Organophosphate Resistance Research Articles

First Report of Mutations Associated With Pyrethroid (L1014F) and Organophosphate (G119S) Resistance in Belgian Culex (Diptera: Culicidae) Mosquitoes.

The emergence of West Nile virus and Usutu virus in Europe poses a significant risk to public health. In the absence of efficient antiviral therapy or vaccine candidates, the only strategy to control these arboviruses is to target the Culex (Diptera: Culicidae) mosquito vector. However, the selection pressure caused by exposure to insecticides for vector control or agricultural pest control can lead to insecticide resistance, thereby reducing the efficacy of insecticide-based vector control interventions. In Culex mosquitoes, two of the most common amino acid substitutions associated with insecticide resistance are the kdr L1014F in voltage gated sodium channels and G119S in acetylcholinesterase. In this study, Culex pipiens biotype pipiens, Culex torrentium, and Culex modestus were sampled from 2019 to 2021 in three distinct environmental habitats (urban, peri-urban, and agricultural) in and around the city of Leuven, Belgium. Individual mosquitoes were screened for two mutations resulting in L1014F and G119S amino acid substitutions. Both mutations were observed in Cx. pipiens and Cx. modestus but not in Cx. torrentium mosquitoes across the four collection sites. Furthermore, multi-resistance or cross-resistance in Cx. pipiens could be a threat in these areas, as both mutations were observed at low frequencies. These results provide the first report of kdr L1014F and ace-1 G119S resistance mutations in Cx. pipiens and Cx. modestus mosquitoes from Belgium, highlighting the importance of mosquito surveillance to design effective arbovirus outbreak control strategies.

Insecticide Resistance and Mechanisms of Culex pipiens Populations in the Mediterranean and Aegean Regions of Turkey During 2017-2018.

Culex pipiens has a significant public health importance since it is an important vector of West Nile virus and Rift Valley fever virus. We, therefore, aimed to determine the insecticide resistance level in Cx. pipiens populations in the Aegean and Mediterranean regions of Turkey. Bioassays have been carried out against Dichlorodiphenyltrichloroethane (DDT) (4%), Malathion (5%), Fenitrothion (1%), Propoxur (0.1%), Bendiocarb (0.1%), Permethrin (0.75%) and Deltamethrin (0.05%). Biochemical analyses have been performed to detect non-specific esterase, mixed function oxidase, glutathione-s-transferase and acetylcholinesterase levels. A knockdown resistance (kdr) (L1014F) and Acetylcholinesterase (Ace-1) (G119S) mutations have been detected by using allele-specific primers and a polymerase chain reaction (PCR) amplification of specific alleles (PASA) diagnostic test was performed for detection of F290V mutation. Bioassay results showed that all Cx. pipiens populations were resistant to DDT, Malathion, Fenitrothion, Bendiocarb, Propoxur and some of the populations have started to gain Permethrin and Deltamethrin resistance. Biochemical analyses results revealed that altered glutathione-s-transferases, P450 monooxygenases, esterase levels might be responsible for DDT, organophosphate, carbamate and pyrethroid resistance in Cx. pipiens populations. Results showed mild to high frequency of L1014F, low frequency of F290V but no Ace-1 G119S mutation within the populations. Additionally, acetylcholinesterase insensitivity was not significantly high within the most of these populations. Overall results may help to fulfil the lacking information in the literature regarding insecticide resistance status and underlying mechanism of Culex pipiens populations of the Mediterranean and Aegean region of Turkey by using all bioassays, molecular tests and biochemical assays.

Bendiocarb and Malathion Resistance in Two Major Malaria Vector Populations in Cameroon Is Associated with High Frequency of the G119S Mutation (Ace-1) and Overexpression of Detoxification Genes.

