closely related to their breeding behaviour. Previous research suggests that oestrous female koalas use bellows to locate unique males to mate with, and that males can similarly use bellows to evaluate the physical attributes of their peers. We tested the behavioural responses of 20 free ranging koalas to bellow recordings collected from small (<6 kg) and large (>8.5 kg) adult male koalas. Individual koala movement was reported by hourly-uploaded GPS coordinates. We report evidence of intra-male competition, with adult males approaching bellow playbacks, particularly those from small-sized males. In contrast, males under three years of age were averse to the playbacks. No patterns in the response of females were detected. Our results provide the strongest evidence yet that bellows are primarily a means by which males occupy and control space during the breeding season. Future studies are required to see if female response to bellows depends on their reproductive status. [ABSTRACT FROM AUTHOR]

Simple Summary: Doxycycline is an antimicrobial used for treating chlamydial infections in various species, including the koala. The dose and route of administration used initially are based on first principles. Therefore, this study investigates the absorption, distribution, metabolism, and excretion of subcutaneous doxycycline injections, and evaluates the suitability of the current dosage regimen for inhibiting chlamydial pathogens. The results suggest that the current doxycycline dosage remained therapeutically effective for up to six days after each dose, with some accumulation over successive doses. All koalas in the study improved clinically and tested negative for chlamydial pathogens post-treatment before being released. This study contributes to determining the optimal dosage of doxycycline to treat chlamydiosis safely and effectively in infected koalas. Six mature, male koalas (Phascolarctos cinereus), with clinical signs of chlamydiosis, were administered doxycycline as a 5 mg/kg subcutaneous injection, once a week for four weeks. Blood was collected at standardised time points (T = 0 to 672 h) to quantify the plasma doxycycline concentrations through high-pressure liquid chromatography (HPLC). In five koalas, the doxycycline plasma concentration over the first 48 h appeared to have two distinct elimination gradients; therefore, a two-compartmental analysis was undertaken to describe the pharmacokinetic (PK) profile. The average ± SD maximum plasma concentration (Cmax) was 312.30 ± 107.74 ng/mL, while the average time ± SD taken to reach the maximum plasma concentration (Tmax) was 1.68 ± 1.49 h. The mean ± SD half-life of the distribution phase (T1/2 α) and the elimination phase (T1/2 β) were 10.51 ± 7.15 h and 82.93 ± 37.76 h, respectively. The average ± SD percentage of doxycycline binding to koala plasma protein was 83.65 ± 4.03% at three different concentrations, with a mean unbound fraction (fu) of 0.16. Using probability of target attainment modelling, doxycycline plasma concentrations were likely to inhibit 90% of pathogens with the doxycycline minimum inhibitory concentration (MIC) of 8.0–31.0 ng/mL, and the reported doxycycline MIC to inhibit Chlamydia pecorum isolates at the area under the curve/minimum inhibitory concentration (AUC/MIC) target of ≥24. All koalas were confirmed to be negative for Chlamydia pecorum using loop-mediated isothermal amplification (LAMP), from ocular and penile urethra swabs, three weeks after the last doxycycline injection. [ABSTRACT FROM AUTHOR]

Simple Summary: Folivore marsupials can metabolise high levels of dietary terpenes compared to eutherian mammals, indicating that they possess highly efficient drug metabolising enzymes. However, two such highly efficient drug metabolising enzymes, cytochrome P450 3A and UDP-glucuronosyltransferase, are involved in terpene metabolism, evidence for inhibitory actions by these dietary terpenes on these enzymes are limited. Therefore, we investigated the effect of cineole, a major component of eucalyptus essential oils, and its derivatives on these hepatic drug metabolising enzymes in mice, rats, and possums. Our findings revealed no major inhibition on these drug metabolising enzymes by cineole and its derivatives at concentrations up to 50 µM. However, species-specific difference in their basal enzymatic activity was observed and may provide an avenue for developing future pest control strategies. Folivore marsupials, such as brushtail possum (Trichosurus Vulpecula) and koala (Phascolarctos cinereus), can metabolise higher levels of dietary terpenes, such as cineole, that are toxic to eutherian mammals. While the highly efficient drug metabolising enzymes, cytochrome P450 3A (CYP3A) and phase II conjugating enzymes (UDP-glucuronosyltransferase, UGT), are involved in the metabolism of high levels of dietary terpenes, evidence for inhibitory actions on these enzymes by these terpenes is scant. Thus, this study investigated the effect of cineole and its derivatives on catalytic activities of hepatic CYP3A and UGT in mice, rats, and possums. Results showed that cineole (up to 50 µM) and its derivatives (up to 25 µM) did not significantly inhibit CYP3A and UGT activities in mice, rats, and possums (both in silico and in vitro). Interestingly, basal hepatic CYP3A catalytic activity in the possums was ~20% lower than that in rats and mice. In contrast, possums had ~2-fold higher UGT catalytic activity when compared to mice and rats. Thus, these basal enzymatic differences may be further exploited in future pest management strategies. [ABSTRACT FROM AUTHOR]

