Modification Of Contact Irritability Associated With Deltamethrin Resistance In Anopheles Sinensis And Preliminary Studies On Its Molecular Mechanism

Anopheles sinensis is one of the main vectors of malaria in China and Southeast Asian countries.Therefore,the key measure to control malaria is An.sinensis management and block it transmission routes.The main method to fight against malaria is insecticide utilization.The pyrethroid insecticides have been recommended by WHO and applied for bed net treatment.Among them,deltamethrin is widely used in China and long-term usage has resulted in serious resistance of mosquitoes,even changed their behaviors,which significantly weakens the effect of existing prevention methods.The resistance to deltamethrin in An.sinensis has been extensively reported,but there is no relevant studies on their behavior changes.In this study,a laboratory susceptible population(WX-LS)and field resistant populations from Yuanyang in Yunnan province(YY-FR),Bishan in Chongqing Municipality(BS-FR),Wuhe in Anhui province(WH-FR)were used as materials.The deltamethrin resistance levels were determined according to the standard recommended by WHO.Species identification and mutation analysis of gene kdr were achieved by molecular experiments.Their contact irritability behavior on deltamethrin-treated nets was determined using WHO cone bioassays.To explore the molecular mechanism of behavior changes,comparative transcriptome analysis between YY-FR and WX-LS was performed.The main results are as follows:(1)Resistance level and kdr mutation detection of An.sinensisThe resistance to deltamethrin was found all populations of YY-FR,BS-FR,WH-FR by using contact tube bioassays.The strongest resistance was found in WH-FR population,then followed by YY-FR and BS-FR.Mutation analysis showed that three mutation type of kdr gene,L1014F(TTT or TTC)and L1014C(TGT),were detected in BS-FR and WH-FR population.However,no mutation was found in YY-FR population.The mutation frequency of kdr gene was 82.88% in BS-FR population,and 100% in WH-FR population.(2)Determinations of contact behavior to deltamethrin in different geographical populationsContact repellency to deltamethrin-impregnated bed nets(DTNs)was evaluated,and contact behavioral changes of field resistance populations to DTNs were compared using a modified WHO cone bioassay.The results showed that the contact irritabilities to deltamethrin were much stronger in laboratory susceptible population than in field resistance populations.The contact irritability in YY-FR population without kdr mutation was significantly decreased,suggesting that the behavior changes were not directly related to kdr mutation.(3)Transcriptome analysis for WX-LS and YY-FR population.The differentially expressed genes(DEG)in WX-LS and YY-FR population were analyzed based on the transcriptome sequencing approach.Totally,4101 DEGs were identified,of which 1843 genes were up-regulated and 2258 genes were down-regulated in YY-FR population.2494 DEGs were annotated by GO terms and mainly enriched in biological process and molecular function categories,including oxidation-reduction process,single organism metabolism process,organic cyclic compound binding,catalytic activity,et al.The KEGG pathways were enriched for up-regulated genes,including eukaryotic ribosomal synthesis,DNA replication,mismatch repair,RNA polymerase,nucleotide resection repair,base resection repair,RNA transport,et al.For down-regulated genes,KEGG pathways were enriched into carbon metabolism,starch and sucrose metabolism,and pyruvate metabolism,suggesting that the weakness of contact behavior of field resistant populations might be related to the reduction of carbohydrate metabolism.