The spread of pyrethroid resistance in malaria vectors is a major threat affecting the performance of current control measures. However, there is still not enough information on the resistance profile of mosquitoes to carbamates and organophosphates which could be used as alternatives. The present study assessed the resistance profile of Anopheles gambiae s.l. to bendiocarb and malathion, at the phenotypic and molecular levels, in different eco-epidemiological settings in Cameroon. Anopheles gambiae s.l. mosquitoes were collected from four eco-epidemiological settings across the country and their susceptibility level to bendiocarb and malathion was determined using WHO tubes bioassays. The ace-1 target site G119S mutation was screened by PCR. Reverse Transcription quantitative PCR 3-plex TaqMan assays were used to quantify the level of expression of eight genes associated with metabolic resistance. Resistance to malathion and/or bendiocarb was recorded in all study sites except in mosquitoes collected in Kaélé and Njombé. The Ace-1 (G119S) mutation was detected in high frequencies (>40%) in Kékem and Santchou. Both An. gambiae and An. coluzzii were detected carrying this mutation. The cytochrome P450s gene Cyp6p3 associated with carbamate resistance and the glutathione S-transferase gene Gste2 associated with organophosphate resistance were found to be overexpressed. Genes associated with pyrethroid (Cyp6m2, Cyp9k1, Cyp6p3) and organochlorine (Gste2, Cyp6z1, Cyp6m2) and cuticle resistance (Cyp4g16) were also overexpressed. The rapid spread of resistance to organophosphates and carbamates could seriously compromise future control strategies based on IRS. It is therefore becoming important to assess the magnitude of bendiocarb and malathion resistance countrywide.

Screening for insecticide resistance in Australian field populations of Bemisia tabaci (Hemiptera: Aleyrodidae) using bioassays and DNA sequencing.

BackgroundSpecies within the Bemisia tabaci cryptic species complex can cause significant crop damage. We used high‐throughput amplicon sequencing to identify the species composition and resistance allele genotypes in field populations from cotton fields in Australia. For selected populations, the resistance phenotype was determined in bioassays and compared with sequencing data.ResultsA metabarcoding approach was used to analyse the species composition in 144 field populations collected between 2013 and 2021. Two mixed AUS I and MEAM1 populations were detected, whereas the remaining 142 populations consisted of MEAM1 only. High‐throughput sequencing of organophosphate and pyrethroid resistance gene amplicons showed that the organophosphate resistance allele F331W was fixed (> 99%) in all MEAM1 populations, whereas the pyrethroid resistance allele L925I in the voltage‐gated sodium channel gene was detected at varying frequencies [1.0%–7.0% (43 populations); 27.7% and 42.1% (two populations); 95%–97.5% (three populations)]. Neither organophosphate nor pyrethroid resistance alleles were detected in the AUS I populations. Pyrethroid bioassays of 85 MEAM1 field‐derived populations detected no resistance in 51 populations, whereas 32 populations showed low frequency resistance, and 2 populations were highly resistant.ConclusionsWe demonstrate that high‐throughput sequencing and bioassays are complementary approaches. The detection of target site mutations and the phenotypic provides a comprehensive analysis of the low‐level resistance to pyrethroids that is present in Australian cotton farms. By contrast, a limited survey of whitefly populations from horticulture found evidence of high‐level resistance against pyrethroids. Furthermore, we found that the F331W allele (linked to organophosphate resistance) is ubiquitous in Australian MEAM1. © 2022 Commonwealth of Australia. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

Warmer temperatures reduce chemical tolerance in the redlegged earth mite (Halotydeus destructor), an invasive winter-active pest.