Simple Summary: The koala population is declining in northern Australia, and a major reason for this is the infection of the immunosuppressive koala retrovirus (KoRV), which is endogenous in many animals. This endogenous virus and its exogenous forms may induce lymphomas and immunodeficiency associated with opportunistic infections, including chlamydia infections. To generate a vaccine, we produced the recombinant surface and transmembrane envelope proteins of the KoRV and immunized goats, rats and mice. In all cases, we obtained antibodies which were able to neutralize the KoRV and recognize defined epitopes in the envelope proteins. However, we and others observed that koalas carrying the endogenous KoRV are tolerant, e.g., are unable to induce an immune response to the virus. Nevertheless, we propose that KoRV-negative animals will produce an antiviral immune response and will be protected when immunized with such a vaccine. This immunization will also reduce the number and severity of opportunistic infections because there is no KoRV-induced immunosuppression. The koala retrovirus (KoRV) is spreading in the koala population from the north to the south of Australia and is also in the process of endogenization into the koala genome. Virus infection is associated with tumorigenesis and immunodeficiency and is contributing to the decline of the animal population. Antibody production is an excellent marker of retrovirus infection; however, animals carrying endogenous KoRV are tolerant. Therefore, the therapeutic immunization of animals carrying endogenous KoRV seems to be ineffective. Using the recombinant transmembrane (TM) envelope protein of the KoRV, we immunized goats, rats and mice, obtaining in all cases neutralizing antibodies which recognize epitopes in the fusion peptide proximal region (FPPR), and in the membrane-proximal external region (MPER). Immunizing several animal species with the corresponding TM envelope protein of the closely related porcine endogenous retrovirus (PERV), as well as the feline leukemia virus (FeLV), we also induced neutralizing antibodies with similar epitopes. Immunizing with the TM envelope protein in addition to the surface envelope proteins of all three viruses resulted in higher titers of neutralizing antibodies. Immunizing KoRV-negative koalas with our vaccine (which is composed of both envelope proteins) may protect these animals from infection, and these may be the starting points of a virus-free population. [ABSTRACT FROM AUTHOR]

Simple Summary: Koalas can be injured by cars and bushfires, and be affected by painful infectious diseases. When koalas undergo surgery to repair broken bones, they require analgesia. Fentanyl is a potent opioid that can be administered during surgery to provide analgesia. This study describes the rate of elimination of fentanyl in koalas' blood when administered as a single intravenous injection and consequently calculates the dose rate to administer a constant rate fentanyl infusion into the koalas' veins to provide short-term pain control. Fentanyl can also be absorbed via the skin into the circulation when applied as a transdermal patch. Although the data for transdermal fentanyl patch absorption is from two koalas only, the results demonstrate that when a patch is applied, pain control is likely to occur 12 h after application to koalas' skin. Fentanyl may provide effective pain control to koalas either as an intravenous infusion or as a transdermal patch. Fentanyl was administered as a single intravenous bolus injection at 5 µg/kg to five koalas and fentanyl plasma concentrations for a minimum of 2 h were quantified by an enzyme-linked immunosorbent assay (ELISA). The median (range) fentanyl elimination half-life and clearance were 0.53 (0.38–0.91) h, and 10.01 (7.03–11.69) L/kg/h, respectively. Assuming an analgesic therapeutic plasma concentration of 0.23 ng/mL (extrapolated from human studies), an intravenous constant infusion rate was estimated at approximately between 1.7 to 2.7 µg/kg/h (using the clearance 95% confidence intervals). A transdermal fentanyl patch was applied to the antebrachium of an additional two koalas for 72 h. Fentanyl plasma concentrations were determined during the patch application and after patch removal at 80 h. The fentanyl plasma concentration was greater than 0.23 ng/mL after 12 to 16 h. While the patch was applied, the maximum fentanyl concentration was approximately 0.7 ng/mL from 32 to 72 h. Fentanyl plasma concentrations increased to 0.89 ng/mL 1 h after the patch was removed, and then decreased to a mean of 0.47 ng/mL at 80 h. The transdermal fentanyl patch is likely to provide some level of analgesia but should be initially co-administered with another faster acting analgesic for the first 12 h. [ABSTRACT FROM AUTHOR]

