Detection of a putative hemolysin operon, hhdBA, of Haemophilus parasuis from pigs with Glasser disease

The aim of the current study was to investigate whether polymerase chain reaction amplification of 16S ribosomal (r)RNA and a putative hemolysin gene operon, hhdBA, can be used to monitor live pigs for the presence of Haemophilus parasuis and predict the virulence of the strains present. Nasal cavity swabs were taken from 30 live, healthy, 1- to 8-week-old pigs on a weekly cycle from a commercial Thai nursery pig herd. A total of 27 of these pigs (90%) tested positive for H. parasuis as early as week 1 of age. None of the H. parasuis-positive samples from healthy pigs was positive for the hhdBA genes. At the same pig nursery, swab samples from nasal cavity, tonsil, trachea, and lung, and exudate samples from pleural/peritoneal cavity were taken from 30 dead pigs displaying typical pathological lesions consistent with Glasser disease. Twenty-two of 140 samples (15.7%) taken from 30 diseased pigs yielded a positive result for H. parasuis. Samples from the exudate (27%) yielded the most positive results, followed by lung, tracheal swab, tonsil, and nasal swab, respectively. Out of 22 positive samples, 12 samples (54.5%) harbored hhdA and/or hhdB genes. Detection rates of hhdA were higher than hhdB. None of the H. parasuis-positive samples taken from nasal cavity of diseased pigs tested positive for hhdBA genes. More work is required to determine if the detection of hhdBA genes is useful for identifying the virulence potential of H. parasuis field isolates.

Natural outbreak of Streptococcus agalactiae (GBS) infection in wild giant Queensland grouper, Epinephelus lanceolatus (Bloch), and other wild fish in northern Queensland, Australia

Ninety-three giant Queensland grouper, Epinephelus lanceolatus (Bloch), were found dead in Queensland, Australia, from 2007 to 2011. Most dead fish occurred in northern Queensland, with a peak of mortalities in Cairns in June 2008. In 2009, sick wild fish including giant sea catfish, Arius thalassinus (Ruppell), and javelin grunter, Pomadasys kaakan (Cuvier), also occurred in Cairns. In 2009 and 2010, two disease epizootics involving wild stingrays occurred at Sea World marine aquarium. Necropsy, histopathology, bacteriology and PCR determined that the cause of deaths of 12 giant Queensland grouper, three wild fish, six estuary rays, Dasyatis fluviorum (Ogilby), one mangrove whipray, Himantura granulata (Macleay), and one eastern shovelnose ray, Aptychotrema rostrata (Shaw), was Streptococcus agalactiae septicaemia. Biochemical testing of 34 S.agalactiae isolates from giant Queensland grouper, wild fish and stingrays showed all had identical biochemical profiles. The 16S rRNA gene sequences of isolates confirmed all isolates were S.agalactiae; genotyping of selected S.agalactiae isolates showed the isolates from giant Queensland grouper were serotype Ib, whereas isolates from wild fish and stingrays closely resembled serotype II. This is the first report of S.agalactiae from wild giant Queensland grouper and other wild tropical fish and stingray species in Queensland, Australia.

Agricultural intensification, priming for persistence and the emergence of Nipah virus: a lethal bat-borne zoonosis

Emerging zoonoses threaten global health, yet the processes by which they emerge are complex and poorly understood. Nipah virus (NiV) is an important threat owing to its broad host and geographical range, high case fatality, potential for human-to-human transmission and lack of effective prevention or therapies. Here, we investigate the origin of the first identified outbreak of NiV encephalitis in Malaysia and Singapore. We analyse data on livestock production from the index site (a commercial pig farm in Malaysia) prior to and during the outbreak, on Malaysian agricultural production, and from surveys of NiV's wildlife reservoir (flying foxes). Our analyses suggest that repeated introduction of NiV from wildlife changed infection dynamics in pigs. Initial viral introduction produced an explosive epizootic that drove itself to extinction but primed the population for enzootic persistence upon reintroduction of the virus. The resultant within-farm persistence permitted regional spread and increased the number of human infections. This study refutes an earlier hypothesis that anomalous El Nino Southern Oscillation-related climatic conditions drove emergence and suggests that priming for persistence drove the emergence of a novel zoonotic pathogen. Thus, we provide empirical evidence for a causative mechanism previously proposed as a precursor to widespread infection with H5N1 avian influenza and other emerging pathogens.

