Mechanism of resistance to pyrethroid insecticides in Aedes albopictus from Anshun City, Guizhou Province

doi:10.12140/j.issn.1000-7423.2026.03.012

Abstract

Abstract:

Objective To investigate the resistance to pyrethroid insecticides and metabolic enzyme levels in Aedes albopictus from Anshun City, Guizhou Province, and to analyze the genotypes and distribution characteristics of the knockdown resistance (kdr) gene, so as to provide a theoretical basis for the chemical control of Ae. albopictus. Methods The larvae of Ae. albopictus were captured from small water containers in residential areas at different locations of Anshun City from July 2023 to December 2024, and then reared in laboratory to F1 to F2 generations. The resistance to deltamethrin, beta-cypermethrin and permethrin was tested in late 3rd-instar and early 4th-instar mosquito larvae of the susceptible strain and field strain with the larval dipping method, and the resistance to the three pyrethroid insecticides was determined in non-blood-fed female adults of the field strain 3 to 5 days following emergence with the tube-test method. The activity of non-specific esterase (NSE), mixed function oxidase (MFO), and glutathione-S-transferase (GST) was measured with a microplate reader in larval mosquitoes without bioassay. Genomic DNA was extracted from individual adult mosquito after bioassay, and partial fragments of the voltage-gated sodium channel (vgsc) gene were amplified using PCR assay. Following sequencing, the sequences were aligned with reference sequences from NCBI to analyze the mutations and frequency distributions of loci V1016, I1532, F1534 and D1763, respectively. The degree of resistance to pyrethroid insecticides in larval mosquitoes was graded using the software SPSS 31.0 professional, and Shapiro-Wilk test and Wilcoxon rank-sum test were performed to determine the distribution of enzyme activity. In addition, the relative importance of metabolic mechanisms and kdr mechanisms in insecticide resistance in Ae. albopictus were predicted by the classification and regression trees (CART) algorithm. Results Larvae of Ae. albopictus exhibited resistance ratios of 38.47, 18.21 and 209.17-fold to deltamethrin, beta-cypermethrin and permethrin, respectively, all appearing a moderate to high resistance level. The mortality rates of adult mosquitoes were 2.04% (2/98), 2.06% (2/97)and 7.00% (7/100), respectively, indicating resistance to all three pyrethroid insecticides. The activities of NSE, MFO, and GST were 356.17 (321.65 to 497.79) nmol α-naphthol/(min·mg pr), 12.88 (11.17 to 18.01) nmol cyt c/mg pr, and 128.70 (116.63 to 153.03) nmol/(min·mg pr) in the field population of Ae. albopictus, respectively, all showing a positively skewed distribution, which were 1.26, 0.45, and 1.65 times higher than those in susceptible strains (Z = ‒4.030, ‒8.191 and ‒7.371, all P values < 0.01). kdr gene mutations were not detected at the locus 1763 in Ae. albopictus from Anshun City, and were detected at loci V1016, I1532, and F1534. At loci 1016 and 1532, wild-type homozygotes were predominant genotypes (65.97%, 98.61%), followed by wild-type/mutant heterozygotes (30.9%, 1.39%), with the lowest proportion seen for the mutant homozygotes (3.13%, 0). There were three alleles at locus 1534, including wild-type (TTC/F), mutant (TCC/S) and mutant (TGC/C), and the mutant-type (TCC/S) had a higher gene frequency than the wild-type (TTC/F). At locus 1534, the mutant homozygote (S/S) had the highest genotype frequency (61.11%), followed by the wild/mutant-type heterozygote (F/S) (28.47%) and wild-type homozygote (F/F) (7.64%), and the mutant homozygote (C/C) had the lowest genotype frequency (2.78%). A total of 10 genotype combinations were detected in the three-locus mutation test, and the T1 genotype (V/V + I/I + S/S) accounted for the highest proportion (60.76%, 175/288), followed by the T2 genotype (V/G + I/I + F/S) (25.69%, 74/288). Results from CART analysis showed that the factors contributing to pyrethroid resistance in Ae. albopictus in Anshun City included mutation at locus 1534 (100%), MFO activity (96.7%), GST activity (66.6%), NSE activity (21.4%), mutation at locus 1016 (16.4%), and mutation at locus 1532 (1.2%). Conclusion Ae. albopictus has developed resistance to pyrethroid insecticides at varying degrees in Anshun City. The NSE, MFO or GST activities have not yet presented typical characteristics of metabolic resistance, and kdr gene mutation is dominated at locus F1534. Emergency of resistance to pyrethroid insecticides in Ae. albopictus results from the synergistic effect of changes in metabolic detoxification enzyme activities and gene mutations.

Key words: Aedes albopictus, Insecticide resistance, Metabolic enzyme, Knockdown resistance gene, Resistance mechanism

CLC Number:

R384.1

WANG Dan, ZHANG Yan, YU Taohong, XIE Pengliang, ZHOU Jingzhu, SHI Weifang, LUO Xiaolong, LIANG Wenqin. Mechanism of resistance to pyrethroid insecticides in Aedes albopictus from Anshun City, Guizhou Province[J]. CHINESE JOURNAL OF PARASITOLOGY AND PARASITIC DISEASES, 2026, 44(3): 393-401.

