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The long-term and frequent use of pyrethroid insecticides has led to the development of pyrethroid resistance in many insect populations around the world. Specific mutations in the voltage-gated sodium channel (VGSC) have been well documented to be responsible for knockdown resistance (kdr) to pyrethroids and dichlorodiphenyltrichloroethane (DDT) in a variety of arthropods. However, reports regarding naturally occurring kdr mutation in field populations of the German cockroach Blattella germanica (Linnaeus) (Dictyoptera: Blattellidae) in China have remained scarce. In this study, a survey was conducted to detect the presence and frequency of kdr mutations in field populations of B. germanica across Neijiang, Sichuan province of China. In addition to the previously reported L993F mutation, a new amino acid substitution L993S was discovered for the first time. Overall, the classical 993F was the dominant allele with frequencies ranging from 61.8 to 89.6%, while the frequencies of the novel L993S mutation were in the range between 2.5 and 15.0%. Notably, high frequencies (50.0–79.2%) of resistant homozygotes were detected in our samples, indicating high levels of pyrethroid resistance in these B. germanica populations. The results suggest that alternative insecticides with a mode of action different from pyrethroids should be considered in the control of German cockroaches in these regions. [ABSTRACT FROM AUTHOR]

Background: Sichuan province is located in the southwest of China, and was previously a malaria-endemic region. Although no indigenous malaria case has been reported since 2011, the number of imported cases is on the rise. Insecticide-based vector control has played a central role in the prevention of malaria epidemics. However, the efficacy of this strategy is gravely challenged by the development of insecticide resistance. Regular monitoring of insecticide resistance is essential to inform evidence-based vector control. Unfortunately, almost no information is currently available on the status of insecticide resistance and associated mechanisms in Anopheles sinensis, the dominant malaria vector in Sichuan. In this study, efforts were invested in detecting the presence and frequency of insecticide resistance-associated mutations in three genes that encode target proteins of several classes of commonly used insecticides. Methods: A total of 446 adults of An. sinensis, collected from 12 locations across Sichuan province of China, were inspected for resistance-conferring mutations in three genes that respectively encode acetylcholinesterase (AChE), voltage-gated sodium channel (VGSC), and GABA receptor (RDL) by DNA Sanger sequencing. Results: The G119S mutation in AChE was detected at high frequencies (0.40–0.73). The predominant ace-1 genotype was GGC/AGC (119GS) heterozygotes. Diverse variations at codon 1014 were found in VGSC, leading to three different amino acid substitutions (L1014F/C/S). The 1014F was the predominant resistance allele and was distributed in all 12 populations at varying frequencies from 0.03 to 0.86. The A296S mutation in RDL was frequently present in Sichuan, with 296SS accounting for more than 80% of individuals in six of the 12 populations. Notably, in samples collected from Chengdu (DJY) and Deyang (DYMZ), almost 30% of individuals were found to be resistant homozygotes for all three targets. Conclusions: Resistance-related mutations in three target proteins of the four main classes of insecticides were prevalent in most populations. This survey reveals a worrisome situation of multiple resistance genotypes in Sichuan malaria vector. The data strengthen the need for regular monitoring of insecticide resistance and establishing a region-customized vector intervention strategy. [ABSTRACT FROM AUTHOR]

The cotton aphid, Aphis gossypii Glover, is one of the most important agricultural insect pests. Pyrethroid and neonicotinoid insecticides have generally shown excellent control of A. gossypii, but many populations of this pest have developed resistance against these classes of insecticides. The success of insecticide resistance management strategies requires detailed knowledge of both phenotype and genotype of the target insect pest. In this study, we attempted to understand the molecular status of insecticide resistance in cotton aphid populations in Xinjiang Uygur Autonomous Region of China, the major cotton planting region of China. In addition to the previously reported M918L mutation, we discovered another substitution (M918V) in the voltage‐gated sodium channel (VGSC). Moreover, we developed a molecular assay that could be used to detect precisely the R81T mutation in the nicotinic acetylcholine receptor (nAChR). This survey revealed that 918L was the predominant VGSC allele with a frequency ranging from 50.0% to 56.7%. Notably, appreciable frequencies (between 10% and 40%) of the resistance 81T allele of the nAChR gene were detected in three investigated populations. The prevalent co‐occurrence of both VGSC 918L/V and nAchR 81T indicates a worrisome situation of multiple resistance to both pyrethroids and neonicotinoids. [ABSTRACT FROM AUTHOR]

