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The Occurrence of Photorhabdus-Like Toxin Complexes in Bacillus thuringiensis

Blackburn, Michael B., Martin, Phyllis A. W., Kuhar, Daniel, Farrar, Robert R., Gundersen-Rindal, Dawn E., Tse, Herman
Photorhabdus, toxicity, quantitative polymerase chain reaction, plasmids, phylogeny, phenotype, multilocus sequence typing, insect larvae, gene expression, entomopathogens, Photorhabdus luminescens, bacterial toxins, Bacillus thuringiensis, operon, sequence homology, Lymantria dispar, Leptinotarsa decemlineata, insecticidal properties, gene expression regulation, pathogenesis
Recently, genomic sequencing of a Bacillus thuringiensis (Bt) isolate from our collection revealed the presence of an apparent operon encoding an insecticidal toxin complex (Tca) similar to that first described from the entomopathogen Photorhabdus luminescens . To determine whether these genes are widespread among Bt strains, we screened isolates from the collection for the presence of tccC , one of the genes needed for the expression of fully functional toxin complexes. Among 81 isolates chosen to represent commonly encountered biochemical phenotypes, 17 were found to possess a tccC . Phylogenetic analysis of the 81 isolates by multilocus sequence typing revealed that all the isolates possessing a tccC gene were restricted to two sequence types related to Bt varieties morrisoni , tenebrionis , israelensis and toumanoffi . Sequencing of the ∼17 kb tca operon from two isolates representing each of the two sequence types revealed >99% sequence identity. Optical mapping of DNA from Bt isolates representing each of the sequence types revealed nearly identical plasmids of ca. 333 and 338 kbp, respectively. Selected isolates were found to be toxic to gypsy moth larvae, but were not as effective as a commercial strain of Bt kurstaki . Some isolates were found to inhibit growth of Colorado potato beetle. Custom Taqman® relative quantitative real-time PCR assays for Tc-encoding Bt revealed both tcaA and tcaB genes were expressed within infected gypsy moth larvae.