Opportunistic bacterial pathogens frequently associated with human clinical infections, including antimicrobial-resistant strains, are infiltrating the microbiomes of wild animals, where they have the potential to negatively impact wildlife health. Bacterial genes conferring resistance to amoxicillin have previously been reported in koala ( Phascolarctos cinereus ) faecal DNA. Koalas are facing several key threats, including wildfires, and affected individuals may receive amoxicillin therapy to treat burn wounds. This study aimed to identify the species of amoxicillin-resistant bacteria in koala gut microbiomes and determine if they are opportunistic pathogens. Faecal samples collected from 98 wild-caught koalas were cultured using amoxicillin-supplemented media to isolate amoxicillin-resistant Gram-negative enteric bacteria. Isolates were screened using 16S rRNA PCR and Sanger sequencing to identify opportunistic pathogenic species, which then underwent whole-genome sequencing and antimicrobial susceptibility testing. Intrinsically amoxicillin-resistant opportunistic pathogens were obtained from 9.2% (9/98) of koala faecal samples and comprised Klebsiella oxytoca (6/98, 6.1%), Klebsiella pneumoniae (1/98, 1.0%) and Citrobacter spp. (2/98, 2.0%). Seven of nine amoxicillin-resistant opportunistic pathogens also exhibited cephalosporin resistance. Four K. oxytoca isolates belonged to lineages associated with human clinical infections, which also have the potential to cause disease in koalas, including fatal systemic infections in pouch young. The presence of amoxicillin- and cephalosporin-resistant strains may also increase the risk of gut dysbiosis and opportunistic infections when penicillins or cephalosporins are required to treat bacterial infections in koalas, highlighting the importance of good antimicrobial stewardship. The study findings demonstrate the One Health perspective of microbial pathogens and the intertwined microbial ecology between humans and wildlife.

Humans have historically been considered the only natural host of typical enteropathogenic Escherichia coli (tEPEC), a cause of human infantile diarrhoea. Recent findings of bat-specific tEPEC in two Australian bat species (Pteropus poliocephalus and Pteropus conspicillatus) revealed that these Pteropus spp. are also tEPEC hosts. tEPEC pathogenicity is associated with key virulence factors including intimin (eae) and the bundle-forming pilus (bfp) operon, which contains the bfpA gene. This study characterised 63 tEPEC isolates from five Australian Pteropus spp.; four from mainland Australia (P. poliocephalus, P. conspicillatus, P. alecto and P. scapulatus) and one from Christmas Island (P. natalis). The 63 tEPEC isolates included 10 novel tEPEC strains and eight previously identified Pteropus tEPEC strains. Faecal DNA samples (n = 386) from the five Australian Pteropus spp. were screened for eae and bfpA genes to identify tEPEC-positive samples. The estimated true prevalence of tEPEC ranged from 14.2% to 37.0% across the five Pteropus spp. Typing of 119 bfpA alleles (63 tEPEC isolates and 56 faecal DNA samples) from the five Pteropus spp. identified 27 bfpA allele types, 24 of which belonged to bat-specific bfpA lineages. Bat-specific bfpA alleles were shared between tEPEC strains, Pteropus spp. and across regions, including the geographically isolated P. natalis endemic to Christmas Island. This study reveals that diverse bat-specific tEPEC strains and bfpA types have evolved in Pteropus spp. and have been circulating among populations for an extensive period, thereby confirming that all Australian Pteropus spp. are natural tEPEC hosts.

