The cotton aphid (Aphis gossypii Glover) (Hemiptera: Aphididae) is a major pest of cotton (Gossypium hirsutum L.) grown in Xinjiang and is an agricultural pest found worldwide. With the extensive use of insecticides, A. gossypii in Xinjiang has developed high levels of resistance to a variety of insecticides. Afidopyropen is a new biogenic insecticide that is effective against insects with piercing-sucking mouthparts and has no cross-resistance with any commonly used insecticides in cotton fields. However, to our knowledge, the risk and stability of resistance of A. gossypii to afidopyropen and its detoxification enzymes under high selection pressure have not yet been elucidated. In this study, resistant A. gossypii specimens were selected with the cotton leaf-dipping method under laboratory conditions, and the realized heritability of A. gossypii resistance to afidopyropen was calculated using domain trait analysis. After 18 successive generations of selection for afidopyropen resistance, a resistant population with a median lethal dose (LC50) of 9.006 mg/L was obtained; the resistance ratios had increased 23.033 times. The realized heritability was 0.063, and a 10-fold increase in resistance required only 20–26 generations when mortality was 60–80%. The 15th generation of the resistant strain had some resistance instability, and glutathione S-transferases played an important role in the detoxification metabolism of afidopyropen. The results of this study provide a basis for application of afidopyropen and management of A. gossypii resistance.

Aphis gossypii Glover (Hemiptera: Aphididae) is a worldwide pest (Cao et al. 2017, Jiang et al. 2018) and is currently the most serious pest of cotton (Gossypium hirsutum L.) in China (Quan et al. 2019). The overall occurrence of A. gossypii in Xinjiang, China was estimated to be as high as 5.33 × 105 individuals/ha in 2022 (Li et al. 2022). Cotton fields are prone to attacks by A. gossypii (Liu 2021a) because of their diverse reproductive patterns, adaptability, and host plant species (Chao et al. 2019, Zhang et al. 2022a). Currently, chemical control is the dominant management strategy for A. gossypii outbreaks (Gao 2010, Ma et al. 2021a). However, long-term use of insecticides has led to a rapid increase in the resistance of A. gossypii (Patima et al. 2019, Zhang 2020, Zhao et al. 2018), especially to insecticides such as organophosphates, carbamates, pyrethroids, and neonicotinoids (Liang et al. 2013, Patima et al. 2019). Therefore, a new target mechanism of action for insecticides against resistant A. gossypii is urgently needed.

Afidopyropen, a novel biogenic insecticide (Li 2020, Shi et al. 2022), interferes with the function of insect chordotonal organs (Jiang et al. 2020), causing insects to lose their senses of gravity, balance, hearing, position, and movement (Gu and Bai 2020, Tan 2019). Afidopyropen can cause insects to immediately stop feeding (Gu and Bai 2018, Kandasamy et al. 2017, Zhang 2020), which in turn can reduce the spread of vector viruses and reduce damage to agricultural crops (Fu et al. 2022). Afidopyropen was registered in China in 2019 (Liu et al. 2022) and has been widely used with significant results in the cotton production areas of China (Shi et al. 2022). Aphis glycines Matsumura, Bemisia tabaci (Gennadius), Diaphorina citri Kuwayama, A. gossypii, and other field pests are sensitive to afidopyropen (Obiratanea et al. 2020, Shi et al. 2022, Zhang et al. 2021).

