Real-time fluorescence loop-mediated isothermal amplification for the diagnosis of hemorrhagic enteritis virus

• A one-step, economical, RealAmp assay for the rapid diagnosis of HEV was developed.
• RealAmp assay and nested PCR assay were performed on the field and experimental samples.
• The coincidence rate of the two methods was 100%.

Suspected cases of hemorrhagic enteritis associated with hemorrhagic enteritis virus (HEV) are becoming more frequent among yellow chickens in the Guangdong Province of China. In this study, we have developed a one-step, ecumenical, real-time fluorescence loop-mediated isothermal amplification (RealAmp) assay for the rapid diagnosis of HEV. The RealAmp assay was performed at 63 °C and reduced the assay time to 15 min, using a simple and portable device, the ESE-Quant Tube Scanner. The detection limit of DNA was 1 fg/μl, and the detection was specific only to HEV. We also used nested PCR to evaluate the application of the RealAmp assay. The coincidence rate of the two methods was 100%. Our data indicated that the RealAmp assay provides a sensitive, specific, and user-friendly diagnostic tool for the identification and quantification of HEV for field diagnosis and in laboratory research.