Antimicrobial-resistant Escherichia coli have now infiltrated the gut microbiomes of wild animals globally, posing a threat to the health and conservation of endangered species. In previous studies of an Australian fruit bat species, the grey-headed flying fox (GHFF; Pteropus poliocephalus), the prevalence of amoxicillin-resistant E. coli was significantly higher (77.4%) in GHFF pups from South Australia (SA) compared to adult GHFF (3.8%). In rehabilitation settings, amoxicillin is one of the most frequently administered antibiotics to GHFF. Our study aimed to determine the prevalence and genetic characteristics of amoxicillin-resistant E. coli in two additional cohorts of young GHFF: hand-reared pups from New South Wales (NSW) undergoing rehabilitation for release back to wild habitats (n=25) and older juveniles from SA in a pre-release facility (n=30), and compare these with previous findings in GHFF. Amoxicillin-resistant E. coli were cultured from GHFF faecal samples using amoxicillin-supplemented media. Isolates confirmed as E. coli by PCR underwent whole-genome sequencing and antimicrobial susceptibility testing. Amoxicillin-resistant E. coli were identified in 40.0% (10 out of 25) of NSW GHFF pups and 3.3% (1 out of 30) of SA juvenile GHFF. Twelve amoxicillin-resistant E. coli isolates were obtained from 11 bats, with all belonging to anthropogenic-associated lineages. Additional resistance to amoxicillin-clavulanic acid, cephalosporins, tetracyclines, fluoroquinolones and trimethoprim-sulfamethoxazole was detected across the 12 isolates. Overall, 33.3% of GHFF E. coli were multidrug-resistant, and 66.7% had extraintestinal pathogenic traits and were classified as opportunistic pathogens. These findings indicate that the prevalence of amoxicillin-resistant E. coli is significantly higher in GHFF pups than in juvenile and adult GHFF, and carriage may not be long-term. Further research is required to examine the prevalence and dynamics of antibiotic-resistant E. coli in GHFF pups as they mature. The infiltration of antibiotic-resistant E. coli into GHFF microbiomes presents yet another threat to this endangered species and highlights the need for good antimicrobial stewardship when treating wildlife.

Abnormal vertebral column curvature is sporadically reported in koalas of the Mount Lofty Ranges, South Australia. This study evaluates the imaging features of 23 koalas from the Mount Lofty Ranges presenting with abnormal vertebral column curvature between 2015 and 2023 using digital radiography and computed tomography (CT). All images were evaluated by four reviewers to assess curve morphology, severity and Cobb angles for both scoliosis and kyphosis. For Cobb angle measurement, radiography performed similarly to CT with good agreement as measured by intraclass correlation coefficient (0.835 and 0.825 respectively) and Lin’s concordance correlation coefficient (>0.85). The apex vertebra was always located between T7 and L6. For both scoliosis and kyphosis apex vertebrae, the thoracolumbar region was the most common location (8/22 and 9/19, respectively). For scoliosis, the caudal thoracic and lumbar regions were equally common (7/22 each), whereas for kyphosis, the caudal thoracic region (7/19) was more frequent than the lumbar region (3/19). Vertebral body rotation was a common component particularly in severely affected individuals, in which complex or ‘S’ shaped curves also occurred. Severity ranged from minimal or mild (6/23) to moderate (5/23) and severe (12/23), with simultaneous kyphosis and scoliosis present most frequently (21/23). As a result, the term, kyphoscoliosis is the most appropriate morphological description for abnormal vertebral curvature in koalas and may have pathophysiologic commonalities with human idiopathic scoliosis. This study is the first to describe imaging features of abnormal vertebral column curvature in koalas and evaluate inter-modality and interobserver agreement between radiography and CT.

ABSTRACT Extreme heat weather events can lead to energy deficits by increasing thermoregulatory energy costs while reducing energy intake because of limits to dissipating the metabolic heat generated by foraging activity. Understanding the consequences of exposure to extreme heat is increasingly important under ongoing climate warming. We measured acceleration forces using collar‐mounted loggers to gain indices of activity and energy expenditure in grey‐headed flying‐foxes (Pteropus poliocephalus) during mild and hot summer conditions. Overall dynamic body acceleration (ODBA) is positively correlated with energy expenditure in a wide range of animals. During the daytime rest phase, air temperatures above 30°C decreased low‐level activity indicative of rest, whereas during the nocturnal active phase, temperatures above 21°C reduced high‐level activity indicative of flight. On hot days (maximum Ta: 40.6°C ± 3.1°C), ODBA increased threefold during the daytime and decreased by one‐third during the first 2 h of the night, compared to milder summer conditions. These patterns indicate that extreme heat simultaneously elevates energy expenditure and constrains foraging activity. Our data support the hypothesis that heat‐induced energetic deficits are an important component of the negative effects of extreme heat exposure. Such energy deficits could reduce survival and population growth, especially as heat events become more frequent and severe under climate change.