Four putative knockdown resistance (kdr) mutations have been documented in the voltage-gated sodium channel (VGSC) gene of Cimex hemipterus from several countries. However, no information regarding kdr mutations in any Chinese tropical bed bug population is available to date. In this study, a double-mutation（M918I + L1014F）kdr allele was identified in six C. hemipterus populations across Guangxi Zhuang Autonomous Region of China. The frequency of this allele was 100% in all the six examined populations. In addition, only two cytochrome c oxidase I (COI) gene haplotypes, with one synonymous nucleotide variation, were identified in a total of 48 individuals from six locations. The fixation and broad geographic distribution of this resistant allele questions the continued use of pyrethroids in the treatment of tropical bed bug infestations. The very low genetic diversity within and among these populations indicates that these bed bugs may have a single origin. [ABSTRACT FROM AUTHOR]

Background: Malaria is a deadly vector-borne disease in tropical and subtropical regions. Although indigenous malaria has been eliminated in Guangxi of China, 473 confirmed cases were reported in the Northern region of neighbouring Vietnam in 2014. Considering that frequent population movement occurs across the China–Vietnam border and insecticide resistance is a major obstacle in disease vector control, there is a need to know the genotype and frequency of insecticide resistance alleles in Anopheles sinensis populations along the China–Vietnam border and to take action to prevent the possible migration of insecticide resistance alleles across the border. Methods: Two hundred and eight adults of An. sinensis collected from seven locations in Guangxi along the China–Vietnam border were used in the investigation of individual genotypes of the AsRDL gene, which encodes the RDL gamma-aminobutyric acid (GABA) receptor subunit in An. sinensis. PCR-RFLP (polymerase chain reaction-restriction fragment length polymorphism) analysis was deployed to genotype codon 345, while direct sequencing of PCR products was conducted to clarify the genotypes for codons 296 and 327 of the AsRDL gene. The genealogical relation of AsRDL haplotypes was analyzed using Network 5.0. Results: Three putative insecticide resistance related mutations (A296S, V327I and T345S) were detected in all the seven populations of An. sinensis in Guangxi along the China–Vietnam border. The resistance-conferring A296S mutation was found to be widely distributed and present at notably high frequencies (78.8% to 100%). Relatively lower frequencies of mutations V327I (26.9% to 53.2%) and T345S (0% to 28.8%) were observed. The V327I or T345S always occurred in the presence of A296S. Evolutionary analysis of 21 AsRDL haplotypes indicated multiple origins of the A296S and V327I mutations. Conclusion: The resistance A296S allele was present at high frequencies in the An. sinensis populations along the China–Vietnam border, indicating a risk of resistance to insecticides targeting RDL. The double mutations (A296S + V327I) may have evolved from alleles carrying the A296S mutation by scaffolding the additional mutation V327I, and A296S allele may have multiple evolutionary origins. These findings will help inform strategies for vector control and malaria prevention. [ABSTRACT FROM AUTHOR]

Abstract: A Helicoverpa armigera population was collected from Shandong province, China. After 15 generations of selection in the laboratory, the H. armigera strain developed more than 20-fold resistance to spinosad. At LD50 level, no significant cross-resistance was found between spinosad and chlorpyrifos, methomyl, avermectin and chlorfenapyr except for fenvalerate with a low cross-resistance of 2.4-fold. However, LD99 values of fenvalerate against the parental and resistant strains were not different significantly. After inhibitors were used, spinosad resistance could be partially suppressed by piperonylbutoxide (PBO) and triphenylphosphate (TPP), but not by diethylmaleate (DEM). Activities of p-nitroanisole O-demethylase (ODM) developed to 8.26-fold compared with the parental strain, but no obvious changes were found in activities of carboxyl esterase (CarE) and glutathione-S-transferase (GST). The results indicated that resistance to spinosad in the cotton bollworm might be associated with an increase in cytochrome P450 monooxygenase. [Copyright &y& Elsevier]

Abstract: We investigated the molecular basis of resistance in a strain of house fly (BJD) from Beijing, China. This strain showed 567-fold resistance to commonly used deltamethrin. Flies were 64-fold resistant to deltamethrin synergized by piperonyl butoxide (PBO). The 5′-flanking sequence of the cytochrome P450 gene CYP6D1 in BJD strain had a 15-bp insert as in the LPR strain. Two mutations (kdr, super-kdr) in the voltage sensitive sodium channel (VSSC) were also detected in the BJD strain. Our results showed that a combination of resistance alleles for CYP6D1 and VSSC existed in deltamethrin resistant house flies in China. [Copyright &y& Elsevier]

