Different Mechanisms of Resistance to Bacillus thuringiensis Toxins in the Indianmeal Moth (original) (raw)
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Journal of Economic Entomology, 2000
We studied mechanisms of resistance to Bacillus thuringiensis insecticidal crystal protein Cry1C in the diamondback moth, Plutella xylostella (L.). Binding assays with midgut brush border membrane vesicles prepared from whole larvae showed no signiÞcant difference between resistant and susceptible strains in binding of radioactively-labeled Cry1C. These results indicate that reduced binding of Cry1C to midgut membrane target sites did not cause resistance to Cry1C. Thus, the mechanism of resistance to Cry1C differs from that observed in several previously reported cases of resistance to Cry1A toxins in diamondback moth. We tested Cry1C toxin and Cry1C crystalline protoxin against resistant and susceptible larvae using leaf disk bioassays. After adjusting for the size difference between Cry1C toxin and protoxin, we found that with resistant larvae, toxin was signiÞcantly more toxic than protoxin. In contrast, with susceptible larvae, no signiÞcant difference in toxicity occurred between Cry1C toxin and protoxin. The resistance ratios for Cry1C were 19 for toxin and 48 for protoxin. These results suggest that reduced conversion of Cry1C protoxin to toxin is a minor mechanism of resistance to Cry1C. Because neither reduced binding nor reduced conversion of protoxin to toxin appear to be major mechanisms, one or more other mechanisms are important in diamondback moth resistance to Cry1C.
Applied and Environmental Microbiology, 2001
We tested toxins of Bacillus thuringiensis against larvae from susceptible, Cry1C-resistant, and Cry1A-resistant strains of diamondback moth ( Plutella xylostella ). The Cry1C-resistant strain, which was derived from a field population that had evolved resistance to B. thuringiensis subsp. kurstaki and B. thuringiensis subsp. aizawai , was selected repeatedly with Cry1C in the laboratory. The Cry1C-resistant strain had strong cross-resistance to Cry1Ab, Cry1Ac, and Cry1F, low to moderate cross-resistance to Cry1Aa and Cry9Ca, and no cross-resistance to Cry1Bb, Cry1Ja, and Cry2A. Resistance to Cry1C declined when selection was relaxed. Together with previously reported data, the new data on the cross-resistance of a Cry1C-resistant strain reported here suggest that resistance to Cry1A and Cry1C toxins confers little or no cross-resistance to Cry1Bb, Cry2Aa, or Cry9Ca. Therefore, these toxins might be useful in rotations or combinations with Cry1A and Cry1C toxins. Cry9Ca was much mor...
Journal of Economic Entomology, 2000
We studied mechanisms of resistance to Bacillus thuringiensis insecticidal crystal protein Cry1C in the diamondback moth, Plutella xylostella (L.). Binding assays with midgut brush border membrane vesicles prepared from whole larvae showed no signiÞcant difference between resistant and susceptible strains in binding of radioactively-labeled Cry1C. These results indicate that reduced binding of Cry1C to midgut membrane target sites did not cause resistance to Cry1C. Thus, the mechanism of resistance to Cry1C differs from that observed in several previously reported cases of resistance to Cry1A toxins in diamondback moth. We tested Cry1C toxin and Cry1C crystalline protoxin against resistant and susceptible larvae using leaf disk bioassays. After adjusting for the size difference between Cry1C toxin and protoxin, we found that with resistant larvae, toxin was signiÞcantly more toxic than protoxin. In contrast, with susceptible larvae, no signiÞcant difference in toxicity occurred between Cry1C toxin and protoxin. The resistance ratios for Cry1C were 19 for toxin and 48 for protoxin. These results suggest that reduced conversion of Cry1C protoxin to toxin is a minor mechanism of resistance to Cry1C. Because neither reduced binding nor reduced conversion of protoxin to toxin appear to be major mechanisms, one or more other mechanisms are important in diamondback moth resistance to Cry1C.
Applied and Environmental Microbiology, 2000
Four subpopulations of a Plutella xylostella (L.) strain from Malaysia (F 4 to F 8 ) 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 F 9 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 F 9 . 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 (F 9 to F 13 ) to give a resistance ratio of >2,400 compared with UNSEL-MEL. Binding studies with 125 I-labeled Cry1Ab and Cry1Ac revealed complete lack of binding to brush border membrane vesicles prepared from Cry1Ac-selected larvae (F 15 ). 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 F 2 progeny from a backcross of F 1 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.
