Transformation Systems

Modification of entomopathogenic fungi has long been contemplated, but rarely reported. A limited number of studies have reported successful insertion of foreign genes into entomopathogenic fungi. A precursor to manipulation of entomopathogenic fungi using molecular techniques has been the development of transformation systems. There are several aims of transforming entomopathogenic fungi. These techniques enable gene disruption methods to be applied, which can lead to greater understanding of the genetics of disease processes, or the ability to introduce DNA into fungi may allow the modification of cell processes, potentially allowing improvements in the use of entomopathogenic fungi for insect control.

The first transformation of an entomopathogenic fungus was reported by Goettel et al. (1990), where M. anisopliae was transformed to be benomyl tolerant using pBENA3, a plasmid containing the benA3 allele from Aspergillus nidulans. Since then, there have been other reports on transformation of the Dueteromycete entomopathogens using a variety of methods. St Leger et al. (1995) used eletroporation and biolistic delivery to transform M. anisopliae with the plasmids (pNOM102 and pBENA3) containing the ß-glucuronidase and benomyl resistance genes. The cotransformants showed normal growth rates and retained their pathogenicity to insects (Bombyx mori). Polyethylene glycol (PEG)-mediated transformation of protoplasts is another method for transformation of entomopathogenic fungi, as used with the P. fumosoroseus and P. lilacinus (Inglis et al. 1992) using benomyl as the selective agent. More recently, a heterologous transformation system for B. bassiana and M. anisopliae was developed based on the use of the A. nidulans nitrate reductase gene (niaD) (Sandhu et al. 2001). The niaD stable mutants of B. bassiana and M. anisopliae were selected by treatment of protoplasts with ethane methane sulfonate (EMS) and regenerated on chlorate medium.

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