Emmanuel Elanga-Ndille

Insecticide resistance management is imperative to sustain the effectiveness of vector control tools. While metabolic detoxification and target-site mutations are well described mechanistically, their combined fitness effects across mosquito life stages and implications for transmission and resistance management are less clear. We investigated biological costs of P450-based resistance (E205D-CYP6P3) and the L995F-kdr target-site mutation, alone and together, in a highly pyrethroid-resistant Anopheles gambiae population from Mangoum, Cameroon. Using field-derived and Kisumu–Mangoum hybrid colonies, we measured fecundity, fertility, larval development, pupation timing, adult longevity, Plasmodium falciparum infection, and multi-generational resistance dynamics under relaxed insecticide pressure. E205D-CYP6P3 carriers showed reduced oviposition success, delayed pupation and increased early adult mortality, indicating clear fitness costs of metabolic resistance; larval development and egg viability were largely unaffected. L995F-kdr produced strong early-life penalties with impaired larval progression and elevated adult mortality, but paradoxically conferred advantages during pupation, revealing stage-specific trade-offs. Concurrent carriage of both mutations generally amplified fecundity and early-survival costs, though some compensatory benefits appeared at pupation. Over successive generations without insecticide, L995F-kdr frequencies declined rapidly and were nearly lost by F12, whereas E205D-CYP6P3 decreased more slowly and persisted at intermediate frequencies, suggesting greater persistence of metabolic resistance. P. falciparum assays indicated higher whole-mosquito infection prevalence in E205D heterozygotes but no effect at the sporozoite stage, implying limited impact on transmission potential. These results show mechanism- and stage-dependent fitness costs in An. gambiae, with target-site resistance more reversible than P450-mediated resistance, and support mechanism-tailored, evolution-informed management including rotations, mixtures and next-generation interventions.

Background: Continued expansion of arboviral diseases poses a serious threat to public health systems. The use of biomarkers for mosquito exposure, such as antibodies against the Aedes Nterm-34 kDa peptide, serves as a proxy for assessing the risk of Aedes-borne diseases. This study aimed to evaluate whether IgG antibodies against the Nterm-34 kDa peptide are associated with the level of human exposure to Aedes mosquito bites and the risk of flavivirus and alphavirus infections. Methods: Three longitudinal surveys were conducted during the rainy, dry, and short rainy seasons in three villages in Bondo, Tanga. The study included individuals aged between 2 to 70 years. A questionnaire was administered to participants to collect information on socio-demographic factors, housing conditions, surrounding environmental features, and mosquito bite prevention measures. In parallel, intravenous blood was collected from each participant for virus detection using RT-qPCR and for quantification of IgG antibodies against the Aedes Nterm-34kDa peptide using an ELISA assay Results: A total of 362 participants were enrolled, with a mean age of 28.3 ± 23.1 years. Alphavirus positivity peaked in the rainy season 19 (5.2%), with lower prevalence during dry 4 (1.1%) and short rainy 9 (2.5%) seasons, whereas flavivirus positivity was lower overall 11 (3.0%) rainy, 3 (0.8%) dry, 5 (1.4%) short rainy). IgG antibody levels against the Aedes 34 kDa peptide were significantly higher in PCR-positive. The level of IgG antibody response to the Nterm-34kDa peptide exhibited seasonal variation, with significantly higher values observed during the rainy season compared to the dry season (Mann-Whitney U test, p < 0.0001). IgG seropositivity was significantly associated with proximity to vegetation ( p < 0.001), presence of garbage pits ( p = 0.005), and recent travel history ( p = 0.018), whereas demographic and housing factors showed no significant associations. Participants positive for dengue demonstrated higher levels of anti-salivary IgG compared with dengue-negative participants ( p = 0.02; non-parametric Mann–Whitney test). Conclusions: This study reveals distinct seasonal dynamics in human IgG antibody responses to the Aedes Nterm-34kDa salivary peptide, reflecting the seasonal transmission patterns of Alphavirus and Flavivirus in northern Tanzania. Elevated seropositivity during the rainy season aligns with increased vector exposure driven by environmental factors near human dwellings. The findings underscore the potential of the Nterm-34kDa peptide as a biomarker for arboviral transmission monitoring and highlight the importance of targeted vector control strategies considering seasonal and microenvironmental variations to mitigate Aedes-borne viral infections in the region.