Systemic toxoplasmosis in captive flying-foxes

Systemic toxoplasmosis caused by Toxoplasma gondii was diagnosed in two juvenile, captive flying-foxes (Pteropus conspicillatus and P. scapulatus), which died following respiratory distress. One animal displayed clinical signs suggestive of neurological disease. This is the first report of this disease in megachiropteran bats and adds to the list of differential diagnoses for both systemic and neurological disease in these animals. The role of captivity in the exposure and development of the disease is discussed.

Immuno-fluorescence staining patterns of leukocyte subsets in the skin of taurine and indicine cattle

The immuno-staining patterns of skin leukocytes were investigated in three breeds of cattle: Holstein–Friesian, Brahman and Santa Gertrudis of similar age before and after tick infestation. The antibodies specific for CD45 and CD45RO reacted with cells in the skin of all Holstein–Friesian cattle but did not react with cells in the skin of any Brahman cattle. The same antibodies reacted with cells from the skin of four (CD45) and seven (CD45RO) of twelve Santa Gertrudis cattle. The antibodies specific for T cells and γδ subset of T cells recognized cells from all three breeds of cattle. The antibody specific for MHC class II molecules labelled cells of mostly irregular shape, presumably dermal dendritic cells and/or macrophages and Langerhans cells. The antibody specific for granulocytes (mAb CH138) reacted with cells only in sections cut from skin with lesions. The antibody specific for CD25+ cells labelled regularly shaped cells that showed a wide range of intensities of staining.

Is Mycoplasma bovis a missing component of the bovine respiratory disease complex in Australia?

Background: Bovine respiratory disease complex (BRDC) is a multi-factorial disease in which numerous factors, such as animal management, pathogen exposure and environmental conditions, contribute to the development of acute respiratory illness in feedlot cattle. The role of specific pathogens in the development of BRDC has been difficult to define because of the complex nature of the disease and the presence of implicated bacterial pathogens in the upper respiratory tract of healthy animals. Mycoplasma bovis is an important pathogen of cattle and recognised as a major contributor to cases of mastitis, caseonecrotic bronchopneumonia, arthritis and otitis media. To date, the role of M.bovis in the development of BRDC of Australian feeder cattle has not been investigated. Methods: In this review, the current literature pertaining to the role of M.bovis in BRDC is evaluated. In addition, preliminary data are presented that identify M.bovis as a potential contributor to BRDC in Australian feedlots, which has not been considered previously. Results and Conclusion: The preliminary results demonstrate detection of M.bovis in samples from all feedlots studied. When considered in the context of the reviewed literature, they support the inclusion of M.bovis on the list of pathogens to be considered during investigations into BRDC in Australia. © 2014 Australian Veterinary Association.

Variation in the hindgut microbial communities of the Florida manatee, Trichechus manatus latirostris over winter in Crystal River, Florida

