Asymptomatic Plasmodium infection among primary schoolchildren and Anopheles-mediated malaria transmission: A cross-sectional study in Ouidah; south-western Benin

Understanding the contribution of asymptomatic Plasmodium carriers in malaria transmission might be helpful to design and implement new control measures. The present study explored the prevalence of asymptomatic and symptomatic Plasmodium infections (asexual and sexual stages) and the contribution of asymptomatic P. falciparum carriers to Anopheles-mediated malaria transmission in Ouidah (Benin). Thick and thin blood smears were examined from finger-prick blood specimens using light microscopy, and the density of both asexual and sexual stages of Plasmodium species was calculated. Infectivity of gametocyte-infected blood samples to Anopheles gambiae was assessed through direct membrane feeding assays. The prevalence of asymptomatic Plasmodium infections was 28.73% (289/1006). All the asymptomatic gametocyte-carriers (19/19), with gametocytaemia ranging from 10 ̶ 1200 gametocytes/μL of blood, were infectious to An. gambiae mosquitoes. The mean oocyst prevalences varied significantly (χ2 = 16.42, df = 7, p = 0.02) among laboratory mosquito strains (6.9 ̶ 39.4%) and near-field mosquitoes (4.9 ̶ 27.2%). Likewise, significant variation (χ2 = 56.85, df = 7, p = 6.39 × 10−10) was observed in oocyst intensity. Our findings indicate that asymptomatic Plasmodium carriers could significantly contribute to malaria transmission. Overall, this study highlights the importance of diagnosing and treating asymptomatic and symptomatic infection carriers during malaria control programmes

Interplay Between Oxytetracycline and the Homozygote kdr (L1014F) Resistance Genotype on Fecundity in Anopheles gambiae (Diptera: Culicidae) Mosquitoes

The insecticide resistance in Anopheles gambiae mosquitoes has remained the major threat for vector control programs but the fitness effects conferred by these mechanisms are poorly understood. To fill this knowledge gap, the present study aimed at testing the hypothesis that antibiotic oxytetracycline could have an interaction with insecticide resistance genotypes and consequently inhibit the fecundity in An. gambiae. Four strains of An. gambiae: Kisumu (susceptible), KisKdr (kdr (L1014F) resistant), AcerKis (ace-1 (G119S) resistant) and AcerKdrKis (both kdr (L1014F) and ace-1 (G119S) resistant) were used in this study. The different strains were allowed to bloodfeed on a rabbit previously treated with antibiotic oxytetracycline at a concentration of 39·10–5 M. Three days later, ovarian follicles were dissected from individual mosquito ovaries into physiological saline solution (0.9% NaCl) under a stereomicroscope and the eggs were counted. Fecundity was substantially lower in oxytetracycline-exposed KisKdr females when compared to that of the untreated individuals and oxytetracycline-exposed Kisumu females. The exposed AcerKis females displayed an increased fecundity compared to their nontreated counterparts whereas they had reduced fecundity compared to that of oxytetracycline-exposed Kisumu females. There was no substantial difference between the fecundity in the treated and untreated AcerKdrKis females. The oxytetracycline-exposed AcerKdrKis mosquitoes had an increased fecundity compared to that of the exposed Kisumu females. Our data indicate an indirect effect of oxytetracycline in reducing fecundity of An. gambiae mosquitoes carrying kdrR (L1014F) genotype. These findings could be useful for designing new integrated approaches for malaria vector control in endemic countries

Phenotypic Insecticide Resistance in Anopheles gambiae (Diptera: Culicidae): Specific Characterization of Underlying Resistance Mechanisms Still Matters

An effective control of malaria vectors requires an extensive knowledge of mechanisms underlying the resistance-phenotypes developed by these vectors against insecticides. We investigated Anopheles gambiae mosquitoes from Benin and Togo for their intensity of insecticide resistance and we discussed the involvement of genotyped mechanisms in the resistance-phenotypes observed. Three- to five-day-old adult mosquitoes emerged from field and laboratory An. gambiae larvae were assayed using WHO tube intensity tests against various doses of deltamethrin: 1× (0.05%); 2× (0.1%); 5× (0.25%); 7.5× (0.375%) and those of pirimiphos-methyl: 0.5× (0.125%); 1× (0.25%). Members of An. gambiae complex were screened in field populations using polymerase chain reaction (PCR) assays. The presence of kdrR(1014F/1014S) and ace-1R(119S) mutations was also investigated using TaqMan and PCR-RFLP techniques, respectively. Anopheles gambiae from field were very resistant to deltamethrin, whereas KisKdr and AcerKdrKis strains displayed 100% mortality rates at 2× the diagnostic dose. In contrast, the field mosquitoes displayed a low resistance-intensity against 1× the diagnostic dose of pirimiphos-methyl, whereas AcerKis and AcerKdrKis strains showed susceptibility at 0.5× the diagnostic dose. Anopheles gambiae s.s., Anopheles coluzzii, and Anopheles arabiensis were identified. Allelic frequencies of kdrR (1014F) and ace-1R (119S) mutations in the field populations varied from 0.65 to 1 and 0 to 0.84, respectively. The field An. gambiae displayed high-resistance levels against deltamethrin and pirimiphos-methyl when compared with those of the laboratory An. gambiae-resistant strains. These results exhibit the complexity of underlying insecticide resistance mechanisms in these field malaria vectors