The ability to navigate is crucial to the survival of many flying animals. Though relatively much less is known about the navigational abilities of bats versus birds, recent progress has been made in understanding the navigational abilities of cave roosting bats, but little is known about those of arboreal roosting flying-foxes, despite their extreme mobility. We use extremely high spatiotemporal resolution GPS tracking to examine the flight behaviour of 11 grey-headed flying-foxes (Pteropus poliocephalus) displaced 16.8 km from their roost. We examined flight metrics of the resulting high-resolution traces to understand whether the displaced animals were aware their location with respect to the roost of capture. We use 7 grey-headed flying-foxes tracked from the roost of capture—as part of a separate, concurrent study—to aid in this comparison. Ten of 11 displaced individuals were detected at the roost of capture within four days of release, but all displaced individuals roosted for at least one night away from the roost of capture. Six individuals returned ‘home’ the next day, and four roosted away from ‘home’ for ≥ one further night. Prior to their return ‘home’, displaced individuals on average flew 2.7 times further and stopped 1.7 more times than reference individuals or displaced animals that had already returned ‘home’. This indicates that displaced individuals expended more effort each night than non-displaced individuals. This suggests that these individuals were attempting to return ‘home’, rather than choosing not to return due to a lack of motivation to home. Flight segments of displaced individuals were higher, less straight, and less likely to be oriented. Flight segments that ended in a point that an individual had previously visited were faster, higher, and straighter than those not known to end in a point previously visited. Our findings suggest that approximately half of the displaced animals were aware of where they were with respect to ‘home’ the night after release, whereas other individuals took at least a further night to orient themselves. While our results are consistent with previous work suggesting that non-echolocating bats may use a large-scale navigational map based on vision, sensory manipulations would be needed to confirm this.

Extreme heat events increasingly challenge the thermoregulatory capacities of wildlife, as the frequency, intensity, and duration of these events rise under climate change. Biologging can reveal the physiological and behavioural responses of wild animals to natural variation in environmental conditions, but few studies have recorded thermoregulatory patterns during extreme heat events. Flying-foxes (Pteropus spp.) are convenient bioindicators of the impacts of extreme heat events on wildlife because they often roost in accessible colonies in trees where population-level consequences, including mass mortalities, can be readily observed. To understand how flying-foxes thermoregulate in response to extreme heat, we used implanted temperature-sensitive transmitters to record the core body temperature (Tb) of 17 free-living, adult male grey-headed flying-foxes (Pteropus poliocephalus) on 142 days across two Austral summers, including six days when air temperature (Ta) exceeded 42 °C and thousands of flying-foxes died. Flying-foxes exhibited daily heterothermy, with a decrease in Tb after dawn (minimum: 35.9 ± 0.1 °C; absolute: 31.1 °C) followed by an increase in Tb during the day (maximum: 38.7 ± 0.2 °C; absolute: 44.3 °C). Above Ta of 29.5 °C, bats allowed Tb to rise above normal levels (i.e., controlled hyperthermia). On extreme heat days (Ta > 42 °C), Tb increased by up to 6 °C to a daily maxima between 40.5 to 42.4 °C, and, consequently, the Ta at which Ta exceeded Tb occurred at 40 °C, 2.5 °C higher than without controlled hyperthermia. Large variation in Tb, as exhibited by grey-headed flying-foxes, reduces the physiological costs of exposure to thermal conditions in summer. Controlled hyperthermia during extreme heat events increases the range of Ta allowing non-evaporative heat loss and hence decreases water loss. The ability to manage elevated Tb and high rates of evaporative water loss will shape the resilience of many mammals to future extreme heat events.