The Florida manatee, Trichechus manatus latirostris, is a hindgut-fermenting herbivore. In winter, manatees migrate to warm water overwintering sites where they undergo dietary shifts and may suffer from cold-induced stress. Given these seasonally induced changes in diet, the present study aimed to examine variation in the hindgut bacterial communities of wild manatees overwintering at Crystal River, west Florida. Faeces were sampled from 36 manatees of known sex and body size in early winter when manatees were newly arrived and then in mid-winter and late winter when diet had probably changed and environmental stress may have increased. Concentrations of faecal cortisol metabolite, an indicator of a stress response, were measured by enzyme immunoassay. Using 454-pyrosequencing, 2027 bacterial operational taxonomic units were identified in manatee faeces following amplicon pyrosequencing of the 16S rRNA gene V3/V4 region. Classified sequences were assigned to eight previously described bacterial phyla; only 0.36% of sequences could not be classified to phylum level. Five core phyla were identified in all samples. The majority (96.8%) of sequences were classified as Firmicutes (77.3 ± 11.1% of total sequences) or Bacteroidetes (19.5 ± 10.6%). Alpha-diversity measures trended towards higher diversity of hindgut microbiota in manatees in mid-winter compared to early and late winter. Beta-diversity measures, analysed through permanova, also indicated significant differences in bacterial communities based on the season.

Equine multinodular pulmonary fibrosis in three horses in Australia

Background Equine multinodular pulmonary fibrosis (EMPF) is a recently described form of interstitial pneumonia associated with the presence of equine herpesvirus type 5 (EHV-5). Since 2007, several case reports from America, Europe and the United Kingdom have further characterised the clinical presentation and laboratory findings of this disease. Case reports Three Thoroughbred broodmares were diagnosed with EMPF. Diagnosis was based on lung histopathology and positive identification of EHV-5 using PCR DNA amplification. There was multiple organ involvement in all three cases, including identification of EHV-5 in hepatic tissue in one case. Two of the three horses died. Treatment with acyclovir was unsuccessful in one horse and one horse survived without antiviral or corticosteroid treatment. Conclusion This case series is, to the authors' knowledge, the first report of EMPF in Australia and adds to the clinical description of the disease.

Quantitative effects of in-line operations on Campylobacter and Escherichia coli through two Australian broiler processing plants

Campylobacter is an important food borne pathogen, mainly associated with poultry. A lack of through-chain quantitative Campylobacter data has been highlighted within quantitative risk assessments. The aim of this study was to quantitatively and qualitatively measure Campylobacter and Escherichia coli concentration on chicken carcasses through poultry slaughter. Chickens (n = 240) were sampled from each of four flocks along the processing chain, before scald, after scald, before chill, after chill, after packaging and from individual caeca. The overall prevalence of Campylobacter after packaging was 83% with a median concentration of 0.8 log10 CFU/mL. The processing points of scalding and chilling had significant mean reductions of both Campylobacter (1.8 and 2.9 log10 CFU/carcase) and E. coli (1.3 and 2.5 log10 CFU/carcase). The concentration of E. coli and Campylobacter was significantly correlated throughout processing indicating that E. coli may be a useful indicator organism for reductions in Campylobacter concentration. The carriage of species varied between flocks, with two flocks dominated by Campylobacter coli and two flocks dominated by Campylobacter jejuni. Current processing practices can lead to significant reductions in the concentration of Campylobacter on carcasses. Further understanding of the variable effect of processing on Campylobacter and the survival of specific genotypes may enable more targeted interventions to reduce the concentration of this poultry associated pathogen.

Variable-number tandem repeats genotyping used to aid and inform management strategies for a bovine Johne's disease incursion in tropical and subtropical Australia

The application of variable-number tandem repeats (VNTR) genotyping of Mycobacterium avium subsp. paratuberculosis isolates to assist in investigating incidents of bovine Johne’s disease in a low-prevalence region of Australia is described in the current study. Isolates from a response to detection of bovine Johne’s disease in Queensland were compared with strains from national and international sources. The tandem application of mycobacterial interspersed repetitive unit (MIRU) and multilocus short sequence repeats (MLSSR) genotyping identified 2 strains, 1 that infected cattle on multiple properties with trace-forward histories from a common infected property, and 1 genotypically different strain recovered from a single property. The former strain showed an identical genotype to an isolate from India. Neither strain showed a genotypic link to regions of Australia with a higher prevalence of the disease. Genotyping has indicated incursions from 2 independent sources. This intelligence has informed investigations into potential routes of entry and the soundness of ongoing control measures, and supported strategy and policy decisions regarding management of Mycobacterium avium subsp. paratuberculosis incursions for Queensland.

