Abstract
The use of conventional chemical insecticides and bacterial toxins to control lepidopteran pests of global agriculture has imposed significant selection pressure leading to the rapid evolution of insecticide resistance. Transgenic crops (e.g., cotton) expressing the Bt Cry toxins are now used world wide to control these pests, including the highly polyphagous and invasive cotton bollworm Helicoverpa armigera. Since 2004, the Cry2Ab toxin has become widely used for controlling H. armigera, often used in combination with Cry1Ac to delay resistance evolution. Isolation of H. armigera and H. punctigera individuals heterozygous for Cry2Ab resistance in 2002 and 2004, respectively, allowed aspects of Cry2Ab resistance (level, fitness costs, genetic dominance, complementation tests) to be characterised in both species. However, the gene identity and genetic changes conferring this resistance were unknown, as was the detailed Cry2Ab mode of action. No cross-resistance to Cry1Ac was observed in mutant lines. Biphasic linkage analysis of a Cry2Ab-resistant H. armigera family followed by exon-primed intron-crossing (EPIC) marker mapping and candidate gene sequencing identified three independent resistance-associated INDEL mutations in an ATP-Binding Cassette (ABC) transporter gene we named HaABCA2. A deletion mutation was also identified in the H. punctigera homolog from the resistant line. All mutations truncate the ABCA2 protein. Isolation of further Cry2Ab resistance alleles in the same gene from field H. armigera populations indicates unequal resistance allele frequencies and the potential for Bt resistance evolution. Identification of the gene involved in resistance as an ABC transporter of the A subfamily adds to the body of evidence on the crucial role this gene family plays in the mode of action of the Bt Cry toxins. The structural differences between the ABCA2, and that of the C subfamily required for Cry1Ac toxicity, indicate differences in the detailed mode-of-action of the two Bt Cry toxins.
Highlights
In recent decades, agriculture has increasingly come to rely on toxins encoded by the Grampositive bacterium Bacillus thuringiensis (Bt) for the production of insect–resistant transgenic crops
Biphasic linkage analysis of a Cry2Ab-resistant H. armigera family followed by exon-primed intron-crossing (EPIC) marker mapping and candidate gene sequencing identified three independent INDEL mutations in an ATP-Binding Cassette transporter subfamily A gene (ABCA2)
A deletion mutation was identified in the same gene of resistant H. punctigera
Summary
Agriculture has increasingly come to rely on toxins encoded by the Grampositive bacterium Bacillus thuringiensis (Bt) for the production of insect–resistant transgenic crops Narrow spectrum insecticides such as the protoxin crystals produced by Bt during sporulation are highly specific for certain insect groups including the Lepidoptera, Diptera and Coleoptera (e.g., [1]). A major reason for the uptake of Bt was the evolution of resistance to chemical insecticides such as organochlorides, synthetic pyrethroids, and organophosphates in pests such as the cotton bollworm Helicoverpa armigera. This species is one of the most damaging and economically important lepidopteran pests known worldwide in a variety of crops, and one of four major pests in the genus Helicoverpa, the others being H. punctigera, H. zea and H. assulta [5,6,7,8]
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