Background: The oriental rat flea, Xenopsylla cheopis, is the most efficient vector of the plague. Pyrethroid insecticides such as cypermethrin, cyhalothrin and deltamethrin have been often used to limit plague transmission via controlling the vector during outbreaks. However, this strategy is threatened by the development of insecticide resistance. Understanding the mechanisms underlying pyrethroid resistance is the prerequisite for successful flea control. Methods: Partial DNA sequences of X. cheopis voltage gated sodium channel (VGSC) gene were amplified from a total of 111 individuals, collected from a natural plague epidemic foci in Baise City, Guangxi Zhuang Autonomous Region of China. These DNA fragments were sequenced. The frequency and distribution of kdr mutations were assessed in four X. cheopis populations. The origin of kdr mutations was investigated by phylogenetic and network analysis. Results: The classical knockdown resistance (kdr) mutation (L1014F) was detected in four field populations at frequencies ranging between 0.021–0.241. The mutant homozygote was observed only in one of the four populations. Seven haplotypes were identified, with two of them carrying the resistance L1014F mutation. Phylogenetic tree and network analysis indicated that the L1014F allele was not singly originated. Based on polymerase chain reaction restriction fragment length polymorphism (PCR-RFLP) profiling, an easy-to-use and accurate molecular assay for screening individual fleas for the L1014F mutation was developed. Conclusions: To our knowledge, this work represents the first report of the L1014F mutation in the plague vector X. cheopis. The incidence of the L1014F allele highlights the need of further studies on the phenotypic effect of this mutation in this plague vector. Early detection and monitoring of insecticide resistance is suggested in order to make effective control strategies in case of plague outbreaks in this region. [ABSTRACT FROM AUTHOR]

The cotton bollworm (Helicoverpa armigera) is a worldwide agricultural pest that infests many important crops. Pyrethroids targeting the voltage-gated sodium channel (VGSC) have been long used in the control of the cotton bollworm. Two amino acid substitutions (D1561V and E1565G) in H. armigera VGSC (HaVGSC) and the presence of a chimeric P450 gene (CYP337B3) have been documented to be associated with pyrethroid resistance. To understand the current occurrence of kdr mutations and the CYP337B3 gene in Chinese H. armigera populations, high-throughput amplicon sequencing was adopted to detect potential nucleotide variations in three fragments of the VGSC gene that cover 10 reported knockdown resistance (kdr) sites in insects, and gene-specific PCR was performed to examine the presence of CYP337B3 gene in H. armigera samples collected across China. The nucleotide variation analysis revealed a wealth of nucleotide variations in not only exons but also introns in the VGSC gene in Chinese H. armigera populations. However, neither previously reported kdr -conferring amino acid replacements nor other non-synonymous mutations were observed in a total of 1439 examined individuals. Population genetic analysis suggested that the H. armigera population in Nanchang, Jiangxi Province (JNC) had a moderate genetic differentiation from other populations, while no significant divergence was observed in other populations in northern and northwestern China. The CYP337B3 was present in all the examined individuals, indicating that CYP337B3 is extensively fixed in H. armigera populations across China. These results support that point mutations in VGSC are not a major factor involved in the current pyrethroid resistance in H. armigera. Instead, CYP337B3 plays a prevalent role in the development of resistance to pyrethroids in H. armigera. [Display omitted] • Amplicon sequencing is used to detect knockdown resistance (kdr) mutation in H. armigera. • Extensive nucleotide variations are observed in the voltage-gated sodium channel (VGSC) gene. • Neither the reported kdr mutations nor other non-synonymous variations are identified in VGSC. • The CYP337B3 gene is commonly fixed in field populations of H. armigera across China. • A low overall genetic differentiation is found among different H. armigera populations. [ABSTRACT FROM AUTHOR]

