Field immobilization using alfaxalone and alfaxalone-medetomidine in free-ranging koalas (Phascolarctos cinereus): a randomized comparative study.

OBJECTIVE: To characterize and compare two intramuscular drug protocols using alfaxalone and alfaxalone-medetomidine combination for the field immobilization of free-ranging koalas. STUDY DESIGN: Blinded, randomized, comparative field study. ANIMALS: A total of 66 free-ranging koalas from the Mount Lofty Ranges, South Australia. METHODS: Koalas were randomly allocated into two groups. Group A animals were given alfaxalone alone at 3.5 mg kg-1. Group AM animals were given alfaxalone 2 mg kg-1 and medetomidine 40 µg kg-1, reversed with atipamezole at 0.16 mg kg-1. Blinded operators recorded heart rate (HR), respiratory rate (fR), cloacal temperature, depth of sedation and times to: first effect, sedation suitable for clinical interventions, first arousal and full recovery. Data were analysed using independent t test, Mann-Whitney U test, chi-square analysis and log-rank test at 5% level of significance. RESULTS: Suitable immobilization for clinical examination and sample collection was achieved in all animals. In groups A and AM, median time to working depth was 6.5 minutes (range: 3.4-15) and 8.1 minutes (range: 4.3-24) and time to complete recovery was 66 minutes (range: 12-138) and 34 minutes (range: 4-84), respectively, following reversal. Time to first effect was significantly shorter in group A (p = 0.013), whereas time to full arousal was significantly shorter in group AM (p = 0.007) probably due to the administration of atipamezole. Maximum HR was 117 ± 28 beats minute-1 in group A, which was a significant increase from baseline values (p < 0.0001), whereas group AM showed a significant tachypnoea of 67 ± 25 (normal fR 10-15; p < 0.0001). CONCLUSIONS AND CLINICAL RELEVANCE: Both the protocols produced immobilization, enabling clinical examination and sample collection; however, protocol AM was more suitable for field work due to shorter recovery times.

SERUM BIOCHEMISTRY OF FREE-RANGING SOUTHERN HAIRY-NOSED WOMBATS (LASIORHINUS LATIFRONS).

Listed as near-threatened by the International Union for Conservation of Nature (IUCN), the southern hairy-nosed wombat (SHNW, Lasiorhinus latifrons) faces threats such as drought, habitat degradation and loss, disease, and persecution because of competition with agriculture. To assist with evaluation of wombat health, this study reports serum biochemical reference intervals (RIs) for wild-caught SHNW from South Australia established from 126 apparently healthy SHNW using a Beckman Coulter AU480® Automated Chemistry Analyzer using RefVal Advisor. Partitioning of RIs for male and female wombats and for the two methods of sampling was performed as appropriate, and additional significant differences (P < 0.05) in biochemical profiles were identified across age class and season examined. A number of differences were observed between male and female wombats, most notably higher creatinine, urea, and sodium in females. Subadult and juvenile wombats had significantly lower total protein, globulin, and ALT activity, and significantly higher ALP activity than adults. Wombats sampled in winter and spring had significantly higher total protein, albumin, potassium, bicarbonate, and enzyme activities (ALP, ALT, AST, GGT, GLDH, lipase), and significantly lower glucose and creatinine when compared to individuals sampled in summer and autumn. Differences in CK activity and anion gap observed between the two methods of sampling likely reflect delay and handling of animals between capture and blood collection. The serum biochemical RIs documented here are considered representative of a population of healthy SHNW, providing a tool for health assessment and monitoring of SHNW health in South Australia and elsewhere.

Diagnostic test accuracy when screening for Haliotid herpesvirus 1 (AbHV) in apparently healthy populations of Australian abalone Haliotis spp.