BACKGROUNDQuantifying how chemical tolerance of pest arthropods varies with temperature is important for understanding the outcomes of chemical control, for measuring and monitoring resistance, and for predicting how pesticide resistance will evolve under future climate change. We studied the redlegged earth mite, Halotydeus destructor (Tucker), a winter‐active invasive agricultural pest in Australia. Using a replicated block experiment, we tested the effect of different thermal conditions on the expression of chemical tolerance to a pyrethroid and two organophosphates. Our chemical bioassays were conducted on two redlegged earth mite populations: one possessed organophosphate resistance, whilst the other was susceptible to pesticides. Mites were first acclimated at cool (4 °C) and warm (14 °C) conditions and then exposed to pesticides in both cool (11 °C) and warm (18 °C) test conditions.RESULTSWarm test conditions generally reduced chemical tolerance to all pesticides relative to cool test conditions. Median lethal dose (LD50) values of mites tested under cool conditions were 1.12–3.57‐fold greater than of mites tested under warm conditions. Acclimation had a variable and small impact on chemical responses. Thermal factors (ratio between test temperatures) were similar between populations for each active ingredient. Despite reduced chemical tolerances under warm test conditions for individual mite populations, resistance factors (ratio between resistant and susceptible mite populations) were relatively consistent.CONCLUSIONOur data provides context for prior theoretical work demonstrating climatically constrained pesticide resistances in Australian redlegged earth mites. Estimates of temperature dependent toxicity measured in this study may be useful in parameterizing models of redlegged earth mite control under an increasingly warm and more variable climate. © 2022 The Authors. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

Associations between acetylcholinesterase-1 mutations and chlorpyrifos resistance in beet armyworm, Spodoptera exigua

Control of the beet armyworm, Spodoptera exigua depends heavily on chemical insecticides. Chlorpyrifos, an acetylcholinesterase (AChE) inhibitor, has been used in beet armyworm control for many years in China. Here we describe high level resistance to chlorpyrifos in a S. exigua strain, FX19-R, which was developed from a field-collected Chinese strain (FX) by selection with chlorpyrifos in the laboratory. FX19-R showed 1001-fold resistance to chlorpyrifos compared with the laboratory reference strain WH-S. The esterase inhibitor triphenyl phosphate (TPP) provided significant but small synergism (only 3.5-fold) for chlorpyrifos and neither of the glutathione s-transferase depletor diethyl maleate and the cytochrome P450s inhibitor piperonyl butoxide provided any detectable synergism, indicating that AChE insensitivity may play the major role in the resistance in FX19-R. Consistent with this, an amino acid substitution, F443Y (F331Y in standard Torpedo californica numbering) in AChE1 was identified in the FX19-R strain and shown to be tightly linked to chlorpyrifos resistance. Precisely homologous substitutions have been associated with organophosphate resistance in other pest species. A novel amino acid substitution, G311S (or G198S in standard numbering), was also identified in the reference strain WH-S. Recombinantly expressed AChE1 proteins carrying the G311S and F443Y substitutions were about 4.2-fold and 210-fold less sensitive to inhibition by chlorpyrifos oxon than wild-type AChE1, respectively. These results enhance our understanding of the mechanisms of chlorpyrifos resistance and provide a basis for resistance management based on monitoring the F443Y and G311S substitutions.

POTENTIAL BREEDING SITES AND ORGANOPHOSPHATE RESISTANCE STATUS OF AEDES SPP. IN YOGYAKARTA

Background: The discovery of Aedes spp. in the breeding sites is the leading cause of the high incidence of Dengue Hemorrhagic Fever (DHF) in Yogyakarta in 2019. This incidence can be calculated based on the Container Index (CI), House Index (HI), and Breteau Index (BI). Efforts to control the density of Aedes spp. The use of organophosphate insecticides in the long term and with the wrong dose will cause resistance. Purpose: This research was conducted to determine the potential breeding sites, the density, and the organophosphate resistance status of Aedes spp. in Yogyakarta. Methods: A literature review with the criteria of referenced journals from 2011-2020, national journals accredited at least Sinta 5 to Sinta 1 and international journals at least Q4 to Q1. Results: This study shows that the potential breeding sites for Aedes spp. in Yogyakarta, in general, are both indoors or outdoors, stagnant clean water, opened, not exposed to direct sunlight, and the duration of not eradicating the mosquito nests > 1 month. Aedes spp. in Yogyakarta has a high population density, so it is at high risk of transmitting dengue cases. The resistance status of Aedes spp. in various regions of Yogyakarta are categorized as resistant to organophosphate insecticides. Conclusion: Literature review requires field research also, implementation of prevention through the mosquito nests eradication program and 3M (Shutting, Draining, and Burying), and the replacement of insecticides types other than organophosphates.