Abstract: Insecticides have been extensively used for house fly control in China, with dichlorvos and deltamethrin being widely used. Knowledge about the current status of insecticide resistance and the underlying genetic changes is crucial for developing effective fly control strategies. The susceptibility to dichlorvos and deltamethrin, and the frequencies of genetic mutations involved in insecticide resistance were studied in five field populations of the house fly collected across China. Bioassay results show that flies exhibit 14- to 28-fold resistance to dichlorvos and 41- to 94-fold resistance to deltamethrin, indicating that dichlorvos and deltamethrin resistance are common in house fly populations in China. Molecular analysis reveals that flies from the five various locations carry resistance alleles at multiple loci and have diverse allelic types, different relative frequencies and combinations of each allele. Four non-synonymous single nucleotide polymorphisms (SNPs) (i.e. V260L, G342A/V, F407Y) in acetylcholinesterase (Ace) and two mutations (W251L/S) in a carboxylesterase (MdαE7) were commonly present in the field house flies. The L1014H rather than L1014F mutation in the voltage sensitive sodium channel gene (Vssc) was widely distributed in Chinese house flies. CYP6D1v1, which confers pyrethroid resistance, was found in all the five tested populations in China, although its frequency in house fly from Shandong province was very low. Our results suggest that resistance monitoring and management of house flies should be customized for a given location. [Copyright &y& Elsevier]

The Asian tiger mosquito, Aedes albopictus, is an important vector of chikungunya, dengue, yellow fever, and Zika viruses. Vector control remains an important means for the prevention and control of vector-borne diseases. The development of insecticide resistance has become a serious threat to the efficacy of insecticide-based control programs. To understand the resistance status and the underlying genetic mechanism in mosquitoes in Guangyuan City of Sichuan Province, China, we investigated the susceptibility of Ae. albopictus to four commonly used insecticides. We found that all the examined populations were susceptible to malathion and propoxur. However, Ae. albopictus populations in Guangyuan showed a possible resistance to the two tested pyrethroids (beta-cypermethrin and deltamethrin). Notably, phenotypic resistance to deltamethrin was detected in 2 of the 7 populations. The potential of resistance to pyrethroids was confirmed by the presence of knockdown resistance (kdr) related mutations in the voltage-gated sodium channel. Four kdr mutations (V1016G, I1532T, F1534L, and F1534S) were identified to be present alone or in combination, and their distribution displayed significant spatial heterogeneity. These findings are helpful for making evidence-based mosquito control strategies and highlight the need to regularly monitor the dynamics of pyrethroid resistance in this city., (Copyright © 2024 by The American Mosquito Control Association, Inc.)

Culex quinquefasciatus and Cx. tritaeniorhynchus are 2 dominant disease vectors in Neijiang City, Sichuan Province, China. Although there is evidence of confirmed resistance against insecticides in mosquito vectors, nothing is known about the existing insecticide resistance-conferring mutations in Cx. quinquefasciatus and Cx. tritaeniorhynchu in this region so far. In this study, the G119S mutation in the acetylcholinesterase (AChE) was detected in Cx. quinquefasciatus at a very low frequency (0.9%) with no resistant homozygotes being observed. Two resistance mutations in the voltage-gated sodium channel (VGSC) (L1014F and L1014S) were found in Cx. quinquefasciatus with frequencies of 88.7% and 8.3%, respectively. By contrast, the AChE F455W mutation was found to be fixed (with a frequency of 100%) in 3 of the 5 studied populations, with an overall frequency being 98.1%. In addition, 1 resistance-conferring VGSC mutation (L1014F) was detected with an overall frequency of 15.2% in Cx. tritaeniorhynchus. These results indicate that the well-recognized insecticide resistance-conferring mutations in both AChE and VGSC are present in the 2 Culex species in Neijiang. The contrasting patterns in the frequency of resistance alleles indicate that species-customized strategies of insecticide resistance management should be considered for the 2 species., (Copyright © 2023 by The American Mosquito Control Association, Inc.)