The accuracy of 3 real-time PCR assays (ORF49, ORF66 and ORF77) and histopathology was evaluated for the purpose of demonstrating or certifying abalone free from Haliotid herpesvirus 1 (AbHV), the causative agent of abalone viral ganglioneuritis. Analytically, all 3 qPCRs showed equivalent limit of detection (20 copies per reaction); however, ORF49 could not detect 2 of the AbHV genotypes. A selection of 1452 archive specimens sourced from apparently healthy abalone populations was screened using all 4 tests. In the absence of a perfect reference standard, a Bayesian latent class analysis was built to estimate diagnostic sensitivity (DSe), diagnostic specificity (DSp) and likelihood ratios of a positive (LR+) and negative test result (LR-) for each individual test and for all possible combinations of test pairs interpreted either in series or in parallel. The pair ORF49/ORF66 interpreted in parallel performed the best both analytically and diagnostically to demonstrate freedom from AbHV in an established population of abalone and to certify individual abalone free from AbHV for trade or movement purposes (DSe = 96.0%, 95% posterior credibility interval [PCI]: 82.6 to 99.9; DSp = 97.7%, 95% PCI: 96.4 to 99.4; LR+ = 41.4, 95% PCI: 27.4 to 148.7; LR- = 0.041, 95% PCI: 0.001 to 0.176). Histopathology showed very poor DSe (DSe = 6.3%, 95% PCI: 2.4 to 13.1) as expected since most infected abalone in the study were likely sub-clinical with limited pathological change. Nevertheless, we recommend histopathology when clinically investigating outbreaks to find potential, new, emerging AbHV genotype(s) that may not be detectable by either ORF49 or ORF66.

Recommended reporting standards for test accuracy studies of infectious diseases of finfish, amphibians, molluscs and crustaceans: the STRADAS-aquatic checklist.

Complete and transparent reporting of key elements of diagnostic accuracy studies for infectious diseases in cultured and wild aquatic animals benefits end-users of these tests, enabling the rational design of surveillance programs, the assessment of test results from clinical cases and comparisons of diagnostic test performance. Based on deficiencies in the Standards for Reporting of Diagnostic Accuracy (STARD) guidelines identified in a prior finfish study (Gardner et al. 2014), we adapted the Standards for Reporting of Animal Diagnostic Accuracy Studies-paratuberculosis (STRADAS-paraTB) checklist of 25 reporting items to increase their relevance to finfish, amphibians, molluscs, and crustaceans and provided examples and explanations for each item. The checklist, known as STRADAS-aquatic, was developed and refined by an expert group of 14 transdisciplinary scientists with experience in test evaluation studies using field and experimental samples, in operation of reference laboratories for aquatic animal pathogens, and in development of international aquatic animal health policy. The main changes to the STRADAS-paraTB checklist were to nomenclature related to the species, the addition of guidelines for experimental challenge studies, and the designation of some items as relevant only to experimental studies and ante-mortem tests. We believe that adoption of these guidelines will improve reporting of primary studies of test accuracy for aquatic animal diseases and facilitate assessment of their fitness-for-purpose. Given the importance of diagnostic tests to underpin the Sanitary and Phytosanitary agreement of the World Trade Organization, the principles outlined in this paper should be applied to other World Organisation for Animal Health (OIE)-relevant species.

EVALUATION OF ETORPHINE AND MIDAZOLAM ANESTHESIA, AND THE EFFECT OF INTRAVENOUS BUTORPHANOL ON CARDIOPULMONARY PARAMETERS IN GAME-RANCHED WHITE RHINOCEROSES (CERATOTHERIUM SIMUM).

Nineteen white rhinoceroses ( Ceratotherium simum ) were anesthetized with 4 mg of etorphine hydrochloride; 35-40 mg of midazolam; and 7,500 international units of hyaluronidase for dehorning purposes at a game ranch in South Africa, to investigate this anesthetic combination. Median time to recumbency was 548 sec (range 361-787 sec). Good muscle relaxation and no muscle rigidity or tremors were observed in 18 animals, and only 1 individual showed slight tremors. In addition, all animals received butorphanol i.v. 5 min after recumbency at the ratio of 10 mg of butorphanol per 1 mg of etorphine. Blood gas and selected physiologic parameters were measured in the recumbent animal, immediately before and 10 min after the administration of butorphanol. Statistically significant improvements were observed in blood gas physiologic and cardiopulmonary parameters 10 min after the administration of butorphanol, with a reduction in arterial partial pressure of carbon dioxide, systolic blood pressure, and heart rate and an increase in pH, arterial partial pressure of oxygen, oxygen saturation, and respiratory rate (all P < 0.005). After i.v. naltrexone reversal, recovery was uneventful, and median time to walking or running was 110 sec (range 71-247 sec). The results indicate etorphine and midazolam combination is an effective alternative anesthetic protocol and produces good muscle relaxation. Furthermore, i.v. butorphanol was associated with improved blood gas values and cardiopulmonary function for at least 10 min postinjection.

