Resistance Allele Frequency Of Pink Bollworm On Bt Cotton In The Yangtze River Valley

Transgenic cotton producing insecticidal proteins from Bacillus thuringiensis (Bt cotton) has effectively suppressed major targeted pests since its commercialization in1996. Bt cotton was commercialized in1997in China, and it had accounted for80%by the year2012across the country, and exceeded90%in the Yangtze River Valley of China. It has brought about substantial economic and ecological benefits by improving yields and reducing insecticide sprays. However, vast plantings of Bt cotton caused continuous selection pressure to targeted pests, which might rapidly lead to pest resistance, and this is the main threat towards the long term use of Bt cotton.The pink bollworm, Pectinophora gossypiella (Saunders), a major pest in the Yangtze River Valley, is also one of the targeted pests on Bt cotton. Continuous resistance monitoring data in recent years showed that field population of pink bollworm in the Yangtze River Valley had entered the early stage of resistance, which severely threatens the sustainable use of Bt cotton. Therefore, it is crucial for the management of pest resistance to keep working on resistance monitoring to learn the variations of resistance level and resistance allele frequency in the field. In this study, resistant and susceptible laboratory strains, and more than10field populations collected from the Yangtze River Valley were adopted, and three resistance monitoring methods were conducted:diet bioassay, single pair crosses and DNA screening, in order to systematically evaluate the resistance level and estimate the resistance allele frequency during2012and2013. The results were as follows:1) Responses to Bt toxin CrylAc of field populations of pink bollwormLaboratory diet bioassays were conducted to test the susceptibility to Bt toxin CrylAc of pink bollworm from12monitoring sites of the Yangtze River Valley. Results showed that the range of LC50was0.19-0.26,0.17-0.35in2012and2013, respectively, and corresponding range of resistance ratio was2.71-3.71,2.43-5.00each. The resistance ratio was below5times in all sites of the2years, and variations between different sites were less than3times. The LC50value was somewhat higher than that in2012as a whole, and the resistance level showed a slightly increase in most single sites, but there was no significant difference between them. Results above indicated that pink bollworm basically remained susceptible to Bt cotton after13years of planting in the Yangtze River Valley. 2) F1screen to estimate the resistance allele frequencySingle pair mating systems were established between field collected insects and individuals of a laboratory resistant strain, and offspring were tested under diagnostic concentration of Bt toxin Cry1Ac to estimate the resistance allele frequency. Results showed that there were2and12positive individuals detected respectively in2012and2013, and the frequency was0.0034(95%confidence interval:0.0009-0.0108) and0.0165(95%confidence interval:0.0096-0.0277) for each year, which showed a slightly increase without significant difference. The average frequency of the two years was0.0101(95%confidence interval:0.0062-0.0163), one magnitude greater than the upper limit (10-3) of the refuge strategy, which implied that more attention should be paid to the Bt resistance risk in the field.3) DNA screening to estimate the resistance allele frequencyDNA based detection mothod was applied to screen the three known cadherin mutational alleles, r1, r2, r3of pink bollworm from cotton fields in the Yangtze River Valley. Screening data showed that there were totally88heterozygotes and12homozygous resistant individuals detected out of10370samples during2012and2013, and three genotypes (rls, r\rl and rlr2) of individuals were identified harboring resistant alleles. Frequency of the three known resistance alleles was calculated as0.0097and0.0030for the year2012and2013, respectively, and the average frequency was0.0101(95%confidence interval:0.0045-0.0065) for two years. Compared with results of the F1screen, results here were close to the former, which proved that the DNA screening technology was applicable for resistance monitoring of pink bollworm.To date, this is the first time that DNA screening method was applied for resistance monitoring of pink bollworm on Bt cotton in China, in combination with conventional bioassays and F1screen methods. Results in this research figured out the susceptibility level of field populations of pink bollworm to Cry1Ac and the resistance allele frequency in the Yangtze River Valley, which would provide gist for the resistance management of pink bollworm.