Beninese Plant Extracts with Antiplasmodial Activity Select New Allele Variants Msp1 and Msp2 in Plasmodium falciparum

Background. Natural medicinal products are commonly used as a remedy against malaria infections in African populations and have become a major source of information for the screening of new and more effective antiplasmodial molecules. Therefore, in vitro studies are needed to validate the efficacy of these medicinal products and to explore the potential effects of such drugs on the genetic diversity of Plasmodium falciparum. The current study has investigated the impact of some Beninese plant extracts with antiplasmodial activity on the genetic diversity of P. falciparum. Method. Five (5) ethanolic plant extracts (Dissotis rotundifolia, Ehretia cymosa Thonn, Hibiscus surattensis L., Cola millenii K. Shum, and Costus afer Ker Gawl) and a compound extracted from Ehretia cymosa Thonn (encoded CpE2) were tested against asexual stage parasites of a culture-adapted strain of P. falciparum. Subsequently, the P. falciparum Msp1 and Msp2 markers were genotyped, and the number of allelic variants and the multiplicity of infection (MOI) were compared between drug-exposed and unexposed parasites. Results. All plant extracts have shown inhibitory activity against asexual P. falciparum and selected new allelic variants of the Msp1 and Msp2 genes compared to unexposed parasites. The newly selected allelic variants were K1_100bp and RO33_300bp of the Msp1 gene and FC27_150bp, FC27_300bp, FC27_400bp, and FC27_600bp of the Msp2 gene. However, there was no significant difference in MOI between drug-exposed and unexposed parasites. Conclusion. Our study highlights a source for the selection of new Msp1 and Msp2 alleles after exposure to antimalarial drugs. These findings pave the way for further studies investigating the true roles of these newly selected alleles in P. falciparum

Putative pleiotropic effects of the knockdown resistance (L1014F) allele on the life-history traits of Anopheles gambiae.

Background Existing mechanisms of insecticide resistance are known to help the survival of mosquitoes following contact with chemical compounds, even though they could negatively affect the life-history traits of resistant malaria vectors. In West Africa, the knockdown resistance mechanism kdrR (L1014F) is the most common. However, little knowledge is available on its effects on mosquito life-history traits. The fitness effects associated with this knockdown resistance allele in Anopheles gambiae sensu stricto (s.s.) were investigated in an insecticide-free laboratory environment. Methods The life-history traits of Kisumu (susceptible) and KisKdr (kdr resistant) strains of An. gambiae s.s. were compared. Larval survivorship and pupation rate were assessed as well as fecundity and fertility of adult females. Female mosquitoes of both strains were directly blood fed through artificial membrane assays and then the blood-feeding success, blood volume and adult survivorship post-blood meal were assessed. Results The An. gambiae mosquitoes carrying the kdrR allele (KisKdr) laid a reduced number of eggs. The mean number of larvae in the susceptible strain Kisumu was three-fold overall higher than that seen in the KisKdr strain with a significant difference in hatching rates (81.89% in Kisumu vs 72.89% in KisKdr). The KisKdr larvae had a significant higher survivorship than that of Kisumu. The blood-feeding success was significantly higher in the resistant mosquitoes (84%) compared to the susceptible ones (34.75%). However, the mean blood volume was 1.36 µL/mg, 1.45 µL/mg and 1.68 µL/mg in Kisumu, homozygote and heterozygote KisKdr mosquitoes, respectively. After blood-feeding, the heterozygote KisKdr mosquitoes displayed highest survivorship when compared to that of Kisumu. Conclusions The presence of the knockdown resistance allele appears to impact the life-history traits, such as fecundity, fertility, larval survivorship, and blood-feeding behaviour in An. gambiae. These data could help to guide the implementation of more reliable strategies for the control of malaria vectors

Genome-wide association studies reveal novel loci associated with pyrethroid and organophosphate resistance in Anopheles gambiae