Association between indoor environmental contamination by Salmonella enterica and contamination of eggs on layer farms.

This study involves longitudinal and point-in-time surveys of Salmonella carriage and environmental contamination on two commercial cage layer farms positive for Salmonella enterica subsp. enterica serovar Typhimurium (flock A age, 32 weeks; flock B age, 34 weeks). Salmonella-positive fecal, egg belt, and dust samples were all unconditionally associated with eggshells testing positive for Salmonella. The odds of an eggshell testing positive for Salmonella were 91.8, 61.5, and 18.2 times higher when fecal, egg belt, and dust samples, respectively, tested positive for Salmonella. The agreement between the culture-based methods and real-time PCR on preenriched broths for detecting Salmonella was almost perfect for eggshell (observed agreement, 99.19%; kappa coefficient, 0.94) and egg belt samples (observed agreement, 95%; kappa coefficient, 0.88), and it was substantial for fecal (observed agreement, 87.14%; kappa coefficient, 0.47) and floor dust samples (observed agreement, 80.61%; kappa coefficient, 0.58). A 1-log increase in the load of Salmonella detected in the fecal, egg belt, and floor dust samples resulted in 35%, 43%, and 45% increases, respectively (P < 0.001), in the odds of an eggshell testing positive for Salmonella. The multilocus variable-number tandem-repeat analysis (MLVA) patterns of the S. Typhimurium strains isolated from flock A were distinct from those of flock B. S. Typhimurium strains detected from human food poisoning cases exhibited an MLVA pattern similar to those of the strains isolated from flocks A and B.

Temperature-bounded development of Dirofilaria immitis larvae restricts the geographical distribution and seasonality of its transmission: case study and decision support system for canine heartworm management in Australia.

Dirofilaria immitis is the causative agent of canine heartworm disease. We used the established heartworm development unit (HDU) principle to map the extrinsic incubation period (EIP) of D. immitis in Australia using historical weather data from 2013-2022. We found weather conditions suitable for EIP completion showed substantial seasonality and geographical variability. Whilst a considerable percentage of the Australian territory showed suitable weather conditions to always support EIP completion (17%), only 2.7% of the 2021 Australian human population lived in this region. Therefore, 97% of the population lived in an area that changed its EIP suitability within the study period. EIP completion is required prior to D. immitis transmission, meaning that infection risk of D. immitis is seasonal and location-dependent, being disrupted each year for most of the human population's dogs. We developed an online, open access tool allowing us to visualise EIP completion across Australia historically and in near real-time. We aim to support veterinarians to make risk-based recommendations for dirofilariosis prevention by using the tool, available at https://heartworm-mapping.adelaide.edu.au/shiny/.

The two-step Fagan's nomogram: ad hoc interpretation of a diagnostic test result without calculation.

In 1975, Fagan published a nomogram to help practitioners determine, without the use of a calculator or computer, the probability of a patient truly having a condition of interest given a particular test result. Nomograms are very useful for bedside interpretations of test results, as no test is perfect. However, the practicality of Fagan's nomogram is limited by its use of the likelihood ratio (LR), a parameter not commonly reported in the evaluation studies of diagnostic tests. The LR reflects the direction and strength of evidence provided by a test result and can be computed from the conventional diagnostic sensitivity (DSe) and specificity (DSp) of the test. This initial computation is absent in Fagan's nomogram, making it impractical for routine use. We have seamlessly integrated the initial step to compute the LR and the resulting two-step nomogram allows the user to quickly interpret the outcome of a test. With the addition of the DSe and DSp, the nomogram, for the purposes of interpreting a dichotomous test result, is now complete. This tool is more accessible and flexible than the original, which will facilitate its use in routine evidence-based practice. The nomogram can be downloaded at: www.adelaide.edu.au/vetsci/research/pub_pop/2step-nomogram/.

