Mycoinsecticide improvement againts Chagas disease vectors

MP. JUÁREZ; N. PEDRINI; R. CRESPO

Valencia, España

Congreso; IX European Multicolloquium of Parasitology; 2004

The first barrier against biological or chemical contact insecticides is the insect cuticle. In this lab we study the chemistry and biochemistry of insect cuticular lipids, and its application to improve or develop ecologically friendly vector control methods. The insect surface is covered by a thin layer of lipids with a main function restricting water loss. They also participate in the absorption of chemicals, regulating the activity of microorganisms, as well as in chemical communication processes. Fungal insecticides invade their host through the cuticle, having a great potential for controlling insects that feed by sucking juices, infection starts whith conidia germination on the insect cuticle. We will focus in the biochemistry of the initial cuticular degradation events and its potential as an alternative control methodology for triatomins. Entomopathogenic fungi are a diverse group including hundreds of species, many of them are under consideration for development as pest control agents, very few are registered currently. Efforts have been made to optimize formulation, production and application of virulent strains against a variety of insect pests. Reducing the time to kill is the aim of current research in this field. Our working hipothesis is based in the wellknown ability of certain microorganisms to utilize hydrocarbons as carbon source for their growth. It was first shown in this laboratory that entomopathogenic fungi have the ability to degrade the hydrocarbons of Triatoma infestans -a complex mixture of  straight and branched chains from 29 to 39 carbons-, it was also proposed the metabolic pathway describing the biochemical interactions between insect host cuticle and fungi, as well as its potential to control a variety of insect pests (Napolitano and Juárez, 1997). We obtained the first evidence of complete catabolism of insect-like hydrocarbons by the entomopathogenic fungi Beauveria bassiana and Metarhizium anisopliae (Deuteromycotina: Hyphomycetes); a variety of intermediate metabolites were also identified employing labelled substrates. In analogy to yeast systems, the first oxidation round is presumably carried out by a cytochrome P-450 enzyme system. The fatty acid or the fatty alcohol,  will traverse the peroxisomal membrane, and, after successive transformations will eventually provide the appropriate fatty acyl-CoA for complete degradation in the peroxisomes,  the site of b-oxidation in fungi. As in yeast systems, peroxisomal proliferation was reported to be related to alkane-grown induction in the fungi B. bassiana (Crespo et al., 2000). Catalase, the peroxisomal marker enzyme, is a tetrameric hemoprotein that, together with other oxidases, decompose excess H2O2 and other reactive oxygen species preventing their damaging effect on cellular components.  The aim of this project was to gain in the understanding of the biochemical interaction between entomopathogenic fungi and their insect host hydrocarbons so to improve their potential efficacy as mycoinsecticides. We report a reduction in the time to kill (< 17%) based on an enhanced hydrocarbon degradation by inducible fungal enzymes. Correspondingly, the specific activity of catalase increased very significantly (>8-fold). An apparent molecular mass of 54.7 kDa was determined for the peroxisomal catalase subunits, after purification to homogeneity by gel filtration and strong anion exchange chromatography. We propose alkane-growth induction to improve micoinsecticide performance, and to measure catalase activity as an appropriate marker of fungal induction. Work supported by funding through the ECLAT network, IC18*CT980366, WHO/TDR Contract A20036, National Agency for Promotion of Science and Technology, Argentina, ANPCYT/FONCYT Contract 08-09653. Napolitano, R., Juárez, M. P., 1997. Entomopathogenous fungi degrade epicuticular hydrocarbons of T.infestans. Arch. Biochem. Biophys 344: 208-214. Crespo, R.,  Juárez, M.P.,  Cafferata, L.F.R., 2000. Biochemical interaction between entomopathogenous fungi and their insect host-like hydrocarbons. Mycologia 92: 528- 536.