Submitted to: Applied and Environmental Microbiology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: August 16, 2011
Publication Date: August 25, 2011
Citation: Martin, P.A., Mongeon, P., Gundersen, D.E., Blackburn, M.B. 2011. Recovery of Bacillus thuringiensis and insect toxic related strains from forest soil. Applied and Environmental Microbiology. 2(7):285-291.
Interpretive Summary: Gypsy moths defoliate thousands of acres of forest and shade trees annually. One method of control is the use of an insect specific bacterial pathogen, Bacillus thuringiensis (Bt). These bacteria have often been applied multiple times in some areas, and there is a question how long these bacteria survive and whether they can reproduce in environments where they have been sprayed. For field identification, most microbiologists use a standard technique in which they isolate bacteria in culture and identify it. An alternative is to detect the presence of Bt bacteria by their ability to kill insects. Using a type of caterpillar that is very sensitive to Bt we isolated bacteria that killed caterpillars in 14 out of the 18 environmental sites sampled in an area where Bt was sprayed for several consecutive years for control of gypsy moth. Most of the bacteria recovered using caterpillar assay were Bt or closely related to Bt. In contrast, by using standard techniques, Bt could only be recovered from one site. Scientists interested in insecticidal bacteria application and environmental persistence will find this new technique accurate and useful.
We attempted to recover Bacillus thuringiensis (Bt) from soil that had been sprayed two years prior with Bt for gypsy moth control. By amplifying the bacteria found in the soil on bacterial agar and feeding this diverse microbial population to tobacco hornworm larvae, 15 spore-forming bacteria from 14 samples were found to kill these larvae. Most of the sporeforming bacteria (14) belonged to the Bacillus cereus group including 3 B. thuringiensis and B. weihenstephanensis. All the Bt strains and the most of the B. cereus strains were closely related (<0.001 substitutions/site in their 16S rDNA) to the strain that was sprayed. The other spore-former was Lysinobacillus fusiformis. All 15 strains were phenotypically distinguishable from one another, as well as from the Bt strain that was applied, by substrate utilization and antibiotic resistance, although 7 strains differed from the Bt strain applied by 5 or less of the 24 traits tested. The toxicity to gypsy moth larvae of the three Bt strains isolated was similar to the applied Bt strain. Thus, amplification of bacteria present in soil in combination a sensitive insect can recover insect toxic strains that are related to an applied strain.