The role of sand flies in Leishmania transmission in Cameroon remains understudied, limiting the implementation of a national control program. To address t

Monitoring the trend of insecticide resistance and understanding associated genetic mechanisms is important for designing efficient malaria vector control strategies. This study was conducted to provide temporal data on insecticide resistance status and mechanisms in the major malaria vector Anopheles gambiae s.l. from Ebolowa, Southern Cameroon. Methods: Larvae of An. gambiae s.l. were collected from typical breeding sites throughout the city and reared to adulthood. Emerging adults were morphologically identified and WHO tube assays were performed to determine their susceptibility to carbamate, organophosphate and pyrethroid insecticides at diagnostic doses. When resistance was observed, its intensity was determined by performing WHO tube tests using 5 and 10 times the concentration of the diagnostic dose. Metabolic resistance mechanisms were investigated using insecticide-synergist assays. Sibling species of the An. gambiae complex were identified using SINE-PCR protocol. TaqMan assay was used to genotype the L1014F and L1014S kdr mutations, and the N1575Y mutation, an amplifier of the resistance conferred by the L1014F mutation. Results: Anopheles coluzzii was by far the dominant (99%) member of the An. gambiae s.l. complex in Ebolowa. The species was fully susceptible to carbamates and organophosphates, but resistant to all pyrethroid insecticides tested. Resistance was of moderate intensity for deltamethrin (mortality: 37%, 70% and 99% for 1×, 5× and 10× insecticide concentration, respectively) but rather of high intensity for permethrin (5% for 1×; 62% for 5× and 75% for 10×) and for alphacypermethrin (4.4% for 1×; 57% for 5× and 80% for 10×). Pre-exposure to the synergist PBO resulted in a full recovery of the susceptibility to delthametrin, but this was not observed for the other two pyrethroids tested. L1014S (kdr-East) and the N1575Y mutations were absent, whereas the L1014F (kdr-West) mutation was present at a high frequency (75%), showing a significant association with resistance to permethrin (OR = 3.8; 95%; CI [1.9–7.4]; p < 0.0001) and alphacypermethrin (OR = 3; 95%; CI [1.6–5.4]; p = 0.0002). Conclusion: The increased resistance of An. gambiae s.l. to pyrethroid insecticides as observed in Ebolowa poses a threat to the efficacy of LLINs used to protect populations from the bites of Anopheles mosquitoes that transmit malaria parasites. The present study further highlights the urgent need to implement resistance management strategies in order to maintain the effectiveness of insecticide-based vector control interventions and prevent a rebound in malaria-related mortality.

Human IgG antibody response to Anopheles gambiae gSG6-P1 salivary peptide was reported to be a pertinent indicator for assessing human exposure to mosquito bites and evaluating the risk of malaria transmission as well as the effectiveness of vector control strategies. However, the applicability of this marker to measure malaria transmission risk where human populations are mostly bitten by secondary vectors in Africa has not yet been evaluated. In this study, we aimed to investigate whether anti-gSG6-P1 antibodies response could be induced in humans living in forest areas in Cameroon where An. gambiae s.l is not predominant. In October 2019 at the pick of the rainy season, blood samples were collected from people living in the Nyabessang in the forest area in the South region of Cameroon. Malaria infection was determined using thick blood smear microscopy and Rapid Diagnostic Test. The level of IgG Anti-gSG6-P1 response as a biomarker of human exposure to Anopheles bite, was assessed using enzyme-linked immunosorbent assay. Mosquitoes were collected using the human landing catches to assess Anopheles density and for the identification of Anopheles species present in that area. IgG antibody response to the gSG6-P1 salivary peptide was detected in inhabitants of Nyabessang with high inter-individual heterogeneity. No significant variation in the level of this immune response was observed according to age and gender. The concentration of gSG6-P1 antibodies was significantly correlated with the malaria infection status and, Plasmodium falciparum-infected individuals presented a significantly higher level of IgG response than uninfected individuals (p = 0.0087). No significant difference was observed according to the use of insecticide treated nets. Out of the 1,442 Anopheles mosquitoes species collected, 849 (58.9%) were identified as An. paludis, 489 (33.91%) as An. moucheti, 28 (4.44%) as An. nili, 22 (2.08%) as An. gambiae s.l and 10 (0.69%) as An. marshallii. Our findings show that IgG response to An. gambiae gSG6-P1 peptide could be detected in humans exposed predominantly to An. moucheti and An. paludis bites. Taken together, the data revealed the potential of the Anti-gSG6-P1 IgG antibody response to serve as a universal marker to assess human exposure to any Anopheles species.