The resistance to insecticides usually develops in insects as a result of a change in the genetic structure that reduces pesticide sensitivity (Zhang 2016); this change is called realized heritability (h2) (Zhao et al. 2022). Resistance risk assessment is the calculation of the potential ability of pests in the field to develop resistance to pesticides (Ijaz et al. 2021, Li et al. 2013, Liu 2021b, Wang 2022). Currently, the threshold trait analysis method is widely used to calculate h2 and perform resistance risk assessments of pests (Tabashnik 1992), such as Phenacoccus solenopsis Tinsley, Laodelphax striatellus (Fallén), Helicoverpa armigera (Hübner), and Plutella xylostella (L.) (Afzal et al. 2022, Roy et al. 2023, Zhang et al. 2022b). For example, the resistance ratio of Oxycarenus hyalinipennis (Costa) was increased 2631.50-fold after the 19th generation of selected resistance to fipronil, with h2 = 0.11; a 10-fold increase in resistance ratios required six to eight generations with a greater risk of resistance when selection intensity was 60–80% (Ijaz and Shad 2022a). These studies provide an opportunity to assess the theoretical basis for predicting the occurrence of insecticide resistance in the field.

Insects gradually recover sensitivity to insecticides when their selection pressure disappears, and the rate of recovery of sensitivity is used to express resistance stability (Wen 2021). Oxycarenus hyalinipennis was selected for resistance to imidacloprid for 19 generations and reached a 146.5-fold resistance ratio. The resistance ratio returned to sensitive levels after eight generations without insecticide selection pressure (Ijaz and Shad 2022b). Phenacoccus solenopsis was selected for resistance to spirotetramat for 13 generations and reached a 328.7-fold resistance ratio, which decreased to 107.7-fold after cessation of selection pressure for four generations, with poor resistance stability (Ejaz and Ali 2017). In L. striatellus populations resistant to buprofezin for 10 generations and reared for 10 consecutive generations without selection pressure, resistance ratios decreased to 576.3-fold; thus, their resistance level was somewhat unstable (Muhammad et al. 2012).

However, the risk of resistance of A. gossypii to afidopyropen and essential enzymes required for detoxification have not yet been reported; thus, a systematic study of the resistance mechanism of A. gossypii to afidopyropen was needed. We selected for afidopyropen resistance for 18 consecutive generations of A. gossypii by using the leaf-dipping method and then assessed resistance risk and stability. The effects of detoxification enzymes and target enzymes of A. gossypii resistant to afidopyropen were investigated. An attempt was made also to find the resistance mechanism of A. gossypii to afidopyropen at the physiological level. Our study provides a theoretical basis for the application of afidopyropen as part of a sustainable pest management program and elucidates the pesticide resistance mechanism of cotton aphids.

Insects and insecticide

In 2017, A. gossypii strains were collected from cotton fields at the teaching experimental site of Shihezi University, Shihezi, China (where no insecticide was applied), followed by 5 yr of continuous rearing on potted cotton plants in incubators isolated by nylon nets without exposure to any chemicals. Aphis gossypii reproduced mainly as solitary females to form a sensitive laboratory strain (G0). The resistant strain was established using the sensitive strain and was propagated by successive selection for afidopyropen resistance. The sensitive and resistant strains were reared in a light incubator (light intensity = 12,000 lux, temperature = 26 ± 1°C, relative humidity = 45–55%, light period:dark period = 16 h:8 h). Afidopyropen (96.1%) was obtained from BASF SE (Ludwigshafen, Germany), and acetone was produced by Shanghai Yuanye Biotechnology Co. (Shanghai, China).

Toxicity bioassays

Toxicity was determined using the maceration method recommended by the Food and Agriculture Organization of the United Nations with slight modifications (Shi et al. 2011). Afidopyropen (0.1 g) was dissolved in 10 ml of acetone and adjusted to 0.01, 0.1, 1, 10, and 100 mg/L by serial dilution with distilled water containing 0.05% (vol/vol) Triton X-100; distilled water containing 0.05% (vol/vol) Triton X-100 was used as the control. Six concentrations were used, and each concentration treatment was repeated three times. Cotton seedlings for the bioassay were grown hydroponically, and 18 cotton seedlings with two true leaves of uniform growth were selected. Each cotton seedling was dipped in the corresponding concentration of insecticide for 5 s and then air-dried naturally. Each cotton seedling was then inoculated with 30 wingless A. gossypii adults of uniform size. Each cotton seedling was covered with a nylon net, and a perforated plastic bottle was placed upside down over the seedlings. The cotton aphids were examined for mortality at 72 h. Mortality was determined by lightly touching the feet and antennae of each cotton aphid with a brush. Control mortality rate was ≥90%, and median lethal concentration (LC50) values were calculated and analyzed using SPSS 20.0 statistical software (IBM, Armonk, NY). The LC50 ratio of resistant lines to sensitive lines was calculated to obtain the resistance ratio.

