Melbourne Veterinary School - Research Publications

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    Antimicrobial Prescribing in Dogs and Cats in Australia: Results of the Australasian Infectious Disease Advisory Panel Survey
    Hardefeldt, LY ; Holloway, S ; Trott, DJ ; Shipstone, M ; Barrs, VR ; Malik, R ; Burrows, M ; Armstrong, S ; Browning, GF ; Stevenson, M (WILEY, 2017-07-01)
    BACKGROUND: Investigations of antimicrobial use in companion animals are limited. With the growing recognition of the need for improved antimicrobial stewardship, there is urgent need for more detailed understanding of the patterns of antimicrobial use in this sector. OBJECTIVES: To investigate antimicrobial use for medical and surgical conditions in dogs and cats by Australian veterinarians. METHODS: A cross-sectional study was performed over 4 months in 2011. Respondents were asked about their choices of antimicrobials for empirical therapy of diseases in dogs and cats, duration of therapy, and selection based on culture and susceptibility testing, for common conditions framed as case scenarios: 11 medical, 2 surgical, and 8 dermatological. RESULTS: A total of 892 of the 1,029 members of the Australian veterinary profession that completed the survey satisfied the selection criteria. Empirical antimicrobial therapy was more common for acute conditions (76%) than chronic conditions (24%). Overall, the most common antimicrobial classes were potentiated aminopenicillins (36%), fluoroquinolones (15%), first- and second-generation cephalosporins (14%), and tetracyclines (11%). Third-generation cephalosporins were more frequently used in cats (16%) compared to dogs (2%). Agreement with Australasian Infectious Disease Advisory Panel (AIDAP) guidelines (generated subsequently) was variable ranging from 0 to 69% between conditions. CONCLUSIONS AND CLINICAL IMPORTANCE: Choice of antimicrobials by Australian veterinary practitioners was generally appropriate, with relatively low use of drugs of high importance, except for the empirical use of fluoroquinolones in dogs, particularly for otitis externa and 3rd-generation cephalosporins in cats. Future surveys will determine whether introduction of the 2013 AIDAP therapeutic guidelines has influenced prescribing habits.
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    Exploration of antibiotic resistance risks in a veterinary teaching hospital with Oxford Nanopore long read sequencing
    Kamathewatta, KI ; Bushell, RN ; Young, ND ; Stevenson, MA ; Billman-Jacobe, H ; Browning, GF ; Marenda, MS ; Luo, Y (PUBLIC LIBRARY SCIENCE, 2019-05-30)
    The Oxford Nanopore MinION DNA sequencing device can produce large amounts of long sequences, typically several kilobases, within a few hours. This long read capacity was exploited to detect antimicrobial resistance genes (ARGs) in a large veterinary teaching hospital environment, and to assess their taxonomic origin, genetic organisation and association with mobilisation markers concurrently. Samples were collected on eight occasions between November 2016 and May 2017 (inclusive) in a longitudinal study. Nanopore sequencing was performed on total DNA extracted from the samples after a minimal enrichment step in broth. Many ARGs present in the veterinary hospital environment could potentially confer resistance to antimicrobials widely used in treating infections of companion animals, including aminoglycosides, extended-spectrum beta-lactams, sulphonamides, macrolides, and tetracyclines. High-risk ARGs, defined here as single or multiple ARGs associated with pathogenic bacterial species or with mobile genetic elements, were shared between the intensive care unit (ICU) patient cages, a dedicated laundry trolley and a floor cleaning mop-bucket. By contrast, a floor surface from an office corridor without animal contact and located outside the veterinary hospital did not contain such high-risk ARGs. Relative abundances of high-risk ARGs and co-localisation of these genes on the same sequence read were higher in the laundry trolley and mop bucket samples, compared to the ICU cages, suggesting that amplification of ARGs is likely to occur in the collection points for hospital waste. These findings have prompted the implementation of targeted intervention measures in the veterinary hospital to mitigate the risks of transferring clinically important ARGs between sites and to improve biosecurity practices in the facility.
