Quantitative isolation of the mycoparasite Verticillium biguttatum from soil

An improved method for the isolation of the mycoparasite Verticillium biguttatum from soil was developed. Initially, soil dilution plating onto potato dextrose agar (PDA) that had been colonized with Rhizoctonia solani anastomosis group (AG)-3 (RPs) was used as a selective medium for V. biguttatum. This method gave 73% recovery of V. biguttatum from brickearth soil in comparison with only 12% recovery using the existing method involving distribution of soil particles onto RPs using an Andersen Air Sampler. Subsequently, acidified RPs (potato yeast dextrose agar at pH 4.0 colonized with R. solani AG-3: ARPs) were developed and recovery of V. biguttatum on this medium was equivalent to that on RPs, but V. biguttatum propagule density could be quantified in only 7 days compared to 10 days on RPs. ARPs were then used as the standard isolation medium. Recovery of V. biguttatum differed with soil type, and highest recovery was obtained on ARPs if the medium had just been overgrown by R. solani AG-3. Of a range of R. solani AGs tested, colonization with AG-3 provided the optimal combination of a high recovery of V. biguttatum with a low contamination by the soil-microbiota. Gliocladium roseum and other fungi were isolated from within V. biguttatum colonies with increased frequency as soil dilution RPs and ARPs aged.