Mitigating disease risk in an endangered pinniped: early hookworm elimination optimizes the growth and health of Australian sea lion pups.

The Australian sea lion (Neophoca cinerea) experiences high pup mortality of seasonally alternating severity, partly attributed to endemic hookworm (Uncinaria sanguinis) infection. To further explore health outcomes of early hookworm elimination, a treatment trial was conducted at Seal Bay Conservation Park, South Australia, over consecutive lower and higher mortality breeding seasons (2019, 19.2%; 2020-1; 28.9%). Pups (n = 322) were stratified into two age cohorts (median 14 d and 24 d recruitment ages) and randomly assigned to treated (topical ivermectin 500 µg/kg) or control (untreated) groups. A younger prepatent cohort <14 d old (median 10 d) was identified a posteriori. A seasonally independent growth benefit resulted from hookworm elimination across all age cohorts. The greatest relative improvements (bodyweight + 34.2%, standard length + 42.1%; p ≤ 0.001) occurred in the month post-treatment, in the youngest prepatent cohort. A significant benefit of lesser magnitude (bodyweight + 8.6-11.6%, standard length + 9.5-18.4%; p ≤ 0.033) persisted up to 3 months across all age cohorts - greatest in the youngest pups. Treatment resulted in immediate improvement in hematological measures of health - decreased anemia and inflammation severity (p ≤ 0.012). These results enhance our understanding of host-parasite-environment interactions within the context of hematological ontogenesis, confirm the seasonally independent benefits of hookworm disease intervention, and further inform conservation recommendations for this endangered species.

Investigation of Weather Triggers Preceding Outbreaks of Acute Bovine Liver Disease in Australia.

Acute bovine liver disease (ABLD) is a hepatic disease affecting cattle sporadically in southern Australia, characterised histologically by striking periportal hepatocellular necrosis. The cause of ABLD is unknown; however, the seasonality and acute presentation of outbreaks suggest mycotoxin involvement. We described the geographical and seasonal occurrence of ABLD reports from 2010 to 2020 in Victoria, Australia, and explored potential weather triggers preceding 26 outbreaks occurring across 23 properties using a case-crossover design. Outbreaks occurred most frequently in autumn/early winter and in herds located along the southern coastal plain of Victoria, and occasionally within the low-lying regions of the Great Dividing Range. Lactating adult dairy cattle represented the most reported cases. We observed a significant association between an increase in average daily dewpoint in the 15 days preceding an ABLD outbreak, suggesting that dew formation may be a key determinant for this disease. Our findings support the etiology of a potent hepatotoxic agent that requires moisture for proliferation and/or toxin production.

The impact of PIT tags on the growth and survival of pythons is insignificant in randomised controlled trial.

Individual identification is fundamental to the study of captive and wild animals but can have adverse impacts if the method of identification is inappropriate for the species or question of interest. We conducted a randomised controlled trial to test whether passive integrated transponder (PIT) tags reduced the growth or survival of pythons. We randomly allocated 200 captive-bred Burmese python (Python bivittatus) hatchlings into two groups, tagged versus untagged. Hatchlings were individually identified using a combination of PIT tags and unique colour patterns, and their mass, snout-vent length (SVL) and body condition measured at 9, 73, 134, 220, 292 and 385 days of age. We recorded the date of all mortalities. Python morphometrics and their rate of change increased or fluctuated non-linearly with age. The impact of PIT tagging on python body mass and body mass growth over the 376 day study period was insignificant. PIT tagging additionally had an insignificant impact on python survival. However, we found minor differences in SVL growth between tagged and untagged pythons. These differences peaked at approximately 0.5 mm/day and appeared to drive similar, but more pronounced, differences between tagged and untagged pythons in their rate of change in body condition; peaking at approximately 3-4 g/day. While we cannot be certain that these small differences are, or are not, biologically meaningful, they nonetheless appear to be short-term and readily resolved. Unsurprisingly, the strongest driver of python growth was their age, with growth rapidly increasing or highest amongst younger snakes for all measures of size. Python sex was associated with their body mass and survival, with higher mass but lower survival amongst females. Python size at hatching did not impact on their growth or survival. Our results confirm that PIT tags are a valuable and effective tool for the identification and tracking of captive pythons, and snakes generally, and meet high safety and animal welfare standards.

