Detection of <i>Phaeomoniella chlamydospora</i> in soil using speciesspecific PCR

Abstract

Phaeomoniella chlamydospora is a fungal pathogen of woody grapevine tissue that causes Petri disease As one potential source of inoculum is infested soil a SDS/phenol/chloroform DNA extraction method and PCR assay utilising speciesspecific primers were evaluated for the ability to detect P chlamydospora in grapevine nursery soil Using a nested PCR approach the assay was able to detect 10² conidia/ml when a spore suspension was added to sterilised soil samples and 50 fg when P chlamydospora genomic DNA was added directly to the reaction In this process preamplification of a 600 bp region of the ribosomal DNA was followed by amplification with primers Pch 1 and Pch 2 to produce a 360 bp speciesspecific product This highly sensitive diagnostic tool will be used in future studies to determine if pathogen propagules are present in soils collected from New Zealand grapevine nurseries