A loop-mediated isothermal amplification-based microfluidic chip for triplex detection of shrimp pathogens.

Abstract

Decapod iridescent virus 1 (DIV1), White spot syndrome virus (WSSV), and Enterocytozoon hepatopenaei (EHP) pose serious threats to the shrimp farming. To date, early detection remains an important way to control the occurrence and diffusion of these pathogens. Here, we developed for the first time, a loop-mediated isothermal amplification (LAMP)-based microfluidic chip detection system, which could detect DIV1, WSSV, and EHP simultaneously. The limits of detection (LoD) of the system were 10 copies/reaction for EHP and DIV1, and 102 copies/reaction for WSSV. The entire detection procedure could be completed rapidly in 40 min at 63°C with 100% specificity and had no cross-reaction with other common shrimp pathogens. This newly established method was further validated using 94 Penaeus vannamei clinical samples, which were comparable to a typical qPCR assay and revealed good stability and reproducibility. These results illustrate that this LAMP microfluidic chip detection system allows rapid triplex pathogen analysis and could satisfy the demands of the field and routine diagnoses in aquaculture.