Journal of Economic Entomology, 1996
Previous results have shown that diamondback moth, Plutella xylostella (L.), populations resistant to toxins from Bacillus thuringiensis subsp. kurstaki were susceptible to toxin CryIC. Use of commercial formulations of B. thuringiensis subsp. ai;:;awaithat contain CrylC has increased recently. Analysis of two commercial formulations by high pressure liquid chromatography showed that CryIC accounted for 26% of the CryI protein in the B. thuringiensis subsp. ai;:;awaiformulation, but did not occur in the B. thuringiensis subsp. kurstaki formulation. CrylAb was the most abundant Cryl protein in the commercial formulations of B. thuringiensis subspp. ai;:;awai and kurstaki. We found resistance to CrylC in a field population of diamondback moth from Hawaii that had been treated with B. thuringiensis subsp. aizawai. Leaf residue bioassays showed that, at 5 d after treatment with CryIC, LCsos for colonies derived from this population in 1993 and 1995 were ""20 times greater than the LC50 for a susceptible laboratory colony. For a nearby population that had not been treated with B. thuringiensis subsp. aizawai, responses to CrylC did not differ significantly from those of the susceptible laboratory colony. Resistance to CrylAb was lower in a CrylC-resistant colony than in a CrylC-susceptible colony that had been selected with B. thuringiensis subsp. kurstaki. These results suggest that the gene(s) conferring resistance to CrylC segregate independently from the gene(s) conferring resistance to CryIAb. In contrast with previous results with colonies derived in 1989, resistance to B. thuringiensis subsp. kurstaki in a colony derived in 1993 from the same field population did not decline when exposure to B. thuringiensis stopped. Thus, stability of resistance is not necessarily a fixed character, even for a specific population and pesticide. Despite substantial resistance to CrylC and B. thuringiensis subsp. kurstaki, resistance to a spore-crystal formulation of B. thuringiensis subsp. ai;:;awaiwas only 2-to 4-fold.
Resistance: A Threat to the Insecticidal Crystal Proteins of Bacillus thuringiensis
The Florida Entomologist, 1995
Insecticidal crystal proteins (also known as δ-endotoxins) synthesized by the bacterium Bacillus thuringiensis Berliner (Bt) are the active ingredient of various environmentally friendly insecticides that are 1) highly compatible with natural enemies and other nontarget organisms due to narrow host specificity, 2) harmless to vertebrates, 3) biodegradable in the environment, and 4) highly amenable to genetic engineering. The use of transgenic plants expressing Bt δ-endotoxins has the potential to greatly reduce the environmental and health costs associated with the use of conventional insecticides. The complex mode of action of Bt is the subject of intensive research. When eaten by a susceptible insect δ-endotoxin crystals are solubilized in the midgut; proteases then cleave protoxin molecules into activated toxin which binds to receptors on the midgut brush border membrane. Part of the toxin molecule inserts into the membrane causing the midgut cells to leak, swell, and lyse; death results from bacterial septicemia. Insecticides formulated with Bt account for less than 1% of the total insecticides used each year worldwide because of high cost, narrow host range, and comparatively low efficacy. Environmental contamination, food safety concerns, and pest resistance to conventional insecticides have caused a steady increase in demand for Bt-based insecticides. The recent escalation of commercial interest in Bt has resulted in more persistent and efficacious formulations. For example, improved Bt-based insecticides have allowed management of the diamondback moth, Plutella xylostella (L.). Unfortunately this has resulted in the evolution of resistance to δ-endotoxins in P. xylostella populations worldwide. The recent appearance of Bt resistance in the field, corroborated by the results of laboratory selection experiments, demonstrates genetically-based resistance in several species of Lepidoptera, Diptera, and Coleoptera. The genetic capacity to evolve resistance to these toxins is probably Florida Entomologist 78(3) September, 1995 ducir la presión de seleccción, minimizando la exposición a Bt e incrementando otros factores de mortalidad, para disminuir la velocidad de adaptación de la plaga a Bt. The bacterium Bacillus thuringiensis Berliner (Bt) is a complex of subspecies characterized by their ability to synthesize crystalline inclusions during sporulation. These crystalline inclusions are comprised of relatively high quantities of one or more glycoproteins known as δ-endotoxins or Cry toxins (Table 1). The toxins produced by Bt play a vital role in the pathogenicity of this bacterium to insects and other invertebrates. The Cry toxins have enormous commercial value as safe, biodegradable