Veterinary Biosciences - Research Publications

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    Bayesian Validation of the Indirect Immunofluorescence Assay and Its Superiority to the Enzyme-Linked Immunosorbent Assay and the Complement Fixation Test for Detecting Antibodies against Coxiella burnetii in Goat Serum
    Muleme, M ; Stenos, J ; Vincent, G ; Campbell, A ; Graves, S ; Warner, S ; Devlin, JM ; Nguyen, C ; Stevenson, MA ; Wilks, CR ; Firestone, SM ; Pasetti, MF (AMER SOC MICROBIOLOGY, 2016-06)
    Although many studies have reported the indirect immunofluorescence assay (IFA) to be more sensitive in detection of antibodies to Coxiella burnetii than the complement fixation test (CFT), the diagnostic sensitivity (DSe) and diagnostic specificity (DSp) of the assay have not been previously established for use in ruminants. This study aimed to validate the IFA by describing the optimization, selection of cutoff titers, repeatability, and reliability as well as the DSe and DSp of the assay. Bayesian latent class analysis was used to estimate diagnostic specifications in comparison with the CFT and the enzyme-linked immunosorbent assay (ELISA). The optimal cutoff dilution for screening for IgG and IgM antibodies in goat serum using the IFA was estimated to be 1:160. The IFA had good repeatability (>96.9% for IgG, >78.0% for IgM), and there was almost perfect agreement (Cohen's kappa > 0.80 for IgG) between the readings reported by two technicians for samples tested for IgG antibodies. The IFA had a higher DSe (94.8%; 95% confidence interval [CI], 80.3, 99.6) for the detection of IgG antibodies against C. burnetii than the ELISA (70.1%; 95% CI, 52.7, 91.0) and the CFT (29.8%; 95% CI, 17.0, 44.8). All three tests were highly specific for goat IgG antibodies. The IFA also had a higher DSe (88.8%; 95% CI, 58.2, 99.5) for detection of IgM antibodies than the ELISA (71.7%; 95% CI, 46.3, 92.8). These results underscore the better suitability of the IFA than of the CFT and ELISA for detection of IgG and IgM antibodies in goat serum and possibly in serum from other ruminants.
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    Performance Evaluation and Validation of Air Samplers To Detect Aerosolized Coxiella burnetii
    Abeykoon, AMH ; Poon, M ; Firestone, SM ; Stevenson, MA ; Wiethoelter, AK ; Vincent, GA ; Uzal, F (AMER SOC MICROBIOLOGY, 2022-10-26)
    Coxiella burnetii, the etiological agent of Q fever, is an intracellular zoonotic pathogen transmitted via the respiratory route. Once released from infected animals, C. burnetii can travel long distances through air before infecting another host. As such, the ability to detect the presence of C. burnetii in air is important. In this study, three air samplers, AirPort MD8, BioSampler, and the Coriolis Micro, were assessed against a set of predetermined criteria in the presence of three different aerosolized C. burnetii concentrations. Two liquid collection media, phosphate-buffered saline (PBS) and alkaline polyethylene glycol (Alk PEG), were tested with devices requiring a collection liquid. Samples were tested by quantitative polymerase chain reaction assay (qPCR) targeting the single-copy com1 gene or multicopy insertion element IS1111. All air samplers performed well at detecting airborne C. burnetii across the range of concentrations tested. At high nebulized concentrations, AirPort MD8 showed higher, but variable, recovery probabilities. While the BioSampler and Coriolis Micro recovered C. burnetii at lower concentrations, the replicates were far more repeatable. At low and intermediate nebulized concentrations, results were comparable in the trials between air samplers, although the AirPort MD8 had consistently higher recovery probabilities. In this first study validating air samplers for their ability to detect aerosolized C. burnetii, we found that while all samplers performed well, not all samplers were equal. It is important that these results are further validated under field conditions. These findings will further inform efforts to detect airborne C. burnetii around known point sources of infection. IMPORTANCE Coxiella burnetii causes Q fever in humans and coxiellosis in animals. It is important to know if C. burnetii is present in the air around putative sources as it is transmitted via inhalation. This study assessed air samplers (AirPort MD8, BioSampler, and Coriolis Micro) for their efficacy in detecting C. burnetii. Our results show that all three devices could detect aerosolized bacteria effectively; however, at high concentrations the AirPort performed better than the other two devices, showing higher percent recovery. At intermediate and low concentrations AirPort detected at a level higher than or similar to that of other samplers. Quantification of samples was hindered by the limit of quantitation of the qPCR assay. Compared with the other two devices, the AirPort was easier to handle and clean in the field. Testing air around likely sources (e.g., farms, abattoirs, and livestock saleyards) using validated sampling devices will help better estimate the risk of Q fever to nearby communities.
