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Validation of qPCR from a crude extract for the rapid detection of white spot syndrome virus
White spot disease (WSD) has posed a serious threat to the China and the global shrimp aquaculture. In order to diagnose white spot syndrome virus (WSSV) early and prevent the spread and outbreak of WSD, it is necessary to establish a highly sensitive WSSV diagnosis method suitable for shrimp farmin...
Autores principales: | , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Springer International Publishing
2022
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9309450/ https://www.ncbi.nlm.nih.gov/pubmed/35910332 http://dx.doi.org/10.1007/s10499-022-00920-9 |
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author | Ma, Chao Tian, Zhuo Yang, Lili Cao, Jijuan |
author_facet | Ma, Chao Tian, Zhuo Yang, Lili Cao, Jijuan |
author_sort | Ma, Chao |
collection | PubMed |
description | White spot disease (WSD) has posed a serious threat to the China and the global shrimp aquaculture. In order to diagnose white spot syndrome virus (WSSV) early and prevent the spread and outbreak of WSD, it is necessary to establish a highly sensitive WSSV diagnosis method suitable for shrimp farming sites. In this study, a pre-amplification qPCR assay from the crude extract of samples heated lysis was established, which was further compared with the universal qPCR assay to verify the shrimp samples. The limit of detection (LOD) of pre-amplification qPCR assay and universal qPCR assay was 2.80 copies and 20.57 copies per reaction at 95% CI, respectively. It had good WSSV specificity and did not show cross-detection of infectious hypodermal and hematopoietic necrosis virus (IHHNV), hepatopancreatic parvovirus (HPV), Enterocytozoon hepatopenaei (EHP), acute hepatopancreas necrosis disease (AHPND), necrotizing hepatopancreatitis bacteria (NHPB), and decapod iridescent virus 1 (DIV1). A total of 36 shrimp samples were detected as WSSV DNA positive by pre-amplification qPCR with crude extract from samples heated lysis and universal qPCR with DNA extraction. The diagnostic sensitivity and specificity were 97.22% (85.5 ~ 99.9%, 95% CI) and 100% (81.5 ~ 100%, 95% CI), respectively. The agreement Kappa value was 0.959 (0.879 ~ 1, 95% CI), and the analysis results were basically consistent. Eliminating the tedious steps of extracting DNA and using pre-amplified qPCR to detect WSSV in shrimp, it is a good choice for aquaculture farms. GRAPHICAL ABSTRACT: [Image: see text] SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s10499-022-00920-9. |
format | Online Article Text |
id | pubmed-9309450 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2022 |
publisher | Springer International Publishing |
record_format | MEDLINE/PubMed |
spelling | pubmed-93094502022-07-25 Validation of qPCR from a crude extract for the rapid detection of white spot syndrome virus Ma, Chao Tian, Zhuo Yang, Lili Cao, Jijuan Aquac Int Article White spot disease (WSD) has posed a serious threat to the China and the global shrimp aquaculture. In order to diagnose white spot syndrome virus (WSSV) early and prevent the spread and outbreak of WSD, it is necessary to establish a highly sensitive WSSV diagnosis method suitable for shrimp farming sites. In this study, a pre-amplification qPCR assay from the crude extract of samples heated lysis was established, which was further compared with the universal qPCR assay to verify the shrimp samples. The limit of detection (LOD) of pre-amplification qPCR assay and universal qPCR assay was 2.80 copies and 20.57 copies per reaction at 95% CI, respectively. It had good WSSV specificity and did not show cross-detection of infectious hypodermal and hematopoietic necrosis virus (IHHNV), hepatopancreatic parvovirus (HPV), Enterocytozoon hepatopenaei (EHP), acute hepatopancreas necrosis disease (AHPND), necrotizing hepatopancreatitis bacteria (NHPB), and decapod iridescent virus 1 (DIV1). A total of 36 shrimp samples were detected as WSSV DNA positive by pre-amplification qPCR with crude extract from samples heated lysis and universal qPCR with DNA extraction. The diagnostic sensitivity and specificity were 97.22% (85.5 ~ 99.9%, 95% CI) and 100% (81.5 ~ 100%, 95% CI), respectively. The agreement Kappa value was 0.959 (0.879 ~ 1, 95% CI), and the analysis results were basically consistent. Eliminating the tedious steps of extracting DNA and using pre-amplified qPCR to detect WSSV in shrimp, it is a good choice for aquaculture farms. GRAPHICAL ABSTRACT: [Image: see text] SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s10499-022-00920-9. Springer International Publishing 2022-07-25 2022 /pmc/articles/PMC9309450/ /pubmed/35910332 http://dx.doi.org/10.1007/s10499-022-00920-9 Text en © The Author(s), under exclusive licence to Springer Nature Switzerland AG 2022 This article is made available via the PMC Open Access Subset for unrestricted research re-use and secondary analysis in any form or by any means with acknowledgement of the original source. These permissions are granted for the duration of the World Health Organization (WHO) declaration of COVID-19 as a global pandemic. |
spellingShingle | Article Ma, Chao Tian, Zhuo Yang, Lili Cao, Jijuan Validation of qPCR from a crude extract for the rapid detection of white spot syndrome virus |
title | Validation of qPCR from a crude extract for the rapid detection of white spot syndrome virus |
title_full | Validation of qPCR from a crude extract for the rapid detection of white spot syndrome virus |
title_fullStr | Validation of qPCR from a crude extract for the rapid detection of white spot syndrome virus |
title_full_unstemmed | Validation of qPCR from a crude extract for the rapid detection of white spot syndrome virus |
title_short | Validation of qPCR from a crude extract for the rapid detection of white spot syndrome virus |
title_sort | validation of qpcr from a crude extract for the rapid detection of white spot syndrome virus |
topic | Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9309450/ https://www.ncbi.nlm.nih.gov/pubmed/35910332 http://dx.doi.org/10.1007/s10499-022-00920-9 |
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