ISOLATION AND CHARACTERIZATION OF CONDITIONAL-LETHAL MUTANTS OF BEAUVERIA-BASSIANA

Temperature-sensitive mutants of Beauveria bassiana GK2016 were isolat ed and characterized. Heat-sensitive mutants that grew at 20 degrees C but not at 30 degrees C were generated using mutagenesis with ultravi olet light and several rounds of selection. After 2160 colonies from a heat-sensitive mutant enriched population were screened, 11 heat-sens itive strains were isolated for further study. Five mutant strains, HS 1, HS2, HS6, HS9, and HS11, were stable and closely resembled the wild -type strain with respect to morphology, growth rate, and enzyme synth esis at 20 degrees C. Characterization of macromolecular synthesis at 30 degrees C using a radiolabelled precursor uptake assay indicated th at three mutants, HS6, HS9, and HS11, had reduced levels of DNA, RNA, and protein synthesis at the nonpermissive temperature. Mutants affect ed in cell division and cell wall synthesis were characterized by micr oscopy. At 30 degrees C, mutant HS6 was defective in cellular compartm entalization and formed long, branched, aseptate mycelia that fragment ed easily. Mutant HS1 was defective in cell wall biosynthesis and at 3 0 degrees C ceased to maintain cell wall integrity and lysed. Mutants HS2 and HS9 possessed temperature-sensitive lesions that could not be specified. None of the mutants were directly affected in either nuclea r division or mitosis as evidenced by the accumulation of Giemsa-stain able mitotic nuclei at 30 degrees C. Bioassays conducted with grasshop pers showed the efficacy of the mutant strains, with the exception of HS11, to be comparable to that of the parent strain, at 20 degrees C. Conversely, at 32 degrees C the mutant strains were uninfective wherea s the parental strain GK2016 was infective. It is our intention to use these strains to examine fundamental aspects of entomopathology by di ssecting fungal growth and development in vitro and extending these ob servations to pathogenesis in insects.