Simple Summary: Faecal cortisol metabolites (FCMs) are a useful, non-invasive tool for the assessment of stress in koalas. However, FCM stability after defecation is a critical issue. Therefore, we exposed faecal pellets of koalas to three different environmental conditions and measured FCMs with three recently evaluated enzyme immunoassays (EIAs). Because water loss over time exerted the greatest influence on FCMs, we strongly recommend collecting freshly defecated pellets in koalas. Faecal material can be a valuable source of information for a range of animal health aspects and can be used to measure faecal cortisol metabolites (FCMs). FCM values can relate to physiological stress responses. However, freshly defecated pellets are not always available and environmental conditions, such as temperature and humidity, might affect faecal pellet consistency and FCM levels. Therefore, the impact of environmental conditions on FCMs needs to be evaluated. We collected 107 pellets from two female and two male koalas, exposed them to three types of treatments, and analysed FCMs in these samples with three enzyme immunoassays (EIAs). After analysis, the original FCM values were mathematically corrected for water loss. Results show that the FCMs were more stable when measured using tetrahydrocorticosterone (50c) and 5α-pregnane-3β,11β,21-triol-20-one (37e) EIAs, and were less stable when measured with the cortisol EIA. With 50c, the FCM values did not vary significantly over time either before or after the adjustment with water in the environment treatment group. For samples kept under constant low (25 °C) and high (35 °C) temperatures, the 50c FCM values did not vary significantly over time, after adjustments were made for water loss. Thus, this study highlights the importance of considering the suitability of faecal field samples for FCM analysis. Because water loss was the main driver of FCM changes, we strongly recommend collecting koala pellets that are freshly defecated, despite the effort and time it might take to collect such pellets. [ABSTRACT FROM AUTHOR]

Chlamydia pecorum, an obligate intracellular pathogen, causes significant morbidity and mortality in livestock and the koala (Phascolarctos cinereus). A variety of C. pecorum gene-centric molecular studies have revealed important observations about infection dynamics and genetic diversity in both koala and livestock hosts. In contrast to a variety of C. pecorum molecular studies, to date, only four complete and 16 draft genomes have been published. Of those, only five draft genomes are from koalas. Here, using whole-genome sequencing and a comparative genomics approach, we describe the first two complete C. pecorum genomes collected from diseased koalas. A de novo assembly of DBDeUG_2018 and MC/MarsBar_2018 resolved the chromosomes and chlamydial plasmids each as single, circular contigs. Robust phylogenomic analyses indicate biogeographical separation between strains from northern and southern koala populations, and between strains infecting koala and livestock hosts. Comparative genomics between koala strains identified new, unique, and shared loci that accumulate single-nucleotide polymorphisms and separate between northern and southern, and within northern koala strains. Furthermore, we predicted novel type III secretion system effectors. This investigation constitutes a comprehensive genome-wide comparison between C. pecorum from koalas and provides improvements to annotations of a C. pecorum reference genome. These findings lay the foundations for identifying and understanding host specificity and adaptation behind chlamydial infections affecting koalas. [ABSTRACT FROM AUTHOR]

Simple Summary: In the 2019–2020 Australian bushfires, Kangaroo Island, South Australia, experienced catastrophic bushfires that burnt approximately half the island, with an estimated 80% of the koala population lost. During and after the fires, koalas presented to a designated triage facility over a span of 10 weeks, with 50.2% during the first 14 days of the bushfire response (304 records available). Burns were observed in 67.4% of koalas, with the majority (60.9%) classified as superficial burns, primarily affecting the limbs and face. Poor body condition was recorded in 74.6% of burnt koalas and dehydration in 77.1%. Negative final outcomes (death or euthanasia, either at triage or at a later date) occurred in 45.6% of koalas and were significantly associated with higher mean burn score, maximum burn severity, number of body regions burnt, poor body condition score, and dehydration severity. The findings of this retrospective study may assist clinicians in the field with decision making when triaging koalas in future fire rescue efforts. In the 2019–2020 Australian bushfires, Kangaroo Island, South Australia, experienced catastrophic bushfires that burnt approximately half the island, with an estimated 80% of the koala population lost. During and after the event, rescued koalas were triaged at a designated facility and a range of initial data were recorded including rescue location and date, sex, estimation of age, body condition and hydration, and assessment of burn severity (n = 304 records available). Koalas were presented to the triage facility over a span of 10 weeks, with 50.2% during the first 14 days of the bushfire response, the majority of which were rescued from regions of lower fire severity. Burns were observed in 67.4% of koalas, with the majority (60.9%) classified as superficial burns, primarily affecting the limbs and face. Poor body condition was recorded in 74.6% of burnt koalas and dehydration in 77.1%. Negative final outcomes (death or euthanasia, at triage or at a later date) occurred in 45.6% of koalas and were significantly associated with higher mean burn score, maximum burn severity, number of body regions burnt, poor body condition score, and dehydration severity. The findings of this retrospective study may assist clinicians in the field with decision making when triaging koalas in future fire rescue efforts. [ABSTRACT FROM AUTHOR]