Resistance to insecticides in Anopheles mosquitoes threatens the effectiveness of malaria control, but the genetics of resistance are only partially understood. We performed a large scale multi-country genome-wide association study of resistance to two widely used insecticides: deltamethrin and pirimiphos-methyl, using sequencing data from An. gambiae and An. coluzzii from ten locations in West Africa. Resistance was highly multi-genic, multiallelic and variable between populations. While the strongest and most consistent association with deltamethrin resistance came from Cyp6aa1, this was based on several independent copy number variants (CNVs) in An. coluzzii, and on a non-CNV haplotype in An. gambiae. For pirimiphos-methyl, signals included Ace1, cytochrome P450s, glutathione S-transferases and the nAChR target site of neonicotinoid insecticides. The regions around Cyp9k1 and the Tep family of immune genes showed evidence of cross-resistance to both insecticides. These locally-varying, multi-allelic patterns highlight the challenges involved in genomic monitoring of resistance, and form the basis for improved surveillance methods

Asymptomatic Plasmodium infection among primary schoolchildren and Anopheles-mediated malaria transmission: A cross-sectional study in Ouidah; south-western Benin

Understanding the contribution of asymptomatic Plasmodium carriers in malaria transmission might be helpful to design and implement new control measures. The present study explored the prevalence of asymptomatic and symptomatic Plasmodium infections (asexual and sexual stages) and the contribution of asymptomatic P. falciparum carriers to Anopheles-mediated malaria transmission in Ouidah (Benin). Thick and thin blood smears were examined from finger-prick blood specimens using light microscopy, and the density of both asexual and sexual stages of Plasmodium species was calculated. Infectivity of gametocyte-infected blood samples to Anopheles gambiae was assessed through direct membrane feeding assays. The prevalence of asymptomatic Plasmodium infections was 28.73% (289/1006). All the asymptomatic gametocyte-carriers (19/19), with gametocytaemia ranging from 10 ̶ 1200 gametocytes/μL of blood, were infectious to An. gambiae mosquitoes. The mean oocyst prevalences varied significantly (χ2 = 16.42, df = 7, p = 0.02) among laboratory mosquito strains (6.9 ̶ 39.4%) and near-field mosquitoes (4.9 ̶ 27.2%). Likewise, significant variation (χ2 = 56.85, df = 7, p = 6.39 × 10−10) was observed in oocyst intensity. Our findings indicate that asymptomatic Plasmodium carriers could significantly contribute to malaria transmission. Overall, this study highlights the importance of diagnosing and treating asymptomatic and symptomatic infection carriers during malaria control programmes

Vectorial competence, insecticide resistance in Anopheles funestus and operational implications for malaria vector control strategies in Benin Republic

Abstract The primary reason for the failure of malaria vector control across endemic regions is the widespread insecticide resistance observed in Anopheles vectors. The most dominant African vectors of malaria parasites are Anopheles gambiae and Anopheles funestus mosquitoes. These species often exhibit divergent behaviours and adaptive changes underscoring the importance of deploying active and effective measures in their control. Unlike An. gambiae, An. funestus mosquitoes are poorly studied in Benin Republic. However, recent reports indicated that An. funestus can adapt and colonize various ecological niches owing to its resistance against insecticides and adaptation to changing breeding habitats. Unfortunately, scientific investigations on the contribution of An. funestus to malaria transmission, their susceptibility to insecticide and resistance mechanism developed are currently insufficient for the design of better control strategies. In an attempt to gather valuable information on An. funestus, the present review examines the progress made on this malaria vector species in Benin Republic and highlights future research perspectives on insecticide resistance profiles and related mechanisms, as well as new potential control strategies against An. funestus. Literature analysis revealed that An. funestus is distributed all over the country, although present in low density compared to other dominant malaria vectors. Interestingly, An. funestus is being found in abundance during the dry seasons, suggesting an adaptation to desiccation. Among the An. funestus group, only An. funestus sensu stricto (s.s.) and Anopheles leesoni were found in the country with An. funestus s.s. being the most abundant species. Furthermore, An. funestus s.s. is the only one species in the group contributing to malaria transmission and have adapted biting times that allow them to bite at dawn. In addition, across the country, An. funestus were found resistant to pyrethroid insecticides used for bed nets impregnation and also resistant to bendiocarb which is currently being introduced in indoor residual spraying formulation in malaria endemic regions. All these findings highlight the challenges faced in controlling this malaria vector. Therefore, advancing the knowledge of vectorial competence of An. funestus, understanding the dynamics of insecticide resistance in this malaria vector, and exploring alternative vector control measures, are critical for sustainable malaria control efforts in Benin Republic