Simultaneous Detection and Differentiation of Vairimorpha spp. and Nosema spp. by Multiplex Polymerase Chain Reaction

摘要

A multiplex polymerase chain reaction (PCR) was developed for the simultaneous detection and differentiation among Vairimorpha spp. and Nosema spp. and identification of Vairimorpha necatrix from Lepidoptera insects. Three sets of primers were selected from different genomic sequences to specifically amplify an 831 bp amplicon within the SSU rRNA gene, specific for both Vairimorpha spp. and Nosema spp. (MSSR primer); a 542 bp amplicon within the SSU rRNA gene, specific for Vairimorpha spp. (VSSU primer); and a 476 bp amplicon within the actin gene, specific for Vairimorpha necatrix (VNAG primer). Using the primers in conjunction with multiplex PCR, it was possible to detect Vairimorpha spp. and Nosema spp. and to differentiate between them. The sensitivity of this PCR assay was approximately 10 spores per milliliter. It is proposed that the multiplex PCR is a sensitive, specific, and rapid tool that can serve as a useful differential diagnostic tool for detecting Vairimorpha spp. and Nosema spp. in Lepidoptera insect.