Production of the fungal biocontrol agent Ulocladium atrum by submerged fermentation: accumulation of endogenous reserves and shelf-life studies

A method was developed for the induction of submerged conidiation of Ulocla dium atrum Preuss (isolate 385) for the first time, using an oatmeal extrac t broth. Two inoculum types were produced by this process: spores and mycel ial fragments. Spore production was stimulated by reducing the broth water potential (Psi) to -2.1 MPa and adding 20 mM calcium chloride. In contrast, mycelial fragments were dominant at -7.0 MPa Psi. Maximum total inoculum ( mycelial fragments and conidia) yields were approximately 2x10(7) ml(-1) af ter 9 days incubation at 25 degreesC at 100 rpm. Biomass from liquid cultur es responded to water-stress by accumulating increased concentrations of en dogenous sugar alcohols (polyols), particularly glycerol. Long-term shelf-l ife studies showed that submerged inoculum from cultures subjected to an in termediate water-stress (-2.1 MPa Psi) and containing enhanced levels of gl ycerol (> 300 mg g(-1) freeze-dried material) retained viability significan tly better (P <0.05) than that from unstressed cultures, when assessed on a gar with fully available water. This level of viability was comparable to t hat of aerial U. atrum spores from a 4-week solid-substrate fermentation on oat grains. However, in contrast to aerial spores, the ability of submerge d biomass to germinate in drier conditions declined significantly after 6 m onths.