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Background: During field surveillance in Gujarat, India, in 2009, scientists found that pink bollworm was surviving in the first generation of Bollgard cotton, which contains a single protein. This survival was observed in four districts of Gujarat. From 2012 to 2014, surveys revealed that pink bollworm larvae had higher survival rates on Bollgard II cotton in Amreli and Bhavnagar districts. In 2016, a pink bollworm outbreak in Maharashtra caused significant losses in cotton production, leading to damage, yield losses, and increased management costs in several regions. The objective of the present study was to determine the reaction of Pectinophora gossypiellato Cry1Ac and Cry2Ab toxins. Results: Neonates of P. gossypiellacollected from six distinct locations in Vidarbha were subjected to seven different concentrations of Cry1Ac. The highest LC50value was recorded in the population collection of Yavatmal (1.362 μg/ml diet) but the lowest LC50value of 0.600 μg/ml diet in Buldana population. The highest MIC50value was recorded in the population collected from Yavatmal (0.743 μg/ml diet) and the lowest MIC50value of 0.413 μg/ml diet in the Buldana population (0.431 μg/ml diet). The highest IC50value was recorded in the population collected from Yavatmal (0.303 μg/ml diet) but the lowest IC50value of 0.127 μg/ml diet in the Buldana population. Neonates of P. gossypiellacollected from six different locations in Vidarbha were exposed to seven different concentrations (9.0, 3.0, 1.0, 0.333, 0.111, 0.037 and 0.012 for Cry1Ac and 27.0, 9.0, 3.0, 1.0, 0.333, 0.111 and 0.037 μg/ml for Cry2Ab) which revealed that the Yavatmal population had the highest LC50value of 94.294 μg/ml diet and the lowest LC50value was recorded in the Amravati population (42.144 μg/ml diet). The highest MIC50value of 27.258 μg/ml diet was recorded in Buldana population, but the lowest MIC50value was recorded in the Amravati population (12.881 μg/ml diet). The highest IC50value of 3.209 μg/ml diet was observed in Yavatmal population, and the lowest one was recorded in the Amravati population (1.574 μg/ml diet). Conclusions: According to the LC50values, all of the P. gossypiellapopulations in Vidarbha were extremely insensitive and had field-evolved resistance to the Btproteins found in the transgenic cotton varieties of Bollgard II.

Background: After the commercialization of insect-resistant transgenic Btcotton Bollgard I & II, India ranks first in the world in cotton production. Cotton insecticide consumption was drastically reduced as nearly 95% of the cotton area was replaced with Bollgard II. However, the benefits of transgenic cotton appear to have been diminished as the pink bollworm, Pectinophora gossypiella(Saunders) (Lepidoptera: Gelechiidae) developed field resistance to Cry1Ac expressing Btcotton in India in 2008. In 2015, there was an unusual survival of P. gossypiellaon Bollgard II cotton in some parts of Gujarat and Maharashtra, which is a cause for concern. Results: In the present study, PCR analysis and gel visualization of BGII resistant field population of P. gossypiellashowed that the presence of r1, r2, r3, r1s, r2s, r3s, and ss mutated cadherin alleles, which produced amplicon sizes of 750 bp, 700 bp, 730 bp, 12,700 bp, 750 bp, 980 bp, 600 bp and 1600 bp, respectively, when seven different types of cadherin allele(s) specific primers were used. The r1 mutant allele was detected in Amaravati, Wardha, Yavatmal, and Nagpur, but not in Akola and Buldhana, using primers r1disfor and Int-540. The r2 mutant allele was detected in Akola, Wardha, and Nagpur, but not in Buldhana, Amaravati, and Yavatmal, using primers r2disback and Cad2366. The r3 mutant allele was detected in all locations using primers r3disback and Cad3221. Amplicons of sizes 750 bp, 700 bp and 730 bp were obtained for r1, r2, and r3 mutant alleles, respectively. The sizes of the amplicons were 1270 bp for r1s, 750 bp for r2s, and 980 bp for r3s.The absence of bands for r1, r2, and r3 cadherin alleles in individuals indicated the possibility of having the ss allele, which was confirmed using primers Cad3324 and Int-651. The presence of ss mutant allele was observed in field-collected P. gossypiellapopulations from BG II cotton in various locations, with a 600 bp and 1600 bp size amplicons produced using the same primers. Conclusions: PCR analysis revealed the presence of r1, r2, r3, r1s, r2s, r3s, and ss mutated cadherin alleles in field-collected populations of Vidarbha which provide evidence to field-evolved resistance of P. gossypiellato BGII cotton.

Background: Compared to Bollgard-I, the utilization of Cry2Ab protein in Bollgard-II cotton cultivars enhances insect control. Field and laboratory studies demonstrated reduction in the numbers of bollworm-infested terminals, squares, and bolls in Bollgard-II cotton lines when compared to both Bollgard-I and non-Bollgard cotton cultivars. This indicates that the combination of Cry2Ab with Cry1Ac increased the overall expression of proteins. The expression of Cry protein varied across different plant parts, such as leaves, bracts, squares, and bolls. As the season progresses, the expression of Cry protein decreased in these plant parts. Leaves exhibited the highest levels of Cry protein expression, followed by squares, flowers, and bolls. Variation in the expression levels of delta endotoxins in different plant parts was one of the contributing factors to the survival of pest populations on Btcotton. Results: Using a commercially available QL 96 ELISA plate kit, the concentration of delta endotoxin in various plant parts at different phenological stages was determined in twelve BG-II cotton hybrids, namely Ajeet-155, JKCH-2245, RCH-3863, NCS-866, MRC-7373, JKCH-99, MRC-7387, NCEH-21, ANKR-3324, NCSI-1904, and NCHB 9902. Cry1Ac and Cry2Ab protein levels were determined from samples of flowers and fruiting parts (Rind, locule, seed, locule wall and seed) at 40, 75, 100 and 125 days after sowing (DAS) over two consecutive years 2018–2019 and 2019–2020. Cry1Ac protein content and expression was the highest at 100DAS in locule, seed and rind; followed by rind and locule wall and seed in green bolls at 125 DAS; followed by seed, locule, rind and flowers at 75 DAS; followed by flowers at 40 DAS in all the tested twelve Bollgard-II hybrids. Cry2Ab protein content and expression was the highest at 125DAS in locule wall and seed and rind, followed by seed, locule and rind in green bolls at 100 DAS, followed by seed, locule, rind and flowers at 75 DAS, then by flowers at 40 DAS in all the tested twelve Bollgard-II hybrids. Cry1Ac protein expression was less in comparison with Cry2Ab. Conclusions: The research findings indicate that the locule of BG-II cotton plants exhibited the highest expression levels of Cry1Ac and Cry2Ab proteins, followed by the seeds, rind, locule wall, and flowers. These results provide valuable insights into the distribution of Cry protein expression in different plant parts, which can contribute to a better understanding of insect control in Btcotton cultivars.