Coronatin-2 from the entomopathogenic fungus Conidiobolus coronatus kills Galleria mellonella larvae and incapacitates hemocytes

Abstract

Coronatin-2, a 14.5 kDa protein, was isolated from culture filtrates of the entomopathogenic fungus Conidiobolus coronatus (Costantin) Batko (Entomophthoramycota: Entomophthorales). After LC-MS/MS (liquid chromatography tandem mass spectrometry) analysis of the tryptic peptide digest of coronatin-2 and a mass spectra database search no orthologs of this protein could be found in fungi. The highest homology was observed to the partial translation elongation factor 1a from Sphaerosporium equinum (protein sequence coverage, 21%), with only one peptide sequence, suggesting that coronatin-2 is a novel fungal protein that has not yet been described. In contrast to coronatin-1, an insecticidal 36 kDa protein, which shows both elastolytic and chitinolytic activity, coronatin-2 showed no enzymatic activity. Addition of coronatin-2 into cultures of hemocytes taken from larvae of Galleria mellonella Linnaeus (Lepidoptera: Pyralidae), resulted in progressive disintegration of nets formed by granulocytes and plasmatocytes due to rapid degranulation of granulocytes, extensive vacuolization of plasmatocytes accompanied by cytoplasm expulsion, and cell disintegration. Spherulocytes remained intact, while oenocytes rapidly disintegrated. Coronatin-2 produced 80% mortality when injected into G. mellonella at 5 µg larva-1. Further study is warranted to determine the relevance of the acute toxicity of coronatin-2 and its effects on hemocytes in vitro to virulence of C. coronatus against its hosts.

Conidiobolus coronatus induces oxidative stress and autophagy response in Galleria mellonella larvae

“…One such species, C. coronatus, is a soil fungus which demonstrates entomopathogenicity towards a number of insects, including G. mellonella, Dendrolimus pini [35] and various collembolans [36]. It is known to produce various mycotoxins [37,38], including a 30 kDa protein believed to be toxic to G. mellonella larvae, coronatin-1 (36 kDa) and coronatin-2 (14.5 kDa) which compromise the function of hemocytes, and the 30-32 kDa serine protease, whose properties resemble subtilisin [35,[37][38][39]. A previous study also has shown that while exposure did not influence the fat bodies, characteristic changes were observed in the Malpighian tubules in G. mellonella larvae after infection [40].…”

Human Pathogenic Entomophthorales

“…The expressions of secondary toxic metabolites such as azoxybenzene dicarboxylic acid and hydroxymethyl azoxybenzene in C. coronatus during culture growth were reported (106). However, their release during infection in insects and mammals has yet to be confirmed (23,106,107).…”

“…They need a traumatic lesion on the host's skin to attach their conidia and for the formation of a germ tube to penetrate the infected tissue using lipolytic and proteolytic enzymes. In fact, the expression of these types of enzymes in cultures of entomophthoralean species has been reported (12,102,106,107). Most of these enzymes have been found to play important roles in infected insects (11,76,99,107).…”

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

“…mellonella is also a model host for human pathogens like Bacillus cereus [11], Candida albicans [12], Listeria monocytogenes [13] or Staphylococcus aureus [14]. This insect is also used in studies of entomopathogenic fungi [15][16][17]. Conidiobolus coronatus (Entomophthorales) is a soil fungus that is pathogenic to insects [18] and sometimes also to humans.…”