Optimizing Inoculum Production Methods for Infesting Soil with Phytophthora Species

Inoculum production is an important part of conducting research with soilborne Phytophthora species. One common method is to incubate Phytophthora cultures in nutrient-amended vermiculite. However, inoculum levels often vary among batches of inoculum, even when production methods remain the same, and incubation typically takes ≥6 weeks, increasing risks for delayed experiments if the resulting inoculum level is too low. A more reliable and rapid method is needed for future studies. Experiments were conducted to determine inoculum levels of Phytophthora cinnamomi and Phytophthora plurivora after incubation in V8 juice-amended vermiculite (standard method); evaluate how inoculum viability was affected by air-drying; develop a modified method that takes less time to produce a vermiculite-based inoculum; and evaluate the effect of storage on inoculum viability. Results showed that the standard method produced inoculum levels from 716 to 1,808 colony forming units (CFUs)/g and that drying to <78% moisture content significantly reduced viability. The modified method used 2-week-old Phytophthora cultures to infest vermiculite at 80% moisture content and produced inoculum levels from 214 to 525 CFU/g. Storage for >1 day generally reduced inoculum viability. Although inoculum levels from the modified method were lower than the standard method, inoculum levels for each isolate were more consistent between trials, and the modified method was 6 to 8 weeks faster. Production with the modified method can also be easily scaled up by infesting a greater volume of vermiculite with additional cultures of Phytophthora. These results are important because they help explain variability in soilborne Phytophthora inoculum production and storage and provide a new method for producing inoculum more quickly.