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Molecular detection of genotype II grass carp reovirus based on nucleic acid sequence-based amplification combined with enzyme-linked immunosorbent assay (NASBA-ELISA).
- Source :
-
Journal of virological methods [J Virol Methods] 2017 May; Vol. 243, pp. 92-97. Date of Electronic Publication: 2017 Feb 04. - Publication Year :
- 2017
-
Abstract
- Grass carp reovirus (GCRV) is the causative agent of the grass carp hemorrhagic disease that has resulted in severe economic losses in the grass carp (Ctenopharyngodon idella) farming industry in China. Early diagnosis and vaccine administration are important priorities for GCRV control. In this study, a nucleic acid sequence-based amplification with enzyme-linked immunosorbent assay (NASBA-ELISA) was developed for to detect genotype II GCRV (GCRV- II). Primers specifically targeting viral RNA genome segment 6 were utilized for amplification in an isothermal digoxigenin-labeling NASBA process, resulting in DIG-labeled RNA amplicons. The amplicons were hybridized to specific biotinylated DNA probes and the products were detected colorimetrically using horseradish peroxidase and a microplate reader. The new method is able to detect GCRV at 14 copies/μL within 5h and had a diagnostic sensitivity and a specificity of 100% when GCRV-II and non-target virus were tested. This NASBA-ELISA was evaluated using a panel of clinical samples (n=103) to demonstrate that it is a rapid, effective and sensitive method for GCRV detection in grass carp aquaculture.<br /> (Copyright © 2017 Elsevier B.V. All rights reserved.)
- Subjects :
- Animals
Carps
China
Diagnostic Tests, Routine methods
Fish Diseases virology
Reoviridae genetics
Reoviridae Infections virology
Sensitivity and Specificity
Time Factors
Colorimetry methods
Enzyme-Linked Immunosorbent Assay methods
Fish Diseases diagnosis
Genotype
Reoviridae isolation & purification
Reoviridae Infections veterinary
Self-Sustained Sequence Replication methods
Subjects
Details
- Language :
- English
- ISSN :
- 1879-0984
- Volume :
- 243
- Database :
- MEDLINE
- Journal :
- Journal of virological methods
- Publication Type :
- Academic Journal
- Accession number :
- 28174075
- Full Text :
- https://doi.org/10.1016/j.jviromet.2017.02.001