Molecular and kinetic characterization of two acetylcholinesterases with particular focus on the roles of two amino acid substitutions (Y390N and F392W) in Bemisia tabaci

Two acetylcholinesterases (AChEs) are present in Bemisia tabaci (BtAChE1 and BtAChE2). A conserved AChE mutation conferring organophosphate (OP) resistance (F392W in BtAChE1) is saturated in field populations despite its potential fitness cost, and a highly conserved amino acid residue forming the backdoor of AChE is substituted with a unique amino acid (Y390N in BtAChE1) in B. tabaci. Thus, the roles and relationships of the two amino acid substitutions in the evolutionary adaptation of B. tabaci remain to be elucidated, and little information is available on the catalytic and molecular properties of BtAChE1 and BtAChE2. To determine which AChE is a more relevant target of OPs and carbamates, the molecular and kinetic properties of BtAChE1 and BtAChE2 were investigated. Both BtAChE1 and BtAChE2 were exclusively expressed in head and thorax but not in abdomen, bound to the membrane via GPI anchoring, and present as dimeric forms. Soluble monomeric form was detected only in BtAChE2. The catalytic activity of baculovirus-expressed BtAChE1 was 19.5-fold higher than that of BtAChE2. The inhibition assay revealed that the F392W mutation in BtAChE1 enhanced resistance to OPs. The artificial substitution of N390 (wild form) to Y (putative ancient form) led to reduced catalytic efficiency and increased inhibition by glycoalkaloids, suggesting that the Y390N substitution in BtAChE1 may have been required for Solanaceae host adaptation. BtAChE1 was proven to function as a main catalytic enzyme for ACh hydrolysis, thus being the main target of OPs and carbamates.

Current status of insecticide resistance in malaria vectors in the Asian countries: a systematic review

Background: The application of insecticides for malaria vector control has led to a global problem, which is the current trend of increased resistance against these chemicals. This study aimed to review the insecticide resistance status was previously determined in Asia and how to implement the necessary interventions. Moreover, the implications of resistance in malaria vector control in this region were studied. Methods: This systematic review was conducted using a predefined protocol based on PRISMA-retrieved articles from four science databases, namely ProQuest, Science Direct, EBSCO, and PubMed in the last ten years (2009 to 2019). The searching process utilized four main combinations of the following keywords: malaria, vector control, insecticide, and Asia. In ProQuest, malaria control, as well as an insecticide, were used as keywords. The following criteria were included in the filter, namely full text, the source of each article, scholarly journal, Asia, and publication date as in the last ten years. Results: There were 1408 articles retrieved during the initial search (ProQuest=722, Science Direct=267, EBSCO=50, PubMed=285, and Scopus=84). During the screening, 27 articles were excluded because of duplication, 1361 based on title and abstract incompatibility with the inclusion criteria, and 20 due to content differences. In the final screening process, 15 articles were chosen to be analyzed. From the 15 articles, it is known that there wasorganochlorine (DDT), organophosphate (malathion), and pyrethroids resistance in several Anopheles species with a less than 80% mortality rate. Conclusion:This review found multiple resistance in several Anopheles includes resistance to pyrethroid.The reports of pyrethroid resistance were quite challenging because it is considered effective in the malaria vector control. Several countries in Asia are implementing an insecticide resistance management (IRM) strategy against malaria vectors following the Global Plan for IRM.

Organophosphate Resistance in Aedes aegypti: Study from Dengue Hemorrhagic Fever Endemic Subdistrict in Riau, Indonesia.