Selection for afidopyropen resistance

Resistance screening for afidopyropen was performed with 1,000 sensitive A. gossypii for 18 consecutive generations, and the concentration of afidopyropen was gradually increased on the basis of the bioassay results of the previous generation. Insect-bearing cotton leaves were impregnated with the afidopyropen treatment solution for 5 s, and mortality was recorded after 72 h of exposure, with mortality maintained at 60–80%; the surviving cotton aphids were transferred to fresh untreated potted cotton plant leaves for reproduction. The resistance selection experiments were conducted under controlled laboratory conditions as described previously, and bioassays were performed for each generation for a total of 18 generations.

Estimation of h2

The threshold trait analysis reported by Tabashnik (1992) and Tabashnik and McGaughey (1994) was used to assess h2 for A. gossypii in response to afidopyropen using the formula
formula
where R is the selection response and indicates the difference between the average phenotypic value of the offspring and average phenotypic value of the entire parental population. R was estimated using the following formula:
formula
where n is the number of generations selected. The selection difference S is the average phenotypic difference between the parent used for screening and the whole parental generation. S was calculated as follows:
formula
where i is the intensity of selection, and δp is the phenotypic standard deviation. The selection intensity (i) was calculated as follows:
formula
where P is the mean percent chance of surviving selection. The phenotypic standard deviation (δp) was estimated as follows:
formula
According to the method of Tabashnik (1992) for the evaluation of afidopyropen, h2 and S can be used to predict the number of generations required for a 10-fold increase in A. gossypii resistance to afidopyropen for a 50–90% A. gossypii lethality rate. Prediction of the rate of development of A. gossypii resistance to afidopyropen is based on mortality (S) and h2 with
formula

Enzyme assay

Enzyme-linked immunosorbent assay kits (Shanghai Yuanye Biological Technology Co.) were used to determine the activity of multifunctional oxidase (MFO), carboxylesterase (CES), glutathione S-transferases (GSTs), and acetylcholinesterase (AChE) in each A. gossypii generation. Seventy adult cotton aphids were selected from the 3rd, 6th, 9th, 12th, 15th, and 18th generations of resistant A. gossypii strains and the sensitive A. gossypii strain and were transferred to a 1.5-ml centrifuge tube with a brush. Physiological saline (100 μl) and was added to each tube, and the cotton aphids were mashed with a grinding rod. That mixture was centrifuged at 4°C and 3,000 × g for 10 min with a high-speed refrigerated centrifuge (Shanghai Anting Scientific Instrument Factory), and the supernatant was obtained and used for three replicates.

Statistical analysis

SPSS 20.0 software was used to perform an analysis of variance, and the least significant difference method was used for determining the significance of the differences. The probability unit regression method (Probit) was used to calculate the linear equation of virulence regression and the LC50 and its 95% confidence interval. Excel software was used for data processing and graphing.

Selection for resistance to afidopyropen

The resistance ratios and LC50 values of the 18 generations of A. gossypii are shown in Table 1 and Fig. 1. The overall trend of the resistance ratios of G0–G15 was steady growth. The most rapid growth was 10.852-fold in G12–G15, but the resistance ratios changed more slowly among other generations, and negative increases were noted for G15–G18, possibly related to the production of resistant A. gossypii and their resulting fitness changes (Table 1). The overall LC50 values of the 18 A. gossypii generations steadily increased (Fig. 1), with a fitted curve of y = 0.0212x2 + 0.0956x + 0.3236, R2 = 0.953. From G6 to G7 and G15 to G16, two abrupt decreases in LC50 values were detected, attributed to the adaptation of A. gossypii to afidopyropen. During resistance selection, LC50 reached a maximum of 9.480 mg/L at G15. The LC50 of the resistant 18th generation was 9.006 mg/L, and the resistance level was moderate.

