Rapid Detection and Quantification of Infectious Hypodermal and Hematopoietic Necrosis Virus in Whiteleg Shrimp Penaeus vannamei Using Real-time Loop-mediated Isothermal Amplification

摘要

A one-step, single tube, real-time loop-mediated isothermal amplification (real-time LAMP) assay was developed to detect a non-structural protein encoding gene of infectious hypodermal and haematopoietic necrosis virus (IHHNV). The real-time LAMP method for IHHNV is simple and rapid with specific amplification within 60 min at 63 deg C employing four primers. This method requires less time than the PCR method and is specific for IHHNV when analysed with PRDV (= WSSV), YHV and TSV DNA/cDNA, and healthy shrimp DNA. Sensitivity analysis revealed this method is capable of detecting as few as 10~2-10~3 copies//iL, suggesting it can be used as an alternative quantitative detection method for IHHNV in diagnostic laboratories.