Ribosomal DNA-targeted oligonucleotide probe and PCR assay specific for Fusarium oxysporum

Abstract

A PCR assay and a hybridisation assay were developed for identification of Fusarium oxysporum. Based on sequence data from the 28S rDNA, two primers (PFO2 and PFO3) and one 20-mer probe (HFO1) were designed. These oligonucleotides were tested with 16 strains of F. oxysporum and 80 strains of 23 other Fusarium species or of eight species from other fungal genera. A PCR procedure for specific amplification of DNA from the F. oxysporum strains using primers PFO2 and PFO3 was set up. Under the conditions defined, no PCR product was obtained with these primers from the 80 other strains. The oligonucleotide probe HFO1 was labelled using a non-radioactive method and evaluated for specificity in dot blot hybridisation experiments with rDNA amplified by PCR. Under optimised conditions, the probe hybridised exclusively with DNA from F. oxysporum strains. Both PCR and hybridisation procedures were validated with 53 additional isolates of F. oxysporum representative of the populations present in four different soils. These specific assays appear to be reliable tools for the identification of F. oxysporum and may have various applications in ecological studies.