Targeted disruption of the polyketide synthase gene pks15 affects virulence against insects and phagocytic survival in the fungus Beauveria bassiana

Abstract

The reducing clade III polyketide synthase genes, including pks15, are highly conserved among entomopathogenic fungi. To examine the function of pks15, we used targeted disruption to investigate the impact of Beauveria bassiana pks15 on insect pathogenesis. Southern analysis verified that the Δpks15 mutant was disrupted by a single integration of the transformation cassette at the pks15 locus. The Δpks15 mutant had a slight reduction in radial growth, and it produced fewer spores. Our insect bioassays indicated the Δpks15 mutant to be significantly reduced in virulence against beet armyworms compared to wild type (WT), which could be partially accounted for by its markedly decreased ability to survive phagocytosis. Total haemocyte count decreased sharply by 50-fold from days 1-3 post-inoculation in insects infected with WT, compared to a 5-fold decrease in the Δpks15 mutant. The mutant also produced fewer hemolymph hyphal bodies than WT by 3-fold. In co-culture studies with amoebae that have phagocytic ability similar to that of insect haemocytes, at 48 h the mortality rate of amoebae engulfing Δpks15 decreased by 72 %, and Δpks15 CFU decreased by 83 % compared to co-culture with WT. Thus, the Δpks15 mutant had a reduced ability to cope with phagocytosis and highly reduced virulence in an insect host. These data elucidate a mechanism of insect pathogenesis associated with polyketide biosynthesis.

Keywords: Acanthamoeba castellanii; Entomopathogenic fungi; Insect pathogenesis; Phagocytosis; Polyketides; Reducing clade III polyketide synthase.