Monitoring Of Resistance And Target Mutation Detection Of Spodoptera Frugiperda To Commonly Used Insecticides In Anhui Province

The fall armyworm Spodoptera frugiperda(J.E.Smith),a deadly agricultural pest native to the Americas,entered Huangshan City of Anhui Province in May 2019 and expanded quickly.Due to constraints such as lack of natural enemies in its native range,it is highly susceptible to outbreaks.The Spodoptera frugiperda has developed resistance to most of insecticides as a result of the long-term,excessive usage of pesticides.The study’s emphasis on improving the detection of target mutations and resistance monitoring guide using of pesticides and hinder the development of resistance.Compared to PCR,loop-mediated isothermal amplification(LAMP)is a practical technology for target mutations in fall armyworm,which is quick,easy,effective,visible,and inexpensive.1.The toxicity of commonly insecticides to the fall armyworm in Anhui ProvinceThis study was to assess the toxicity of frequently utilized organophosphorus and carbamate insecticides,as well as other novel insecticides,in Anhui Province during 2022.The study found that fall armyworm did not show a significant increase in susceptibility to organophosphorus and carbamate insecticides when compared to relatively sensitive strains.However,the toxicity activity was lower than other common pesticides,like emamectin benzoate,spinetoram,and chlorantraniliprole.Resistance to indoxacarb and spinetoram in Spodoptera frugiperda field populations was less than 2.0-fold.Spodoptera frugiperda in Fuyang and Bengbu areas was 3.69-fold and 6.08-fold resistant to emamectin benzoate,whereas resistance to chlorantraniliprole was 5.11-fold,7.33-fold and 10.52-fold in Bengbu,Hefei and Fuyang,respectively.Therefore,resistance monitoring needs to be strengthened when using insecticides such as emamectin benzoate and chlorantraniliprole for control of Spodoptera frugiperda.2.Detection of target site mutations of commonly insecticides by PCR techniqueIn Anhui Province,the PCR technique was employed to identify mutations in related target sites to Spodoptera frugiperda with acetylcholinesterase(AChE),ryanodine receptor(RyR),voltage-gated sodium channel(VGSC),nicotinic acetylcholine(nAChRs)α6subunit,and glutamate-gated chloride channel(GluCl)in the field of 2022.The sequencing analysis showed the presence of heterozygous mutations in the ace-1 gene at the A201 S,homozygous and heterozygous mutations at the F290 V.Field populations had a higher frequency of these mutations in the ace-1 gene compared to relatively sensitive strains.The presence of the I4734 M heterozygous mutation in RyR was also detected on Spodoptera frugiperda in the Hefei area,with an occurrence frequency of 1/60.Additionally,the Spodoptera frugiperda that invaded Anhui Province were found to have mutations associated with resistance to organophosphorus,carbamate,and diamide insecticides.However,they did not carry mutations associated with resistance to emamectin benzoate,indoxacarb,and spinetoram.3.Identify of ace-1 at the sites of A201 and F290 of the fall armyworm using LAMPIn this study,a rapid detection method based on LAMP technology was established to detect the mutation frequency of organophosphorus and carbamate insecticide targets.The method was used to detect the mutations A201 S and F290 V in the Spodoptera frugiperda ace-1 gene.To amplify the Spodoptera frugiperda ace-1 gene,two pairs of specific primers were designed for the A201 S and F290 V.The amplification was conducted using LAMP and the reaction systems were observed in a water bath at 68°C for 85 minutes.The results were highly specific and sensitive,capable of detecting samples as low as 1 pg/μL.The results of the LAMP,which used extracted DNA samples of Spodoptera frugiperda in the field,were found to be consistent with those obtained from the PCR method.The accuracy rate of the LAMP method was 100%,making it a suitable option for detecting organophosphorus and carbamate insecticide target mutations in Spodoptera frugiperda in the field.This method is not only simple and rapid,but also have strong specificity.4.Identify of RyR at the sites of I4743 and G4891 of the fall armyworm using LAMPThis study utilized LAMP technology to quickly identify low-level target mutations of Spodoptera frugiperda to diamide insecticides.The researchers screened for LAMP primer sets that could detect the I4734 L,I4734V,and I4734 M mutations on RyR associated with diamide insecticide resistance.The selection of these primer sets was based on LAMP technology.To detect the presence of one or more morphological mutations at the I4734 site in RyR,different BIP primers were designed for the different mutation types of I4734 L,I4734V,and I4734 M.Additionally,three primers(FIP,F3,and B3)were designed for the same position.The colour change is observed when the LAMP reaction system is incubated in a water bath at 65°C for 65 min.The LAMP method can detect the mutation of the G4891 E site on RyR.The mutation of the G4891 E site on RyR can be detected using the LAMP method.This is accomplished by designing two pairs of specific primers at the G4891 E site and amplifying the LAMP.After 85 minutes of water bath at 68°C,the mutant sample solution changes from pink to yellow,while the unchanged sample and water control remain unchanged in color.The LAMP method is a stable,time-efficient,simple,and sensitive technique that can detect one or more morphological mutations at the I4734 site simultaneously.This makes it suitable for rapid detection of resistance to diamide insecticides in the field and provides a basis for integrated control of Spodoptera frugiperda...