|Mitchell, Ashaki - Teddi|
|Blackburn, Michael - Mike|
|Farrar, Robert - Bob|
Submitted to: Applied and Environmental Microbiology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 8/9/2006
Publication Date: 6/15/2008
Citation: Goh, H.G., Gelman, D., Martin, P.A.W., Shropshire, A.D., Rowley, D.L., Blackburn, M.B., Choi, I.Y., Farrar, R.R. 2007. Insecticidal effect of new strains of Bacillus thuringiensis on the diamondback moth, Putella xylostella. Côté, J.C. Otvos, I.S., Schwartz, J.-L. and Vincent C. (eds.) Proceedings of the 6th Pacific Rim Conference on the biotechnology of Bacillus thuringiensis and its environmental impact. Érudit, Montréal, 141 pp. Interpretive Summary: The diamondback moth (DBM), Plutella xylostella, is a major agricultural insect pest that only attacks plants in the family Crucifera (e.g., cabbage, cauliflower, kale, turnip and Brussels sprouts). Host plants are damaged by larval mining and feeding. Bacillus thuringiensis (Bt) is a rod-shaped bacterium that occurs naturally in the soil and on plants. It has been found to be a very effective, environmentally-safe insect-specific biopesticide. In conjunction with spore formation, Bt produces crystal proteins that are toxic to certain groups of insects including moths. To be effective, Bt must be eaten by the insect larva and the proteinaceous crystal dissolved and digested in the gut to release the toxic protein(s) that will destroy the gut and kill the larva. A Bt strain (var. kurstaki) effective in killing DBM has been commercialized, but DBM is reported to be the only pest to evolve Bt resistance in open field populations. This study was conducted to identify and select other strains of Bt that are highly toxic to DBM and thus, would be good candidates for use as biopesticides against this serious pest. Eight Insect Biocontrol Laboratory (IBL) strains were found to cause high levels of larval mortality after two-four days of DBM feeding. When these strains were tested on a strain of DBM that was resistant to the commercial Bt, three of the strains (IBL 425, 24 and 194) were found to be highly toxic to the DBM. These strains should be useful for the development of insect-specific biopesticides for use against the DBM. We have partially sequenced some of the genes that are present in the toxic IBL strains. When the active genes are completed sequenced, using molecular techniques, plants can be protected from DBM attack by engineering them to express these insect-specific Bt toxins. This information will be of use to scientists and pest managers.
Technical Abstract: Twenty-eight strains of Bacillus thuringiensis (Bt) grown on T-3 agar were screened for toxicity against the diamondback moth (DBM), Plutella xylostella. Samples of Bt whole culture preparations (spores and crystals), solublized crystals (pH-treated Bt)] and spores alone were each applied to artificial diet prior to placing 2nd instar DBM larvae on the diet. Eight of the 28 Bt strains tested, caused high levels of larval mortality after 2-3 days of feeding. Whole culture Bt preparations were more toxic than samples that contained spores alone or samples that contained primarily dissolved crystal protein. With the addition of any one of these three preparations of Bt, DBM mortality increased in a dose-dependent manner. Preparations of pH-treated Bt were not stable to boiling and therefore, a heat-stable exotoxin was not responsible for the observed toxicity. Spores that were damaged (shaken with beads) were no longer insecticidal, indicating that spores must be intact to cause mortality. When the various Bt strains were tested on Dipel-resistant DBM, the LC50 was lowest for IBL 425, followed by IBL 24 and IBL 194. Thus, IBL 425 was the most promising for development as a bioinsecticide against DBM. Seven of the eight strains (not IBL 24) were found to have a cry1 gene (cry1Aa). IBL 425 also had a cry1B gene. Experiments are underway to determine the sequence of the cry genes present in our active Gt strains.