POTENTIAL USE OF PCR-AMPLIFIED RIBOSOMAL INTERGENIC SEQUENCES IN THE DETECTION AND DIFFERENTIATION OF VERTICILLIUM WILT PATHOGENS

Verticillium wilt pathogens are widespread throughout the world and an important factor in most agricultural economies. The ribosomal genes from the two major pathogenic species, V. albo-atrum and V. dahliae, are essentially identical, but small differences were identified in the internal transcribed spacers (ITS 1 and ITS 2) of the two species. A cluster of three non-homologous nucleotides was observed in ITS 1 and a cluster of two in ITS 2. These differences permitted the synthesis of oligonucleotides that hybridized differentially with the rDNA of the two species and allowed for an efficient, fungus-specific amplification of either DNA sequence by a polymerase chain reaction (PCR). The results illustrate the effective use of intergenic sequences for the detection of fungus in a crop such as alfalfa and also suggest that PCR-amplified intergenic sequences may provide sensitive probes for the differentiation of closely related species even when the mature rRNAs are too homologous or contain no exploitable sequence differences. © 1991.