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There has been a renewed interest for larviciding during the recent decade. Although biological larvicides are considered not to be harmful to non-target organisms, there is still not sufficient data on the effect of new long-lasting larvicide formulations such as VectoMax G combining Bacillus thuringiensis israelensis and Lysinibacullus sphaericus on the environment especially on non-target organisms. The present study aimed to assess the possible influence of VectoMax G on the diversity and abundance of the aquatic fauna cohabiting with mosquito larvae in breeding habitats during a larviciding trial in the city of Yaoundé.Twelve districts of the city of Yaoundé divided into 6 intervention and 6 control sites were chosen for the study. In each district 4 semi-permanent or permanent aquatic habitats were followed. VectoMax G application was done once every two weeks during 6 months and aquatic organisms were collected 48 h after each treatment. All collected organisms were brought to the laboratory for identification. Physico-chemical parameters were recorded as well.A high diversity of the zooplankton was recorded in the intervention areas with 28 species collected against 14 species in the control areas. Cladocerans were the most represented group in both sites while Ostracods were found only in control sites. A total of 19 macro-invertebrates species were recorded in the control areas vs 16 species in the intervention areas. Gasteropods were the most represented groups of macro-invertebrates. Vertebrates such as larvivorous fishes and amphibians larvae were also found in approximately similar densities in both sites.The study indicated no significant influence of larviciding with VectoMax G on the diversity and abundance of the non-target aquatic fauna in the city of Yaoundé.

The versatility of cytochrome P450 reductase (CPR) in transferring electrons to P450s from other closely related species has been extensively exploited, e.g., by using An. gambiae CPR (AgCPR), as a homologous surrogate, to validate the role of An. funestus P450s in insecticide resistance. However, genomic variation between the AgCPR and An. funestus CPR (AfCPR) suggests that the full metabolism spectrum of An. funestus P450s might be missed when using AgCPR. To test this hypothesis, we expressed AgCPR and AfCPR side-by-side with CYP6P9a and CYP6P9b and functionally validated their role in the detoxification of insecticides from five different classes. Major variations were observed within the FAD- and NADP-binding domains of AgCPR and AfCPR, e.g., the coordinates of the second FAD stacking residue AfCPR-Y456 differ from that of AgCPR-His456. While no significant differences were observed in the cytochrome c reductase activities, when co-expressed with their endogenous AfCPR, the P450s significantly metabolized higher amounts of permethrin and deltamethrin, with CYP6P9b-AfCPR membrane metabolizing α-cypermethrin as well. Only the CYP6P9a-AfCPR membrane significantly metabolized DDT (producing dicofol), bendiocarb, clothianidin, and chlorfenapyr (bioactivation into tralopyril). This demonstrates the broad substrate specificity of An. funestus CYP6P9a/-b, capturing their role in conferring cross-resistance towards unrelated insecticide classes, which can complicate resistance management. [ABSTRACT FROM AUTHOR]

Deciphering the evolutionary forces controlling insecticide resistance in malaria vectors remains a prerequisite to designing molecular tools to detect and assess resistance impact on control tools. Here, we demonstrate that a 4.3kb transposon-containing structural variation is associated with pyrethroid resistance in central/eastern African populations of the malaria vector Anopheles funestus. In this study, we analysed Pooled template sequencing data and direct sequencing to identify an insertion of 4.3kb containing a putative retro-transposon in the intergenic region of two P450s CYP6P5-CYP6P9b in mosquitoes of the malaria vector Anopheles funestus from Uganda. We then designed a PCR assay to track its spread temporally and regionally and decipher its role in insecticide resistance. The insertion originates in or near Uganda in East Africa, where it is fixed and has spread to high frequencies in the Central African nation of Cameroon but is still at low frequency in West Africa and absent in Southern Africa. A marked and rapid selection was observed with the 4.3kb-SV frequency increasing from 3% in 2014 to 98% in 2021 in Cameroon. A strong association was established between this SV and pyrethroid resistance in field populations and is reducing pyrethroid-only nets' efficacy. Genetic crosses and qRT-PCR revealed that this SV enhances the expression of CYP6P9a/b but not CYP6P5. Within this structural variant (SV), we identified putative binding sites for transcription factors associated with the regulation of detoxification genes. An inverse correlation was observed between the 4.3kb SV and malaria parasite infection, indicating that mosquitoes lacking the 4.3kb SV were more frequently infected compared to those possessing it. Our findings highlight the underexplored role and rapid spread of SVs in the evolution of insecticide resistance and provide additional tools for molecular surveillance of insecticide resistance. Author summary: Anopheles funestus is an important malaria transmitting mosquito whose control relies on pyrethroid insecticides. The ability to control malaria transmitting mosquitoes is threatened by the emergence of insecticide resistance. Developing effective strategies to prevent, slow, or overcome resistance require an understanding of the underlying genetic mechanisms. In this study, we identified a transposon containing insertion in Anopheles funestus mosquitoes in Central/East Africa but absent in other regions. Tracking its distribution and frequency over the years revealed a strong selection with frequencies reaching fixation in less than 3 years. Phenotype/genotype association studies revealed a strong correlation between pyrethroid resistance and genetic variant genotype but negative correlation with plasmodium parasite infection in mosquitoes. These findings shed light on the often-overlooked role of structural variations in the evolution of insecticide resistance, offering valuable insights and tools for monitoring and addressing this critical issue in malaria control. [ABSTRACT FROM AUTHOR]