Previous research has shown that the Koala (Phascolarctos cinereus) prefers larger trees, potentially making this a key factor influencing koala habitat quality. Generally, tree height is considered at regional scales which may overlook variation at patch or local scales. In this study, we aimed to derive a set of parameters to assist in classifying koala habitat in terms of tree height, which can then be used as an overlay for existing habitat maps. To determine canopy height variation within a specific forest community across a broad area in eastern Australia, we used freely available Airborne Laser Scanning (ALS) data and adopted a straightforward approach by extracting maximum-height ALS returns within a total of 288 30 m × 30 m "virtual" ALS plots. Our findings show that while maximum tree heights generally fall within published regional-scale parameters (mean height 33.2 m), they vary significantly between subregions (mean height 28.8–39.0 m), within subregions (e.g., mean height 21.3–29.4 m), and at local scales, the tree heights vary in response to previous land-use (mean height 28.0–34.2 m). A canopy height dataset useful for habitat management needs to recognise and incorporate these variations. To examine how this information might be synthesised into a usable map, we used a wall-to-wall canopy height map derived from ALS to investigate spatial and nonspatial clustering techniques that capture canopy height variability at both intra-subregional (100s of hectares) and local (60 hectare) scales. We found that nonspatial K-medians clustering with three or four height classes is suited to intra-subregional extents because it allows for simultaneous assessment and comparison of multiple forest community polygons. Spatially constrained clustering algorithms are suited to individual polygons, and we recommend the use of the Redcap algorithm because it delineates contiguous height classes recognisable on a map. For habitat management, an overlay combining these height classification approaches as separate attributes would provide the greatest utility at a range of scales. In addition to koala habitat management, canopy height maps could also assist in managing other fauna; identifying forest disturbance, regenerating forest, and old-growth forest; and identifying errors in existing forest maps. [ABSTRACT FROM AUTHOR]

Simple Summary: Koalas (Phascolarctos cinereus) are one of Australia's most charismatic native small marsupial species, but populations are rapidly declining throughout Australia as they continue to face increasing pressure from a changing ecosystem. In response to stressors, koalas will use their hypothalamic–pituitary–adrenal (HPA) axis to produce catecholamines and glucocorticoids (e.g., cortisol). In this pilot study, we set out to quantify faecal and fur cortisol metabolites in wild rescued koalas undergoing wildlife rehabilitation. Sampling occurred sporadically over four months (the start of September 2018 to the end of December 2018), and was performed on three rescued koalas (Maree, Tai, and Solstice) being held at the rehabilitation centre. Results of this study show that between the three koalas, the highest recorded faecal cortisol result was 241 ng/g, and the lowest recorded faecal cortisol result was 4 ng/g, whereas the highest recorded fur cortisol result was 1.75 ng/g, and the lowest recorded fur cortisol result was 0.10 ng/g. Statistically, there was a significant difference between all three koalas and their faecal cortisol responses, as well as their fur cortisol responses. Statistically for Maree and Solstice, there was a significant difference in their faecal cortisol response between days when a stressor was recorded, and days when a stressor was not recorded. However, statistically for Tai, this was not the case, as there was no significant difference in his faecal cortisol response between days when a stressor was recorded, and days when a stressor was not recorded. In summary, the hypothesis that faecal glucocorticoids and fur glucocorticoids between koalas will differ based on individual responses to stressors was true as a whole but individually, this hypothesis was true for Maree and Solstice, but untrue for Tai. Measuring faecal and fur glucocorticoids is the first step in understanding how koalas undergoing wildlife rehabilitation respond to stressors. Koalas (Phascolarctos cinereus) are one of Australia's most charismatic native small marsupial species. Unfortunately, populations of koalas are rapidly declining throughout Australia as they continue to face increasing pressure from a changing ecosystem. All wildlife species to some degree will use their hypothalamic–pituitary–adrenal (HPA) axis in response to stress. Depending on the duration of activation, the stress response can lead to either acute or chronic side effects and is modulated through the neuroendocrine stress system with the release of catecholamines and glucocorticoids (e.g., cortisol). It is well known that rehabilitation sanctuaries are inherently stressful for all animals, in particular for rescued wild koalas, as it is an unfamiliar environment where the animals cannot predict or control what will happen to them. In this pilot study, we set out to quantify faecal and fur cortisol metabolites in wild rescued koalas undergoing wildlife rehabilitation. Absolute levels of acute and chronic stress were indexed non-invasively, with faecal samples taken to evaluate acute stress, and fur samples taken to evaluate chronic stress. Sampling occurred sporadically over four months (the start of September 2018 to the end of December 2018), and was performed on three rescued koalas (Maree, Tai, and Solstice) being held at the rehabilitation centre. Results of this study show that between the three koalas, the highest recorded faecal cortisol result was 241 ng/g, and the lowest recorded faecal cortisol result was 4 ng/g, whereas the highest recorded fur cortisol result was 1.75 ng/g, and the lowest recorded fur cortisol result was 0.10 ng/g. Statistically, there was a significant difference between all three koalas and their faecal cortisol responses, as well as their fur cortisol responses. Statistically for Maree and Solstice, there was a significant difference in their faecal cortisol response between days when a stressor was recorded, and days when a stressor was not recorded. However, statistically for Tai, this was not the case, as there was no significant difference in his faecal cortisol response between days when a stressor was recorded, and days when a stressor was not recorded. In summary, the hypothesis that faecal glucocorticoids and fur glucocorticoids between koalas will differ based on individual responses to stressors was true as a whole, but individually, this hypothesis was true for Maree and Solstice, but untrue for Tai. The use of biological samples such as faeces and fur to obtain readings of glucocorticoids is a method of measuring absolute levels of physiological stress that is still evolving for koalas, and there is no current glucocorticoid baseline with which to compare the results of this study; although, measuring faecal and fur glucocorticoids is the first step in understanding how koalas undergoing wildlife rehabilitation respond to stressors. [ABSTRACT FROM AUTHOR]