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    Estimation of a Within-Herd Transmission Rate for African Swine Fever in Vietnam
    Le, VP ; Lan, NT ; Canevari, JT ; Villanueva-Cabezas, JP ; Padungtod, P ; Trinh, TBN ; Nguyen, VT ; Pfeiffer, CN ; Oberin, MV ; Firestone, SM ; Stevenson, MA (MDPI, 2023-02)
    We describe results from a panel study in which pigs from a 17-sow African swine fever (ASF) positive herd in Thái Bình province, Vietnam, were followed over time to record the date of onset of ASF signs and the date of death from ASF. Our objectives were to (1) fit a susceptible-exposed-infectious-removed disease model to the data with transmission coefficients estimated using approximate Bayesian computation; (2) provide commentary on how a model of this type might be used to provide decision support for disease control authorities. For the outbreak in this herd, the median of the average latent period was 10 days (95% HPD (highest posterior density interval): 2 to 19 days), and the median of the average duration of infectiousness was 3 days (95% HPD: 2 to 4 days). The estimated median for the transmission coefficient was 3.3 (95% HPD: 0.4 to 8.9) infectious contacts per ASF-infectious pig per day. The estimated median for the basic reproductive number, R0, was 10 (95% HPD: 1.1 to 30). Our estimates of the basic reproductive number R0 were greater than estimates of R0 for ASF reported previously. The results presented in this study may be used to estimate the number of pigs expected to be showing clinical signs at a given number of days following an estimated incursion date. This will allow sample size calculations, with or without adjustment to account for less than perfect sensitivity of clinical examination, to be used to determine the appropriate number of pigs to examine to detect at least one with the disease. A second use of the results of this study would be to inform the equation-based within-herd spread components of stochastic agent-based and hybrid simulation models of ASF.
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    A cross-sectional survey of risk factors for the presence of Coxiella burnetii in Australian commercial dairy goat farms
    Hou, KW ; Wiethoelter, AK ; Stevenson, MA ; Soares Magalhaes, RJ ; Lignereux, L ; Caraguel, C ; Stenos, J ; Vincent, G ; Aleri, JW ; Firestone, SM (WILEY, 2022-07)
    The largest Australian farm-based outbreak of Q fever originated from a dairy goat herd. We surveyed commercial dairy goat farms across Australia by testing bulk tank milk (BTM) samples using a commercial indirect enzyme-linked immunosorbent assay and two quantitative polymerase chain reactions (PCRs). Of the 66 commercial dairy goat herds on record, managers from 61 herds were contacted and 49 provided BTM samples. Five of the surveyed herds were positive on at least one of the diagnostic tests, thus herd-level apparent prevalence was 10% (95% confidence interval [CI] 4 to 22). True prevalence was estimated to be 3% (95% credible interval: 0 to 18). Herd managers completed a questionnaire on herd management, biosecurity and hygiene practices and risk factors were investigated using multivariable logistic regression. Herds with >900 milking does (the upper quartile) were more likely to be Coxiella burnetii positive (odds ratio = 6.75; 95% CI 1.65 to 27.7) compared with farms with ≤900 milking does. The odds of BTM positivity increased by a factor of 2.53 (95% CI 1.51 to 4.22) for each order of magnitude increase in the number of goats per acre. C. burnetii was not detected in samples from the majority of the Australian dairy goat herds suggesting there is an opportunity to protect the industry and contain this disease with strengthened biosecurity practices. Intensification appeared associated with an increased risk of positivity. Further investigation is required to discriminate the practices associated with an increased risk of introduction to disease-free herds, from practices associated with maintenance of C. burnetii infection in infected dairy goat herds.
