Rapid Detection of Duck Enteritis Virus with MIRA, MIRA-qPCR, and MIRA-LFD Assays

Duck viral enteritis (DVE) is an acute and highly contagious disease that affects waterfowl such as ducks, geese and swans. Duck enteritis virus (DEV) is the pathogen, causing huge economic losses to waterfowl farming in recent years. Establishing a rapid, simple, and visual detection should facilitate the early identification of DEV. After the amplification primers and reaction conditions were optimized, three multienzyme isothermal rapid amplification (MIRA) methods-basic MIRA, MIRA-quantitative PCR (MIRA-qPCR) and MIRA-lateral flow dipstick (MIRA-LFD)-were established to detect DEV. Specificity analyses showed that the three MIRA methods specifically detected DEV, with no cross-reaction with fowl adenovirus serotype 4, novel goose astrovirus, Muscovy duck reovirus, avian influenza virus subtype H9, or duck circovirus. The basic MIRA reaction was completed in 30 min at 35 °C, requiring only a pair of primers. Detection with MIRA-qPCR or MIRA-LFD was completed within 20 min, and the limits of detection were 1 × 10¹ copies/μL for both. MIRA-LFD required no specialized instruments, and the results could be viewed directly with the naked eye. Compared with the traditional PCR, MIRA assays are simple, rapid, and effective and therefore more suitable for the field detection of DEV.