Topical ivermectin is a highly effective seal 'spot-on': A randomised trial of hookworm and lice treatment in the endangered Australian sea lion (Neophoca cinerea).

The Australian sea lion (Neophoca cinerea) is an endangered and declining otariid species, with a high rate of pup mortality associated with endemic hookworm (Uncinaria sanguinis) infection a suspected contributor to this decline. Injected ivermectin is an effective treatment for Uncinaria sp. in otariids, with optimal outcomes achieved by the early treatment of pups prior to disease development. This randomised controlled trial evaluated the effectiveness of the novel use of a topical ivermectin formulation against hookworm infection and lice (Antarctophthirus microchir) infestation, in comparison with injected ivermectin. During the 2017 breeding season at Dangerous Reef, South Australia, pups ≤ 70 cm in standard length (≤ 2 weeks of age; n = 85) were randomised to single dose topical (500 µg/kg spot-on; n = 27) or injected (200 µg/kg subcutaneous; n = 29) ivermectin treatment groups, or to an untreated control group (n = 29). Topical ivermectin was highly effective for U. sanguinis elimination, and not significantly different to the injected formulation (estimated effectiveness 96.4% and 96.8%, respectively; P > 0.05). Its application resulted in an 81.6% reduction and 62.7% additional clearance for A. microchir infestation by 15-24 days post-treatment, compared with untreated control pups (also not significantly different to injected ivermectin; 83.1% and 59.4%, respectively; P > 0.05). Treatment with either ivermectin formulation significantly ameliorated increases in inflammatory markers detected in the blood of untreated control pups - peripheral blood eosinophil counts (persisting to 36-41 days post-recruitment P < 0.05) and increased plasma protein concentrations (15-24 days post-recruitment; P < 0.05). Further, an initial short-term decrease in body condition in the control group was not observed in either of the treatment groups. This study demonstrates that topical ivermectin is an effective antiparasitic treatment in N. cinerea. It offers an alternative administration method for ivermectin delivery to a young pup cohort in this species, and an alternative, minimally invasive management tool for species conservation.

INFECTION PRESSURE IS NECESSARY, BUT NOT SUFFICIENT BY ITSELF, TO EXPLAIN TOXOPLASMA GONDII SEROPREVALENCE IN INTERMEDIATE HOST SPECIES.

Parasite infection pressure is suggested to be a strong driver of transmission within ecosystems. We tested if infection pressure drives seroprevalence in intermediate host species for Toxoplasma gondii. We defined Toxoplasma infection pressure to intermediate host species as the combined influence of cat abundance, environmental conditions, and its prevalence in the cat population. We sampled and tested 2 species of rodent and collated information on Toxoplasma seroprevalence in koalas, wallabies, kangaroos, and sheep. All species were sampled using equivalent methods, within a 2-yr period, and from adjacent regions of low and high Toxoplasma infection pressure. The seroprevalence of Toxoplasma in kangaroos scaled with infection pressure, but we observed no statistical difference in seroprevalence for any other species between these 2 regions. Within the region of low infection pressure, Toxoplasma seroprevalence did not differ between species. However, within the region of high Toxoplasma infection pressure, we observed large variation in seroprevalence between species. Our results demonstrate that infection pressure is not sufficient by itself, but merely necessary, to drive Toxoplasma seroprevalence in intermediate host species. Where Toxoplasma seroprevalence in an intermediate host species is already low, further reducing infection pressure will not necessarily further decrease seroprevalence in those species. This has important ramifications for the mitigation of parasite infections and suggests that reductions in Toxoplasma infection pressure, intended to reduce infections, may be most effective and applicable to species that are known to experience high rates of infection.