Background: The first dengue outbreak in Sao Tome and Principe was reported in 2022. Entomological investigations were undertaken to establish the typology of Aedes larval habitats, the distribution of Ae. aegypti and Ae. albopictus, the related entomological risk and the susceptibility profile of Ae. aegypti to insecticides, to provide evidence to inform the outbreak response. Methodology/Principal findings: Entomological surveys were performed in all seven health districts of Sao Tome and Principe during the dry and rainy seasons in 2022. WHO tube and synergist assays using piperonyl butoxide (PBO) and diethyl maleate (DEM) were carried out, together with genotyping of F1534C/V1016I/V410L mutations in Ae. aegypti. Aedes aegypti and Ae. albopictus were found in all seven health districts of the country with high abundance of Ae. aegypti in the most urbanised district, Agua Grande. Both Aedes species bred mainly in used tyres, discarded tanks and water storage containers. In both survey periods, the Breteau (BI > 50), house (HI > 35%) and container (CI > 20%) indices were higher than the thresholds established by WHO to indicate high potential risk of dengue transmission. The Ae. aegypti sampled were susceptible to all insecticides tested except dichlorodiphenyltrichloroethane (DDT) (9.2% mortality, resistant), bendiocarb (61.4% mortality, resistant) and alpha-cypermethrin (97% mortality, probable resistant). A full recovery was observed in Ae. aegypti resistant to bendiocarb after pre-exposure to synergist PBO. Only one Ae. aegypti specimen was found carrying F1534C mutation. Conclusions/Significance: These findings revealed a high potential risk for dengue transmission throughout the year, with the bulk of larval breeding occurring in used tyres, water storage and discarded containers. Most of the insecticides tested remain effective to control Aedes vectors in Sao Tome, except DDT and bendiocarb. These data underline the importance of raising community awareness and implementing routine dengue vector control strategies to prevent further outbreaks in Sao Tome and Principe, and elsewhere in the subregion. Author summary: During the first dengue outbreak in Sao Tome and Principe reported in 2022, entomological investigations were undertaken to establish the typology of Aedes larval habitats, the distribution of Ae. aegypti and Ae. albopictus, the related entomological risk and the susceptibility profile of Ae. aegypti to insecticides to inform the outbreak response. Surveys revealed the presence of Ae. aegypti and Ae. albopictus in all seven health districts of the country with high abundance of Ae. aegypti in the most urbanised district, Agua Grande. Both Aedes species bred mainly in used tyres, discarded tanks and water storage containers suggesting good waste management and improved water supply system could help to reduce Aedes densities and the risk of dengue transmission. Analyses also revealed that most of the insecticides tested remain effective to control Aedes vectors in Sao Tome, except dichlorodiphenyltrichloroethane and bendiocarb. These findings revealed a high potential risk for dengue transmission throughout the year and underline the importance of raising community awareness and implementing routine dengue vector control strategies to prevent further outbreaks in Sao Tome and Principe, and elsewhere in the subregion. [ABSTRACT FROM AUTHOR]