Simple Summary: Koala populations in Australia are declining and the species is vulnerable to extinction. In the past decade, grazing livestock emerged anecdotally as a threat to koala survival; cattle and horses were reported to have trampled koalas to death when encountered on the ground. In this study, we investigated the scale, frequency, and outcome of livestock-inflicted incidents to koalas via an online survey, and analysed koala admission records from Queensland wildlife hospitals and a wildlife rescue group (Wildlife Victoria) in Victoria. The results provide evidence of both livestock-inflicted injuries and deaths to koalas, especially as these have been confirmed by witness statements. The outcomes for the koala victims of the incidents were severe, which had a 75% death rate. The reported frequency of livestock–koala incidents was low but increasing, with 72 cases (0.14% out of 50,873 admissions) in Queensland wildlife hospitals during 1997–2019, and 59 cases (0.8% of 7017 rescue records) in Wildlife Victoria during 2007–2019, but it is recognised that this was likely to be under-reported. Future research is encouraged to explore the causes of livestock–koala incidents and to develop management strategies to minimise the livestock threat to koalas. Koala populations in Australia are declining due to threats such as chlamydiosis, wild dog predation and vehicle collision. In the last decade, grazing livestock emerged anecdotally as a threat to koala survival in areas where koala habitat and livestock grazing land overlap. This is the first study investigating the significance of livestock-inflicted injuries and deaths in koala populations over a large spatial and temporal scale. We investigated the outcome, scale, and frequency of livestock–koala incidents via an online survey and analysed koala admission records in Queensland wildlife hospitals and a wildlife rescue group (Wildlife Victoria) in Victoria. The results provide evidence of both livestock-inflicted injuries and deaths to koalas, especially as these have been confirmed by witness statements. The outcomes for the koala victims of the incidents were severe with a 75% mortality rate. The reported frequency of livestock–koala incidents was low but increasing, with 72 cases (0.14% out of 50,873 admissions) in Queensland wildlife hospitals during 1997–2019, and 59 cases (0.8% of 7017 rescue records) in Wildlife Victoria during 2007–2019. These incidents were likely to be under-reported due to the remoteness of the incident location, possible mis-diagnoses by veterinarians and the possible reluctance of farmers to report them. Future research is encouraged to explore the mechanics and causes of livestock–koala incidents and to develop management strategies to minimise the livestock threat to koalas. [ABSTRACT FROM AUTHOR]

A significant threat to koala populations is infection from Chlamydia, which results in disease and death. Wild koalas with Chlamydia infections are admitted to wildlife hospitals and treated with antibiotics; however, up to 50% of koalas that present to wildlife hospitals do not survive. A major contributor to high mortality is the development of reproductive cysts, resulting in female infertility and euthanasia. However, the diagnosis of reproductive disease is limited to ultrasound with no further investigations. This communication highlights reports of histological and microbiological findings, the accuracy of ultrasound to necropsy reports and other possible causes for reproductive cyst development previously reported in other hosts. Our conclusions identify a significant knowledge gap in the aetiology of koala reproductive cysts and highlight the urgent need for future investigations. [ABSTRACT FROM AUTHOR]

Simple Summary: Koala habitat is threatened by urbanisation, agricultural activities and by increased temperatures and droughts caused by climate change. Habitat loss may cause stress in koalas, which, in turn, may exacerbate disease occurrence. Stress is associated with an increase of glucocorticoids, of which cortisol is the main one in most mammals. Cortisol is heavily metabolized and excreted via the faeces, where its metabolites can be measured, without interfering with the animal. However, before a link between events causing stress in koalas can be established, baseline levels in stress-free koalas need to be established. Our study has established which diagnostic procedure is best suited to measure these metabolites and has evaluated the physiological baselines levels in male and female koalas and in koalas during the breeding and non-breeding season. The Koala (Phascolarctos cinereus) is an endemic marsupial inhabiting four states of Australia. Urbanisation, declining habitat, drought and fires are threatening the survival of this flagship species. These threats may cause acute and chronic stress in koalas, which might also be associated with occurrence of infectious diseases in koala populations. Stress may induce an increase in cortisol reflected in increased faecal cortisol metabolite (FCM) values. To be able to use faecal cortisol metabolites to measure stress levels in this species, our aim was to determine baseline values for males and females during breeding and non-breeding season. A total of 351 defecations were collected fortnightly, twice a day, for 12 months from koalas at a wildlife facility in South East Queensland. Samples were analysed with three different enzyme immunoassays (EIAs): a cortisol, 5α-pregnane-3β,11β,21-triol-20-one (37e) and tetrahydrocorticosterone (50c) EIA. The latter, which also reacts with tetrahydrocortisol, the main metabolite in koala faeces, was found to have the highest biological sensitivity and, therefore, is the most suitable EIA to measure stress levels in koalas. Utilising this EIA, we found significant differences (p < 0.05) in FCM values between males and females, breeding and non-breeding season, and between morning and evening samples. Values of faecal cortisol metabolites established in stress-free koalas in this study can serve as a reference for future studies in koalas. [ABSTRACT FROM AUTHOR]

