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Detection and characterisation of canine astrovirus, canine parvovirus and canine papillomavirus in puppies using next generation sequencing.

ABSTRACT: Gastroenteritis in young animals is a clinical presentation with many infectious and non- infectious aetiologies. We used next generation sequencing (NGS) to investigate the possible infectious causes of gastroenteritis in puppies from a dog kennel in Victoria, Australia. The near complete genome of a canine astrovirus was obtained from pooled faecal samples, and was found to be 94.7% identical with a canine astrovirus detected in the United Kingdom in 2012. The phylogenetic analysis of the capsid gene found similarities to those of canine astroviruses identified in Italy in 2005 and in UK and Hungary in 2012, but distant from that of a canine astrovirus previously identified in Australia in 2012. Thus, different serotypes of canine astrovirus are likely circulating in Australia. The close relationship to European astroviruses also suggested that there had been recent movements of ancestor canine astroviruses between Australia and Europe. NGS also detected other infections in the puppies including several canine papillomaviruses and a canine parvovirus (vaccine strain) as well as a very low level of campylobacter. Canine astrovirus was the probable cause of diarrhoea in these puppies, with the possible involvement of campylobacter bacteria. NGS was effective as a non-targeted method to determine the likely infectious cause of gastroenteritis.

SUBMITTER: Bhatta TR

PROVIDER: S-EPMC6418273 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:BackgroundVector-borne diseases (VBDs) impact both human and veterinary medicine and pose special public health challenges. The main bacterial vector-borne pathogens (VBPs) of importance in veterinary medicine include Anaplasma spp., Bartonella spp., Ehrlichia spp., and Spotted Fever Group Rickettsia. Taxon-targeted PCR assays are the current gold standard for VBP diagnostics but limitations on the detection of genetically diverse organisms support a novel approach for broader detection of VBPs. We present a methodology for genetic characterization of VBPs using Next-Generation Sequencing (NGS) and computational approaches. A major advantage of NGS is the ability to detect multiple organisms present in the same clinical sample in an unsupervised (i.e. non-targeted) and semi-quantitative way. The Standard Operating Procedure (SOP) presented here combines industry-standard microbiome analysis tools with our ad-hoc bioinformatic scripts to form a complete analysis pipeline accessible to veterinary scientists and freely available for download and use at https://github.com/eltonjrv/microbiome.westernu/tree/SOP .ResultsWe tested and validated our SOP by mimicking single, double, and triple infections in genomic canine DNA using serial dilutions of plasmids containing the entire 16 S rRNA gene sequence of (A) phagocytophilum, (B) v. berkhoffii, and E. canis. NGS with broad-range 16 S rRNA primers followed by our bioinformatics SOP was capable of detecting these pathogens in biological replicates of different dilutions. These results illustrate the ability of NGS to detect and genetically characterize multi-infections with different amounts of pathogens in a single sample.ConclusionsBloodborne microbiomics & metagenomics approaches may help expand the molecular diagnostic toolbox in veterinary and human medicine. In this paper, we present both in vitro and in silico detailed protocols that can be combined into a single workflow that may provide a significant improvement in VBP diagnostics and also facilitate future applications of microbiome research in veterinary medicine.

Dataset Information

Detection and characterisation of canine astrovirus, canine parvovirus and canine papillomavirus in puppies using next generation sequencing.

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