BACKGROUND:Reference intervals (RIs) are an essential tool for assessment of clinical pathology data of animals, and are particularly important for monitoring the health status of free-ranging and captive wildlife, such as koalas (Phascolarctos cinereus). OBJECTIVES:The purpose of this study was to: (1) provide comprehensive serum biochemistry reference intervals based on clinically healthy South Australian koalas from two populations, Mount Lofty Ranges (MLR) and Kangaroo Island (KI); and (2) identify any factors that can affect biochemical analytes, including koala retrovirus (KoRV) and Chlamydia pecorum subclinical infection status, age, sex, and population. METHODS:Serum biochemistry analytes were determined in 206 clinically healthy South Australian koalas caught from the wild in 2016 and 2018 using a Cobas 8000 Chemistry Analyzer and analyzed using Reference Value Advisor and SPSS v28 Statistical software. RESULTS:Biochemical reference intervals were established. Also, clinically and statistically significant differences in analytes were found based on age for alkaline phosphatase and phosphate, and albumin: globulin ratio, globulins, and total protein, most likely associated with physiological bone growth and immunological development, respectively, as observed in other species. Statistically significant differences between animals subclinically positive for KoRV and Chlamydia pecorum, were found for glucose and gamma glutamyl transferase respectively; however, these were marginal, and their reference intervals were similar. CONCLUSIONS:This study is the first to describe serum biochemical reference intervals for clinically healthy South Australian koalas of known Chlamydia and KoRV infection status. It represents an important tool to assist health assessments of koalas by veterinarians, as well as research and population monitoring.

In the 2019–2020 summer, wildfires decimated the Australian bush environment and impacted wildlife species, including koalas ( Phascolarctos cinereus ) and grey headed flying fox pups (Pteropid bats, Pteropus poliocephalus ). Consequently, hundreds of koalas and thousands of bat pups entered wildlife hospitals with fire-related injuries/illness, where some individuals received antimicrobial therapy. This study investigated the dynamics of antimicrobial resistance (AMR) in pre-fire, fire-affected and post-fire koalas and Pteropid bat pups. PCR and DNA sequencing were used to screen DNA samples extracted from faeces (koalas and bats) and cloacal swabs (koalas) for class 1 integrons, a genetic determinant of AMR, and to identify integron-associated antibiotic resistance genes. Class 1 integrons were detected in 25.5% of koalas (68 of 267) and 59.4% of bats (92 of 155). Integrons contained genes conferring resistance to aminoglycosides, trimethoprim and beta-lactams. Samples were also screened for bla TEM (beta-lactam) resistance genes, which were detected in 2.6% of koalas (7 of 267) and 25.2% of bats (39 of 155). Integron occurrence was significantly higher in fire-affected koalas in-care compared to wild pre-fire koalas ( P < 0.0001). Integron and bla TEM occurrence were not significantly different in fire-affected bats compared to pre-fire bats ( P > 0.05), however, their occurrence was significantly higher in fire-affected bats in-care compared to wild fire-affected bats ( P < 0.0001 and P = 0.0488 respectively). The observed shifts of AMR dynamics in wildfire-impacted species flags the need for judicious antibiotic use when treating fire-affected wildlife to minimise unwanted selective pressure and negative treatment outcomes associated with carriage of resistance genes and antibiotic resistant bacteria.

The African leopard (Panthera pardus pardus) has lost a significant proportion of its historical range, notably in north-western Africa and South Africa. Recent studies have explored the genetic diversity and population structure of African leopards across the continent. A notable genetic observation is the presence of two divergent mitochondrial lineages, PAR-I and PAR-II. Both lineages appeared to be distributed widely, with PAR-II frequently found in southern Africa. Until now, no study has attempted to date the emergence of either lineage, assess haplotype distribution, or explore their evolutionary histories in any detail. To investigate these underappreciated questions, we compiled the largest and most geographically representative leopard data set of the mitochondrial NADH-5 gene to date. We combined samples (n = 33) collected in an altitudinal transect across the Mpumalanga province of South Africa, where two populations of leopard are known to be in genetic contact, with previously published sequences of African leopard (n = 211). We estimate that the maternal PAR-I and PAR-II lineages diverged approximately 0.7051 (0.4477–0.9632) million years ago (Ma). Through spatial and demographic analyses, we show that while PAR-I underwent a mid-Pleistocene population expansion resulting in several closely related haplotypes with little geographic structure across much of its range, PAR-II remained at constant size and may even have declined slightly in the last 0.1 Ma. The higher genetic drift experienced within PAR-II drove a greater degree of structure with little haplotype sharing and unique haplotypes in central Africa, the Cape, KwaZulu-Natal and the South African Highveld. The phylogeographic structure of PAR-II, with its increasing frequency southward and its exclusive occurrence in south-eastern South Africa, suggests that this lineage may have been isolated in South Africa during the mid-Pleistocene. This hypothesis is supported by historical changes in paleoclimate that promoted intense aridification around the Limpopo Basin between 1.0–0.6 Ma, potentially reducing gene flow and promoting genetic drift. Interestingly, we ascertained that the two nuclear DNA populations identified by a previous study as East and West Mpumalanga correspond to PAR-I and PAR-II, respectively, and that they have come into secondary contact in the Lowveld region of South Africa. Our results suggest a subdivision of African leopard mtDNA into two clades, with one occurring almost exclusively in South Africa, and we identify the potential environmental drivers of this observed structure. We caution that our results are based on a single mtDNA locus, but it nevertheless provides a hypothesis that can be further tested with a dense sample of nuclear DNA data, preferably whole genomes. If our interpretation holds true, it would provide the first genetic explanation for the smaller observed size of leopards at the southernmost end of their range in Africa.