Serological evidence of exposure to a coronavirus antigenically related to severe acute respiratory syndrome virus (SARS-CoV-1) in the Grey-headed flying fox (Pteropus poliocephalus).

Many infectious pathogens can be transmitted by highly mobile species, like bats that can act as reservoir hosts for viruses such as henipaviruses, lyssaviruses and coronaviruses. In this study, we investigated the seroepidemiology of protein antigens to Severe acute respiratory syndrome virus (SARS-CoV-1) and Middle eastern respiratory syndrome virus (MERS-CoV) in Grey-headed flying foxes (Pteropus poliocephalus) in Adelaide, Australia sampled between September 2015 and February 2018. A total of 301 serum samples were collected and evaluated using a multiplex Luminex binding assay, and median fluorescence intensity thresholds were determined using finite-mixture modelling. We found evidence of antibodies reactive to SARS-CoV-1 or a related antigen with 42.5% (CI: 34.3%-51.2%) seroprevalence but insufficient evidence of reactivity to MERS-CoV antigen. This study provides evidence that the Grey-headed flying foxes sampled in Adelaide have been exposed to a SARS-like coronavirus.

Fractional seroprevalence rates in common prey species can cause more than half of feral cats to be exposed to Toxoplasma gondii annually.

Rodents comprise a major component of cat (Felis catus) diets in many ecosystems, and life cycle diagrams of Toxoplasma gondii typically depict small rodents as quintessential intermediate hosts. Counter-intuitively, small rodents often experience a lower T. gondii seroprevalence than do larger sympatric mammals. This observation has repeatedly caused confusion about the relative importance of small rodents to the ecology of T. gondii. To address this confusion, we modified the Reed-Frost epidemic model to develop a simple binomial equation to model T. gondii transmission from prey to feline predators. This equation takes into account variations in prey seroprevalence and the frequency with which they are consumed by felids. Even when T. gondii seroprevalence in prey is < 1%, computation reveals that the risk of feline exposure to T. gondii can easily exceed 50 % annually. For example, if cats eat an average of 1 mouse per day, a seroprevalence of 0.2 % (1/500) in mice will cause 51.9 % of cats to be exposed to T. gondii annually. Our simple equation demonstrates that both prey seroprevalence and the rate at which prey are consumed are of approximately equal importance to the ecology of T. gondii. When inferring the importance of various prey species to the ecology of T. gondii, researchers must consider the predation and dietary habits of felids from within their study system. Our simple binomial equation could also be used to predict T. gondii exposure rates of humans or other carnivorous animals from various dietary sources or be applied to other predator-prey parasite life cycles.

Investigation of risk factors associated with sub-optimal holding survival in southern rock lobster (Jasus edwardsii) in Australia.

Southern rock lobster (SRL1, Jasus edwardsii) are caught from the wild waters of southern Australia and form an important commodity economically for the fisheries industry in Australia. Between landing and export, SRL are held in specialised tanks within holding facilities for varying time periods before being exported to China. During the 2015-16 fishing season a lower stock survival rate was reported by some holding facilities when compared to previous fishing seasons. An investigation was undertaken to identify possible causes and favouring factors using a retrospective, single cohort, observational study. A custom questionnaire was built and trialled to collect qualitative and quantitative information on facilities infrastructure and capacity (10 questions), water systems (16 questions), SRL stock and health management (12 questions). Onsite visits and interviews of consenting holding facilities, with active holding operations, were conducted. Facilities were categorised into different capacity and management profiles based on a multivariate factor analysis. The association between facility profiles and perceived sub-optimal survival was then explored using simple logistic regression models. Out of the 83 license holders on record, 63 were in operation at the time of survey and 52 (83%) consented to participate. Perceived sub-optimal survival was reported in 22 (42%) facilities. The capacity, bio-filtration and water management practices across the surveyed facilities was highly variable but did not appear to be associated with survival. However, 'Intensive holding' facilities compared to the 'Extensive holding' facilities were significantly associated with an increased risk of sub-optimal survival (OR = 7.0, 95% CI: 2.1-26.13, P = 0.002). 'Intensive holding' facilities were more likely to hold higher annual tonnage sourced from distant, diverse and higher number of sources, to multiple handle and stock SRL in crates (as opposed to free swimming in tanks), and hold them for longer time periods. Holding practices are highly diverse across the SRL industry with little evidence of impact on survival, however, intensive and large scale holding practices appeared to be at higher risk of sub-optimal survival. A longitudinal and finer scale study is warranted to identify which one(s) of the stock management practices directly impact SRL survival during holding.