Simple Summary: In the summer of 2019–2020, a series of more than 15,000 bushfires raged across Australia in a catastrophic event called Australia's Black Summer. An estimated 3 billion native animals, and whole ecosystems, were impacted by the bushfires, with many endangered species pushed closer to extinction. Zoos Victoria was part of a state-led bushfire response to assist wildlife, alongside government, non-government organisations, and key partners. Here, we detail the role of Zoos Victoria in wildlife triage and welfare, threatened species evacuation and recovery, media and communications, and fundraising during and after the fires. We provide case studies on the triage, care, release, and monitoring of injured koalas (Phascolarctos cinereus); the evacuation and return of endangered eastern bristlebirds (Dasyornis brachypterus) and brush-tailed rock wallabies (Petrogale penicillata); and the development of nutritionally suitable supplementary food and emergency feeding of critically endangered mountain pygmy-possums (Burramys parvus). We share our strategies for future resilience and readiness for similar catastrophic events, as well as the development of triage protocols, emergency response kits, emergency enclosures, captive breeding programs, and nature-based healing for communities directly affected by the fires. We hope that by outlining these contributions from a zoo-based conservation organisation, other zoos and wildlife organisations, both nationally and internationally, may be assisted or encouraged to commit resources and build expertise to assist wildlife in catastrophic events. Modern zoos are increasingly taking a leading role in emergency management and wildlife recovery. In the face of climate change and the predicted increase in frequency and magnitude of catastrophic events, zoos provide specialised expertise to assist wildlife welfare and endangered species recovery. In the 2019–2020 Australian bushfire season, now called Australia's Black Summer, a state government-directed response was developed, assembling specialised individuals and organisations from government, non-government organisations, research institutions, and others. Here, we detail the role of Zoos Victoria staff in wildlife triage and welfare, threatened species evacuation and recovery, media and communications, and fundraising during and after the fires. We share strategies for future resilience, readiness, and the ability to mobilise quickly in catastrophic events. The development of triage protocols, emergency response kits, emergency enclosures, and expanded and new captive breeding programs is underway, as are programs for care of staff mental health and nature-based community healing for people directly affected by the fires. We hope this account of our response to one of the greatest recent threats to Australia's biodiversity, and steps to prepare for the future will assist other zoos and wildlife organisations around the world in preparations to help wildlife before, during, and after catastrophic events. [ABSTRACT FROM AUTHOR]

The koala (Phascolarctos cinereus) is an arboreal folivorous marsupial endemic to Australia. Anthropogenic activities and climate change are threats to this species' survival and are potential stressors. A suitable non-invasive method is needed to objectively detect stress in koalas. Under conditions of stress, the concentration of the hormone cortisol in plasma or in saliva is elevated, and this would provide a convenient measure; however, collecting blood or saliva from wild animals is both practically difficult and stressful, and so likely to confound any measurement. In contrast, measurement of cortisol metabolites in faeces provides a practical and non-invasive method to objectively measure stress in koalas. Unfortunately, the identity of the main faecal cortisol metabolites of koalas is unknown. In this study, we have used both untargeted liquid chromatography–mass spectrometry (LC-MS) and enzyme immunoassays (EIAs) to identify several faecal cortisol metabolites in two koalas, one female (18 months old, 4.1 kg) and one male (4 years old, 6.95 kg) upon administration of hydrocortisone (cortisol) sodium succinate. The LC-MS analysis identified tetrahydrocortisol along with several other isomers as cortisol metabolites. After a survey of five enzyme immunoassays, we found that two metabolites, tetrahydrocortisol and 3β-allotetrahydrocortisol, could be detected by EIAs that used antibodies that were raised against their structurally similar corticosterone counterparts, tetrahydrocorticosterone and 3β-allotetrahydrocorticosterone, respectively. While the 3β-allotetrahydrocortisol metabolite was detected in the faeces of only one of the two animals studied, tetrahydrocortisol was detected in both. These results ultimately indicate that tetrahydrocortisol is likely the main faecal cortisol metabolite in koalas, and we demonstrate that it can be measured by an EIA (50c) that was originally developed to measure tetrahydrocorticosterone. [ABSTRACT FROM AUTHOR]