Torpor is widespread among bats presumably because most species are small, and torpor greatly reduces their high mass-specific resting energy expenditure, especially in the cold. Torpor has not been recorded in any bat species larger than 50 g, yet in theory could be beneficial even in the world’s largest bats (flying-foxes; Pteropus spp.) that are exposed to adverse environmental conditions causing energy bottlenecks. We used temperature telemetry to measure body temperature in wild-living adult male grey-headed flying-foxes ( P. poliocephalus ; 799 g) during winter in southern Australia. We found that all individuals used torpor while day-roosting, with minimum body temperature reaching 27°C. Torpor was recorded following a period of cool, wet and windy weather, and on a day with the coldest maximum air temperature, suggesting it is an adaptation to reduce energy expenditure during periods of increased thermoregulatory costs and depleted body energy stores. A capacity for torpor among flying-foxes has implications for understanding their distribution, behavioural ecology and life history. Furthermore, our discovery increases the body mass of bats known to use torpor by more than tenfold and extends the documented use of this energy-saving strategy under wild conditions to all bat superfamilies, with implications for the evolutionary maintenance of torpor among bats and other mammals.

Enteropathogenic Escherichia coli (EPEC) is an important cause of diarrhoeal disease in human infants. EPEC strains are defined by the presence of specific virulence factors including intimin (encoded by the eae gene) and bundle forming pili (Bfp). Bfp is encoded by the bfp operon and includes the bfpA gene for the major pilus subunit. By definition, Bfp are only present in typical EPEC (tEPEC), for which, humans are considered to be the only known natural host. This study detected tEPEC in faecal samples from a wild Australian fruit bat species, the grey-headed flying-fox (Pteropus poliocephalus). Whole genome sequencing of 61 E. coli isolates from flying-foxes revealed that 21.3 % (95%CI: 13 %-33 %) were tEPEC. Phylogenetic analyses showed flying-fox tEPEC shared evolutionary lineages with human EPEC, but were predominantly novel sequence types (9 of 13) and typically harboured novel bfpA variants (11 of 13). HEp-2 cell adhesion assays showed adherence to human-derived epithelial cells by all 13 flying-fox tEPEC, indicating that they all carried functional Bfp. Using an EPEC-specific duplex PCR, it was determined that tEPEC comprised 17.4 % (95%CI: 13 %-22 %) of 270 flying-fox E. coli isolates. Furthermore, a tEPEC-specific multiplex PCR detected the eae and bfpA virulence genes in 18.0 % (95%CI: 8.0 %-33.7 %) of 506 flying-fox faecal DNA samples, with occurrences ranging from 1.3 % to 87.0 % across five geographic areas sampled over a four-year period. The identification of six novel tEPEC sequence types and five novel bfpA variants suggests flying-foxes carry bat-specific tEPEC lineages. However, their close relationship with human EPEC and functional Bfp, indicates that flying-fox tEPEC have zoonotic potential and that dissemination of flying-fox tEPEC into urban environments may pose a public health risk. The consistent detection of tEPEC in flying-foxes over extensive geographical and temporal scales indicates that both wild grey-headed flying-foxes and humans should be regarded as natural tEPEC hosts.

