Characterization of two genes encoding Bacillus thuringiensis insecticidal crystal proteins toxic to Coleoptera species

Abstract

Bacilus thuringiensis EG2838 and EG4961 are highly toxic to Colorado potato beetle larvae, and only strain EG4961 is toxic to southern corn rootworm larvae. To investigate the cause of the different insecticidal activities of EG2838 and EG4961, cryIII-type genes toxic to coleopterans were cloned from each strain. The crylIIB gene, cloned as part of an 8.0-kb EcoRI fragment of EG2838 DNA, encoded a crystal protein (CryIIIB) of 74,237 Da. The cryIIIB2 gene, cloned as part of an 8.3-kb PstI-Asp718 fragment of EG4961 DNA, encoded a crystal protein (CryIIIB2) of 74,393 Da that was 94% identical to CryIlIB. Analysis of the transcriptional start sites showed that cryIIIB and cryIIIB2 were initiated from a conserved region located within 130 nucleotides upstream from the translation start sites of both genes. Although the CryIIIB and CryIIIB2 proteins were similar in sequence, they displayed distinct insecticidal activities: CryIIIB was one-third as toxic as CryIIIB2 to Colorado potato beetle larvae, and CryHIB2, but not CryIIIB, was toxic to southern corn rootworm larvae. Genes encoding crystal proteins of approximately 32 and 31 kDIa were located adjacent to the cryIIIB and cryIIIB2 genes, respectively. The 32-and 31-kDa crystal proteins failed to enhance the insecticidal activities of CryIIIB and CryIIIB2.

Cloning and characterization of a novel Cry1A toxin fromBacillus thuringiensiswith high toxicity to the Asian corn borer and other lepidopteran insects

“…The specificity of toxicity makes B. thuringiensis insecticidal proteins desirable alternative insecticides to synthetic chemicals for the control of pests in agriculture, forestry, and public health (Schnepf et al, 1998). Bioinsecticides derived from B. thuringiensis are widely used for the control of insects in the orders Lepidoptera, Diptera and Coleoptera and can be used to control insects in the order Hymenoptera as well as other types of pests including nematodes and mites (Chungjatupornchai et al, 1988;Donovan et al, 1992;Lee & Gill, 1997;Garcia-Robles et al, 2001;Bischof et al, 2006;Ruiz et al, 2006).…”

Evaluation of dietary effects of transgenic corn pollen expressing Cry3Bb1 protein on a non‐target ladybird beetle, Coleomegilla maculata

“…The cry3Bb1 gene encodes the insecticidal Cry3Bb1 protein, which has insecticidal activity against Coleoptera species such as corn rootworms, Diabrotica spp. and the Colorado potato beetle, Leptinotarsa decemlineata (Say) (Donovan et al, 1992). Recent field trials have demonstrated that this transgenic corn event expressing Cry3Bb1 protein is highly efficacious against larvae of Diabrotica species (Monsanto Company, unpublished data) and can provide corn growers an effective tool for managing corn rootworms.…”

Scientific Opinion on application (EFSA‐GMO‐CZ‐2008‐54) for placing on the market of genetically modified insect resistant and herbicide tolerant maize MON 88017 for cultivation under Regulation (EC) No 1829/2003 from Monsanto

“…(1) protection against certain coleopteran target pests belonging to the genus Diabrotica such as the larvae of Western corn rootworm (Diabrotica virgifera virgifera), Northern corn rootworm (Diabrotica barberi), Southern corn rootworm (Diabrotica undecimpunctata howardi) and Mexican corn rootworm (Diabrotica virgifera zea) by the introduction of a part of a B. thuringiensis subsp. kumamotoensis gene, encoding the insecticidal Cry3Bb1 protein (Donovan et al, 1992;Galitsky et al, 2001;Vaughn et al, 2005;Gray et al, 2007). The mode of action of the Cry3Bb1 protein and other Cry proteins is to bind selectively to specific receptors on the epithelial surface of the midgut of larvae of susceptible insect species, leading to death of larvae through pore formation, cell burst and subsequently septicemia (Broderick et al, 2006(Broderick et al, , 2009OECD, 2007;Bravo and Soberón, 2008;Kaiser-Alexnat, 2009;Raymond et al, 2009;Soberón et al, 2009;Sayed et al, 2010;Van Frankenhuyzen et al, 2010;Sanahuja et al, 2011).…”