Analysis of knockdown resistance gene mutations in pyrethroid insecticide resistant populations of Aedes aegypti in Yunnan Province
LAN Xue-mei, YANG Ming-dong, YANG Rui, JIANG Jin-yong
Yunnan Institute of Parasitic Diseases,Yunnan Provincial Center of Arborvirus Research, Yunnan Provincial Key Laboratory of Vector-borne Diseases Control and Research, Academician Workstation of Professor Jin Ningyi, Training Base of International Scientific Exchange and Education in Tropical Diseases for South and Southeast Asia, Yunnan Institute of Parasitic Diseases Innovative Team of Key Techniques for Vector Borne Disease Control and Prevention (Developing), Pu'er 665099, China
Abstract：To detect the knockdown resistance (kdr) gene mutations against permethrin and cypermethrin insecticides in Ae.aegypti mosquitoes in key areas of dengue fever in Yunnan Province, with the aim of the elucidating the relationship between the resistance phenotype and kdr gene mutations, we collected Ae.aegypti from Jinghong City, Mengla County, Menghai County, Ruili City and Gengma County, and bioassayed them with permethrin and cypermethrin insecticides after species identification. Individual genomic DNA was extracted. Allele-specific PCR (AS-PCR) amplification and analysis of kdr gene fragments, genotypes and gene frequencies of kdr mutations in resistant and sensitive phenotypes were performed. AS-PCR successfully detected V1016G and F1534C mutations in the 580 female wild Ae.aegypti by direct sequencing of the sodium channel gene. The mutation frequency of V1016G was 99.83% (579/580), and that of F1534C was 46.38% (269/580). The mutation frequencies of V1016G and F1534C were statistically significant (χ2=421.338, P=0.000, P<0.05). Among the five Ae.aegypti populations, the V1016G mutation rate reached 100.00% in all regions except Ruili City; the mutation rate of F1534C in Jinghong was the highest at 84.16%, and that in Gengma was the lowest at 5.66%. The mutation rates of V1016G and F1534C in both resistant and sensitive phenotypes were statistically significant (χ2=7.298, P=0.007, P<0.05; χ2=14.010, P=0.000, P<0.05). Among the 580 samples, 65 samples had both V1016G and F1534C site mutations, and the mutation rate was11.21%. The simultaneous mutation rates in different regions ranged from 0 to 33.66%, The maximum mutation rate in Jinghong was 33.66%. V1016G and F1534C mutations were found in the Ae.aegypti populations in key areas of dengue fever in Yunnan Province. The V1016G mutation rate was high and this mutation was widely distributed. The population of Ae.aegypti in Jinghong had the highest rate of F1534C mutation. The sensitive phenotype and resistant phenotype V1016G and F1534C mutation rates differed, thus suggesting a relationship between resistance and mutation.
兰学梅, 杨明东, 杨锐, 姜进勇. 云南省埃及伊蚊对拟除虫菊酯类抗性群体的击倒抗性基因突变分析[J]. 中国人兽共患病学报, 2020, 36(12): 993-999.
LAN Xue-mei, YANG Ming-dong, YANG Rui, JIANG Jin-yong. Analysis of knockdown resistance gene mutations in pyrethroid insecticide resistant populations of Aedes aegypti in Yunnan Province. Chinese Journal of Zoonoses, 2020, 36(12): 993-999.