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    Determination of the minimum protective dose of a glycoprotein-G-deficient infectious laryngotracheitis virus vaccine delivered via eye-drop to week-old chickens
    Korsa, MG ; Devlin, JM ; Hartley, CA ; Browning, GF ; Coppo, MJC ; Quinteros, JA ; Loncoman, CA ; Onasanya, AE ; Thilakarathne, D ; Diaz-Mendez, A ; Cao, Y (PUBLIC LIBRARY SCIENCE, 2018-12-06)
    Infectious laryngotracheitis (ILT) is an upper respiratory tract disease of chickens that is caused by infectious laryngotracheitis virus (ILTV), an alphaherpesvirus. This disease causes significant economic loses in poultry industries worldwide. Despite widespread use of commercial live attenuated vaccines, many poultry industries continue to experience outbreaks of disease caused by ILTV. Efforts to improve the control of this disease have resulted in the generation of new vaccine candidates, including ILTV mutants deficient in virulence factors. A glycoprotein G deletion mutant vaccine strain of ILTV (ΔgG ILTV), recently licenced as Vaxsafe ILT (Bioproperties Pty Ltd), has been extensively characterised in vitro and in vivo, but the minimum effective dose required to protect inoculated animals has not been determined. This study performed a vaccination and challenge experiment to determine the minimum dose of ΔgG ILTV that, when delivered by eye-drop to seven-day-old specific pathogen-free chickens, would protect the birds from a robust challenge with a virulent field strain of virus (class 9 ILTV). A dose of 10(3.8) plaque forming units was the lowest dose capable of providing a high level of protection against challenge, as measured by clinical signs of disease, tracheal pathology and virus replication after challenge. This study has shown that the ΔgG ILTV vaccine strain is capable of inducing a high level of protection against a virulent field virus at a commercially feasible dose. These results lay the foundations upon which a commercial vaccine can be developed, thereby offering the potential to provide producers with another important tool to help control ILTV.
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    Veterinary Students' Knowledge and Perceptions About Antimicrobial Stewardship and Biosecurity-A National Survey
    Hardefeldt, L ; Nielsen, T ; Crabb, H ; Gilkerson, J ; Squires, R ; Heller, J ; Sharp, C ; Cobbold, R ; Norris, J ; Browning, G (MDPI, 2018-06-01)
    A better understanding of veterinary students’ perceptions, attitudes, and knowledge about antimicrobial stewardship and biosecurity could facilitate more effective education of future veterinarians about these important issues. A multicenter cross-sectional study was performed by administering a questionnaire to veterinary students expected to graduate in 2017 or 2018 in all Australian veterinary schools. Four hundred and seventy-six of 1246 students (38%) completed the survey. Many students were unaware of the high importance of some veterinary drugs to human medicine, specifically enrofloxacin and cefovecin (59% and 47% of responses, respectively). Fewer than 10% of students would use appropriate personal protective equipment in scenarios suggestive of Q fever or psittacosis. Students expected to graduate in 2018 were more likely to select culture and susceptibility testing in companion animal cases (OR 1.89, 95% CI 1.33–2.69, p < 0.001), and were more likely to appropriately avoid antimicrobials in large animal cases (OR 1.75, 95% CI 1.26–2.44, p = 0.001) than those expected to graduate in 2017. However, 2018 graduates were less likely to correctly identify the importance rating of veterinary antimicrobials for human health (OR 0.48, 95% CI 0.34–0.67, p < 0.001) than 2017 graduates. Students reported having a good knowledge of antimicrobial resistance, and combating resistance, but only 34% thought pharmacology teaching was adequate and only 20% said that teaching in lectures matched clinical teaching. Efforts need to be made to harmonize preclinical and clinical teaching, and greater emphasis is needed on appropriate biosecurity and antimicrobial stewardship.
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    The performance of three immune assays to assess the serological status of cattle experimentally exposed to Mycoplasma bovis
    Schibrowski, ML ; Barnes, TS ; Wawegama, NK ; Vance, ME ; Markham, PF ; Mansell, PD ; Marenda, MS ; Kanci, A ; Perez-Casal, J ; Browning, GF ; Gibson, JS ; Mahony, TJ (MDPI, 2018-03-08)
    © 2018 by the authors. Mycoplasma bovis is associated with several clinical syndromes of cattle. Currently, limited information is available on the sensitivity (Se) and specificity (Sp) of serological assays used for the detection of M. bovis-specific antibodies. Consequently, it is difficult to critically evaluate the outcomes of studies that use these assays. Therefore, the current study used bovine sera sourced from M. bovis exposure studies from three countries to estimate the Se and Sp of two commercial M. bovis enzyme-linked immunosorbent assays (ELISA), BIO K302 and BIO K260, and Western blotting. Western blotting had the highest Se estimate of 74% (95% confidence interval (CI): 16-98%), compared to the BIO K302: 47% (95% CI: 10-87%) and BIO K260: 28% (95% CI: 1-92%). However, for Sp, the BIO K302: 96% (95% CI: 87-99%) and the BIO K260: 100% (95% CI: 93-100%) out-performed Western blotting: 88% (95% CI: 56-98%). Western blotting was the best assay for detecting seroconversion, correctly identifying 61% (95% CI: 29-86%) of exposed animals compared to 35% for BIO K302 (95% CI: 21-54%) and 8% for BIO K260 (95% CI: 0-87%). While none of the methods assessed had high Se and Sp, the availability of these estimates will aid in the interpretation of studies that use these assays. The results of this study highlight the difficulties encountered when using serology to detect exposure to M. bovis in cattle.
