Abstract
BackgroundMink enteritis virus (MEV) causes mink viral enteritis, an acute and highly contagious disease whose symptoms include violent diarrhea, and which is characterized by high morbidity and mortality. Nanoparticle-assisted polymerase chain reaction (nanoPCR) is a recently developed technique for the rapid detection of bacterial and viral DNA. Here we describe a novel nanoPCR assay for the clinical detection and epidemiological characterization of MEV.ResultsThis assay is based upon primers specific for the conserved region of the MEV NS1 gene, which encodes nonstructural protein 1. Under optimized conditions, the MEV nanoPCR assay had a detection limit of 8.75 × 101 copies recombinant plasmids per reaction, compared with 8.75 × 103 copies for conventional PCR analysis. Moreover, of 246 clinical mink samples collected from five provinces in North-Eastern China, 50.8% were scored MEV positive by our nanoPCR assay, compared with 32.5% for conventional PCR. Furthermore no cross reactivity was observed for the nanoPCR assay with respect to related viruses, including canine distemper virus (CDV) and Aleutian mink disease parvovirus (AMDV). Phylogenetic analysis of four Chinese wild type MEV isolates using the nanoPCR assay indicated that they belonged to a small MEV clade, named “China type”, in the MEV/FPLV cluster, and were closely clustered in the same location.ConclusionsOur results indicate that the MEV China type clade is currently circulating in domestic minks in China. We anticipate that the nanoPCR assay we have described here will be useful for the detection and epidemiological and pathological characterization of MEV.Electronic supplementary materialThe online version of this article (doi:10.1186/s12917-014-0312-6) contains supplementary material, which is available to authorized users.
Highlights
Mink enteritis virus (MEV) causes mink viral enteritis, an acute and highly contagious disease whose symptoms include violent diarrhea, and which is characterized by high morbidity and mortality [4]
Three primer pairs with fragment lengths of 194 bp, 163 bp and 389 bp, respectively, were compared, and based on gel quantification analysis by ImageJ 1.46r software, primer pair No 1 (P1 and P2) was selected for use in conventional polymerase chain reaction (PCR) and nanoPCR assays
Based on the results obtained with different annealing temperatures, primer volumes and plasmid DNA volumes for the MEV nanoPCR assay, an optimal 12 μL reaction volume was established, containing 6.0 μL of 2× nanobuffer, 0.6 μL each of the upstream and downstream primers (10 μmol/L), 1.0 μL of extracted DNA or standard plasmid, 0.2 μL of Taq DNA polymerase (5 U/μL) and ddH2O up to 12 μL
Summary
Mink enteritis virus (MEV) causes mink viral enteritis, an acute and highly contagious disease whose symptoms include violent diarrhea, and which is characterized by high morbidity and mortality. Nanoparticle-assisted polymerase chain reaction (nanoPCR) is a recently developed technique for the rapid detection of bacterial and viral DNA. We describe a novel nanoPCR assay for the clinical detection and epidemiological characterization of MEV. Mink enteritis virus (MEV), a member of the genus Parvovirus within the family Parvoviridae, and a subspecies of the feline parvovirus (FPV), is a single-stranded DNA virus with a genome length of approximately 5,094 nt [1,2,3]. MEV causes mink viral enteritis, an acute and highly contagious disease whose symptoms include violent diarrhea, and which is characterized by high morbidity and mortality [4]. The disease has since been reported in a number of other countries worldwide [2], including China [7], and poses a serious economic threat to the global mink fur farming industry [8].
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