Resistance risk assessment and the cross-resistance of Grapholita molesta to avermectin

Abstract:【Objective】Grapholita molesta is a major pest of many kinds of fruit crops in the world and China. Due to its characteristics of boring, hiding and generations overlapping, G. molesta is difficult to control. With the climate warming, and the change of cultivation mode and management technology in pear orchards, the damage of G. molesta has been increasing year by year in several major fruit species in China, including peaches, pears and apples. Currently, chemical pesticides are still one of the most effective measure to control G. molesta. Long-term frequent and non-standard use of pesticides has led to a gradual decline in the effectiveness of chemical control to G. molesta, and G. molesta has developed varying degrees of resistance to some pesticides in pear orchards. Avermectin, a high efficiency and low toxicity pesticide used in pear orchards frequently, was used for 5-7 times in a year to control many kinds of pests, such as G. molesta, Aphis citricola Van der Goot, Tetranychus viennensis Zacher, and so on. Over the past 20 years, the application concentration of avermectin has increased by 40 times in pear orchards due to its high dose and exceessive usage times. Studies on avermectin mainly focus onits toxicity, control efficacy, management and efficience in pear orchards. The change patterns of resistance and sensitiveness were studied with lethal concentration of 50% of avermectin to G. molesta in this paper. By studying the resistance and sensitivity changes, it is expected to obtain change patterns of resistance, development speed, resistance heritability of G. molesta to avermectin, and its interaction resistance with other pesticides that were also used in pear orchards, such as imidacloprid, chlorphenicol benzamide, lambda-cyhalothrin and so on. The objective of this experiment was to obtain theoretical basis for reasonable use avermectin to delay the development of its resistance in pear orchards.【Methods】In order to measure control efficiency of avermectin to G. molesta neonate larvae, the survival rate and damage rate of G. molesta neonate larvae were measured with fruitlets as the sample. The G. molesta neonate larvae and larvae were fed in an artificial intelligence incubator under the following conditions: (25±1) ℃, 70%-80% relative humidity, 3000-4000 lx illumination and 15 h//9 h (L/D) photoperiod. The test agent was 92.00% avermectin, 96.0% Lambda- cyhalothrin, 96% imidacloprid and 96% chlorantraniliprole. Fruit dipping method was used to measure the resistance development of G. molesta to avermectin. These methods included: (1) the young fruits of apple with same variety, consistent size, and good appearance were wash with pure water, dried in air, and soaked in the pesticide solution for 10 seconds. (2) The young fruits were taken out from pesticide solution, excessive pesticide solution was absorbed with a filter paper, and they were placed on a plastic container with a wet filter paper and a lid at the bottom. (3) The paper containing 50 G. molesta ready-to- hatch eggs was placed on the young fruit gently, then the egg was managed to contact the fruit, and the relative humidity in the container was maintained above 90%. (4) Tween-80 aqueous solution with a mass fraction of 0.02% was taken as the control. Each process was repeated for three times. (5) The survival rate of G. molesta neonate larvae was surveyed in 78 h after laying eggs. Survey method was determined by observing carefully pest morphological characteristic and damage symptoms of G. molesta neonate larvae on apple and damage rate in 78 h. According to mortality of G. molesta neonate larvae, the toxicity equation and LC50 were calculated with SPPS. Two strains of G. molesta neonate larvae were obtained from field populations, which were resistance-selection by LC50 of avermectin. The field population was collected from pear orchards in Yanhu District, Yuncheng City, Shanxi province in 2022. Two strains were field resistant strain and field control strain. Field resistant strain was selected with LC50 of avermectin, and the toxicity of earlier generation of G. molesta neonate larvae was measured for six times. Field control strain was fed with apples, which were not touched by any pesticides. The comparative strain G. molesta was susceptible strain that was reared 100 generations continuously in the lab. The toxicity of different strains of G. molesta was measured in 2 generations, 4 generations and 6 generations, respectively. The resistance ratio (RR) and sensitivity level of different strains G. molesta were calculated with resistance multiple formulas. In order to obtain cross-resistance of different strains of G. molesta, Lambda-cyhalothrin, imidacloprid and chlorflubenzamide that were used frequently in pear orchards were selected to assess the cross- resistance to field resistant strain and field control strain of G. molesta. Experimental data were analyzed with Duncan’s new multiple range test (p＜0.05).【Results】The LC50 of avermectin of field population of G. molesta was 1.092 mg ·L- 1 , and its resistance ratio was 4.608 times higher than the susceptible strain, with low sensitivity. The resistance level of field population selected with LC50 avermectin at second generation was up to low resistance from low sensitivity. The LC50 of field population increased continuously with the increase of selection generations when the resistance ratios were 11.966 and 20.304 times at fourth generation and sixth generation respectively, which increased to the medium resistance level. The sensitivity of the field population without exposure to avermectin in- creased gradually. The resistance ratio of the field population decreased to 2.802 times at fourth generations, which was sensitivity. The toxicity of the field population of G. molesta without exposure to avermectin decreased from first generation to sixth generation and its sensitivity was further improved. The results also showed that the continuous use of avermectin in field populations of G. molesta caused a rapid increase in the level of resistance to avermectin. The sensitivity of strains of G. molesta without continued exposure to avermectin increased sensitivity to avermectin. The field population of G. molesta was selected with LC50 avermectin for 6 generations, and its actual heritability of resistance was h2 = 0.186. The actual heritability of F0-F2 and F4-F6 selection stage was 0.242 and 0.196, respectively. In the case of resistance heritability of 0.186, the resistance of G. molesta to avermectin increased by 10 times in about 4- 9 generations. The resistance ratio of the 6th generation resistant strain of G. molesta to lambda- cyhalothrin was 8.487, which was a cross- resistance between them. The resistance ratio to chlorantraniliprole was 3.940, and its resistance ratio also increased. The resistance ratio to imidacloprid was 1.487 with no cross- resistance. The resistance ratios of the field population control strain to lambda-cyhalothrin, imidacloprid and chlorantraniliprole were 1.688, 0.962 and 1.243, respectively, and there was no cross-resistance among them.【Conclusion】G. molesta could develop resistance to avermectin rapidly. The low- sensitive field population of G. molesta could decrease sensitiveness with no exposure to avermectin. The medium- resistant strain had cross- resistance to lambda- cyhalothrin, and the low-sensitive strain had no cross-resistance to lambda-cyhalothrin, imidacloprid and chlorantraniliprole. The development of resistance of G. molesta to avermectin can be slowed down by interval medication or rotative application of avermectin and its non- cross- resistance pesticides in pear orchards. Therefore, avermectin should be used in pear orchards with an interval, and growers should reduce or limit the use frequency of lambda-cyhalothrin and chlorantraniliprole to avoid or delay the emergence and development of resistance, and ensure the control efficacy of avermectin and other pesticides, which were used in pear orchards frequently.