Cuticular fatty acids of Galleria mellonella (Lepidoptera) inhibit fungal enzymatic activities of pathogenic Conidiobolus coronatus

Abstract

The entomopathogenic fungus Conidiobolus coronatus produces enzymes that may hydrolyze the cuticle of Galleria mellonella. Of these enzymes, elastase activity was the highest: this figure being 24 times higher than NAGase activity 553 times higher than chitinase activity and 1844 times higher than lipase activity. The present work examines the differences in the hydrolysis of cuticles taken from larvae, pupae and adults (thorax and wings), by C. coronatus enzymes. The cuticles of the larvae and adult thorax were the most susceptible to digestion by proteases and lipases. Moreover, the maximum concentration of free N-glucosamine was in the hydrolysis of G. mellonella thorax. These differences in the digestion of the various types of cuticle may result from differences in their composition. GC-MS analysis of the cuticular fatty acids isolated from pupae of G. mellonella confirmed the presence of C 8:0, C 9:0, C 12:0, C 14:0, C 15:0, C 16:1, C 16:0, C 17:0, C 18:1, C 18:0, with C 16:0 and C 18:0 being present in the highest concentrations. Additional fatty acids were found in extracts from G. mellonella imagines: C 10:0, C 13:0, C 20:0 and C 20:1, with a considerable dominance of C 16:0 and C 18:1. In larvae, C 16:0 and C 18:1 predominated. Statistically significant differences in concentration (p≤0.05) were found between the larvae, pupae and imago for each fatty acid. The qualitative and quantitative differences in the fatty acid composition of G. mellonella cuticle occurring throughout normal development might be responsible for the varied efficiency of fungal enzymes in degrading larval, pupal and adult cuticles.

The interaction between cuticle free fatty acids (FFAs) of the cockroaches Blattella germanica and Blatta orientalis and hydrolases produced by the entomopathogenic fungus Conidiobolus coronatus

“…The lipases present in the C. coronatus enzyme cocktail demonstrated less hydrolytic activity against the cuticle samples than the proteases and chitinases; FFAs were only released from the oothecae of both species and thoraces of B. orientalis. Similar results have been noted against the previously described four fly species and G. mellonella [28,47]. It appears that in C. coronatus, lipases play a lesser role in the development of an infection to that of proteases and chitinases, in contrast with the pivotal role of lipolytic activity during M. anisopliae infection [27].…”

“…Our findings indicate that B. orientalis and B. germanica are not susceptible to infection by C. coronatus, but not to infection by other entomopathogenic https://doi.org/10.1371/journal.pone.0235785.g004 [35,[40][41][42] The mycelia of C. coronatus cultivated in vitro secrete a plethora of enzymes, however, the activities of fungal enzymes measured in vitro are not necessarily correlated with their importance in the infection process occurring in nature [15,[43][44][45][46]. The enzymatic cocktail released by C. coronatus mycelia degrades cuticle samples from susceptible insects far more effectively than those from resistant species and/or developmental stages [11,28,47]. Similar differences were observed in the present study for B. orientalis and B. germanica, particularly regarding the digestion of oothecae proteins by fungal proteases; this might indicate higher levels of total protein in B. germanica oothecae than in B. orientalis, or of proteins susceptible to digestion by C. coronatus proteases.…”

“…In contrast to the oothecae, the two cockroach species released similar, low amounts of amino acids, suggesting their protein content was low in wings and had similar protein compositions. C. coronatus proteases have been found to be highly effective against the wings proteins of four fly species (L. sericata, C. vicina, C. vomitoria, M. domestica) and those of G. mellonella [28,47]. It could suggest a lower content of degradable proteins in the cockroach wings compared to other insects we have studied in terms of efficiency of cuticular protein digestion by C. coronatus proteases.…”

“…Correlation between the concentration of compounds identified in the cuticle of two cockroach species and the efficiency of C. coronatus proteases, chitinases and lipases in degrading the cockroach cuticle. PLOS ONE demonstrated greater efficiency against both cockroach species compared to four fly species [28] and the wax moth [47] suggesting higher levels of chitin in cockroach cuticles. N-glucosamine was released from oothecae incubated with C. coronatus enzymatic cocktail containing chitinases, thus confirming the presence of chitin.…”

Heat shock proteins (HSP 90, 70, 60, and 27) in Galleria mellonella (Lepidoptera) hemolymph are affected by infection with Conidiobolus coronatus (Entomophthorales)

“…Our previous studies showed that G. mellonella larvae are susceptible for C. coronatus infection, with the majority of treated larvae dying 24 or 48 hours following contact with the fungal pathogen (79 and 92%, respectively). In the control group comprising larvae which had no contact with the fungus, mortality was much lower (16%) [38].…”