Fig. 1.

Development of resistance to afidopyropen in the susceptible strain of A. gossypii. For G0, the LC50 data are from Wang et al. (2023 ). The experimental data were obtained at almost the same time.

Fig. 1.

Development of resistance to afidopyropen in the susceptible strain of A. gossypii. For G0, the LC50 data are from Wang et al. (2023 ). The experimental data were obtained at almost the same time.

Close modal
Table 1.

Selection of resistance to afidopyropen in A. gossypii.

Selection of resistance to afidopyropen in A. gossypii.
Selection of resistance to afidopyropen in A. gossypii.

h2 of A. gossypii to afidopyropen

Based on the characteristics of A. gossypii resistance to afidopyropen, the resistance screening was divided into two stages. The resistance increased slowly in G0–G9 and increased faster in G9–G18 (Table 2). In the first nine generations of the selection (G0–G9), h2 = 0.064. In the subsequent nine generations (G9–G18), h2 = 0.041, which was not significantly different from the earlier h2 value. This result may be related to the number of generations of A. gossypii and its resistance stability. Some differences in h2 of resistance to insecticides have been observed at various stages of a pest resistance screening, and the higher the h2, the faster the rate of development of resistance at that stage. In the 18 generations of resistant strains, the heritability of the resistant populations was relatively greater and the rate of development was relatively faster in the first G0–G9 generations than in the subsequent generations.

Table 2.

Realized heritability of A. gossypii resistance to afidopyropen.

Realized heritability of A. gossypii resistance to afidopyropen.
Realized heritability of A. gossypii resistance to afidopyropen.

Risk assessment of A. gossypii resistance to afidopyropen

The projected rate of development of resistance to afidopyropen is directly proportional to the selection intensity and h2 (Fig. 2). At 60–80% afidopyropen selection intensity, a 10-fold increase in resistance required 19.8–26.4 generations when the slope was 0.477 and h2 = 0.063. When h2 = 0.069 and h2 = 0.039, a 10-fold increase in resistance required 18–24 and 31.9–42.6 generations, respectively.

Fig. 2.

Effect of realized heritability (h2) on the number of generations of A. gossypii required for a 10-fold increase in the LC50 of afidopyropen (slope = 0.477) at different selection intensities (i).

Fig. 2.

Effect of realized heritability (h2) on the number of generations of A. gossypii required for a 10-fold increase in the LC50 of afidopyropen (slope = 0.477) at different selection intensities (i).

Close modal

The projected rate of resistance evolution is inversely proportional to the slope (Fig. 3). At 60–80% afidopyropen selection intensity, a 10-fold increase in resistance required 19.8–26.4 generations when h2 = 0.063 and the slope was 0.477; when the slope was 0.577 and 0.677, a 10-fold increase in resistance required 16.4–21.8 and 13.9–18.6 generations, respectively. The results indicate that the rate of resistance development accelerated with the increase in genetic power at the same insecticide lethality, and the rate of A. gossypii development of afidopyropen resistance decreased with the increase in insecticide usage and application frequency under stable genetic conditions.

Fig. 3.

Effect of realized heritability (h2) on the number of generations of A. gossypii required for a 10-fold increase in the LC50 of afidopyropen (h2 = 0.063) at different selection intensities (i).

Fig. 3.

Effect of realized heritability (h2) on the number of generations of A. gossypii required for a 10-fold increase in the LC50 of afidopyropen (h2 = 0.063) at different selection intensities (i).

