A Rapid and Sensitive Loop-Mediated Isothermal Amplification Assay for the Detection of Mulberry Mosaic Dwarf-Associated Virus.

Mulberry mosaic dwarf-associated virus (MMDaV), a member of the genus Mulcrilevirus in the family Geminiviridae, poses a serious threat to mulberry cultivation and growth in China. Early and accurate diagnosis is a prerequisite for the effective prevention and control of MMDaV-related disease. In this study, a loop-mediated isothermal amplification (LAMP)-based detection method was established for the specific identification of MMDaV. Three pairs of specific primers were designed targeting the nucleotide sequences of the MMDaV V2 gene. The optimized LAMP reaction was performed at a constant temperature of 65 °C for 60 min, and the established method exhibited high specificity with no cross-reactivity observed against other tested geminiviruses. The LAMP method achieved a detection limit of 200 pg of target MMDaV DNA, which was tenfold more sensitive than conventional PCR. Furthermore, the amplification results could be directly visualized via distinct color change. Collectively, the developed LAMP method enables efficient, sensitive, and specific isothermal detection of MMDaV and holds great promise for rapid diagnosis of mulberry viral disease.