Introgression of a disrupted cadherin gene enables susceptibleHelicoverpa armigerato obtain resistance toBacillus thuringiensistoxin Cry1Ac

Abstract

A disrupted allele (r1) of a cadherin gene (Ha_BtR) is genetically associated with incompletely recessive resistance to Bacillus thuringiensis toxin Cry1Ac in a Cry1Ac-selected strain (GYBT) of Helicoverpa armigera. The r1 allele of Ha_BtR was introgressed into a susceptible SCD strain by crossing the GYBT strain to the SCD strain, followed by repeated backcrossing to the SCD strain and molecular marker assisted family selection. The introgressed strain (designated as SCD-r1, carrying homozygous r1 allele) obtained 438-fold resistance to Cry1Ac, >41-fold resistance to Cry1Aa and 31-fold resistance Cry1Ab compared with the SCD strain; however, there was no significant difference in susceptibility to Cry2Aa between the integrated and parent strains. It confirms that the loss of function mutation of Ha_BtR alone can confer medium to high levels of resistance to the three Cry1A toxins in H. armigera. Reciprocal crosses between the SCD and SCD-r1 strains showed that resistance to Cry1Ac in the SCD-r1 strain was completely recessive. Life tables of the SCD and SCD-r1 strains on artificial diet in the laboratory were constructed, and results showed that the net replacement rate (R0) did not differ between the strains. The toxicity of two chemical insecticides, fenvalerate and monocrotophos, against the SCD-r1 strain was not significantly different from that to the SCD strain. However, larval development time of the SCD-r1 strain was significantly longer than that of the SCD strain, indicating a fitness cost of slower larval growth is associated with Ha_BtR disruption in H. armigera.

Diverse genetic basis of field-evolved resistance to Bt cotton in cotton bollworm from China

“…From 10 isofemale lines that yielded five or more survivors in the F 2 screen, we generated 10 resistant strains for further analysis (Materials and Methods). Sequencing of the cadherin gene revealed that three of these resistant strains from Anyang in northern China (AY9, AY16, and AY27) had the recessive cadherin resistance allele r 1 that was previously identified and characterized from the laboratoryselected SCD-r1 strain (23).…”

“…The cadherin r 1 allele originally identified from a laboratory-selected strain derived from Gaoyang of northern China in 2001 (22,23,26) was the most common resistance allele detected in fieldselected populations from northern China. This allele accounted for the resistance detected in 38% (three of eight) of resistant strains from Anyang of northern China (Table 1), which is about 300 km southwest of Gaoyang.…”

“…In the laboratoryselected SCD-r1 strain of H. armigera derived from China, the recessive r 1 allele of the cadherin gene Ha_BtR has a deletion mutation causing a more than 400-fold increase in the concentration of Cry1Ac needed to kill 50% of larvae (LC 50 ) compared with the nearly isogenic susceptible SCD strain that lacks this allele (23). Bt cotton producing Cry1Ac was commercialized in China in 1997 and has accounted for more than 90% of cotton planted in northern China since 2004 (24), and planting of Bt cotton has been limited in northwestern China (14,25).…”

Efficacy of genetically modified Bt toxins against insects with different genetic mechanisms of resistance

“…We used diet surface overlay bioassays to test second instars that had been starved for 4 h. 18 Toxins were diluted with PBS (0.01 M, pH 7.4). We put 900 μl of liquid artificial diet in each well of a 24-well plate.…”

“…14,15 Disruption of Bt toxin binding to midgut receptors is the most common general mechanism of insect resistance. 9 Mutations in the genes encoding midgut cadherins that bind Cry1Ac are linked with resistance in at least three lepidopteran pests of cotton, [16][17][18] but such cadherin mutations are not the primary cause of many other cases of field-and laboratory-selected resistance. 9,19,20 Although some aspects of the mode of action of Bt toxins remain unresolved, extensive evidence shows that after Cry1A protoxins are ingested by larvae, they are solubilized in the gut and cleaved by mid-gut proteases such as trypsin to yield activated 60-kD monomeric toxins that bind with membrane-associated receptors.…”