[1] 董学书,蔡福昌,周红宁,等.云南省边境口岸蚊类调查[J].中国媒介生物学及控制杂志2004,15(2):142-145. [2] 李春敏,董学书,杨明东.云南省埃及伊蚊地理分布与季节消长[J].中国媒介生物学及控制杂志,2018,29(4):388-392. DOI:10.11853/j.issn.1003.8280.2018.04.000 [3] 刘永华,尹小雄,杨召兰,等.云南省瑞丽市2013年登革热暴发的流行病学分析[J].中国媒介生物学及控制杂志,2014,25(6):524-526.DOI:10.11853/j.issn.1003.4692.2014.06.010 [4] 李华昌,杨贵荣.云南临沧市首次本地感染登革热流行病学调查[J].中国热带医学,2009, 9(10):2013-2014. [5] Wen S,Ma D,Lin Y,et al.Complete genome characterization of the 2017 dengue outbreak in Xishuangbanna, a Border City of China, Burma and Laos[J].Front Cell Infect Microbiol,2018,8:148.DOI: 10.3389/fcimb.2018.00148 [6] 王建羽,兰学梅,杨锐,等.云南省耿马县埃及伊蚊对6种常用卫生杀虫剂的抗药性研究[J].中国媒介生物学及控制杂志,2017,28(5):444-446. DOI:10.11853/j.issn.1003. 8280. 2017.05.008 [7] 兰学梅,郑宇婷,董朝良,等.云南省瑞丽市埃及伊蚊和白纹伊蚊对常用杀虫剂的抗药性调查[J].中国媒介生物学及控制杂志,2017,28(6):572-575. DOI:10.11853/j.issn.1003.8280. 2017.06.014 [8] 董学书,周红宁,龚正达.云南蚊类志(下卷)[M].昆明:云南科技出版社,2010:62-64. [9] Li C-X,Kaufman PE,Xue R-D,et al.Relationship between insecticide resistance and kdr mutations in the dengue vector Aedes aegypti in Southern China[J].Parasit Vectors,2015,8:325.DOI:10.1186/s13071-015-0933-z [10] Wuliandari JR, Lee SF, White VL,et al.Association between three mutations, F1565C, V1023G and S996P, in the voltage-sensitive sodium channel gene and knockdown resistance in Aedes aegypti from Yogyakarta, Indonesia[J]. Insects,2015,6(3):658-685. DOI:10.3390/insects6030658 [11] Du Y, Nomura Y, Zhorov BS,et al.Sodium channel mutations and pyrethroid resistance in Aedes aegypti[J]. Insects,2016,7(4). DOI:10.3390/insects7040060 [12] 师灿南. 景洪市登革热媒介伊蚊对常用杀虫剂的抗药性及机制初步研究[D].北京:中国疾病预防控制中心,2017. [13] Hitoshi Kawada S Z M O.Co-occurrence of point mutations in the voltage-gated sodium channel of pyrethroid-resistant Aedes aegypti populations in Myanmar[J].PLoS Negl Trop Dis,2014 Jul 31;8(7):e3032.DOI: 10.1371/journal.pntd.0003032 [14] Stenhouse SA,Plernsub S,Yanola J,et al.Detection of the V1016G mutation in the voltage-gated sodium channel gene of Aedes aegypti(Diptera:Culicidae)by allele-specific PCR assay,and its disturbution and effect on deltamethrin resistance in Thailand[J].Parasite Vectors,2013,6(1):253.DOI: 10.1186/1756-3305-6-253 [15] Marcombe S, Fustec B, Catte J, et al.Distribution of insecticide resisitance and mechanisms involved in the arbovirus vector Aedes aegypti in Laos and implication for vector control[J]. PLoS Negl Trop Dis,2019,13(12):e0007852.DOI:10.1371/journal.pntd.0007852 [16] Kawada H, Higa Y, Komagata O,et al.Widespread distribution of a newly found point mutation in voltage-gated sodium channel in pyrethroid-resistant Aedes aegypti populations in Vietnam[J]. PLoS Negl Trop Dis, 2009,3(10):e527. DOI: 10.1371/journal.pntd.0000527 [17] Srisawat R, Komalamisra N, Eshita Y, et al.Point mutations in domain II of the voltage-gated sodium channel gene in deltamethrin-resistant Aedes aegypti (Diptera: Culicidae)[J]. Appl Entomol Zool, 2010, 45(2): 275-282. DOI: 10.1303/aez.2010.275 [18] Srisawat R, Komalamisra N, Apiwathnasorn C, et al.Field-collected permethrin-resistant Aedes aegypti from central Thailand contain point mutations in the domain IIS6 of the sodium channel gene (KDR)[J]. Southeast Asian J Trop Med Public Health,2012,43(6):1380-1386. [19] Chang C, Shen WK, Wang TT, et al.A novel amino acid substitution in a voltage-gated sodium channel is associated with knockdown resistance to permethrin in Aedes aegypti[J]. Insect Biochem Mol Biol,2009,39(4):272-278. DOI: 10.1016/j.ibmb.2009.01.001 [20] Ishak IH, Jaal Z, Ranson H, et al.Contrasting patterns of insecticide resistance and knockdown resistance (kdr) in the dengue vectors Aedes aegypti and Aedes albopictus from Malaysia[J]. Parasite Vectors,2015,8:181.DOI:10.1186/s13071-015-0797-2 [21] Kushwah RB, Dykes CL, Kapoor N,et al.Pyrethroid-resistance and presence of two knockdown resistance (kdr) mutations, F1534C and a novel mutation T1520I, in Indian Aedes aegypti[J]. PLoS Negl Trop Dis,2015,9(1):e3332.DOI: 10.1371/journal.pntd.0003332 [22] Chung HH, Cheng IC, Chen YC,et al.Voltage-gated sodium channel intron polymorphism and four mutations comprise six haplotypes in an Aedes aegypti population in Taiwan[J]. PLoS Negl Trop Dis, 2019,13(3):e0007291.DOI: 10.1371/journal.pntd.0007291
