Application and development of a TaqMan-based real-time PCR assay for rapid detection of snakehead vesiculovirus

Abstract Snakehead vesiculovirus (SHVV) is one of the primary pathogens responsible for viral diseases in the snakehead fish. A TaqMan-based real-time PCR assay was established for the rapid detection and quantification of SHVV in this study. Specific primers and fluorescent probes were designed for...

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Bibliographic Details
Published in:FEMS microbiology letters 2024-01, Vol.371
Main Authors: Gong, Cuiping, Zhu, Panpan, Ye, Jiaxin, Lou, Jianfeng, Zhang, Liwen, Liu, Xiaodan, Kong, Weiguang
Format: Article
Language:English
Subjects:
Animals
Fish Diseases - diagnosis
Iridoviridae - genetics
Perciformes - genetics
Real-Time Polymerase Chain Reaction
Reproducibility of Results
Rhabdoviridae Infections
Sensitivity and Specificity
Vesiculovirus - genetics
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Summary:Abstract Snakehead vesiculovirus (SHVV) is one of the primary pathogens responsible for viral diseases in the snakehead fish. A TaqMan-based real-time PCR assay was established for the rapid detection and quantification of SHVV in this study. Specific primers and fluorescent probes were designed for phosphoprotein (P) gene, and after optimizing the reaction conditions, the results indicated that the detection limit of this method could reach 37.1 copies, representing a 100-fold increase in detection sensitivity compared to RT-PCR. The specificity testing results revealed that this method exhibited no cross-reactivity with ISKNV, LMBV, RSIV, RGNNV, GCRV, and CyHV-2. Repetition experiments demonstrated that both intra-batch and inter-batch coefficients of variation were not higher than 1.66%. Through in vitro infection experiments monitoring the quantitative changes of SHVV in different tissues, the results indicated that the liver and spleen exhibited the highest viral load at 3 poi. The TaqMan-based real-time PCR method established in this study exhibits high sensitivity, excellent specificity, and strong reproducibility. It can be employed for rapid detection and viral load monitoring of SHVV, thus providing a robust tool for the clinical diagnosis and pathogen research of SHVV. Application and development of a TaqMan-based real-time PCR assay for detecting for rapid detection of snakehead vesiculovirus.
ISSN:1574-6968
1574-6968
DOI:10.1093/femsle/fnae018