Close modal

Stability of afidopyropen resistance in resistant A. gossypii strains

The resistance level of the 15th generation of the resistant cotton aphid strain had a significant decreasing trend, and the LC50 values of G15 for ST4-resistant strains decreased from 9.480 to 6.188 mg/L, and the resistance ratios decreased by 8.42-fold (Table 3). However, the resistance to afidopyropen was still moderate and relatively stable.

Changes in enzyme activities of resistant and sensitive strains

The activities of the four enzymes in the resistant (G3, G6, G9, G12, G15, and G18) and sensitive (G0) strains of A. gossypii were determined (Fig. 4). The CES activities of the screened resistant strains were greater than those of the sensitive strain, indicating that CES played a role in detoxification metabolism in the resistant strains; the maximum viability CES activity was 13.195 IU/L in G3. The MFO activities of most of the screened resistant strains were greater than those of the sensitive strain, indicating that MFO played a role in detoxification metabolism in the resistant strains; the maximum viability MFO activity was 136.169 IU/L in G12. With the increase in the number of selected generations, the resistance of A. gossypii to afidopyropen increased and the viability of GSTs also increased; the role of GSTs in detoxification metabolism was the strongest in G18. AChE activity did not differ significantly among the resistant and sensitive strains, and AChE did not play a substantial role in the development of resistance to afidopyropen in A. gossypii.

Fig. 4.

Detoxifying enzyme and target enzyme activity in strains of A. gossypii. Bars indicate the mean ± SE (P < 0.05). Bars with different letters indicate significant differences between enzyme activities. For G0, the enzyme activities are from Wang et al. (2023 ). The experimental data were obtained at almost the same time.

Fig. 4.

Detoxifying enzyme and target enzyme activity in strains of A. gossypii. Bars indicate the mean ± SE (P < 0.05). Bars with different letters indicate significant differences between enzyme activities. For G0, the enzyme activities are from Wang et al. (2023 ). The experimental data were obtained at almost the same time.

Close modal
Table 3.

Resistance stability of A. gossypii without afidopyropen.

Resistance stability of A. gossypii without afidopyropen.
Resistance stability of A. gossypii without afidopyropen.

Correlation of resistance level with enzyme activity

The correlation of the level of resistance with detoxification enzyme activity in A. gossypii was analyzed, and the LC50 values revealed a highly significant positive correlation with the activity of GSTs, with a correlation coefficient of 0.959; a positive but not a significant correlation with MFO and AChE; and a nonsignificant negative correlation with CES (Table 4). Thus, changes in the viability of GSTs were closely related to changes in the resistance of A. gossypii to afidopyropen, and GSTs may play an important role in the development of A. gossypii resistance to afidopyropen.

Table 4.

Correlation between resistance level and enzyme activity in strains of A. gossypii.

Correlation between resistance level and enzyme activity in strains of A. gossypii.
Correlation between resistance level and enzyme activity in strains of A. gossypii.

Long-term use of a specific insecticide results in resistance selection pressure on pests, leading to the development of resistance. Usually, laboratory-based resistance selection is used to determine the growth rate and resistance risk of targeted pests. Our results indicate that A. gossypii subjected to afidopyropen resistance selection for 18 generations exhibited a 23-fold increase in resistance. In a previous study (Muhammad et al. 2017), P. solenopsis underwent resistance selection for afidopyropen for 23 generations and exhibited a 26,652-fold rapid increase in resistance. Wang et al. (2021) reported that when A. gossypii was selected for resistance to afidopyropen for 20 generations, the resistance multiplicity increased 40-fold. We observed that the resistance of A. gossypii to afidopyropen increased slowly at the beginning of the resistance selection process (G0–G4) and that the resistance ploidy increased 4.0-fold, which may be related to the low number of selection generations and low selection pressure of A. gossypii to afidopyropen. With the increase in the number of selections and the selection pressure, the resistance of A. gossypii to afidopyropen developed faster and the resistance ploidy increased from 15.4-fold to 23.0-fold from G14 to G18, indicating the increased adaptability of A. gossypii to afidopyropen.