Background: Cameroon is considering the implementation of indoor residual spraying (IRS) as a complementary measure to control malaria in the context of high pyrethroid resistance in major malaria vectors. Non-pyrethroid insecticide classes such as organophosphates and carbamates may be utilized in IRS due to widespread pyrethroid resistance. However, the success of this strategy depends on good knowledge of the resistance status of malaria vectors to carbamates and organophosphates. Here, we assessed the susceptibility profile of Anopheles gambiae sensu lato with respect to carbamates and organophosphate and the distribution of the molecular mechanism underlying resistance to these insecticides. Methods: Anopheles gambiae s.l. mosquitoes were collected from nine settings across the country and bio-assayed with bendiocarb, propoxur and pirimiphos-methyl. The Ace-1 target-site G119S mutation was genotyped using a TaqMan assay. To investigate the polymorphism in the Ace-1 gene, a region of 924 base pairs in a sequence of the gene was amplified from both live and dead females of An. gambiae exposed to bendiocarb. Results: Pirimiphos-methyl induced full mortality in An. gambiae s.l. from all study sites, whereas for carbamates, resistance was observed in four localities, with the lowest mortality rate recorded in Mangoum (17.78 ± 5.02% for bendiocarb and 18.61 ± 3.86% for propoxur) in the southern part of Cameroon. Anopheles coluzzii was found to be the predominant species in the northern tropical part of the country where it is sympatric with Anopheles arabiensis. In the localities situated in southern equatorial regions, this species was predominant in urban settings, while An. gambiae was the most abundant species in rural areas. The G119S Ace-1 target-site mutation was detected only in An. gambiae and only in the sites located in southern Cameroon. Phylogenetic analyses showed a clustering according to the phenotype. Conclusion: The occurrence of the Ace-1 target-site substitution G119S in An. gambiae s.l. populations highlights the challenge associated with the impending deployment of IRS in Cameroon using carbamates or organophosphates. It is therefore important to think about a resistance management plan including the use of other insecticide classes such as neonicotinoids or pyrrole to guarantee the implementation of IRS in Cameroon. Graphical

The specific immune response to the Anopheles salivary peptide could be a pertinent and complementary tool to assess the risk of malaria transmission and the effectiveness of vector control strategies. This study aimed to obtain first reliable data on the current state of the Anopheles gSG6-P1 biomarker for assess the level of exposure to Anopheles bites in high malaria endemic areas in Cameroon. Blood smears were collected from people living in the neighborhoods of Youpwe (suburban area, continental) and Manoka (rural area, Island), both areas in the coastal region of Cameroon. Malaria infection was determined using thick blood smear microscopy, whereas the level of specific IgG response to gSG-P1 peptide was assessed by enzyme-linked immunosorbent assay from the dried blood spots. Of 266 (153 from Youpwe, 113 from Manoka) malaria endemic residents (mean age: 22.8±19.8 years, age range: 6 months–94 years, male/female sex ratio: 1/1.2, with Manoka mean age: 23.71±20.53, male/female sex ratio:1/1.13 and Youpwe mean age: 22.12±19.22, male/female sex ratio 1/0.67) randomly included in the study, Plasmodium infection prevalence was significantly higher in Manoka than in Youpwe (64.6% vs 12,4%, p = 0.0001). The anti-gSG6-P1 IgG response showed a high inter-individual heterogeneity and was significantly higher among individuals from Manoka than those from Youpwe (p = 0.023). Malaria infected individuals presented a higher anti-gSG6-P1 IgG antibody response than non-infected (p = 0.0004). No significant difference in the level of specific IgG response to gSG-P1 was observed according to long lasting insecticidal nets use. Taken together, the data revealed that human IgG antibody response to Anopheles gSG-P1 salivary peptide could be also used to assess human exposure to malaria vectors in Central African region. This finding strengthens the relevance of this candidate biomarker to be used for measuring human exposure to malaria vectors worldwide.

