A Proteomic approach into biological control of sugarcane grubs

Metarhizium anisopliae is a naturally occurring biological control of many insects including the greyback canegrub (Dermolepida albohirtum), a sugarcane pest in Australia. While there have been some gene-based approaches into identifying determinants for biological control and developing improved strains, our study provides a comparative proteomics approach into identifying key proteins produced by Metarhizium anisopliae during infection of greyback canegrubs. At the same time, we have developed a proteomic map for the greyback canegrub larvae responsible for significant losses of the crop. Pathogenicity related proteins were identified by both a liquid culture and solid culture approach. Solid culture approach is thought to give a more realistic view of infection process compared to liquid culture. In order to identify novel target proteins, differential displays of proteomic maps of healthy/infected cuticle (HC/IC) as well as healthy/infected whole grub (HWG/IWG) were produced and analysed using Progenesis image analysis software. Proteomic map of healthy fungus (Metarhizium anisopliae) (MY) was also generated successfully. A total of 156 protein spots on HWG, 88 unique spots on IWG, 17 spots from HC and 14 spots from MY were analysed by mass spectrometry. Of these, 61 protein spots from HWG, 40 protein spots from IWG, 15 protein spots from HC and 9 protein spots from MY were confirmed by identification using mass spectrometry. Among the identified proteins were different forms of actin and tropomyosin, an ATP binding protein, arginine kinase, formate dehydrogenalase, enolase, tara like protein isoform and heat shock proteins. Further identification for most of the protein spots has been hindered due to the limited number of suitable/accessible databases. Metarhizium anisopliae has been successfully transformed to benomyl resistance using pBENA3, a plasmid containing the benA3 allele from Aspergillus nidulans using particle bombardment as a preparation for introducing genes encoding the identified pathogenesis factors into Metarhizium.