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    Detection and biochemical characterization of insecticide resistance in Helicoverpa armigera (Hubner) (Lepidoptera: Noctuidae)
    (Department of Agricultural Entomology, College of Agriculture,Vellanikkara, 2024-03-15) Harsha Thomas.; Ranjith, M T
    Helicoverpa armigera (Hübner) (Lepidoptera: Noctuidae), popularly known as tomato fruit borer or cotton bollworm, or gram caterpillar is a highly polyphagous insect pest causing severe damage to economically important agricultural crops worldwide. It has been reported to cause yield loss to the tune of 50 to 90 per cent in various crops (Ahmed et al., 1986; Lingappa and Yelshetty, 1994) and the extent of damage caused by this pest in various crops under different agro- national importan to reduce H. armigera menace in various crop ecosystems. However, the indiscriminate use of chemical insecticides has accelerated the field-evolved resistance in H. armigera to the major group of insecticides, thus making the management all the more difficult. H. armigera is a major threat to tomato and chilli, which are widely grown in Palakkad district in Kerala. Although farmers apply new generation insecticides on a large scale to manage this pest, it has often been claimed that control is not possible when these insecticides are applied in recommended doses. Hence, screening of resistance in H. armigera to commonly used insecticides and elucidating the biochemical mechanisms of resistance is necessary to develop better alternatives that could be feasible and effective for sustainable pest management. With these objectives, the and biochemical analysis of insecticide resistance in Helicoverpa armigera (Hübner) (Lepidoptera: Noctuidae) was undertaken at the Department of Agricultural Entomology, College of Agriculture, Vellanikkara, Kerala Agricultural University, Thrissur during the period from 2022 to 2024. Two field populations of H. armigera were collected from the tomato-growing belts of Palakkad, Kerala, and Kolar, Karnataka, and were screened for field-evolved resistance against various insecticides viz., chlorpyriphos, quinalphos, lambda-cyhalothrin, flubendiamide, chlorantraniliprole, emamectin benzoate, and spinosad. A susceptible population (ISS) was collected from IARI, New Delhi and maintained at AINPVPM laboratory. The biochemical mechanism conferring insecticide resistance was analyzed for the three major detoxifying enzymes, carboxylesterase, cytochrome P450, and glutathione S-transferase. Subsequently, as a confirmatory test, synergistic bioassay at 1:2, 1:4, and 1:6 ratios was conducted to confirm the involvement of enzymes in resistance. The major synergists used for the study were piperonylbutoxide (PBO) inhibiting cytochrome P450, triphenyl phosphate (TPP) which inhibits carboxyl esterase, and diethyl maleate (DEM) which inhibits glutathione S-transferase. In H. armigera field population collected from Kerala (PKT1), a high level of resistance was marked for the insecticides, chlorpyriphos ( ), quinalphos ( ), and lambda-cyhalothrin ( ), however a low level of resistance was recorded in PKT1 against chlorantraniliprole ( ) and emamectin benzoate (21- ) whereas, flubendiamide ( ) and spinosad ( ) marked a negligible resistance ratio. The H. armigera field population collected from Karnataka (KAT2), on the other hand, exhibited a high level of resistance towards chlorpyriphos ( ), quinalphos ( ) lambda-cyhalothrin ( ), and chlorantraniliprole ( ), whereas a low level of resistance was noted against the insecticide, spinosad ( ). The insecticides emamectin benzoate ( - ) and flubendiamide ( ) displayed a minimal resistance ratio. The biochemical analysis revealed that Kerala population (PKT1) showed a high carboxylesterase activity (16.545 µmol/min/mg) followed by glutathione S-transferase (0.569 µmol/min/mg). Whereas, the level of cytochrome P450 enzyme (0.592 pmol/min/mg) in the PKT1 remained lower than the susceptible population (0.773 pmol/min/mg). The field population collected from Karnataka (KAT2) exhibited significantly high levels of carboxylesterase (26.83 µmol/min/mg) and cytochrome P450 (1.293 pmol/min/mg) when compared to the susceptible population [with low carboxyl esterase (10.622 µmol/min/mg) and cytochrome P450 (0.494 pmol/min/mg)] indicating their involvement in conferring resistance to pyrethroids (lambda-cyhalothrin), organophosphates (quinalphos), and diamides. Conversely, the levels of GST remained low (0.133 µmol/min/mg) in KAT2. To elucidate the resistance mechanism mediated by metabolic detoxifying enzymes, a synergistic bioassay was conducted only for PBO and TPP as the activity of GST remained significantly low. Synergistic bioassay gave a better result at a 1:4 (insecticide: synergist) ratio. Kerala population did not show a significant synergism for chlorpyriphos and quinalphos in combination with PBO and TPP. However, lambda-cyhalothrin was synergized with a significant synergistic ratio (SR) of 96.39 by PBO and 53.596 by TPP. This indicates that resistance conferred to lambda-cyhalothrin was primarily due to enzymes particularly carboxylesterase, whereas resistance to organophosphate was not attributed to any detoxifying enzymes rather, other mechanisms might have involved. The organophosphate and pyrethroid insecticides on the other hand were synergised effectively by PBO and TPP in Karnataka population. The synergistic ratios obtained for chlorpyriphos with PBO at 1:6 ratio was125.428, while the synergism by TPP did not drop the LC50 to the fiducial limit in case of susceptible population.For quinalphos, a significant synergism was obtained at a 1:4 ratio by both PBO and TPP with SR valued at 122.524 and 134.576. The most effective and significant synergism was observed for lambda- cyhalothrin with PBO (SR=386.909) and TPP (148.375) at 1:4 ratio. This indicates the involvement of detoxifying enzymes especially cytochrome P450 and carboxylesterase in conferring resistance towards organophosphate and pyrethroids. The lack of significant synergism of Chlorantraniliprole by PBO, and TPP suggests that other mechanisms might have involved, particularly mutations in the ryanodine receptor (RyR) gene (Jouraku et al., 2019). It can be concluded from the present study that H. armigera infesting tomato in both Kerala and Karnataka is showing a varying level of resistance towards both conventional and new generation insecticides. The biochemical analysis reveals that different mechanisms are involved in both the geographical populations towards conferring resistance to these insecticides. Hence, it is important to keep a track on the status of resistance development in H. armigera consistently for the effective pest management and modification of management tools. This helps to minimize economic losses to the farmers and reduces the environmental impact.