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Insecticide-based interventions such as IRS and LLINs have significantly reduced malaria transmission globally. However, their sustainability is increasingly threatened by insecticide resistance. While insecticide and pesticide use are known resistance drivers, the role of environmental contaminants remains underexplored. This study investigated the impact of environmental contaminants on insecticide resistance by sampling Anopheles larvae from six sites in Ghana, including petroleum spill sites, mining areas and industrial zones. WHO bioassays revealed resistance to clothianidin (Mortality rate (MR) = 54% – 80%) and chlorfenapyr (MR = 80% − 84%) in all site categories. Interestingly, high-intensity resistance to pirimiphos methyl (MR = 10x = 1.7%) was detected in vectors from Obuasi. High-intensity pyrethroid resistance [deltamethrin (MR = 10x = 79–92%); permethrin (MR = 10x = 74–95%)] was observed across all sites, with varying frequencies of kdr mutations ( L995F , V402L , I1527T , P1874L ; 0.12–0.98) were observed across all sites. High allele frequency of Ace-1 (0.62) was observed in Obuasi. Chemical analyses of water from breeding habitats revealed significant associations between heavy metals and insecticide resistance ( P  = 0.04). Anopheles coluzzii (81.7%) was the dominant species across all sites. These findings provide evidence that environmental contaminants may contribute to insecticide resistance. There is an urgent need for enhanced surveillance and resistance management strategies for effective malaria vector control.

Aedes aegypti , the primary vector of dengue, has developed widespread resistance to insecticides, posing a threat to the efficacy of vector control programs. This study assessed the susceptibility status of Ae. aegypti across Malaysian dengue hotspots and characterized knockdown resistance ( kdr ) mutations, including the first detection of T1520I. Adult female mosquitoes were collected from seven localities across five Malaysian states. Susceptibility to deltamethrin (0.03%), permethrin (0.4%), pirimiphos-methyl (60 mg/m 2 ), and malathion (5%) was evaluated using bioassay protocols. Knockdown times (KT 50 , KT 95 ) and resistance ratios (RR) were estimated using probit analysis, and the voltage-gated sodium channel gene was sequenced to detect kdr mutations. All field populations remained susceptible to malathion, with mortality ≥ 95%, though one Johor strain (ABS) showed suspected resistance (95%). Pirimiphos-methyl resistance was widespread, with mortality as low as 6% (ABS). Whereas resistance to pyrethroids was pronounced, with deltamethrin mortality ranging from 22% (ABS) to 95% (AA), and permethrin from 0% (TMB, ABS) to 95% (AA). Knockdown assays revealed high resistance ratios, particularly for permethrin (TMB: RR 50 =9.98, RR 95 =14.98; ABS: RR 95 =41.46). Sequencing identified multiple kdr mutations, including S989P, V1016G, F1534C, A1007G, and the novel detection of T1520I in Malaysian populations. F1534C was strongly associated with permethrin resistance, while V1016G and S989P predominated under deltamethrin exposure. Aedes aegypti populations in Malaysia exhibit widespread pyrethroid resistance and emerging organophosphate resistance, underpinned by multiple kdr mutations. The first detection of T1520I underscores the evolving genetic basis of resistance. These findings highlight the urgent need for integrated resistance management, including molecular surveillance and insecticide rotation, to sustain effective dengue control.

Resistance to pyrethroids is common in Aedes aegypti populations. Mutations in the voltage gated sodium channel have an influence on the insecticide resistance (IR) phenotype. In much of the Western hemisphere, two of these knockdown resistance ( kdr) mutations, V1016I and F1534C, result in six kdr genotype combinations in field populations. Strong pyrethroid IR and the failure of permethrin treated uniforms have been linked to the presence of the homozygous double kdr genotype (IICC) but the effects of the other five kdr combinations have not been rigorously examined, particularly with regard to operational efficacy. To better understand the impacts of these common kdr genotypes, we isolated three kdr haplotypes (VF, VC, & IC) from a field collected strain to produce six Ae. aegypti isoline strains with all the common V1016I/F1534C kdr genotypes. We then characterized the effects of these genotypes by CDC bottle bioassay and topical application and found increasing resistance to permethrin and deltamethrin as the number of IC haplotypes increased. Neither enzymatic activity nor malathion resistance increased with increasing pyrethroid resistance. We then assessed the operational impacts of these kdr genotypes. Field and wind tunnel spray of a pyrethrin formulation showed that even moderate resistance could significantly reduce knockdown and mortality. Studies with a synergized pyrethroid formulation showed effective recovery of mortality against all genotypes except for the IICC. In human bite studies, one or two IC haplotypes compromised the efficacy of permethrin treated fabrics. This study demonstrates that kdr mutations have distinct phenotypic effects in both the laboratory and operational interventions, and that the strength of pyrethroid resistance varies with the number of IC haplotypes present. Assessing kdr genotype is therefore critical for understanding IR in Ae. aegypti .

