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A novel multiplex RT- qPCR method based on dual-labelled probes suitable for typing all known genotypes of viral haemorrhagic septicaemia virus.
- Source :
- Journal of Fish Diseases; Apr2016, Vol. 39 Issue 4, p467-482, 16p
- Publication Year :
- 2016
-
Abstract
- Viral haemorrhagic septicaemia ( VHS) is a notifiable fish disease, whose causative agent is a rhabdovirus isolated from a wide range of fish species, not only in fresh but also in marine and brackish waters. Phylogenetic studies have identified four major genotypes, with a strong geographical relationship. In this study, we have designed and validated a new procedure - named binary multiplex RT- qPCR (bm RT- qPCR) - for simultaneous detection and typing of all four genotypes of VHSV by real-time RT- PCR based on dual-labelled probes and composed by two multiplex systems designed for European and American/Asiatic isolates, respectively, using a combination of three different fluorophores. The specificity of the procedure was assessed by including a panel of 81 VHSV isolates covering all known genotypes and subtypes of the virus, and tissue material from experimentally infected rainbow trout, resulting in a correct detection and typing of all strains. The analytical sensitivity was evaluated in a comparative assay with titration in cell culture, observing that both methods provided similar limits of detection. The proposed method can be a powerful tool for epidemiological analysis of VHSV by genotyping unknown samples within a few hours. [ABSTRACT FROM AUTHOR]
Details
- Language :
- English
- ISSN :
- 01407775
- Volume :
- 39
- Issue :
- 4
- Database :
- Complementary Index
- Journal :
- Journal of Fish Diseases
- Publication Type :
- Academic Journal
- Accession number :
- 113528262
- Full Text :
- https://doi.org/10.1111/jfd.12381