A large-scale metagenomic survey dataset of the post-weaning piglet gut lumen

BackgroundEarly weaning and intensive farming practices predispose piglets to the development of infectious and often lethal diseases, against which antibiotics are used. Besides contributing to the build-up of antimicrobial resistance, antibiotics are known to modulate the gut microbial composition. As an alternative to antibiotic treatment, studies have previously investigated the potential of probiotics for the prevention of postweaning diarrhea. In order to describe the post-weaning gut microbiota, and to study the effects of two probiotics formulations and of intramuscular antibiotic treatment on the gut microbiota, we sampled and processed over 800 faecal time-series samples from 126 piglets and 42 sows.ResultsHere we report on the largest shotgun metagenomic dataset of the pig gut lumen microbiome to date, consisting of >8 Tbp of shotgun metagenomic sequencing data. The animal trial, the workflow from sample collection to sample processing, and the preparation of libraries for sequencing, are described in detail. We provide a preliminary analysis of the dataset, centered on a taxonomic profiling of the samples, and a 16S-based beta diversity analysis of the mothers and the piglets in the first 5 weeks after weaning.ConclusionsThis study was conducted to generate a publicly available databank of the faecal metagenome of weaner piglets aged between 3 and 9 weeks old, treated with different probiotic formulations and intramuscular antibiotic treatment. Besides investigating the effects of the probiotic and intramuscular antibiotic treatment, the dataset can be explored to assess a wide range of ecological questions with regards to antimicrobial resistance, host-associated microbial and phage communities, and their dynamics during the aging of the host

Characterisation of Erysipelothrix rhusiopathiae isolates from pigs associated with vaccine breakdowns

Swine erysipelas vaccines are routinely used to protect pigs against peracute and acute/urticarial forms of Erysipelothrix. Between 1995 and 1998, 34 swine herds across four Australian states experienced vaccine failure. Forty-four isolates of Erysipelothrix rhusiopathiae of serovars 2, 1a, 1b and 1b × 21 were recovered from 15 of these 34 vaccine breakdown herds. These isolates were characterised by restriction fragment length polymorphism (RFLP) analyses using RsaI and AluI on whole cell DNA and for the presence of plasmid DNA. Results were compared with those of 20 isolates from 16 herds unaffected by vaccine breakdown and 13 isolates representing 10 reference strains. The majority of breakdown herds possessed isolates of serovar 2 (9/15 herds), followed by serovar 1a (5 herds). No geographic predominance of a single serovar was evident. The identification of 10 RsaI profiles from whole cell DNA among the 44 isolates from 15 breakdown herds indicated that a single, new clonal lineage of E. rhusiopathiae was not responsible for vaccine failure. RsaI RFLP analyses detected a further 14 distinct profiles among 20 field strains unassociated with vaccine breakdowns, and none matched profiles of the 10 serovar reference strains for serovars 1a, 1b, 2 or 21. This technique is recommended for epidemiological studies of E. rhusiopathiae strains. Crown Copyright © 2006

Demonstration that Australian Pasteurella multocida isolates from sporadic outbreaks of porcine pneumonia are non-toxigenic (toxA(-)) and display heterogeneous DNA restriction endonuclease profiles compared with toxigenic isolates from herds with progress

Capsular types A and D of Pasteurella multocida cause economic losses in swine because of their association with progressive atrophic rhinitis (PAR) and enzootic pneumonia, There have been no studies comparing whole-cell DNA profiles of isolates associated with these two porcine respiratory diseases. Twenty-two isolates of P. multocida from diseased pigs in different geographic localities within Australia were characterised genotypically by restriction endonuclease analysis (REA) with the enzyme CfoI, Seven of 12 P. multocida isolates from nasal swabs from pigs in herds where PAR was either present or suspected displayed a capsular type D phenotype, These were shown to possess the toxA gene by polymerase chain reaction (PCR) and Southern hybridisation, and further substantiated by production of cytotoxin in vitro. The CfoI profile of one of these seven isolates, which was from the initial outbreak of PAR in Australia (in Western Australia, WA), was identical with profiles of all six other toxigenic isolates from sporadic episodes in New South Wales (NSW), The evidence suggests that the strain involved in the initial outbreak was responsible for the spread of PAR to the eastern states of Australia. Another 10 isolates, representing both capsular types A and D, were isolated exclusively from porcine lung lesions after sporadic outbreaks of enzootic pneumonia in NSW and WA. CfoI restriction endonuclease profiles of these isolates revealed considerable genomic heterogeneity. Furthermore, none of these possessed the toxA gene. This suggests that P. multocida strains with the toxA gene do not have a competitive survival advantage in the lower respiratory tract or that toxin production does not play a role in the pathology of pneumonic lesions, or both. REA with polyacrylamide gel electrophoresis and silver staining was found to be a practical and discriminatory tool for epidemiological tracing of P. multocida outbreaks associated with PAR or pneumonia in pigs

