title: Construction of an improved mycoinsecticide overexpressing a toxic protease.
abstract: Mycoinsecticides are being used for the control of many insect pests as an environmentally acceptable alternative to chemical insecticides. A key aim of much recent work has been to increase the speed of kill and so improve commercial efficacy of these biocontrol agents. This might he achieved by adding insecticidal genes to the fungus, an approach considered to have enormous potential for the improvement of biological pesticides. We report here the development of a genetically improved entomopathogenic fungus. Additional copies of the gene encoding a regulated cuticle-degrading protease (Pr1) from Metarhizium anisopliae were inserted into the genome of M. anisopliae such that Pr1 was constitutively overproduced in the hemolymph of Manduca sexta, activating the prophenoloxidase system. The combined toxic effects of Pr1 and the reaction products of phenoloxidase caused larvae challenged with the engineered fungus to exhibit a 25% reduction in time of death and reduced food consumption by 40% compared to infections by the wild-type fungus. In addition, infected insects were rapidly melanized, and the resulting cadavers were poor substrates for fungal sporulation. Thus, environmental persistence of the genetically engineered fungus is reduced, thereby providing biological containment.
authors.affiliation: Boyce Thompson Institute, Cornell University, Ithaca NY 14853, USA.
authors.forename: R
authors.initials: R
authors.lastname: St Leger
authors.forename: L
authors.initials: L
authors.lastname: Joshi
authors.forename: M J
authors.initials: MJ
authors.lastname: Bidochka
authors.forename: D W
authors.initials: DW
authors.lastname: Roberts
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D018076:DNA, Complementary; D004789:Enzyme Activation; D004792:Enzyme Precursors / Q000378:metabolism; D005656:Fungal Proteins / Q000235:genetics* / Q000633:toxicity; D005658:Fungi / Q000235:genetics*; D007306:Insecticides; D007915:Lepidoptera / Q000201:enzymology; D008969:Molecular Sequence Data; D014442:Monophenol Monooxygenase / Q000378:metabolism; D010572:Pest Control, Biological; D012697:Serine Endopeptidases / Q000235:genetics* / Q000633:toxicity; D014158:Transcription, Genetic
chemical_list: D018076:DNA, Complementary; D004792:Enzyme Precursors; D005656:Fungal Proteins; D007306:Insecticides; D014442:Monophenol Monooxygenase; D012697:Serine Endopeptidases; C093192:pr1 protein, Metarhizium anisopliae