Growing reports of diverse antibiotic resistance genes in wildlife species around the world symbolises the extent of this global One Health issue. The health of wildlife is threatened by antimicrobial resistance in situations where wildlife species develop disease and require antibiotics. Chlamydial disease is a key threat for koalas in Australia, with infected koalas frequently entering wildlife hospitals and requiring antibiotic therapy, typically with chloramphenicol or doxycycline. This study investigated the occurrence and diversity of target chloramphenicol and doxycycline resistance genes (cat and tet respectively) in koala urogenital and faecal microbiomes. DNA was extracted from 394 urogenital swabs and 91 faecal swabs collected from koalas in mainland Australia and on Kangaroo Island (KI) located 14 km off the mainland, before (n = 145) and during (n = 340) the 2019-2020 wildfires. PCR screening and DNA sequencing determined 9.9% of samples (95%CI: 7.5% to 12.9%) carried cat and/or tet genes, with the highest frequency in fire-affected KI koalas (16.8%) and the lowest in wild KI koalas sampled prior to fires (6.5%). The diversity of cat and tet was greater in fire-affected koalas (seven variants detected), compared to pre-fire koalas (two variants detected). Fire-affected koalas in care that received antibiotics had a significantly higher proportion (p < 0.05) of cat and/or tet genes (37.5%) compared to koalas that did not receive antibiotics (9.8%). Of the cat and/or tet positive mainland koalas, 50.0% were Chlamydia-positive by qPCR test. Chloramphenicol and doxycycline resistance genes in koala microbiomes may contribute to negative treatment outcomes for koalas receiving anti-chlamydial antibiotics. Thus a secondary outcome of wildfires is increased risk of acquisition of cat and tet genes in fire-affected koalas that enter care, potentially exacerbating the already significant threat of chlamydial disease on Australia's koalas. This study highlights the importance of considering impacts to wildlife health within the One Health approach to AMR and identifies a need for greater understanding of AMR ecology in wildlife.

5.Ultimately, the illegal wildlife trade is minimised thus improving conservation and welfare outcomes.

The leopard (Panthera pardus) is facing the threat of continued population decline across its range. In order to inform more effective conservation management programs, genetic information is needed from leopard populations that persist in previously unstudied, isolated and highly fragmented protected areas. The aim of this study was to explore the population structure and genetic diversity of leopard populations across the Mpumalanga province of South Africa. We collected a total of 33 leopard samples from four major locations along a west to east transect across the province. We analysed 17 polymorphic microsatellites and two regions of the mitochondrial genome (NADH-5 and Cytb) to determine the genetic structure of the leopard population in the province. We also calculated genetic diversity indices and explored gene flow in the region. We found that while there is gene flow occurring across the province, the population was genetically structured. We identified two major population units that we describe as 'West Mpumalanga' and 'East Mpumalanga'. Gene flow was moderate between the two populations and we found very high genetic diversity levels compared to other leopard populations previously studied in South Africa. From a conservation perspective, our results show that gene flow is still occurring across seemingly isolated leopard populations that exist in fragmented landscapes, highlighting the importance of all leopard populations in South Africa. Management authorities need to focus conservation efforts on maintaining corridors between regions that are suitable for leopard occupancy and work closely with human settlements to minimise human-leopard conflicts.