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    Reproduction of respiratory mycoplasmosis in calves by exposure to an aerosolised culture of Mycoplasma bovis
    Kanci, A ; Wawegama, NK ; Marenda, MS ; Mansell, PD ; Browning, GF ; Markham, PF (Elsevier, 2017-10-01)
    © 2017 Mycoplasma bovis is an important pathogen of cattle, causing pneumonia, arthritis and otitis media in young calves, and mastitis in lactating cows, resulting in increased morbidity and, in some instances, mortality. The objective of this study was to evaluate the survival of a M. bovis isolate following nebulisation and to establish whether respiratory disease similar to that seen in the field could be induced in calves by exposing them to an aerosolised culture of M. bovis. A group of eight M. bovis-free calves 14–28 days old were exposed to an aerosolised culture of a field isolate of M. bovis that had originally been recovered from a joint lesion in a calf. Three weeks after aerosol exposure necropsies were conducted on all calves. Lung lesions were seen in 7 of 8 calves exposed to the aerosol of M. bovis, whilst calves exposed to the culture medium alone did not develop lesions. Two calves in the infected group had detectable concentrations of serum antibody against M. bovis on day 7 post infection and 4 calves had detectable concentrations of serum antibody against M. bovis on day 21 post infection when tested by MilA IgG ELISA. M. bovis was reisolated from the upper trachea of 6 of the 8 infected calves. The infection method described here appeared to induce lung lesions typical of naturally occurring disease associated with infection with M. bovis and should be applicable to testing the safety and efficacy of attenuated vaccine candidates to control disease caused by this pathogen.
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    Cross-sectional study of antimicrobials used for surgical prophylaxis by bovine veterinary practitioners in Australia
    Hardefeldt, LY ; Browning, GF ; Thursky, KA ; Gilkerson, JR ; Billman-Jacobe, H ; Stevenson, MA ; Bailey, KE (BMJ PUBLISHING GROUP, 2017-10-01)
    Antimicrobials are widely used in veterinary practices, but there has been no investigation of antimicrobial classes used or the appropriateness of their use in bovine practice. This study investigated antimicrobial use for surgical prophylaxis in bovine practice in Australia. A cross-sectional study of veterinarian antimicrobial usage patterns was conducted using an online questionnaire. Information solicited included respondent's details, the frequency with which antimicrobials were used for specific surgical conditions (including the dose, timing and duration of therapy) and details of practice antimicrobial use policies and sources of information about antimicrobials. In total, 212 members of the Australian veterinary profession working in bovine practice completed the survey. Antimicrobials were always or frequently used by more than 75 per cent of respondents in all scenarios. Generally, antimicrobial drug choice was appropriate for the reported surgical conditions. Procaine penicillin and oxytetracycline accounted for 93 per cent of use. However, there was a wide range of doses used, with underdosing and inappropriate timing of administration being common reasons for inappropriate prophylactic treatment. There was very low use of critically important antimicrobials (3.3 per cent of antimicrobials reported). Antimicrobial use guidelines need to be developed and promoted to improve the responsible use of antimicrobials in bovine practice.
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    Natural recombination in alphaherpesviruses: Insights into viral evolution through full genome sequencing and sequence analysis
    Loncoman, CA ; Vaz, PK ; Coppo, MJC ; Hartley, CA ; Morera, FJ ; Browning, GF ; Devlin, JM (ELSEVIER SCIENCE BV, 2017-04-01)
    Recombination in alphaherpesviruses was first described more than sixty years ago. Since then, different techniques have been used to detect recombination in natural (field) and experimental settings. Over the last ten years, next-generation sequencing (NGS) technologies and bioinformatic analyses have greatly increased the accuracy of recombination detection, particularly in field settings, thus contributing greatly to the study of natural alphaherpesvirus recombination in both human and veterinary medicine. Such studies have highlighted the important role that natural recombination plays in the evolution of many alphaherpesviruses. These studies have also shown that recombination can be a safety concern for attenuated alphaherpesvirus vaccines, particularly in veterinary medicine where such vaccines are used extensively, but also potentially in human medicine where attenuated varicella zoster virus vaccines are in use. This review focuses on the contributions that NGS and sequence analysis have made over the last ten years to our understanding of recombination in mammalian and avian alphaherpesviruses, with particular focus on attenuated live vaccine use.
