Rapid detection of white spot syndrome virus in Penaeus vannamei based on real-time enzymatic recombinase amplification

White spot syndrome virus (WSSV) has caused high mortality in cultured shrimps, and lead to huge economic losses. Due to the lack of effective therapeutics for WSSV, a rapid, efficient, and sensitive detection assay will be beneficial to the prevention of the white spot disease (WSD) in shrimps. In the present study, an enzymatic recombinase amplification (ERA) assay was established and evaluated for the VP28 gene of WSSV (WSSV-ERA), which could be completed at a constant temperature of 42 degrees C within 30 min. The optimal primers and probe of WSSV-ERA were highly specific for WSSV and exhibited no cross-reaction with Vibrio parahaemolyticus causing acute hepatopancreatic necrosis disease (VpAHPND), infectious hypodermal and hematopoietic necrosis virus (IHHNV), Enterocytozoon hepatopenaei (EHP) and healthy shrimp genomic DNA samples. WSSV-ERA could be performed in only 11.91 +/- 0.12 min, with the detection limit of 2.3 copies/mu L. Compared with the WSSV-ERA, nested PCR could obtain the similar sensitivity in nearly 4 h, while quantitative real-time PCR (qPCR) need 22 +/- 0.39 min. Moreover, WSSV-ERA did not suffer from the preference of background shrimp DNA (about 700 ng). It has been also demonstrated that the WSSV-ERA assay could detect pathogens accurately and rapidly using seventy-nine samples from shrimp farms. All these results indicated that WSSV-ERA would provide an alternative assay for simple, rapid and sensitive diagnosis of WSSV and could be helpful to control WSSV infection in shrimp industry.

Automated summary

White spot syndrome virus (WSSV), a major threat to shrimp farming, lacks effective therapeutics. This study developed an enzymatic recombinase amplification (ERA) assay for rapid and sensitive detection of WSSV. The assay showed high specificity, short detection time, and accuracy in detecting WSSV in shrimp farms, providing an alternative method for controlling WSSV infection.