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Applied and Environmental Microbiology, April 2000, p. 1509-1516, Vol. 66, No. 4
0099-2240/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.

Genetic and Biochemical Approach for Characterization of Resistance to Bacillus thuringiensis Toxin Cry1Ac in a Field Population of the Diamondback Moth, Plutella xylostella

Ali H. Sayyed,1 Robert Haward,1 Salvador Herrero,2 Juan Ferré,2 and Denis J. Wright1,*

Department of Biology, Imperial College of Science, Technology and Medicine, Silwood Park, Ascot, Berkshire SL5 7PY, United Kingdom,1 and Departament de Genètica, Universitat de València, 46100 Burjassot (València), Spain2

Received 21 September 1999/Accepted 13 January 2000

Four subpopulations of a Plutella xylostella (L.) strain from Malaysia (F4 to F8) were selected with Bacillus thuringiensis subsp. kurstaki HD-1, Bacillus thuringiensis subsp. aizawai, Cry1Ab, and Cry1Ac, respectively, while a fifth subpopulation was left as unselected (UNSEL-MEL). Bioassays at F9 found that selection with Cry1Ac, Cry1Ab, B. thuringiensis subsp. kurstaki, and B. thuringiensis subsp. aizawai gave resistance ratios of >95, 10, 7, and 3, respectively, compared with UNSEL-MEL (>10,500, 500, >100, and 26, respectively, compared with a susceptible population, ROTH). Resistance to Cry1Ac, Cry1Ab, B. thuringiensis subsp. kurstaki, and B. thuringiensis subsp. aizawai in UNSEL-MEL declined significantly by F9. The Cry1Ac-selected population showed very little cross-resistance to Cry1Ab, B. thuringiensis subsp. kurstaki, and B. thuringiensis subsp. aizawai (5-, 1-, and 4-fold compared with UNSEL-MEL), whereas the Cry1Ab-, B. thuringiensis subsp. kurstaki-, and B. thuringiensis subsp. aizawai-selected populations showed high cross-resistance to Cry1Ac (60-, 100-, and 70-fold). The Cry1Ac-selected population was reselected (F9 to F13) to give a resistance ratio of >2,400 compared with UNSEL-MEL. Binding studies with 125I-labeled Cry1Ab and Cry1Ac revealed complete lack of binding to brush border membrane vesicles prepared from Cry1Ac-selected larvae (F15). Binding was also reduced, although less drastically, in the revertant population, which indicates that a modification in the common binding site of these two toxins was involved in the resistance mechanism in the original population. Reciprocal genetic crosses between Cry1Ac-reselected and ROTH insects indicated that resistance was autosomal and showed incomplete dominance. At the highest dose of Cry1Ac tested, resistance was recessive while at the lowest dose it was almost completely dominant. The F2 progeny from a backcross of F1 progeny with ROTH was tested with a concentration of Cry1Ac which would kill 100% of ROTH moths. Eight of the 12 families tested had 60 to 90% mortality, which indicated that more than one allele on separate loci was responsible for resistance to Cry1Ac.

* Corresponding author. Mailing address: Department of Biology, Imperial College, Silwood Park, Ascot, Berkshire SL5 7PY, United Kingdom. Phone: 44 207 594248. Fax: 207 594239. E-mail: d.wright{at}bio.ic.ac.uk.

Applied and Environmental Microbiology, April 2000, p. 1509-1516, Vol. 66, No. 4
0099-2240/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.


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