The modern resurgence of the common (Cimex lectularius L.) and tropical bed bugs (C. hemipterus [F.]) is thought to be primarily due to insecticide resistance. While there are many reports on insecticide resistance mechanisms in C. lectularius, such information in C. hemipterus is limited. We examined dichloro-diphenyl-trichloroethane (DDT), malathion, deltamethrin, permethrin, lambda-cyhalothrin resistance, and the underlying mechanisms in several C. hemipterus strains (Australia: Queensland [QLD-AU]; Malaysia: Kuala Lumpur [KL-MY], Tanjung Tokong [TT-MY], Christian [CH-MY], and Green Lane [GL-MY]). We used a surface contact method, synergism studies (utilizing piperonyl butoxide [PBO], S,S,S-tributyl phosphorotrithioate [DEF], and diethyl maleate [DEM]), and molecular detection of kdr mutations. Results demonstrated that all C. hemipterus strains possessed high resistance to DDT and the pyrethroids and moderate to high resistance to malathion. Synergism studies showed that deltamethrin resistance in all strains was significantly (P < 0.05) inhibited by PBO. In contrast, deltamethrin resistance was not affected in DEF or DEM. Similar findings were found with lambda-cyhalothrin resistance. Malathion resistance was significantly (P < 0.05) reduced by DEF in all strains. Resistance to DDT was not affected by DEM in all strains. Multiple kdr mutations (M918I, D953G, and L1014F) were detected by molecular analyses. TT-MY strain was found with individuals possessing three kdr mutation combinations; D953G + L1014F (homozygous susceptible: M918), M918I + D953G + L1014F (heterozygous resistant: I918), and M918I + D953G + L1014F (homozygous resistant: I918). Individuals with M918I + D953G + L1014F (homozygous resistant: I918) survived longer on deltamethrin (>12 h) than those (≤1 h) with other combinations. M918I + L1014F mutations most likely conferred super-kdr characteristic toward pyrethroids and DDT in C. hemipterus.

Introduction The Plasmodium transmission-blocking endosymbiont Microsporidia MB was previously identified in Anopheles gambiae s.l., but its association with the carriage of the genotypes of the L1014F kdr mutation, as well as the ecological factors driving its geographical distribution remain understudied. Methods Adult mosquitoes were field-collected using human landing catches (HLCs) across 60 villages in the Covè, Ouinhi, and Zangnanado communes of southern Benin. After morphological identification, a sub-sample of An. gambiae s.l. were molecularly speciated, and genotypied for the L1014F kdr mutation by Polymerase Chain Reaction (PCR). Enzyme-Linked Immunosorbent Assay (ELISA) and qPCR were also used to assess infection with Plasmodium falciparum sporozoites and Microsporidia MB, respectively. The environmental variables that drive the habitat suitability for Microsporidia MB were also assessed using Maximum Entropy (MaxEnt) modelling. Results The An. gambiae complex ( N  = 1040) was composed of 93.7% An. coluzzii , 4.4% An. gambiae s.s., 0.2% An. gambiae s.s./ coluzzii hybrids, while the rest failed to amplify. Infection prevalence with Microsporidia MB was 1.6% (95% CI: 0.7–3.3) in An. coluzzii and 2.2% (95% CI: 0.1–13.2) in An. gambiae s.s. The P. falciparum sporozoite rate was 2% (95% CI: 1.2–3.1, N  = 974) in An. coluzzii , and null in An. gambiae s.s. ( N  = 46). None of the mosquitoes infected with Microsporidia MB were infected with P. falciparum . The frequency of the L1014F kdr mutation was 75.1% (95% CI: 73.1–76.9) in An. coluzzii and 91.3% (95% CI: 83.1–95.9) in An. gambiae s.s. Microsporidia MB was absent in kdr -SS mosquitoes but was present in low proportions in both kdr -RS and kdr -RR mosquitoes (1.9%, 95% CI: 0.6–5.1). The mean load of Microsporidia MB DNA was higher in kdr -RR (23.23 ng/µl, 95% CI: 18.77–28.48) compared to kdr -RS (10.13 ng/µl, 95% CI: 7.36–13.91) mosquitoes. The elevation and soil contributed to explain, at 78% and 20% respectively, the habitat suitability for Microsporidia MB. Conclusion In this study, we demonstrated that An. gambiae s.l. mosquitoes bearing the L1014F kdr resistant allele was associated with a higher prevalence and load of Microsporidia MB than their susceptible counterparts. Moreover, the geographical distribution of Microsporidia MB was found to be influenced by certain environmental conditions, which warrant further large-scale investigations.