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    A high prevalence of beak and feather disease virus in non-psittacine Australian birds
    Amery-Gale, J ; Marenda, MS ; Owens, J ; Eden, PA ; Browning, GF ; Devlin, JM (MICROBIOLOGY SOC, 2017-07-01)
    PURPOSE: Beak and feather disease virus (BFDV) is a circovirus and the cause of psittacine beak and feather disease (PBFD). This disease is characterized by feather and beak deformities and is a recognized threat to endangered Psittaciformes (parrots and cockatoos). The role that non-psittacine birds may play as reservoirs of infection is unclear. This study aimed to begin addressing this gap in our knowledge of PBFD. METHODOLOGY: Liver samples were collected from birds presented to the Australian Wildlife Health Centre at Zoos Victoria's Healesville Sanctuary for veterinary care between December 2014 and December 2015, and tested for BFDV DNA using polymerase chain reaction coupled with sequencing and phylogenetic analyses.Results/Key findings. Overall BFDV was detected in 38.1 % of 210 birds. BFDV was detected at high prevalence (56.2 %) in psittacine birds, in the majority of cases without any observed clinical signs of PBFD. We also found that BFDV was more common in non-psittacine species than previously recognized, with BFDV detected at 20.0 % prevalence in the non-psittacine birds tested, including species with no clear ecological association with psittacines, and without showing any detectable clinical signs of BFDV infection. CONCLUSION: Further research to determine the infectivity and transmissibility of BFDV in non-psittacine species is indicated. Until such work is undertaken the findings from this study suggest that every bird should be considered a potential carrier of BFDV, regardless of species and clinical presentation. Veterinary clinics and wildlife rehabilitation facilities caring for birds that are susceptible to PBFD should reconsider biosecurity protocols aimed at controlling BFDV.
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    Comparative Metabolomics of Mycoplasma bovis and Mycoplasma gallisepticum Reveals Fundamental Differences in Active Metabolic Pathways and Suggests Novel Gene Annotations
    Masukagami, Y ; De Souza, DP ; Dayalan, S ; Bowen, C ; O'Callaghan, S ; Kouremenos, K ; Nijagal, B ; Tull, D ; Tivendale, KA ; Markham, PF ; McConville, MJ ; Browning, GF ; Sansom, FM ; Dorrestein, PC (AMER SOC MICROBIOLOGY, 2017-09-01)
    Mycoplasmas are simple, but successful parasites that have the smallest genome of any free-living cell and are thought to have a highly streamlined cellular metabolism. Here, we have undertaken a detailed metabolomic analysis of two species, Mycoplasma bovis and Mycoplasma gallisepticum, which cause economically important diseases in cattle and poultry, respectively. Untargeted gas chromatography-mass spectrometry and liquid chromatography-mass spectrometry analyses of mycoplasma metabolite extracts revealed significant differences in the steady-state levels of many metabolites in central carbon metabolism, while 13C stable isotope labeling studies revealed marked differences in carbon source utilization. These data were mapped onto in silico metabolic networks predicted from genome wide annotations. The analyses elucidated distinct differences, including a clear difference in glucose utilization, with a marked decrease in glucose uptake and glycolysis in M. bovis compared to M. gallisepticum, which may reflect differing host nutrient availabilities. The 13C-labeling patterns also revealed several functional metabolic pathways that were previously unannotated in these species, allowing us to assign putative enzyme functions to the products of a number of genes of unknown function, especially in M. bovis. This study demonstrates the considerable potential of metabolomic analyses to assist in characterizing significant differences in the metabolism of different bacterial species and in improving genome annotation. IMPORTANCE Mycoplasmas are pathogenic bacteria that cause serious chronic infections in production animals, resulting in considerable losses worldwide, as well as causing disease in humans. These bacteria have extremely reduced genomes and are thought to have limited metabolic flexibility, even though they are highly successful persistent parasites in a diverse number of species. The extent to which different Mycoplasma species are capable of catabolizing host carbon sources and nutrients, or synthesizing essential metabolites, remains poorly defined. We have used advanced metabolomic techniques to identify metabolic pathways that are active in two species of Mycoplasma that infect distinct hosts (poultry and cattle). We show that these species exhibit marked differences in metabolite steady-state levels and carbon source utilization. This information has been used to functionally characterize previously unknown genes in the genomes of these pathogens. These species-specific differences are likely to reflect important differences in host nutrient levels and pathogenic mechanisms.