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    Validation of an Indirect Immunofluorescence Assay and Commercial Q Fever Enzyme-Linked Immunosorbent Assay for Use in Macropods
    Tolpinrud, A ; Stenos, J ; Chaber, A-L ; Devlin, JM ; Herbert, C ; Pas, A ; Dunowska, M ; Stevenson, MA ; Firestone, SM ; Barrs, VR (AMER SOC MICROBIOLOGY, 2022-07-20)
    Kangaroos are considered to be an important reservoir of Q fever in Australia, although there is limited knowledge on the true prevalence and distribution of coxiellosis in Australian macropod populations. Serological tests serve as useful surveillance tools, but formal test validation is needed to be able to estimate true seroprevalence rates, and few tests have been validated to screen wildlife species for Q fever. In this study, we modified and optimized a phase-specific indirect immunofluorescence assay (IFA) for the detection of IgG antibodies against Coxiella burnetii in macropod sera. The assay was validated against the commercially available ID Screen Q fever indirect multispecies enzyme-linked immunosorbent assay (ELISA) kit (IDVet, Grabels, France) to estimate the diagnostic sensitivity and specificity of each assay, using Bayesian latent class analysis. A direct comparison of the two tests was performed by testing 303 serum samples from 10 macropod populations from the east coast of Australia and New Zealand. The analysis indicated that the IFA had relatively high diagnostic sensitivity (97.6% [95% credible interval [CrI], 88.0 to 99.9]) and diagnostic specificity (98.5% [95% CrI, 94.4 to 99.9]). In comparison, the ELISA had relatively poor diagnostic sensitivity (42.1% [95% CrI, 33.7 to 50.8]) and similar diagnostic specificity (99.2% [95% CrI, 96.4 to 100]) using the cutoff values recommended by the manufacturer. The estimated true seroprevalence of C. burnetii exposure in the macropod populations included in this study ranged from 0% in New Zealand and Victoria, Australia, up to 94.2% in one population from New South Wales, Australia.
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    Identifying scenarios and risk factors for Q fever outbreaks using qualitative analysis of expert opinion
    Tan, TS-E ; Hernandez-Jover, M ; Hayes, LM ; Wiethoelter, AK ; Firestone, SM ; Stevenson, MA ; Heller, J (WILEY, 2022-06)
    Q fever is an important zoonotic disease perceived to be an occupational hazard for those working with livestock. Outbreaks involving large numbers of people are uncommon, but the increasing case incidence coupled with changing environmental and industry conditions that promote transmission of Q fever has raised concerns that large and serious outbreaks could become more frequent. The aim of this study was to use expert opinion to better understand how large Q fever outbreaks might occur in an Australian context and to document factors believed to be drivers of disease transmission. Focus groups were conducted with human and animal health professionals across several Australian states. All discussions were recorded, transcribed verbatim and imported into NVIVO for thematic analysis. Four anthropogenic risk factors (disease awareness, industry practices, land use, human behaviour) and three ecological risk factors (physical environment, agent dissemination, animal hosts) emerged from the data. Analysis of expert opinions pointed to the existence of numerous scenarios in which Q fever outbreaks could occur, many of which depict acquisition in the wider community outside of traditional at-risk occupations. This perception of the expansion of Q fever from occupational-acquisition to community-acquisition is driven by greater overarching economic, political and socio-cultural influences that govern the way in which people live and work. Findings from this study highlight that outbreaks are complex phenomena that involve the convergence of diverse elements, not just that of the pathogen and host, but also the physical, political and socioeconomic environments in which they interact. A review of the approaches to prevent and manage Q fever outbreaks will require a multisectorial approach and strengthening of community education, communication and engagement so that all stakeholders become an integrated part of outbreak mitigation and response.
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    Using farmer observations for animal health syndromic surveillance: Participation and performance of an online enhanced passive surveillance system
    Pfeiffer, C ; Stevenson, M ; Firestone, S ; Larsen, J ; Campbell, A (ELSEVIER, 2021-03)
    The challenge of animal health surveillance is to provide the information necessary to appropriately inform disease prevention and control activities within the constraints of available resources. Syndromic surveillance of farmers' disease observations can improve animal health data capture from extensive livestock farming systems, especially where data are not otherwise being systematically collected or when data on confirmed aetiological diagnoses are unavailable at the disease level. As it is rarely feasible to recruit a truly random sample of farmers to provide observational reports, directing farmer sampling to align with the surveillance objectives is a reasonable and practical approach. As long as potential bias is recognised and managed, farmers who will report reliably can be desirable participants in a surveillance system. Thus, one early objective of a surveillance program should be to identify characteristics associated with reporting behaviour. Knowledge of the demographic and managerial characteristics of good reporters can inform efforts to recruit additional farms into the system or aid understanding of potential bias of system reports. We describe the operation of a farmer syndromic surveillance system in Victoria, Australia, over its first two years from 2014 to 2016. Survival analysis and classification and regression tree analysis were used to identify farm level factors associated with 'reliable' participation (low non-response rates in longitudinal reporting). Response rate and timeliness were not associated with whether farmers had disease to report, or with different months of the year. Farmers keeping only sheep were the most reliable and timely respondents. Farmers < 43 years of age had lower response rates than older farmers. Farmers with veterinary qualifications and those working full-time on-farm provided less timely reports than other educational backgrounds and farmers who worked part-time on-farm. These analyses provide a starting point to guide recruitment of participants for surveillance of farmers' observations using syndromic surveillance, and provide examples of strengths and weaknesses of syndromic surveillance systems for extensively-managed livestock. Once farm characteristics associated with reliable participation are known, they can be incorporated into surveillance system design in accordance with the objectives of the system.