The role of detoxification enzymes in pyrethroid resistance intensity among malaria vectors remains a critical area of research. This study evaluated the role of detoxifying enzymes in driving resistance intensity in Anopheles mosquitoes from high insecticide resistance sites in Accra, Ghana. Larvae were collected from Tema, Abossey Okai, and Dansoman, and bioassays were performed on 3–5 days old adult females by exposing them to deltamethrin at discriminating concentrations (1× = 0.05%, 5× = 0.25%, and 10× = 0.5%) to assess resistance intensity. A piperonyl butoxide (PBO) synergist assay was used to test the involvement of cytochrome P450s , while qRT-PCR quantified expression of detoxification genes ( CYP6P1 , CYP9K1 , CYP6M2 , CYP6P3 , CYP4G16 , GSTE2 , and CYP6Z1 ). kdr mutations ( L995F , L995S ) were genotyped. High-intensity resistance was observed across all sites [deltamethrin 10× MR = 75–91%]. Pre-exposure to PBO significantly increased mortality (Tema: 13–56%; Abossey Okai: 20–91%; Dansoman: 34–88%, P  < 0.001), however, complete susceptibility was not restored. The L995F mutation was present at similar frequencies in resistant and susceptible mosquitoes. Transcriptomic profiling revealed concentration-dependent and site-specific expression: Tema; CYP9K1 , CYP6M2 , CYP6P1 , and CYP6P3 were significantly overexpressed (FC = 43.71–1222.98, P  < 0.05), while CYP4G16 expression increased with insecticide concentration. In Abossey Okai, CYP9K1 , CYP6P1 , CYP6M2 , and CYP6P3 were overexpressed (FC = 5.54–162.84). Mosquitoes from Dansoman showed generally low expression, however, CYP6M2 and CYP6P3 were overexpressed (FC = 120.80–292.68). These findings may suggest the dominant role of metabolic resistance, particularly P450 -mediated detoxification in driving high pyrethroid resistance intensity in Ghana.

Monitoring the ecology and insecticide resistance of Anopheles mosquitoes, which transmit malaria, is important for developing effective malaria control strategies. This study characterized the present larval breeding habitats of Anopheles mosquitoes in South Korea; additionally, we investigated the frequency of insecticide resistance in Anopheles in the Ganghwagun, Gyeonggi-do province, near the demilitarized zone (DMZ), of South Korea. While larvae of An. sineroides and An. lindesayi were found near forests in Naega-myeon and Kukhwa-ri, respectively, An. sinensis larvae were predominantly found in irrigation canal near rice fields in Seonhaeng-ri, alongside An. belenrae . Seven genotypes of knockdown resistant ( kdr ) allele in the voltage-gated sodium channel of An. sinensis were introduced, revealing a new kdr genotype. Notably, almost all of An. sinensis captured in this study was shown mutant genotypes with homozygous or heterozygous resistant alleles of acetylcholinesterase ( ace -1). Moreover, the predominant presence of mosquitoes harboring mutations in more than one insecticidal target was observed in An. sinensis . In addition, An. belenrae was identified to possess kdr and ace -1 mutation. Taken together, our findings revealed the multiple insecticide resistance of An. sinensis, with larval habitat near the DMZ of South Korea, 2024.