Seroprevalence of three paramyxoviruses; Hendra virus, Tioman virus, Cedar virus and a rhabdovirus, Australian bat lyssavirus, in a range expanding fruit bat, the Grey-headed flying fox (Pteropus poliocephalus).

Habitat-mediated global change is driving shifts in species' distributions which can alter the spatial risks associated with emerging zoonotic pathogens. Many emerging infectious pathogens are transmitted by highly mobile species, including bats, which can act as spill-over hosts for pathogenic viruses. Over three years, we investigated the seroepidemiology of paramyxoviruses and Australian bat lyssavirus in a range-expanding fruit bat, the Grey-headed flying fox (Pteropus poliocephalus), in a new camp in Adelaide, South Australia. Over six, biannual, sampling sessions, we quantified median florescent intensity (MFI) antibody levels for four viruses for a total of 297 individual bats using a multiplex Luminex binding assay. Where appropriate, florescence thresholds were determined using finite mixture modelling to classify bats' serological status. Overall, apparent seroprevalence of antibodies directed at Hendra, Cedar and Tioman virus antigens was 43.2%, 26.6% and 95.7%, respectively. We used hurdle models to explore correlates of seropositivity and antibody levels when seropositive. Increased body condition was significantly associated with Hendra seropositivity (Odds ratio = 3.67; p = 0.002) and Hendra virus levels were significantly higher in pregnant females (p = 0.002). While most bats were seropositive for Tioman virus, antibody levels for this virus were significantly higher in adults (p < 0.001). Unexpectedly, all sera were negative for Australian bat lyssavirus. Temporal variation in antibody levels suggests that antibodies to Hendra virus and Tioman virus may wax and wane on a seasonal basis. These findings suggest a common exposure to Hendra virus and other paramyxoviruses in this flying fox camp in South Australia.

Spatial Analysis of a Cat-Borne Disease Reveals That Soil pH and Clay Content Are Risk Factors for Sarcocystosis in Sheep.

Cat-borne parasites and their associated diseases have substantial impacts on human, livestock, and wildlife health worldwide. Despite this, large and detailed datasets that allow researchers to study broad-scale trends in the ecology of cat-borne diseases are either difficult to obtain or non-existent. One condition that is easily detected at slaughter is macroscopic sarcocystosis, a cat-borne parasitosis of sheep (Ovis aries). We conducted a cross-sectional study to describe the geographic distribution of sarcocystosis in sheep throughout South Australia and investigate ecosystem characteristics associated with the presence of disease. Data were obtained from two slaughterhouses which processed 3,865,608 sheep from 4,204 farms across 385,468 km2 of South Australia's land mass for the period 2007-2017. A Poisson point process model was developed to quantify environmental characteristics associated with higher densities of sarcocystosis-positive farms. Sarcocystosis was highly clustered on a large island off of the Australian coast and the density of sarcocystosis-positive farms increased in areas of low soil pH (intensity ratio: 0.86, 95% CI: 0.78, 0.95) and high clay content. We hypothesize that region was confounded by, and predominately acted as a proxy for, cat density. Our results have broader implications regarding the health, welfare, economic, and conservation impacts of other cat-borne parasitosis, such as toxoplasmosis.

Intra- and inter-laboratory agreement of the disc diffusion assay for assessing antimicrobial susceptibility of porcine Escherichia coli.