Plasmin activity in the porcine airways is enhanced during experimental infection with Mycoplasma hyopneumoniae, is positively correlated with proinflammatory cytokine levels and is ameliorated by vaccination

In Mycoplasma hyopneumoniae (Mhp) infection of swine, the host immune response is considered a major driver of lung pathology; however the underlying inflammatory mechanisms are not well understood. The serine protease plasmin is being increasingly recognised as a significant player in inflammatory processes. Here we compare plasmin activity in tracheobronchial lavage fluid (TBLF) from pigs experimentally challenged with Mhp that were either unvaccinated (n=10), or vaccinated with the commercial vaccine Suvaxyn® M.hyo (n=10). TBLF collected immediately prior to challenge and at 21d and 35d post-challenge was also assayed for levels of proinflammatory cytokines (TNF-α, IL-1β and IL-6), and for bacterial load (by qPCR). Clinical signs, pathology, cytokine analyses and qPCR all indicated that vaccinated pigs had significantly reduced disease relative to unvaccinated animals. Plasmin activity increased significantly in TBLF collected at 21d post-challenge compared to pre-challenge TBLF in unvaccinated (P0.05). A significant correlation was observed between bacterial load and plasmin activity in the 21d (r=0.66; P<0.01) and the 35d post-challenge samples, (r=0.62; P<0.01). Plasmin activity was also significantly correlated with levels of TNF-α, IL-1β and IL-6 at 21d (r=0.78, P<0.0001; r=0.77, P<0.0001; r=0.64, P<0.005) and with TNF-α and IL-1β at 35d post-challenge (r=0.77, P<0.0001; r=0.74, P<0.0005). Our results indicate that plasminogen is activated to plasmin in the respiratory tract of pigs as part of the host inflammatory response to Mhp infection and that this effect is ameliorated by vaccination. © 2013

Development and validation of a quantitative PCR assay using multiplexed hydrolysis probes for detection and quantification of Theileria orientalis isolates and differentiation of clinically relevant subtypes

Copyright © 2015, American Society for Microbiology. All Rights Reserved. Theileria orientalis is an emerging pathogen of cattle in Asia, Australia, and New Zealand. This organism is a vector-borne hemoprotozoan that causes clinical disease characterized by anemia, abortion, and death, as well as persistent subclinical infections. Molecular methods of diagnosis are preferred due to their sensitivity and utility in differentiating between pathogenic and apathogenic genotypes. Conventional PCR (cPCR) assays for T. orientalis detection and typing are laborious and do not provide an estimate of parasite load. Current real-time PCR assays cannot differentiate between clinically relevant and benign genotypes or are only semiquantitative without a defined clinical threshold. Here, we developed and validated a hydrolysis probe quantitative PCR (qPCR) assay which universally detects and quantifies T. orientalis and identifies the clinically associated Ikeda and Chitose genotypes (UIC assay). Comparison of the UIC assay results with previously validated universal and genotype-specific cPCR results demonstrated that qPCR detects and differentiates T. orientalis with high sensitivity and specificiy. Comparison of quantitative results based on percent parasitemia, determined via blood film analysis and packed cell volume (PCV) revealed significant positive and negative correlations, respectively. One-way analysis of variance (ANOVA) indicated that blood samples from animals with clinical signs of disease contained statistically higher concentrations of T. orientalis DNA than animals with subclinical infections. We propose clinical thresholds to assist in classifying high-, moderate-, and low-level infections and describe how parasite load and the presence of the Ikeda and Chitose genotypes relate to disease