Background: Anopheles sinensis is a dominant vector for malaria transmission in Asian countries. Voltage-gated sodium channel (VGSC) mutation-mediated knock-down resistance (kdr) has developed in many A. sinensis populations because of intensive and long-term use of pyrethroids. Our previous study showed that multiple mutations at position 1014 of the VGSC were heterogeneously distributed in A. sinensis populations across Sichuan, China., Methods: To understand resistance genotypes at the haplotype level and reconstruct the phylogenetic relationship of VGSC haplotypes, a cost-effective next-generation sequencing (NGS)-based amplicon sequencing approach was established to clarify haplotypes containing codon 1014 of the VGSC gene from a total of 446 adults collected in 12 locations of Sichuan, China., Results: Nineteen (19) haplotypes were identified, including 11 wild 1014L, 6 resistance 1014F, and 2 resistance 1014C haplotypes. We found that resistance haplotypes of A. sinensis VGSC were widely distributed at frequencies ranging from 3.67 to 92.61%. The frequencies of the 1014C haplotype in the southeast of Sichuan (Luzhou, Guangan, and Suining) were relatively higher than those in other sampling locations. Phylogenetic analyses support that kdr-type mutation at position 1014 is not singly originated and resistance 1014C haplotypes evolve from TTT-encoding 1014F., Conclusions: A cost-effective next-generation sequencing (NGS)-based amplicon sequencing approach has been established in this study. The data revealed the patchy distribution of VGSC resistance haplotypes with overall high frequencies in Sichuan, China. Phylogenetic analyses support multiple origins and sequential evolution (1014L → 1014F → 1014C) for kdr-type mutations in A. sinensis., (© 2021. The Author(s).)

Background: In South Asia, the epidemiology of malaria is complex, and transmission mainly occurs in remote areas near international borders. Vector control has been implemented as a key strategy in malaria prevention for decades. A rising threat to the efficacy of vector control efforts is the development of insecticide resistance, thus it is important to monitor the type and frequency of insecticide resistant alleles in the disease vectors such as An. sinensis along the China-Vietnam border. Such information is needed to synthesize effective malaria vector control strategies., Methods: A total of 208 adults of An. sinensis, collected from seven sites in southwest Guangxi along the China-Vietnam border, were inspected for the resistance-conferring G119S mutation in acetylcholinesterase (AChE) by PCR-RFLP (polymerase chain reaction restriction fragment length polymorphism) and kdr mutations in the voltage-gated sodium channel (VGSC) by sequencing. In addition, the evolutionary origin of An. sinensis vgsc gene haplotypes was analyzed using Network 5.0., Results: The frequencies of mutant 119S of AChE were between 0.61-0.85 in the seven An. sinensis populations. No susceptible homozygote (119GG) was detected in three of the seven sites (DXEC, LZSK and FCGDX). Very low frequencies of kdr (0.00-0.01) were detected in the seven populations, with most individuals being susceptible homozygote (1014LL). The 1014F mutation was detected only in the southeast part (FCGDX) at a low frequency of 0.03. The 1014S mutation was distributed in six of the seven populations with frequencies ranging from 0.04 to 0.08, but absent in JXXW. Diverse haplotypes of 1014L and 1014S were found in An. sinensis along the China-Vietnam border, while only one 1014F haplotype was detected in this study. Consistent with a previous report, resistant 1014S haplotypes did not have a single origin., Conclusions: The G119S mutation of AChE was present at high frequencies (0.61-0.85) in the An. sinensis populations along the China-Vietnam border, suggesting that the vector control authorities should be cautious when considering carbamates and organophosphates as chemicals for vector control. The low frequencies (0.00-0.11) of kdr in these populations suggest that pyrethroids remain suitable for use against An. sinensis in these regions.

Background: Malaria is one of the most serious vector-borne diseases in the world. Vector control is an important measure for malaria prevention and elimination. However, this strategy is under threat as disease vectors are developing resistance to insecticides. Therefore, it is important to monitor mechanisms responsible for insecticide resistance. In this study, the presence of G119S mutation in the acetyl cholinesterase-encoding gene (ace-1) was investigated in nine Anopheles sinensis populations sampled across Guangxi Zhuang Autonomous Region China., Methods: PCR-RFLP (polymerase chain reaction-restriction fragment length polymorphism) method was used to genotype each individual adult of An. sinensis. Direct sequencing of PCR products was performed to verify the accuracy of PCR-RFLP genotyping result. Population genetics analysis was conducted using Genepop programme., Results: The frequencies of susceptible homozygotes, heterozygotes and resistant homozygotes in the nine populations ranged between 0-0.296, 0.143-0.500 and 0.333-0.857, respectively. Overall, a high frequency (0.519-0.929) of mutant 119S allele was observed and the genotype frequency of the ace-1 gene of An. sinensis was at Hardy-Weinberg equilibrium in each of the nine examined populations., Conclusion: The G119S mutation has become fixed and is widespread in An. sinensis field populations in Guangxi, China. These findings are useful in helping design strategies for An. sinensis control.