Rapid and sensitive detection of Taura syndrome virus (TSV) in shrimp based on an isothermal enzymatic recombinase amplification (ERA) assay

Taura syndrome in shrimp, caused by Taura syndrome virus (TSV), is a disease listed by the World Organisation for Animal Health (OIE) that has been identified in five continents, causing serious economic losses in years. Due to the lack of specific drugs available to treat Taura syndrome, an effective TSV detection method is urgently needed to ensure pathogen prediction and disease control. In this study, we developed and evaluated an isothermal enzymatic recombinase amplification-based assay (TSV-ERA) that was able to diagnose TSV within 20 min at an optimal temperature of 42 degrees C. In the reaction system, the optimal concentrations of the primer and the probe were 500 nM and 120 nM, respectively. When the plasmid standards were used as a detection template, the check-out time of TSV-ERA was 12 min (21.76 +/- 0.60 Ct), and the limit of detection (LOD) was 3.6 x 101 copies/mu L. Compared to the sensitivity of the TSV detection method recommended by the OIE, TSV-ERA was equal to real-time PCR (qPCR) and higher than one-step PCR. This assay was highly specific for TSV and had no cross-reactivity with Enterocytozoon hepatopenaei (EHP), white spot syndrome virus (WSSV), Vibrio parahaemolyticus strain causing acute hepatopancreatic necrosis disease (VpAHPND), infectious hypodermal and hematopoietic necrosis virus (IHHNV), and the genomic DNA of specific pathogen-free (SPF) shrimp. Compared with the common detection methods, the TSV-ERA assay was able to shorten the detection time and decrease the detection temperature while sustaining accurate sensitivity. Based on these results, our TSV-ERA detection method is simple, rapid, and effective, and has great potential in the detection of TSV.