PRODUCTION AND MANIPULATION OF INDIVIDUAL MICROSCLEROTIA OF VERTICILLIUM-DAHLIAE FOR USE IN STUDIES OF SURVIVAL

A method for the laboratory culture of Verticillium dahliae and the harvest, isolation, and manipulation of single microsclerotia (MS) is outlined. MS produced in large numbers and stored at several temperatures maintained high, uniform viability for 12 mo or more. Thus, a single lot of MS could be used in long-term experiments on the effect of microbial interaction on MS survival. The MS were harvested from semisolid media by wet sieving, mixed with sand to dry without clumping, and sorted into size classes by dry sieving. After experimentation, single MS were transferred on the tip of a sterile needle into one of 25 squares marked on an agar plate, and germination and colony growth were monitored. The lethal effect of the treatment was reflected by the number of dead MS. Sublethal effects could also be measured as a reduction in colony growth because larger MS (>75 mu m) exhibited faster, more synchronous germination and produced larger colonies than smaller MS.