Transformation of the entomopathogenic fungusMetarhizium flavovirideto high resistance to benomyl

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Abstract
A Brazilian isolate of Metarhizium flavoviride, which has been developed as a biocontrol agent against the grasshopper Rhammatocerus schistocercoides, was stably transformed to be resistant to benomyl (beta-tubulin gene). Highly resistant transformants were obtained which grew in benomyl concentrations greater than 30 times (200 µg mL-1) the concentration that inhibits wild type proliferation. These transformants were mitotically stable after 20 successive transfers on non-selective media. No significant differences in conidia yield were observed between stably transformed strains and wild type (CG423). Chymoelastase (Pr1) secretion was greater in some transformants than in the wild type. In the presence of benomyl, appressoria differentiation occurred at similar rates in CG423 and transformants. However, the percentage of conidial germination in the transformants was higher than in the wild type, indicating the potential use of these transformants along with benomyl. Additionally, the resistance levels of the transformants observed in the present study demonstrate the potential use of these transformants for assessing the persistence of a particular isolate in fields without this fungicide.Key words: Metarhizium flavoviride, genetic transformation, benomyl resistance, biocontrol.