Higher h2 indicates a higher risk of resistance in the next generation, with a positive correlation of h2 with risk of resistance (Zhao et al. 2022). An h2 value of 0.1325 was obtained when 10 generations of Spodoptera litura (F.) was selected for indoxacarb resistance (Wang et al. 2008). When A. gossypii was selected for sulfoxaflor resistance for 25 generations, its resistance increased 13.44-fold. The h2 value was 0.0304 for generations G0–G15 of sulfoxaflor-resistant lines, which had a low risk of resistance, and was 0.1484 for G15–G25 (An et al. 2020). In our study, h2 was 0.069 in G0–G10, and the risk of resistance was relatively higher and rate of resistance development in the early generations was faster; whereas, h2 was 0.039 in the G10–G18 generations and the risk of resistance was relatively low. The results of this study are consistent with the findings of Roy et al. (2023), Afzal et al. (2018), and Ijaz et al. (2016) and in contrast with the findings of Mudassir and Ali (2022) and An et al. (2020). After 18 generations of selection for fluxametamide resistance in P. xylostella, a 10-fold increase in resistance required 27–40 generations when the mortality rate was 60–80% (Roy et al. 2023). A 10-generation selection for resistance to indoxacarb in S. litura predicted that 8–10 generations were required for a 100-fold increase in resistance at 80–90% selection intensity (Wang et al. 2008), with a very high risk of resistance. In our study, at 60–80% selection pressure, a 10-fold increase in the resistance of A. gossypii to afidopyropen required 19.8–26.4 generations (slope = 0.447, h2 = 0.063), with little risk of resistance. Therefore, we postulate that h2 of the 18th generation of afidopyropen-resistant A. gossypii is not high; however, under selection pressure, the multiplicity of resistant A. gossypii gradually accelerates, and field pest management should avoid the use of a single insecticide and mass application of insecticides.

When an insecticide ceases to act on a pest population, the proportion of resistant individuals in the population gradually decreases over time, showing an overall decreasing level of resistance. The rate of decline in population resistance is expressed in terms of resistance stability (Wang and Han 2010, Yan and Wang 2020). Laodelphax striatellus resistant to triflumezopyrim was screened up to the 16th generation, and 5 successive generations were then reared without exposure to the insecticide. The resistance multiplicity decreased by 2.04 times; therefore, L. striatellus is stable in its resistance to triflumezopyrim, which poses some difficulties for resistance management (Zhang et al. 2022b). The resistance of O. hyalinipennis to chlorfenapyr was screened up to the seventh generation, then rearing with the insecticide was stopped for five generations. The resistance ratios decreased by 81.98 times, and the resistance level was extremely unstable (Saif and Sarfraz 2017). The resistance of S. litura to spinosad was screened up to the 12th generation. Rearing with spinosad was then stopped for four generations, with a 259.5-fold decrease in resistance ratios and a highly unstable resistance level (Adeel and Shoaib 2014). In our study, after culling the resistant lines for 15 consecutive generations, the resistance ratios decreased 8.42-fold when rearing with afidopyropen was stopped for 5 generations. The level of resistance decreased rapidly, indicating that A. gossypii resistance to afidopyropen is somewhat unstable in the absence of selection pressure. This finding is important for afidopyropen resistance management, but the level of resistance also did not decrease from the midlevel resistance stage to the low-level resistance stage. Therefore, when A. gossypii develops some resistance to afidopyropen, attention must be directed to the management of A. gossypii in the field.