Dengue hemorrhagic fever (DHF) is a significant health problem. The high number of cases requires preventions, including controlling the dengue vector, Aedes aegypti mosquito. One of the control methods is the use of insecticides containing organophosphate. This study aims to detect organophosphate resistance in Aedes aegypti from DHF endemic subdistrict, Riau, Indonesia by a sensitivity test of temephos and 5% malathion and measuring the activity of non-specific alpha and beta esterase enzymes. This observational study determined Aedes aegypti resistance from larvae to adult in one DHF endemic subdistrict in Riau, Indonesia. The bioassay was used for temephos sensitivity of Aedes aegypti larvae. The LC99 value was analyzed using probit and compared with the diagnostic value from WHO. The WHO susceptibility test was conducted to determine 5% malathion resistance from adult mosquitoes. The mortality of less than 90% was declared as resistant. Measurement of alpha and beta esterase levels used Lee's microplate assay technique based on visual identification and absorbance value (AV). The results showed that Aedes aegypti were resistant to temephos. It also showed that adult mosquitoes were resistant to 5% malathion. Based on the alpha esterase activity test, it was found that most of the mosquitoes showed very sensitive meanwhile, based on the beta esterase activity test, most of the mosquitoes were moderate resistance. This study suggests that Aedes aegypti population from DHF endemic subdistrict in Riau, Indonesia are indicated to develop resistance to organophosphate.

Biochemical and molecular mechanisms of acaricide resistance in Dermanyssus gallinae populations from Turkey

The poultry red mite, Dermanyssus gallinae, is the most important blood sucking ectoparasite of egg laying hens and causes economic losses in poultry farms worldwide. Although various management methods exist, the control of poultry red mites (PRMs) mainly relies on acaricides such as pyrethroids and organophosphates (OPs) in many regions of the world. However, repeated use of these synthetic chemicals has resulted in resistance development causing control failure of PRM. In this study, we investigated acaricide resistance mechanisms of Turkish PRM populations. First, we obtained the COI sequence from 30 PRM populations from different regions in Turkey and identified four different COI haplotypes. Toxicity assays showed that four field-collected PRM populations were highly resistant to the pyrethroid alpha-cypermethrin, with resistance ratios (RRs) varying between 100- and 400-fold, while two of these populations had a RR of more than 24-fold against the OP acaricide phoxim. Biochemical assays showed a relatively higher activity of glutathione-S-transferases and carboxyl-cholinesterases, two well-known classes of detoxification enzymes, in one of these resistant populations. In addition, we also screened for mutations in the gene encoding the voltage-gated sodium channel (vgsc) and acetylcholinesterase 1 (ace-1), the target-site of pyrethroids and OPs, respectively. In all but two PRM populations, at least one vgsc mutation was detected. A total of four target-site mutations, previously associated with pyrethroid resistance, M918T, T929I, F1534L, F1538L were found in domain II and III of the VGSC. The T929I mutation was present in the vgsc of almost all PRM populations, while the other mutations were only found at low frequency. The G119S/A mutation in ace-1, previously associated with OP resistance, was found in PRM for the first time and present in fourteen populations. Last, both alive and dead PRMs were genotyped after pesticide exposure and supported the possible role of target-site mutations, T929I and G119S, in alpha-cypermethrin and phoxim resistance, respectively. To conclude, our study provides a current overview of resistance levels and resistance mutations in Turkish PRM populations and might aid in the design of an effective resistance management program of PRM in Turkey.

Frequencies and mechanisms of pesticide resistance in Tetranychus urticae field populations in China.