Simple Summary: Koala retrovirus (KoRV) is a significant threat to koalas across Australia. Koalas in northern koala populations (from New South Wales and Queensland) have KoRV inserted into their DNA and inherited to their offspring. Southern koala populations (from Victoria and South Australia) have KoRV infection spread through close contact of koalas. As such, there are koalas within South Australia that are not infected with KoRV. Accurate diagnosis of the infection of each koala is therefore fundamental for disease studies. Previous studies have shown differences in prevalence of different KoRV genes in the Mount Lofty Ranges Koala population; therefore, clarification is necessary. This study uses a large cohort (n = 216) and defines the diagnostic regions of the KoRV genome within the South Australian population. Using multiple molecular techniques, it demonstrates strong evidence for two clear groupings of koalas: KoRV positive and KoRV negative. Within this study, a population of 41% were shown to be KoRV positive and 57% were KoRV negative, with 2% inconclusive. This differentiation is of great importance when examining the clinical importance of KoRV infection within southern koalas. Koala retrovirus, a recent discovery in Australian koalas, is endogenised in 100% of northern koalas but has lower prevalence in southern populations, with lower proviral and viral loads, and an undetermined level of endogenisation. KoRV has been associated with lymphoid neoplasia, e.g., lymphoma. Recent studies have revealed high complexity in southern koala retroviral infections, with a need to clarify what constitutes positive and negative cases. This study aimed to define KoRV infection status in Mount Lofty Ranges koalas in South Australia using RNA-seq and proviral analysis (n = 216). The basis for positivity of KoRV was deemed the presence of central regions of the KoRV genome (gag 2, pol, env 1, and env 2) and based on this, 41% (89/216) koalas were positive, 57% (124/216) negative, and 2% inconclusive. These genes showed higher expression in lymph node tissue from KoRV positive koalas with lymphoma compared with other KoRV positive koalas, which showed lower, fragmented expression. Terminal regions (LTRs, partial gag, and partial env) were present in SA koalas regardless of KoRV status, with almost all (99.5%, 215/216) koalas positive for gag 1 by proviral PCR. Further investigation is needed to understand the differences in KoRV infection in southern koala populations. [ABSTRACT FROM AUTHOR]

Simple Summary: Koala retrovirus (KoRV) is a major pathogen of koala. Toll-like receptors (TLRs) are important innate immune component that are evolutionary conserved and play a crucial role in the early defense against invading pathogens. The expression profile of TLRs in KoRV infection in koalas is not characterized yet. Therefore, in this study, we characterized TLR expression patterns in koalas infected with KoRV-A only vs. KoRV-A with KoRV-B and/or -C. Using qRT-PCR, we measured TLR2–10 and TLR13 mRNA expression in peripheral blood mononuclear cells (PBMCs) and/or tissues from captive koalas in Japanese zoos. We observed variations in TLR expression in koalas with a range of subtype infection profiles (KoRV-A only vs. KoRV-A with KoRV-B and/or -C). The findings of this study might improve our current understanding of koala's immune response to KoRV infection. Toll-like receptors (TLRs), evolutionarily conserved pattern recognition receptors, play an important role in innate immunity by recognizing microbial pathogen-associated molecular patterns. Koala retrovirus (KoRV), a major koala pathogen, exists in both endogenous (KoRV-A) and exogenous forms (KoRV-B to J). However, the expression profile of TLRs in koalas infected with KoRV-A and other subtypes is yet to characterize. Here, we investigated TLR expression profiles in koalas with a range of subtype infection profiles (KoRV-A only vs. KoRV-A with KoRV-B and/or -C). To this end, we cloned partial sequences for TLRs (TLR2–10 and TLR13), developed real-time PCR assays, and determined TLRs mRNA expression patterns in koala PBMCs and/or tissues. All the reported TLRs for koala were expressed in PBMCs, and variations in TLR expression were observed in koalas infected with exogenous subtypes (KoRV-B and KoRV-C) compared to the endogenous subtype (KoRV-A) only, which indicates the implications of TLRs in KoRV infection. TLRs were also found to be differentially expressed in koala tissues. This is the first report of TLR expression profiles in koala, which provides insights into koala's immune response to KoRV infection that could be utilized for the future exploitation of TLR modulators in the maintenance of koala health. [ABSTRACT FROM AUTHOR]