The Grey-headed flying fox ( Pteropus poliocephalus ) is an endemic Australian fruit bat, known to carry pathogens with zoonotic potential. We recently showed these bats harbour the bacterial pathogens Klebsiella pneumoniae and closely related species in the K. pneumoniae species complex ( Kp SC). However, the dynamics of Klebsiella transmission and gene flow within flying fox colonies were not explored and remain poorly understood. Here we report a high-resolution genomic comparison of 39 Kp SC isolates from Greyheaded flying foxes. Illumina whole genome sequences (n=39) were assembled de novo and the Kleborate genotyping tool was used to infer sequence types (STs). Oxford Nanopore sequences were generated for 13 isolates (one for each distinct ST) in order to generate high-quality completed reference genomes. Read mapping and variant calling was used to identify single nucleotide variants (SNVs) within each ST, using the relevant reference genome. In silico genome-scale metabolic models were generated to predict and compare substrate usage to 59 previously published Kp SC models for isolates from human and environmental sources, which indicated no distinction on the basis of metabolic capabilities. High-resolution genome comparisons identified five putative strain transmission clusters (four intra- and one inter-colony, n=2-15 isolates each, ≤25 pairwise SNVs). Inter-colony transmission of Klebsiella africana was found between two flying fox populations located within flying distance. The 13 completed genomes harboured 11 plasmids, all of which showed 37-98% coverage (mean 73%) and ≥95% identity to those previously reported from human-associated Kp SC. Comparison of plasmids from different flying fox associated Kp SC indicated an interspecies horizontal plasmid transmission between K. pneumoniae and K. africana for a 98 kbp plasmid, pFF1003. These data indicate that Kp SC are able to transmit directly via flying fox populations or indirectly via a common source, and that these isolates can harbour plasmids with similarity to those found in human derived Kp SC, indicating gene flow is occurring between isolates from Grey-headed flying fox Kp SC and human clinical isolates.

We describe our experiences collecting blood from Australian Rattus. We found uniform anatomy of the external jugular vein between Australian and exotic Rattus species. Understanding where the maxillary and linguofacial veins join to form the external jugular vein is critical to venepuncture. After locating this union, we consistently achieved successful venepuncture of the external jugular vein; yielding large blood volumes. All other routes of venepuncture yielded minute blood volumes or were unsuccessful. We provide recommendations for venepuncture in Australian rodents and encourage others to share their experiences; such reports facilitate sampling rarely sampled species and promote animal welfare.

Bats are important reservoirs for viruses of public health and veterinary concern. Virus studies in Australian bats usually target the families Paramyxoviridae, Coronaviridae and Rhabdoviridae, with little known about their overall virome composition. We used metatranscriptomic sequencing to characterise the faecal virome of grey-headed flying foxes from three colonies in urban/suburban locations from two Australian states. We identified viruses from three mammalian-infecting (Coronaviridae, Caliciviridae, Retroviridae) and one possible mammalian-infecting (Birnaviridae) family. Of particular interest were a novel bat betacoronavirus (subgenus Nobecovirus) and a novel bat sapovirus (Caliciviridae), the first identified in Australian bats, as well as a potentially exogenous retrovirus. The novel betacoronavirus was detected in two sampling locations 1,375 km apart and falls in a viral lineage likely with a long association with bats. This study highlights the utility of unbiased sequencing of faecal samples for identifying novel viruses and revealing broad-scale patterns of virus ecology and evolution.

1. Types of biological evidence for DNA analyses for species identification 2. Stable isotopes 3.

Urban-living wildlife can be exposed to metal contaminants dispersed into the environment through industrial, residential, and agricultural applications. Metal exposure carries lethal and sublethal consequences for animals; in particular, heavy metals (e.g. arsenic, lead, mercury) can damage organs and act as carcinogens. Many bat species reside and forage in human-modified habitats and could be exposed to contaminants in air, water, and food. We quantified metal concentrations in fur samples from three flying fox species (Pteropus fruit bats) captured at eight sites in eastern Australia. For subsets of bats, we assessed ectoparasite burden, haemoparasite infection, and viral infection, and performed white blood cell differential counts. We examined relationships among metal concentrations, environmental predictors (season, land use surrounding capture site), and individual predictors (species, sex, age, body condition, parasitism, neutrophil:lymphocyte ratio). As expected, bats captured at sites with greater human impact had higher metal loads. At one site with seasonal sampling, bats had higher metal concentrations in winter than in summer, possibly owing to changes in food availability and foraging. Relationships between ectoparasites and metal concentrations were mixed, suggesting multiple causal mechanisms. There was no association between overall metal load and neutrophil:lymphocyte ratio, but mercury concentrations were positively correlated with this ratio, which is associated with stress in other vertebrate taxa. Comparison of our findings to those of previous flying fox studies revealed potentially harmful levels of several metals; in particular, endangered spectacled flying foxes (P. conspicillatus) exhibited high concentrations of cadmium and lead. Because some bats harbor pathogens transmissible to humans and animals, future research should explore interactions between metal exposure, immunity, and infection to assess consequences for bat and human health.