The two-spotted spider mite Tetranychus urticate is an important agricultural pest worldwide. It is extremely polyphagous and has developed resistance to many pesticides. Here, we assessed the pesticide resistance of seven field populations of T. urticae in China, their target site mutations and the activities of their detoxification enzymes. The results showed that abamectin and the traditional pesticides pyridaben, profenofos and bifenthrin had higher resistance or lower toxicity than more recently developed pesticides including chlorfenapyr, spinetoram, cyflumetofen, cyenopyrafen, bifenazate and B-azolemiteacrylic. The frequency of point mutations related to abamectin resistance, G314D in the glutamate-gated chloride channel 1 (GluCl1) and G326E in GluCl3, ranged 47%-70% and 0%-97%, respectively. The frequency of point mutations in A1215D and F1538I of the voltage-gated sodium channel gene (VGSC), which may increase resistance to pyrethroids, ranged 88%-100% and 10%-100%, respectively. For target sites related to organophosphate resistance, mutation frequencies ranged 25%-92% for G119S and 0%-23% for A201S in the acetycholinesterase gene (Ace). Mutation G126S in the bifenazate resistance-related cytochrome b gene (Cytb) was observed in three of the seven T. urticae populations. Higher activities of detoxification enzymes (P450, GST, CarEs and UGTs) were observed in two T. urticae populations, with significant difference in the XY-SX population. These results provide useful information on the status of pesticide resistance of T. urticae in China and suggest that T. urticae field populations may have multiple resistance mechanisms.

A whole transcriptomic approach provides novel insights into the molecular basis of organophosphate and pyrethroid resistance in Anopheles arabiensis from Ethiopia

The development of insecticide resistance in malaria vectors is of increasing concern in Ethiopia because of its potential implications for vector control failure. To better elucidate the specificity of resistance mechanisms and to facilitate the design of control strategies that minimize the likelihood of selecting for cross-resistance, a whole transcriptomic approach was used to explore gene expression patterns in a multi-insecticide resistant population of Anopheles arabiensis from Oromia Region, Ethiopia. This field population was resistant to the diagnostic doses of malathion (average mortality of 71.9%) and permethrin (77.4%), with pools of survivors and unexposed individuals analyzed using Illumina RNA-sequencing, alongside insecticide susceptible reference strains. This population also demonstrated deltamethrin resistance but complete susceptibility to alpha-cypermethrin, bendiocarb and propoxur, providing a phenotypic basis for detecting insecticide-specific resistance mechanisms. Transcriptomic data revealed overexpression of genes including cytochrome P450s, glutathione-s-transferases and carboxylesterases (including CYP4C36, CYP6AA1, CYP6M2, CYP6M3, CYP6P4, CYP9K1, CYP9L1, GSTD3, GSTE2, GSTE3, GSTE4, GSTE5, GSTE7 and two carboxylesterases) that were shared between malathion and permethrin survivors. We also identified nineteen highly overexpressed cuticular-associated proteins (including CYP4G16, CYP4G17 and chitinase) and eighteen salivary gland proteins (including D7r4 short form salivary protein), which may be contributing to a non-specific resistance phenotype by either enhancing the cuticular barrier or promoting binding and sequestration of insecticides, respectively. These findings provide novel insights into the molecular basis of insecticide resistance in this lesser well-characterized major malaria vector species.

Preliminary characterisation of known pesticide resistance alleles in Spodoptera frugiperda (Lepidoptera: Noctuidae) in its invasive Australian range

