Pre-penetration Events During Infection of Host Cuticle by Metarhizium Anisopliae

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Journal of Invertebrate Pathology





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The early involvement of cuticle degrading enzymes in infection was suggested by the presence of chymoelastase protease (Prl), esterase, and N-acetylglucosaminidase in ungerminated spores of Metarhizium anisopliae. Enzymes were released from spores by washing in water or dilute buffer solutions. This, together with the degradation of enzyme substrates by intact spores, indicates that enzyme activities are located at the surface of spores, and probably parallels the availability or release of enzymes under natural conditions. Levels of enzymes on conidia from infected Manduca sexta larvae were higher than those harvested from Sabauraud dextrose agar (SDA), indicating that environmental conditions in which spores develop can preadapt them for the pathogenic life style. Cell-bound enzymes were released from differentiating germlings in varying degrees by water, salt solutions, detergents, and mercaptoethanol. Release of Pr1 and esterase activities by salt solutions suggests that their binding to cell walls involves ionic bonds. By contrast, release of N-acetylglucosaminidase required the disruption of membranes. We suggest that binding of enzymes to fungal walls could, in part, explain localized action of cuticle degrading enzymes during host penetration. The esterase activity in conidia is represented by a single major activity (pI 7) and in infection structures by multiple isoenzymes, all characterizable as β-esterases. Esterase activites were greatest against short and intermediate length fatty acids with little activity above C10. The effects of surface topography on appressorium formation was studied using M. sexta cuticles and plastic replicas of the cuticles. Appressoria were only produced after extensive growth over the microfolds of the cuticle surface of early (1 day) fifth instar larvae. By contrast germination on the comparatively flat surface of 5-day fifth instar larvae allows appressorium formation close to the conidium. The results are discussed in the context of cuticle degradation during prepenetration fungal growth, and the effects of host morphology on pathogen behavior.

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