植物病理学および微生物学ジャーナル

概要

Rapid detection and quantification of Fusarium udum in soil and plant samples using real-time PCR

Sukumar Mesapogu, Bandamaravuri Kishore Babu, Achala Bakshi, Somasani S. Reddy, Sangeeta Saxena, Alok K. Srivastava and Dilip K. Arora

Real-time PCR based detection assay was developed for Fusarium udum, causing vascular wilt of pigeonpea. The Histone-3 gene of F. udum was targeted to design species-specific primers and probe. A comparative study was undertaken to develop a reliable and reproducible procedure for detection and quantification of F. udum from diverse samples. The sensitivity and specificity of oligonucleotides were evaluated through dot blot hybridization, standard and real-time PCR assays. The probe HFUSP showed high degree of sensitivity for DNA obtained from pure cultures of F. udum to that of environmental DNA samples. The qPCR assay specifically differentiated the F. udum from closely related species of Fusarium, other test microbes and environmental samples. The single melting curve at 84.17 and a monomorphic band of 200 bp indicates the specificity and authenticity of the PCR assays. Thus, real-time PCR assay can be used as a rapid and effective procedure that can detect minute amounts of F. udum from complex environments. Therefore, the real-time PCR assay demonstrated in the present study can be successfully used for detection and monitoring of F. udum for early infection and disease epidemiology of vascular wilt of pigeonpea.