Functional Analysis Of Three Adhesin Genes(adh1-3)in Beauveria Bassiana

As a classical entomopathogenic fungus,Beauveria bassiana has been widely used in the biological control of forestry and agricultural pests.Adherence of conidia to insect integument is a crucial step for initiation of fungal infection through cuticular penetration in fungal insect-pathogenic lifecycle.After successful attachment,conidia germinate and form germ tubes under appropriate conditions,followed by hyphal invasion into host body via cuticular penetration as a result of the actions of extracellular(secreted)enzymes required for cuticle degradation and/or the mechanic pressure of appressoria formed at the tips of germ tubes and colonization of host hemocoel,where fungal cells proliferate by yeast-like budding until host death from mummification.It was previously reported that,in Metarhizium anisopliae,conidial adherence to insect cuticle relies upon the Metarhizium adhesin 1(Mad1)rather than Mad2,another adhesin crucial for the fungal adherence to plant root surface.Mad1 and Mad2 have since been considered as respective players in fungal insect pathogenicity and plant root colonization.This study seeks to characterize three adhesins in B.bassiana,including Adh1 homologous to Mad1,Adh2 homologous to Mad2,and Adh3 known as filamentous hemagglutinin/adhesin and virulence factor in some animal-pathogenic bacteria.The main results are summarized as follows.Among the three adhesins,only Adh2 was found to play an active role in sustaining virulence and some phenotypes related to biological control potential of B.bassiana.Disruption of adh2 resulted in decreased conidial adherence to insect wing cuticle.Disruption of adh2 resulted in significantly attenuated virulence via normal cuticle infection or cuticle-bypassing infection(injection)of the Galleria mellonella larva,reduced blastospore production in a insect hemolymph-mimicking broth,largely reduced conidiation capacity,compromised conidial quality indicative of reductions in viability,hydrophobicity and UV resistance,but no growth defects on rich and scant media under normal and stressful culture conditions.The main phenotypic changes correlated well with transcriptional repression of developmental activator genes(brl A and aba A)required for aerial conidiation and submerged blastospore production and of key hydrophobin genes(hyd1 and hyd2)essential for hydrophobin biosynthesis and assembly into rodlet bundles of conidial coat to aid the adherence.In contrast,disruption of adh1 or adh3 caused insignificant changes in the mentioned phenotypes.These findings offer novel insight into essential/nonessential roles of the adhesin genes in fungal insect-pathogenic lifecycleand also suggest a likelihood that Adh1/Mad1 and Adh2/Mad2 orthologues could have evolved along distinct trajectories leading to the adaptation of the Beauveria and Metarhizium lineages to different host spectra and habitats.