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    The use of social network analysis to examine the transmission of Salmonella spp. within a vertically integrated broiler enterprise
    Crabb, HK ; Allen, JL ; Devlin, JM ; Firestone, SM ; Stevenson, MA ; Gilkerson, JR (ACADEMIC PRESS LTD- ELSEVIER SCIENCE LTD, 2018-05)
    To better understand factors influencing infectious agent dispersal within a livestock population information is needed on the nature and frequency of contacts between farm enterprises. This study uses social network analysis to describe the contact network within a vertically integrated broiler poultry enterprise to identify the potential horizontal and vertical transmission pathways for Salmonella spp. Nodes (farms, sheds, production facilities) were identified and the daily movement of commodities (eggs, birds, feed, litter) and people between nodes were extracted from routinely kept farm records. Three time periods were examined in detail, 1- and 8- and 17-weeks of the production cycle and contact networks were described for all movements, and by commodity and production type. All nodes were linked by at least one movement during the study period but network density was low indicating that all potential pathways between nodes did not exist. Salmonella spp. transmission via vertical or horizontal pathways can only occur along directed pathways when those pathways are present. Only two locations (breeder or feed nodes) were identified where the transmission of a single Salmonella spp. clone could theoretically percolate through the network to the broiler or processing nodes. Only the feed transmission pathway directly connected all parts of the network.
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    Traditional Salmonella Typhimurium typing tools (phage typing and MLVA) are sufficient to resolve well-defined outbreak events only
    Crabb, HK ; Allen, JL ; Devlin, JM ; Firestone, SM ; Stevenson, M ; Wilks, CR ; Gilkerson, JR (ACADEMIC PRESS LTD- ELSEVIER SCIENCE LTD, 2019-12-01)
    Between 1991 and 2014 the per capita notification rate of salmonellosis in Australia increased from 31.9 to 69.7 cases per 100,000 people. Salmonella Typhimurium accounted for nearly half the human cases until the end of 2014. In this study, we used cluster analysis tools to compare S. Typhimurium isolates from a chicken-meat study with those reported to the National Enteric Pathogen Surveillance System (NEPSS) from the coincident human and non-human populations. There was limited phage type diversity within all populations and a lack of specificity of MLVA profiling within phage types. The chicken-meat study isolates were not significantly clustered with the human cases and at least 7 non-human sources, based on typing profiles (PT/MLVA combination), could be implicated as a source of human cases during the same period. In the absence of a strong surveillance system representative of all putative sources, MLVA and phage typing alone or in combination are insufficient to identify the source of human cases.
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    A cross-sectional study to quantify the prevalence of avian influenza viruses in poultry at intervention and non-intervention live bird markets in central Vietnam, 2014
    Chu, D-H ; Stevenson, MA ; Nguyen, LV ; Isoda, N ; Firestone, SM ; Nguyen, TN ; Nguyen, LT ; Matsuno, K ; Okamatsu, M ; Kida, H ; Sakoda, Y (WILEY, 2017-12)
    In Vietnam, live bird markets are found in most populated centres, providing the means by which fresh poultry can be purchased by consumers for immediate consumption. Live bird markets are aggregation points for large numbers of poultry, and therefore, it is common for a range of avian influenza viruses to be mixed within live bird markets as a result of different poultry types and species being brought together from different geographical locations. We conducted a cross-sectional study in seven live bird markets in four districts of Thua Thien Hue Province in August and December, 2014. The aims of this study were to (i) document the prevalence of avian influenza in live bird markets (as measured by virus isolation); and (ii) quantify individual bird-, seller- and market-level characteristics that rendered poultry more likely to be positive for avian influenza virus at the time of sale. A questionnaire soliciting details of knowledge, attitude and avian influenza practices was administered to poultry sellers in study markets. At the same time, swabs and faecal samples were collected from individual poultry and submitted for isolation of avian influenza virus. The final data set comprised samples from 1,629 birds from 83 sellers in the seven live bird markets. A total of 113 birds were positive for virus isolation; a prevalence of 6.9 (95% CI 5.8-8.3) avian influenza virus-positive birds per 100 birds submitted for sale. After adjusting for clustering at the market and individual seller levels, none of the explanatory variables solicited in the questionnaire were significantly associated with avian influenza virus isolation positivity. The proportions of variance at the individual market, seller and individual bird levels were 6%, 48% and 46%, respectively. We conclude that the emphasis of avian influenza control efforts in Vietnam should be at the individual seller level as opposed to the market level.