Development of a Rapid Multiplex PCR Assay To Genotype Pasteurella multocida Strains by Use of the Lipopolysaccharide Outer Core Biosynthesis Locus

Pasteurella multocida is a Gram-negative bacterial pathogen that is the causative agent of a wide range of diseases in many animal species, including humans. A widely used method for differentiation of P. multocida strains involves the Heddleston serotyping scheme. This scheme was developed in the early 1970s and classifies P. multocida strains into 16 somatic or lipopolysaccharide (LPS) serovars using an agar gel diffusion precipitin test. However, this gel diffusion assay is problematic, with difficulties reported in accuracy, reproducibility, and the sourcing of quality serovar-specific antisera. Using our knowledge of the genetics of LPS biosynthesis in P. multocida, we have developed a multiplex PCR (mPCR) that is able to differentiate strains based on the genetic organization of the LPS outer core biosynthesis loci. The accuracy of the LPS-mPCR was compared with classical Heddleston serotyping using LPS compositional data as the "gold standard." The LPS-mPCR correctly typed 57 of 58 isolates; Heddleston serotyping was able to correctly and unambiguously type only 20 of the 58 isolates. We conclude that our LPS-mPCR is a highly accurate LPS genotyping method that should replace the Heddleston serotyping scheme for the classification of P. multocida strains.

Cross-protection study of the nine serovars of Haemophilus paragallinarum in the Kume haemagglutinin scheme

The cross-protection and haemagglutination-inhibition antibodies present in chickens vaccinated with one of the nine currently recognized Kume haemagglutinin serovars of Haemophilus paragallinarum were investigated. The results confirmed the widely accepted dogma that serogroups A, B, and C represent three distinct immunovars. Within Kume serogroup A, there was generally good cross-protection among all four serovars. However, within Kume serogroup C, there was evidence of a reduced level of cross protection between some of the four serovars. The haemagglutination-inhibition antibody levels generally showed the same trend as with the cross-protection results. This study suggests that some apparent field failures of infectious coryza vaccines may be due to a lack of cross-protection between the vaccine strains and the field strains. Our results will help guide the selection of strains for inclusion in infectious coryza vaccines.

Neuro-angiostrongylosis in wild Black and Grey-headed flying foxes (Pteropus spp)

Objective: To identify nematodes seen in histological sections of brains of flying foxes (fruit bats) and describe the associated clinical disease and pathology. Proceedures: Gross and histological examination of brains from 86 free-living flying foxes with neurological disease was done as part of an ongoing surveillance program for Australian bat lyssavirus. Worms were recovered, or if seen in histological sections, extracted by maceration of half the brain and identified by microscopic examination. Histological archives were also reviewed. Results: There was histological evidence of angiostrongylosis in 16 of 86 recently submitted flying foxes with neurological disease and in one archival case from 1992. In 10 flying foxes, worms were definitively identified as Angiostrongylus cantonensis fifth-stage larvae. A worm fragment and third stage larvae were identified as Angiostrongylus sp, presumably A cantonensis, in a further three cases. The clinical picture was dominated by paresis, particularly of the hindlimbs, and depression, with flying foxes surviving up to 22 days in the care of wildlife volunteers. Brains containing fifthstage larvae showed a moderate to severe eosinophilic and granulomatous meningoencephalitis (n = 14), whereas there was virtually no inflammation of the brains of bats which died when infected with only smaller, third-stage larvae (n = 3). There was no histological evidence of pulmonary involvement. Conclusion: This is the first report of the recovery and identification of A cantonensis from free-living Australian wildlife. While angiostrongylosis is a common cause of paresis in flying foxes, the initial clinical course cannot be differentiated from Australian bat lyssavirus infection, and wildlife carers should be urged not to attempt to rehabilitate flying foxes with neurological disease.