The Ae. albopictus mosquito has gained global attention due to its ability to transmit viruses, including the dengue and zika. Mosquito control is the only effective way to manage dengue fever, as no effective treatments or vaccines are available. Insecticides are highly effective in controlling mosquito densities, which reduces the chances of virus transmission. However, Ae. albopictus has developed resistance to pyrethroids in several provinces in China. Pyrethroids target the voltage-gated sodium channel gene ( VGSC ), and mutations in this gene may result in knockdown resistance ( kdr ). Correlation studies between resistance and mutations can assist viruses in managing Ae. albopictus , which has not been studied in Guizhou province. Nine field populations of Ae. albopictus at the larval stage were collected from Guizhou Province in 2022 and reared to F1 to F2 generations. Resistance bioassays were conducted against permethrin, beta-cypermethrin, and deltamethrin for both larvae and adults of Ae. albopictus . Kdr mutations were characterized by PCR and sequencing. Additionally, the correlation between the kdr allele and pyrethroid resistance was analyzed. All nine populations of Ae. albopictus larvae and adults were found to be resistant to three pyrethroid insecticides. One kdr mutant allele at codon 1016, one at 1532 and three at 1534 were identified with frequencies of 13.86% (V1016G), 0.53% (I1532T), 58.02% (F1534S), 11.69% (F1534C), 0.06% (F1534L) and 0.99% (F1534P), respectively. Both V1016G and F1534S mutation mosquitoes were found in all populations. The kdr mutation F1534S was positively correlated with three pyrethroid resistance phenotypes (OR > 1, P  < 0.05), V1016G with deltamethrin and beta-cypermethrin resistance (OR > 1, P  < 0.05) and F1534C only with beta-cypermethrin resistance (OR > 1, P  < 0.05). Current susceptibility status of wild populations of Ae. albopictus to insecticides and a higher frequency of kdr mutations from dengue-monitored areas in Guizhou Province are reported in this paper. Outcomes of this study can serve as data support for further research and development of effective insecticidal interventions against Ae. albopictus populations in Guizhou Province.

Background Aedes aegypti is the main vector of arboviral diseases worldwide. The species invaded and became established in southern Iran in 2020. Insecticide-based interventions are primarily used for its control. With insecticide resistance widespread, knowledge of resistance mechanisms is vital for informed deployment of insecticidal interventions, but information from Iranian Ae. aegypti is lacking. Methods Fifty-six Ae. aegypti specimens were collected from the port city of Bandar Lengeh in Hormozgan Province in the South of Iran in 2020 and screened for kdr mutations. The most common kdr mutations in Latin America and Asia (V410L, S989P, V1016G/I and F1534C), especially when present in combinations, are highly predictive of DDT and pyrethroid resistance were detected. Phylogenetic analyses based on the diversity of S989P and V1016G/I mutations were undertaken to assess the phylogeography of these kdr mutations. Results Genotyping all four kdr positions of V410L, S989P, V1016G/I and F1534C revealed that only 16 out of the 56 (28.57%) specimens were homozygous wild type for all kdr mutation sites. Six haplotypes including VSVF (0.537), VSVC (0.107), LSVF (0.016), LSIF (0.071), VPGC (0.257) and LPGC (0.011) were detected in this study. For the first time, 11 specimens harbouring the V410L mutation, and 8 samples with V1016I mutation were found. V410L and V1016I were coincided in 8 specimens. Also, six specimens contained 1016G/I double mutation which was not reported before. Conclusions The relatively high frequency of these kdr mutations in Iranian Ae. aegypti indicates a population exhibiting substantial resistance to pyrethroid insecticides, which are used widely in control operations and household formulations. The detection of the 410L/1016I kdr mutant haplotype in Iranian Ae. aegypti suggests possible convergence of invasive populations from West Africa or Latin America. However, as Iran has very limited maritime/air connections with those African countries, a Latin American origin for the invasive Ae. aegypti in Iran is more plausible. Graphical abstract