Reliable assessment of the susceptibility of animal bacterial pathogens to antimicrobials is of paramount importance in the fight against antimicrobial resistance. This work aims to estimate the repeatability (intra-laboratory agreement) and reproducibility (inter-laboratory agreement) of the disc diffusion assay in veterinary laboratories to understand further if the assay has a role in the surveillance of antimicrobial resistance in animals. Seven major veterinary laboratories from all States in Australia participated, and each tested the same panel of isolates five times at three to four-week intervals, against six antimicrobial agents using Clinical and Laboratory Standards Institute protocols. The panel consisted of twenty different isolates from porcine Escherichia coli from clinical cases and a single reference strain (ATCC 25922). Laboratories were blinded to the identity of the isolates, replicates, and to each other. In total, 4200 inhibition zone diameters (mm) were collected, and analysed descriptively, graphically, and with linear mixed models. Regardless of the laboratories and isolate/antimicrobial combinations, the overall very major error rate (proportion of isolates classified as susceptible when actual status is resistant) was 1.6%; the major error rate (proportion of isolates classified as resistant when actual status is susceptible) was 1.6%; and the 'minor error' rate (proportion of isolates with intermediate susceptibility that measure fully susceptible or resistant or vice versa) was 2.4%. The variation between repeated measurements ranged between 4.4-7.2 mm depending on the antimicrobial agent assessed. The reproducibility was always more variable than the repeatability, which suggested some laboratory effects. The repeatability coefficient of disc diffusion was lowest for tetracycline (4.4 mm, 95% CI: 3.8-5.0 mm) and ampicillin (4.6 mm, 95% CI: 4.2-5.2 mm) and highest for trimethoprim-sulfamethoxazole (6.6 mm, 95% CI: 5.9-7.4 mm). The reproducibility coefficient of disc diffusion was lowest for gentamicin (5.4, 95% CI: 4.0-7.2) and highest for trimethoprim-sulfamethoxazole (7.2 mm, 95%CI: 4.5-11.7 mm). The precision of the disc diffusion assay was deemed satisfactory for use in a national surveillance program for clinical porcine E. coli isolates. However, measurement variation of the disc diffusion assay is of concern for isolates with marginal susceptibility or resistance due to increased risk of misclassification.

Diagnostic accuracy of phenotypic assays for determining antimicrobial resistance status in Staphylococcus pseudintermedius isolates from canine clinical cases.

This study evaluated the diagnostic test accuracy of disc diffusion relative to broth-microdilution for clinical Staphylococcus pseudintermedius isolated from dogs in Australia (n = 614). Accuracy of disc diffusion and broth-microdilution for oxacillin relative to mecA real-time PCR was also assessed. Each isolate had paired minimum inhibitory concentration and zone diameter values for ten antimicrobial agents. Data was dichotomised using Clinical and Laboratory Standards Institute susceptible and resistant clinical breakpoints. Test accuracy was reported using relative diagnostic sensitivity (RSe), specificity (RSp), likelihood ratio pairs, diagnostic odds ratio, and area-under-the receiver-operating characteristic (ROC AUC) analysis. Disc diffusion was found to have high test accuracy for most antimicrobials (ROC AUC range: 0.96 - 0.99) except rifampicin (ROC AUC = 0.80). The RSp of disc diffusion was high for all antimicrobials (range, 97.1%-100%). However, RSe was considerably variable (range, 35.7%-98.8%), particularly for amoxicillin-clavulanic acid (51.5%, 95% CI, 38.9%, 64.0%), cefoxitin (35.7%, 95% CI, 12.8%, 64.9%), and cephalothin (43.6%, 95% CI, 27.8%, 60.4%). When disc diffusion and broth-microdilution were compared to mecA real-time PCR, the overall accuracy of both assays was similar (ROC AUC, 0.99 respectively). However, the RSe for broth-microdilution (96.1%, 95% CI, 88.9%, 99.2%) was significantly higher than for disc diffusion (86.8%, 95% CI, 77.1%, 93.5%) (McNemars mid-p value 0.01). Overall, these findings demonstrate that for most antimicrobials, disc diffusion performed according to CLSI guidelines can be used to differentiate clinical S. pseudintermedius isolates that might otherwise be assessed by broth-microdilution, provided consideration is given to the performance estimates reported here.