Figures/Tables 8

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检测位点 Detection site	扩增片段 Amplicon	引物名称 Primer name	扩增方向 Amplification direction	引物序列（5'→3'） Primer sequence（5'→3'）
V1016	vgsc基因第Ⅱ结构域 Domain Ⅱ of vgsc gene	aegSCF 20	正向 Forward	GACAATGTGGATCGCTTCCC
V1016	vgsc基因第Ⅱ结构域 Domain Ⅱ of vgsc gene	aegSCR 21	反向 Reverse	GCAATCTGGCTTGTTAACTTG
I1532/F1534	vgsc基因第Ⅲ结构域 Domain Ⅲ of vgsc gene	aegSCF 7	正向 Forward	GAGAACTCGCCGATGAACTT
I1532/F1534	vgsc基因第Ⅲ结构域 Domain Ⅲ of vgsc gene	aegSCR 7	反向 Reverse	GACGACGAAATCGAACAGGT
D1763	vgsc基因第Ⅳ结构域 Domain Ⅳ of vgsc gene	albSCF 6	正向 Forward	TCGAGAAGTACTTCGTGTCG
D1763	vgsc基因第Ⅳ结构域 Domain Ⅳ of vgsc gene	albSCR 8	反向 Reverse	AACAGCAGGATCATGCTCTG

杀虫剂 Insecticide	品系 Strain	LC₅₀/mg⋅L^-1（95%CI） LC₅₀/mg⋅L^-1 (95% CI)	毒力回归方程 Toxicity regression equation	抗性倍数 Resistance ratio（R/S）	抗性级别 Resistance status
溴氰菊酯 Deltamethrin	敏感品系 Susceptible strain	0.000 45（0.000 39~0.000 50）	y = 10.05 + 3.00x	1	-
溴氰菊酯 Deltamethrin	野外品系 Field strain	0.017 31（0.014 25~0.021 00）	y = 3.18 + 1.81x	38.47	中抗Moderate resistance
高效氯氰菊酯 Beta-cypermethrin	敏感品系 Susceptible strain	0.000 73（0.000 67~0.000 79）	y = 14.57 + 4.64x	1	-
高效氯氰菊酯 Beta-cypermethrin	野外品系 Field strain	0.013 29（0.012 21~0.014 45）	y = 7.68 + 4.09x	18.21	中抗Moderate resistance
氯菊酯 Permethrin	敏感品系 Susceptible strain	0.001 29（0.001 15~0.001 43）	y = 11.37 + 3.94x	1	-
氯菊酯 Permethrin	野外品系 Field strain	0.269 83（0.198 63~0.359 48）	y = 0.73 + 1.29x	209.17	高抗High resistance

杀虫剂 Insecticide	蚊虫数 No. mosquito	等位基因数量（占比/%） Number of alleles (Percentage/%)		基因型数量（占比/%） Number of genotypes (Percentage/%)
杀虫剂 Insecticide	蚊虫数 No. mosquito	野生型 Wildtype GTA/V	突变型 Mutant GGA/G	野生型纯合子 Wildtype homozygote V/V	野生/突变型杂合子 Wild/mutant heterozygote V/G	突变型纯合子 Mutant homozygote G/G
溴氰菊酯 Deltamethrin	98	169（86.22）	27（13.78）	72（73.47）	25（25.51）	1（1.02）
高效氯氰菊酯 Beta-cypermethrin	94	157（83.51）	31（16.49）	63（67.02）	31（32.98）	0（0）
氯菊酯 Permethrin	96	143（74.48）	49（25.52）	55（57.29）	33（34.38）	8（8.33）
合计 Total	288	469（81.42）	107（18.58）	190（65.97）	89（30.90）	9（3.13）

杀虫剂 Insecticides	蚊虫数 No. mosquito	等位基因数量（占比/%） Number of alleles (Percentage/%)		基因型数量（占比/%） Number of genotypes (Percentage/%)
杀虫剂 Insecticides	蚊虫数 No. mosquito	野生型 Wildtype ATC/I	突变型 Mutant ACC/T	野生型纯合子 Wildtype homozygote I/I	野生/突变型杂合子 Wild/mutant heterozygote I/T	突变型纯合子 Mutant homozygote T/T
溴氰菊酯 Deltamethrin	98	195（99.49）	1（0.51）	97（98.98）	1（1.02）	0（0）
高效氯氰菊酯 Beta-cypermethrin	94	187（99.47）	1（0.53）	93（98.94）	1（1.06）	0（0）
氯菊酯 Permethrin	96	190（98.96）	2（1.04）	94（97.92）	2（2.08）	0（0）
合计 Total	288	572（99.31）	4（0.69）	284（98.61）	4（1.39）	0（0）

杀虫剂 Insecticide	蚊虫数 No. mosquito	等位基因数量（占比/%） Number of alleles (Percentage/%)			基因型数量（占比/%） Number of genotypes (Percentage/%)
杀虫剂 Insecticide	蚊虫数 No. mosquito	野生型 Wildtype TTC/F	突变型 Mutant TCC/S	突变型 Mutant TGC/C	野生型纯合子 Wildtype homozygote F/F	野生/突变型杂合子 Wild/mutant heterozygote F/S	突变型纯合子 Mutant homozygote C/C	突变型纯合子 Mutant homozygote S/S
溴氰菊酯 Deltamethrin	98	32（16.33）	160（81.63）	4（2.04）	2（2.04）	28（28.57）	2（2.04）	66（67.35）
高效氯氰菊酯 Beta-cypermethrin	94	37（19.68）	151（80.32）	0（0）	4（4.26）	29（30.85）	0（0）	61（64.89）
氯菊酯 Permethrin	96	57（29.69）	123（64.06）	12（6.25）	16（16.67）	25（26.04）	6（6.25）	49（51.04）
合计 Total	288	126（21.87）	434（75.35）	16（2.78）	22（7.64）	82（28.47）	8（2.78）	176（61.11）