AbstractFall armyworm (Spodoptera frugiperda) has recently been detected in Australia. Globally, S. frugiperda is reported to be resistant to insecticides, including those permitted for its control by the Australian Pesticides and Veterinary Medicines Authority. Consequently, an understanding of the insecticide resistance status of newly migrated S. frugiperda into Australia, as well as an ability to accurately identify newly hatched larvae, would facilitate sustainable management. To aid identification, we developed a real‐time polymerase chain reaction diagnostic assay that can identify S. frugiperda species, irrespective of life stage, and distinguish between the rice and corn strains. We then screened S. frugiperda individuals from Queensland, Western Australia, the Northern Territory and New South Wales for mutations causing target site insensitivity in the acetylcholinesterase (AChE) gene, the voltage‐gated sodium channel gene and the ryanodine receptor gene. In the populations tested, we found that mutation at positions A210S and F290V in AChE were common. These mutations are known to be associated with carbamate and organophosphate resistance in S. frugiperda outside Australia. In contrast, no mutation was found in gene voltage‐gated sodium channel causing pyrethroid resistance or the ryanodine receptor gene associated with diamide resistance. As S. frugiperda is new to Australia, the wide distribution of mutations in the AChE gene associated with organophosphate/carbamate resistance suggests that the original migrated S. frugiperda moths may have carried the mutations from outside Australia. As resistance genes have been detected, it is important to continue to monitor the resistance status of Australian S. frugiperda and further integrate complementary bioassay when available. Finally, as pesticide resistance is possible, it should be additionally considered when making pest management decisions and any spray failure should be followed by a pesticide from a different chemical group.

Insecticide resistance of Dengue vectors in South East Asia: a systematic review.

The insecticides used widely has led to resistance in the vector and impose a challenge to vector control operation. This review aims to analyse the distribution of insecticide resistance of dengue vectors in South East Asia and to describe the mechanism of insecticide resistance. Literature search for articles published on 2015 to 2019 from PubMed, Scopus and ProQuest was performed. Total of 37 studies included in the final review from the initial 420 studies. Pyrethroid resistance was concentrated on the west coast of Peninsular Malaysia and Northern Thailand and scattered at Java Island, Indonesia while organophosphate resistance was seen across the Java Island (Indonesia), West Sumatera and North Peninsular Malaysia. Organochlorine resistance was seen in Sabah, Malaysia and scattered distribution in Nusa Tenggara, Indonesia. V1016G, S989P, F1269C gene mutation in Aedes Aegypti were associated with Pyrethroid resistance in Singapore and Indonesia. In Malaysia, over-expressed with monooxygenase P450 genes (CYP9J27, CYP6CB1, CYP9J26 and CYP9M4) Glutathione S-transferases, carboxylesterases commonly associated with pyrethroids resistance in Aedes Aegypti and CYP612 overexpressed in Aedes Albopictus. The genetic mutation in A302S in Aedes Albopictus was associated with organochlorine resistance in Malaysia. Rotation of insecticide, integration with synergist and routine assessment of resistance profile are recommended strategies in insecticide resistance management.

Common substitution mutation F348Y of acetylcholinesterase gene contributes to organophosphate and carbamate resistance in Cimex lectularius and C. hemipterus

Bed bug control highly depends on insecticides with a limited number of modes of action, especially since the global prevalence of pyrethroid resistance. De facto insecticide options against bed bugs in Japan are acetylcholinesterase inhibitors (AChEis) that consist of organophosphates and carbamates. However, the status of AChEi resistance and the mechanisms involved have not been ascertained. An amino acid substitution mutation, F348Y (or F331Y in standard numbering), occurring at an acyl-binding site of the paralogous AChE gene (p-Ace), was identified among AChEi-resistant colonies of both common and tropical bed bugs (Cimex lectularius and C. hemipterus, respectively). This mutation was genetically associated with propoxur and fenitrothion resistance in F348Y-segregating colonies of C. hemipterus. Inhibition of heterologously expressed C. lectularius p-Ace with insecticides revealed that the sensitivities of F348Y-carrying AChE decreased by orders of 10- to more than 100-fold for diazoxon, carbaryl, fenitroxon, paraoxon, chlorpyrifos-methyl, malaoxon, azamethiphos, methyl-paraoxon, and propoxur. In contrast, the mutant AChE showed a slightly decreased degree of sensitivity for dichlorvos and almost unchanged sensitivity for metoxadiazone. Further studies are needed to ascertain whether the practical efficacies of dichlorvos and metoxadiazone are ensured against F348Y-carrying bed bugs and whether other resistance mechanisms are involved.

