Studies On Entomopathogenic Fungi: Evaluations Of Germination Protocols For Assesssing Conidial Quality And Modified Atmosphere Packaging For Enhancing High-Temperature Shelf Life
Conidia of the entomopathogenic fungi Beauveria bassiana and Metarhizium anisopliae are the most common active ingredients in commercially available mycoinsecticides, and longer shelf-lives under non-refrigerated conditions could increase their limited market acceptance. Contrasting results from commonly used germination protocols constitute an important impediment to consistent, universally comparable efficacy and shelf-life studies. A series of laboratory experiments investigated the underlying causes of this problem. Lower viabilities due to imbibitional damage were observed with decreasing water activity (aw) and decreasing temperature of the water/surfactant solutions used in preparing conidial suspensions. Dry conidia of M. anisopliae (strain CB-10) and M. acridum (strain IMI-330189), in contrast to those of Beauveria bassiana (strain GHA), were highly susceptible to imbibitional damage. Germination of the driest M. anisopliae conidia was drastically reduced to 66% after immersion at 25 degree C and to less than 1% at 0.5 degree C. Significant loss of viability was prevented when conidia were immersed in warm water (ca. 34 degree C) or when slowly rehydrated by humidification prior to immersion. Inclusion/exclusion of debilitated conidia (conidia exhibiting delayed germination or hypersensitivity to imbibitional damage) in germination counts was identified as an additional factor contributing to variability in quality assessments. Germination protocols based on fast rehydration and shorter incubation times are recommended as a means of discounting debilitated conidia in viability determinations. In shelf-life experiments, longevities of B. bassiana conidia were similar and significantly greater in storage containers flushed with CO2, N2, H2, or He, than in containers flushed with high concentrations of O2 . In atmospheres with 20% CO2/80% N2, viability measured after fast rehydration and incubation for 24 h remained high (87%) after a 16-month storage period at 25 degree C. Active packaging systems comprising aluminized bags and commercially available O2- and moisture-absorbing materials, consistently preserved viability at levels of 8089% for six months at 40 degree C and two months at 50 degree C. Results suggest that optimal aw for long-term storage of conidia under anaerobic conditions are lower than previously reported from studies of storage in the presence of O2. To our knowledge, these are the longest high-temperature survival times for B. bassiana conidia reported to date.
dissertation or thesis