Insecticide resistance genes are often associated with pleiotropic effects on various mosquito life-history traits. However, very little information is available on the impact of insecticide resistance on blood feeding process in mosquitoes. Here, using two recently detected DNA-based metabolic markers in the major malaria vector, An. funestus, we investigated how metabolic resistance genes could affect the blood meal intake. After allowing both the field F1 and lab F8 Anopheles funestus strains to feed on the human arm for 30 minutes, we assessed the association between key parameters of blood meal process including, probing time, feeding duration, blood feeding success, blood meal size, and markers of glutathione S-transferase (L119F-GSTe2) and cytochrome P450 (CYP6P9a_R)—mediated metabolic resistance. None of the parameters of blood meal process was associated with L119F-GSTe2 genotypes. By contrast, for CYP6P9a_R, homozygous resistant mosquitoes were significantly more able to blood-feed than homozygous susceptible (OR = 3.3; CI 95%: 1.4–7.7; P = 0.01) mosquitoes. Moreover, the volume of blood meal ingested by CYP6P9a-SS mosquitoes was lower than that of CYP6P9a-RS (PCYP6P9a gene is inked with the feeding success and blood meal size of An. funestus. However, no correlation was found in the expression of CYP6P9a and that of genes encoding for salivary proteins involved in blood meal process. This study suggests that P450-based metabolic resistance may influence the blood feeding process of Anopheles funestus mosquito and consequently its ability to transmit malaria parasites.

Background:Cameroonian authorities have recently established irrigated rice projects across the country. The assessment of the impact of such projects in the transmission of vector-borne diseases, such as malaria, imposes to compare the situation before and after their implementation in a given locality. In Bankeng, a village in a forest area in Cameroon, no preliminary data on malaria transmission indicators was collected before the implantation of irrigated rice farming. To attempt to catch up for this shortcoming 6 months after the first rice seedlings were sown., the present study aimed to generate data which could serve as a baseline to assess the impact of irrigated rice farming in Bankeng. Methods: A cross-sectional study was performed at the end of the rainy season in July 2018. Mosquitoes were sampled by night collections on human volunteers, identified morphologically and members of different complexes further sorted to species using molecular tools.Plasmodiuminfectious status was determined by Taqman genotyping. To assess resistance profile to insecticides, WHO bioassays were performed using two-to-five days old femalesAn. gambiaes.l. reared from larval collections in Bankeng. Furthermore, the molecular basis of resistance were investigated Results:An. gambiaes.l represented 98% of the 1087 mosquitoes collected withAnopheles gambiaeas the predominant species. The total human biting rate was 44.5 bites/person/night. Entomological inoculation rate was 3.8 ib/p/n. The BankengAn. gambiaepopulation exhibited a high level of resistance to almost all insecticides except to organophosphates with a high frequency of L1014F kdr mutation (93.9%) and a 6-fold over-expression ofCYP6P3P450 gene. Conclusion: In the absence of preliminary data before the implementation of the irrigated rice fields; the present study provides interesting data which could help for the future assessment of the impact of irrigated rice cultivation on malaria transmission in the locality of Bankeng.

Insecticide resistance genes are often associated with pleiotropic effects on various mosquito life-history traits. However, very little information is available on the impact of insecticide resistance, especially metabolic resistance, on blood feeding process in mosquitoes. Here, using two recently detected DNA-based metabolic markers in the major malaria vector, An. funestus , we investigated how metabolic resistance genes could affect blood meal intake. After allowing both field F1 and lab F8 Anopheles funestus strains to feed on human arm for 30 minutes, we assessed the association between key parameters of blood meal process including, probing time, feeding duration, blood feeding success and blood meal size, and markers of glutathione S-transferase ( L119F-GSTe2 ) and cytochrome P450 ( CYP6P9a _R) - mediated metabolic resistance. None of the parameters of blood meal process was associated with L119F-GSTe2 genotypes. In contrast, for CYP6P9a _R, homozygote resistant mosquitoes were significantly more able to blood-feed than homozygote susceptible (OR = 3.3; CI 95%: 1.4-7.7; P =0.01) mosquitoes. Moreover, the volume of blood meal ingested by CYP6P9a-SS mosquitoes was lower than that of CYP6P9a-RS (PCYP6P9a gene affects the feeding success and blood meal size of An. funestus . However, no correlation was found in the expression of CYP6P9a and that of genes encoding for salivary proteins involved in blood meal process. This study suggests that P450-based metabolic resistance may increase the blood feeding ability of malaria vectors and potential impacting their vectorial capacity.