Simple Summary: Chlamydia is a major pathogen of the Australian marsupial, the koala (Phascolarctos cinereus). One approach to improving this situation is to develop a vaccine. Human Chlamydia research suggests that an effective anti-chlamydial response will involve a balance between a cell-mediated Th1 response and a humoral Th2 responses, involving systemic IgG and mucosal IgA. Characterization of koalas with chlamydial disease suggests that increased expression for similar immunological pathways and monitoring of koalas' post-vaccination can be successful and subsequently lead to improved vaccines. These findings offer optimism that a chlamydial vaccine for wider distribution to koalas is not far off. Chlamydia is a significant pathogen for many species, including the much-loved Australian marsupial, the koala (Phascolarctos cinereus). To combat this situation, focused research has gone into the development and refinement of a chlamydial vaccine for koalas. The foundation of this process has involved characterising the immune response of koalas to both natural chlamydial infection as well as vaccination. From parallels in human and mouse research, it is well-established that an effective anti-chlamydial response will involve a balance of cell-mediated Th1 responses involving interferon-gamma (IFN-γ), humoral Th2 responses involving systemic IgG and mucosal IgA, and inflammatory Th17 responses involving interleukin 17 (IL-17) and neutrophils. Characterisation of koalas with chlamydial disease has shown increased expression within all three of these major immunological pathways and monitoring of koalas' post-vaccination has detected further enhancements to these key pathways. These findings offer optimism that a chlamydial vaccine for wider distribution to koalas is not far off. Recent advances in marsupial genetic knowledge and general nucleic acid assay technology have moved koala immunological research a step closer to other mammalian research systems. However, koala-specific reagents to directly assay cytokine levels and cell-surface markers are still needed to progress our understanding of koala immunology. [ABSTRACT FROM AUTHOR]

Koala retrovirus (KoRV) poses a major threat to koala health and conservation, and currently has 10 identified subtypes: an endogenous subtype (KoRV-A) and nine exogenous subtypes (KoRV-B to KoRV-J). However, subtype-related variations in koala immune response to KoRV are uncharacterized. In this study, we investigated KoRV-related immunophenotypic changes in a captive koala population (Hirakawa zoo, Japan) with a range of subtype infection profiles (KoRV-A only vs. KoRV-A with KoRV-B and/or -C), based on qPCR measurements of CD4, CD8b, IL-6, IL-10 and IL-17A mRNA expression in unstimulated and concanavalin (Con)-A-stimulated peripheral blood mononuclear cells (PBMCs). Although CD4, CD8b, and IL-17A expression did not differ between KoRV subtype infection profiles, IL-6 expression was higher in koalas with exogenous infections (both KoRV-B and KoRV-C) than those with the endogenous subtype only. IL-10 expression did not significantly differ between subtype infection profiles but did show a marked increase—accompanying decreased CD4:CD8b ratio—in a koala with lymphoma and co-infected with KoRV-A and -B, thus suggesting immunosuppression. Taken together, the findings of this study provide insights into koala immune response to multiple KoRV subtypes, which can be exploited for the development of prophylactic and therapeutic interventions for this iconic marsupial species. [ABSTRACT FROM AUTHOR]

To determine Phascolarctid gammaherpesviruses (PhaHV) infection in South Australian koala populations, 80 oropharyngeal swabs from wild-caught and 87 oropharyngeal spleen samples and swabs from euthanased koalas were tested using two specific PCR assays developed to detect PhaHV-1 and PhaHV-2. In wild-caught koalas, active shedding of PhaHV was determined by positive oropharyngeal samples in 72.5% (58/80) of animals, of which 44.8% (26/58) had PhaHV-1, 20.7% (12/58) PhaHV-2 and 34.5% (20/58) both viral subtypes. In the euthanased koalas, systemic infection was determined by positive PCR in spleen samples and found in 72.4% (63/87) of koalas. Active shedding was determined by positive oropharyngeal results and found in 54.0% (47/87) of koalas. Koalas infected and actively shedding PhaHV-1 alone, PhaHV-2 alone or shedding both viral subtypes were 48.9% (23/47), 14.9% (7/47) and 36.2% (17/47), respectively. Only 45.9% (40/87) were not actively shedding, of which 40.0% (16/40) of these had systemic infections. Both wild-caught and euthanased koalas actively shedding PhaHV-2 were significantly more likely to be actively shedding both viral subtypes. Active shedding of PhaHV-2 had a significant negative correlation with BCS in the euthanased cohort, and active shedding of PhaHV-1 had a significant positive relationship with age in both wild-caught and euthanased cohorts. [ABSTRACT FROM AUTHOR]

Koala populations are currently declining and under threat from koala retrovirus (KoRV) infection both in the wild and in captivity. KoRV is assumed to cause immunosuppression and neoplastic diseases, favoring chlamydiosis in koalas. Currently, 10 KoRV subtypes have been identified, including an endogenous subtype (KoRV-A) and nine exogenous subtypes (KoRV-B to KoRV-J). The host's immune response acts as a safeguard against pathogens. Therefore, a proper understanding of the immune response mechanisms against infection is of great importance for the host's survival, as well as for the development of therapeutic and prophylactic interventions. A vaccine is an important protective as well as being a therapeutic tool against infectious disease, and several studies have shown promise for the development of an effective vaccine against KoRV. Moreover, CRISPR/Cas9-based genome editing has opened a new window for gene therapy, and it appears to be a potential therapeutic tool in many viral infections, which could also be investigated for the treatment of KoRV infection. Here, we discuss the recent advances made in the understanding of the immune response in KoRV infection, as well as the progress towards vaccine development against KoRV infection in koalas.