pesticides. The specificity of Bt toxicity is highly desirable in integrated pest management (IPM) programs, particularly in sensitive aquatic and forest ecosystems where other life forms, including many beneficial and nontarget insects, must be conserved (May 1993). The selective toxicity, rapid environmental degradation, and vertebrate safety of Bt-based insecticides provide growers and the public with environmentally friendly and effective alternatives to conventional insecticides (Meadows 1993). Advances in biotechnology and genetic engineering, as well as the proteinaceous nature of the Cry toxins, led to the selection of the cry genes as the primary insect-resistance genes transferred into, and expressed in, plants and microbes (
Biochemical Journal, 2009
The bacterium Bacillus thuringiensis produces ICPs (insecticidal crystal proteins) that are deposited in their spore mother cells. When susceptible lepidopteran larvae ingest these spore mother cells, the ICPs get solubilized in the alkaline gut environment. Of approx. 140 insecticidal proteins described thus far, insecticidal protein Cry1Ac has been applied extensively as the main ingredient of spray formulation as well as the principal ICP introduced into crops as transgene for agricultural crop protection. The 135 kDa Cry1Ac protein, upon ingestion by the insect, is processed successively at the N- and C-terminus by the insect midgut proteases to generate a 65 kDa bioactive core protein. The activated core protein interacts with specific receptors located at the midgut epithilium resulting in the lysis of cells and eventual death of the larvae. A laboratory-reared population of Helicoverpa armigera displayed 72-fold resistance to the B. thuringiensis insecticidal protein Cry1Ac. ...
Evolution of Bacillus thuringiensis Cry toxins insecticidal activity
Microbial Biotechnology, 2013
Insecticidal Cry proteins produced by Bacillus thuringiensis are use worldwide in transgenic crops for efficient pest control. Among the family of Cry toxins, the three domain Cry family is the better characterized regarding their natural evolution leading to a large number of Cry proteins with similar structure, mode of action but different insect specificity. Also, this group is the better characterized regarding the study of their mode of action and the molecular basis of insect specificity. In this review we discuss how Cry toxins have evolved insect specificity in nature and analyse several cases of improvement of Cry toxin action by genetic engineering, some of these examples are currently used in transgenic crops. We believe that the success in the improvement of insecticidal activity by genetic evolution of Cry toxins will depend on the knowledge of the rate-limiting steps of Cry toxicity in different insect pests, the mapping of the specificity binding regions in the Cry toxins, as well as the improvement of mutagenesis strategies and selection procedures.
Variations in Susceptibility to Bacillus thuringiensis Cry Toxins �
2013
Bacillus thuringiensis strains isolated from Latin American soil samples that showed toxicity against three Spodoptera frugiperda populations from different geographical areas (Mexico, Colombia, and Brazil) were characterized on the basis of their insecticidal activity, crystal morphology, sodium dodecyl sulfate-polyacrylamide gel electrophoresis of parasporal crystals, plasmid profiles, and cry gene content. We found that the different S. frugiperda populations display different susceptibilities to the selected B. thuringiensis strains and also to pure preparations of Cry1B, Cry1C, and Cry1D toxins. Binding assays performed with pure toxin demonstrated that the differences in the toxin binding capacities of these insect populations correlated with the observed differences in susceptibility to the three Cry toxins analyzed. Finally, the genetic variability of the three insect populations was analyzed by random amplification of polymorphic DNA-PCR, which showed significant genetic diversity among the three S. frugiperda populations analyzed. The data presented here show that the genetic variability of S. frugiperda populations should be carefully considered in the development of insect pest control strategies, including the deployment of genetically modified maize in different geographical regions.
Cross-Resistance to Bacillus thuringiensis Toxin CryIF in the Diamondback Moth (Plutella xylostella)
Applied and Environmental Microbiology
Selection with Bacilus thuringiensis subsp. kurstaki, which contains CrylA and Cryll toxins, caused a >200-fold cross-resistance to CryIF toxin from B. thuringiensis subsp. aizawai in the diamondback moth, PluteUla xylosteUla. CrylE was not toxic, but CryIB was highly toxic to both selected and unselected larvae. The results show that extremely high levels of cross-resistance can be conferred across classes of Cryl toxins of B.