Dynamics of organophosphate and carbamate resistance in Anopheles gambiae s. l. populations from south and north Benin, West Africa

The current study was aimed to investigate on dynamics of propoxur resistance in Anopheles gambiae s.l. populations from N’ dali district in northern Benin (West Africa) and also to investigate on dynamics of malathion resistance in Anopheles gambiae s. l. populations from Toffo district in southern Benin. Larvae and pupae of Anopheles gambiae s. l . mosquitoes were collected from the breeding sites in Borgou and Atlantic departments in 2015 and 2019. WHO susceptibility tests were conducted on unfed female mosquitoes aged 2-5 days old. WHO bioassays were performed with impregnated papers with propoxur 0.1% and with malathion 5%. PCR techniques were used to detect species and Ace-1 mutations in 2015. Anopheles gambiae s. l. populations from N’dali were resistant to propoxur in 2015 and were still remained resistant to this product in 2019. Regarding Anopheles gambiae s. l. populations from Toffo, they were susceptible to malathion in 2015 whereas the malathion resistance status of these mosquitoes requires further investigation in 2019. PCR revealed that all specimens tested were Anopheles gambiae s. s. The presence of Ace-1R at very low frequency (0.01) was observed in Anopheles gambiae s. l. populations from both districts. This study shows that propoxur resistance detected in An. gambiae s. l. populations from N’ dali needs to be monitored for insecticide resistance in this area.

Dynamics of organophosphate and carbamate resistance in Anopheles gambiae s. l. populations from south and north Benin, West Africa

Dynamics of organophosphate and carbamate resistance in Anopheles gambiae s. l. populations from south and north Benin, West Africa

Cover Caption

The fall armyworm, Spodoptera frugiperda, is a major pest native to the Americas that has recently invaded the Old World. A whole‐genome sequencing (WGS) approach was used to identify mutations associated with resistance to insecticides and Bt proteins in 150 individuals of S. frugiperda collected from China, Malawi, Uganda and Brazil. This approach revealed three amino acid substitutions (A201S, G227A and F290V) of the acetylcholinesterase‐1 (ace‐1) in the four geographical populations of S. frugiperda, which are known to be associated with organophosphate resistance. A novel Cry1F resistance allele in the ABCC2 gene was identified from the Brazilian population but absent in the invasive populations (see pages 627‐638). Photo provided by Prof. Run‐Zhi Zhang.

Functional Analysis of SlGSTE12 in Pyrethroid and Organophosphate Resistance in Spodoptera litura.

Glutathione S-transferase genes in the epsilon group were reported to function in insecticide resistance. SlGSTE12 was validated to be overexpressed in pyrethroid- and organophosphate-resistant populations of Spodoptera litura compared to a susceptible population. A functional study of heterologously expressed SlGSTE12 showed that Km and Vmax for 1-chloro-2,4-dinitrobenzene (CDNB) conjugating activity were 0.70 ± 0.18 mmol L-1 and 90.6 ± 9.4 nmol mg-1 min-1, respectively. β-Cypermethrin and cyhalothrin showed much weaker inhibition of SlGSTE12 activity to CDNB conjugation than fenvalerate, chlorpyrifos, and phoxim. Ultrahigh-performance liquid chromatography analysis showed that SlGSTE12 had significant metabolism activity to fenvalerate and phoxim both in vitro and in Escherichia coli, especially to chlorpyrifos, and slight metabolism activity toward cyhalothrin only in vitro. Silencing of SlGSTE12 by RNAi increased the mortality to fenvalerate, cyhalothrin, and chlorpyrifos significantly. SlGSTE12 also had a significant antioxidant ability against cumene hydroperoxide. Our study suggested that SlGSTE12 could metabolize phoxim, fenvalerate, cyhalothrin, and especially chlorpyrifos. SlGSTE12 might also participate in pyrethroid and organophosphate resistance by antioxidant activity.