Detoxification metabolic enzymes have developed as a result of long-term adaptation of insects to the external environment (Wen 2021). Esterases (ESTs) can reduce the effective concentration of toxic substances (Coppin et al. 2012). MFO plays an important role in the oxidative metabolism of insecticides (Wei and Tang 1999). P450 is an important component of MFO and has an important function in oxidative metabolism (Zhang et al. 2002). GSTs protect insects from nucleophiles and ensure normal organismal metabolism (Enayati et al. 2005). For example, the biochemical mechanism of CES is important for the resistance detoxification of deltamethrin in H. armigera and of imidacloprid in Rhopalosiphum padi (L.) (El-Latif and Subrahmanyan 2010; Zhang 2016). CES and CYP450 belong to an important enzyme pathway, with CES involved in the detoxification metabolism of thiamethoxam and sulfoxaflor in A. gossypii (Ma et al. 2019, 2021b). Many studies have shown that CES is a key detoxification enzyme for neonicotinoids and organophosphorus insecticides in A. gossypii (Cui et al. 2010, Guo et al. 2014, Li 2015). MFOs are important enzymes and are involved in H. armigera metabolism of fenvalerate, Empoasca pirisuga (Goethe) metabolism of thiamethoxam, and Nilaparvata lugens (Stål) metabolism of pymetrozine (Mu and Wang 1989, Wang et al. 2021, Zhang 2019). The results of previous studies have shown that P450s are important enzymes in B. tabaci resistant to afidopyropen (Wang et al. 2022b). In our study, we determined changes in the metabolic enzyme activities of moderately resistant A. gossypii strains and found that GSTs increased with the number of selection generations, and all resistant strains were significantly different from the control. GSTs are important for the resistance of P. xylostella to chlorantraniliprole, Sitobion avenae (F.) to pirimicarb, and Spodoptera frugiperda (J.E. Smith) to metabolized indoxacarb (Hu 2017, Wang et al. 2020, Wu et al. 2020). Therefore, GSTs may play an important role in the resistance of A. gossypii to afidopyropen.

Elevated levels of pest resistance to insecticides have been correlated with increased enzyme activity (Armstrong and Suckling 1990). A positive correlation was observed between MFO activity and fenvalerate resistance (r = 0.98) and between EST and fenvalerate resistance (r = 0.97) by screening fenvalerate-resistant populations of S. litura (Cheema et al. 2020). The resistance level of Oedaleus asiaticus Bey-Bienko to the highly effective beta-cypermethrin in the field was highly significantly correlated (r = 0.932) with CES activity (Gao et al. 2022). The resistance level of Myzus persicae (Sulzer) to acetamiprid in the field was significantly positively correlated (r = 0.998) with AChE activity (Cai et al. 2021). A correlation analysis of the LC50 with the enzyme lineage of resistant strains revealed a highly significant positive correlation between GSTs and LC50 (P < 0.01), indicating the importance of GSTs in the development of resistance to afidopyropen in A. gossypii; however, further confirmation of this finding is needed.

Overall, our study revealed the risk of resistance and changes in the enzyme activities of A. gossypii resistant to afidopyropen. We screened afidopyropen-resistant A. gossypii strains to provide a theoretical basis for predicting the risk of A. gossypii becoming resistant to afidopyropen in the field. The relative stability of resistance was clarified in the G15 strains. Field management is necessary for rational use of afidopyropen, and it should be used for years. During the screening of A. gossypii for resistance to afidopyropen, a key enzyme line (GSTs) was identified, which is very different from the key enzyme lines for resistance to afidopyropen in the field previously reported by us and may be related to the different types of insecticides used in the field for long periods of time (Wang et al. 2023). Our study provides a theoretical reference for the development of resistance mechanisms and adaptations by A. gossypii to afidopyropen and reveals the resistance mechanism of A. gossypii to afidopyropen at the physiological level.

This work was supported by the National Natural Science Foundation of China (grant 31660519). Hui Zha and Xiaoli Wang designed the experiments and wrote the manuscript. Hui Zha and Xingchen Ren performed the experiments and analyzed the data. Hui Zha participated in the data analysis and discussion of the results. All authors read and approved the final version of the manuscript. The authors declare that they have no known competing financial interests or personal relationships that could have influenced their contributions to this study and the results reported herein.

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Author notes

2

Co–first authors.