Resting Behavior of Malaria Vectors in Ghana and Its Implication on Vector Control.

Background: In Sub-Saharan Africa, there is widespread use of long-lasting insecticidal nets (LLINs) and Indoor residual spraying (IRS) to help control the density of malaria vectors and decrease the incidence of malaria in communities. An understanding of the interactions between increased insecticide use and resting behaviour patterns of malaria mosquitoes is important for an effective vector control programme. This study was carried out to investigate the resting behavior, host preference and infection with Plasmodium falciparum of malaria vectors in Ghana in the context of increasing insecticide resistance in malaria vectors in sub-saharan Africa.Methods: Indoor and outdoor resting Anopheline mosquitoes were sampled during the dry and rainy seasons in five sites that were in 3 ecological landscapes [Sahel savannah (Kpalsogou, Pagaza, Libga), Coastal savannah (Anyakpor) and Forest (Konongo) zones] using pyrethrum spray catches (PSC), mechanical aspiration (Prokopack) for indoor collections, pit shelter and Prokopack for outdoor collections. PCR based molecular diagnostics were used to determine mosquito speciation, genotype for knockdown resistance mutations (L1014S and L1014F), G119S Ace-1 mutation, specific host blood meal origins and sporozoite infection in field collected mosquitoes.Results: Anopheles gambiae s. l. was the predominant species (89.95%, n = 1,718), followed by An. rufipes (8.48%, n=162), and An. funestus s. l. (1.57%, n = 30). Sibling species of the Anopheles gambiae revealed An. coluzzii accounted for 63% (95% CI: 57.10 – 68.91), followed by An. gambiae s. s [27% (95% CI: 21.66 – 32.55)], and An. arabiensis [9% (95% CI: 6.22 – 13.57)]. The mean resting density of An. gambiae s. l. was higher outdoors (79.63%; 1,368/1,718) than indoors (20.37%; 350/1,718) (z = -4.815, p< 0.0001). The kdr west L1014F and the Ace-1 mutations were highest in indoor resting An. coluzzii and An. gambiae in the sahel-savannah sites compared to the forest and coastal savannah sites. Overall, the blood meal analyses revealed a large proportion of the malaria vectors preferred feeding on humans (70.2 %) than animals (29.8%) in all sites. The sporozoite rates was only detected in indoor resting An. coluzzii from the sahel savannah (5.0%) and forest (2.5%) zones.Conclusion: The study reports high outdoor resting densities of An. gambiae and An. coluzzii with high kdr west mutation frequencies, and persistence of malaria transmission indoors despite the use of LLINs and IRS. Continuous monitoring of changes in resting behavior of mosquitoes and implementation of complementary malaria control interventions are needed to target outdoor resting Anopheles mosquitoes in Ghana.

Anopheline Diversity in Indonesia: An Evaluation of Animal-Baited Sampling Techniques

Indonesia has rich Anopheline (Diptera: Culicidae) mosquito species living in various types of ecosystems. The study was conducted to profile and compare Anopheles diversity, equitability, and dominance in various ecosystems using different animal-based sampling techniques. The present study analyzed a subset of data collected from a nation-wide vector and animal reservoirs survey in 2016. Analyses were restricted to three ecosystem types (forest, nonforest, and coastal areas) in Java and Sumatera Islands. A total of 5,477 Anopheles were collected by using animal-baited (n = 1,909) and animal-baited trap nets (n = 1,978), consisting of 23 Anopheline species. Overall, Anopheles vagus was the most abundant species, followed by An. subpictus and An. barbirostris. Among the three ecosystems, the forest had a higher diversity index (H′ = 1.98), but each ecosystem has its specific predominant species. Compared with the animal-baited method, the Anopheles abundance collected by animal-baited trap nets was two-fold higher. Ecosystem, elevation, and sampling methods were associated with the abundance of female Anopheles (P-value &lt; 0.001). Our findings revealed that Anopheles were found in a different ecosystem, indicating the potential of malaria transmission. This suggests that improved malaria vector surveillance is essential in all types of ecosystem. Furthermore, the study suggested that animal-baited trap nets could be used as the standard method of outdoor resting sampling in Indonesia in addition to the traditional human landing collection approach.

Is outdoor-resting behaviour in malaria vectors consistent? Short report from northern Ghana.

Background Recent reports of a change in the resting behaviour of malaria vectors, from predominantly indoor resting to outdoor resting following blood feeding, have been attributed to selection pressure from use of vector control tools such as indoor residual spraying and long-lasting insecticide-treated nets. Recent studies have observed vectors resting predominantly outdoors in settings where anti-vector tools are extensively deployed. This present study examined if the outdoor resting behaviour in the vector population, is random or indicative of a consistent preference of one resting site over the other. Methods Mark-release-recapture (MRR) experiments were conducted with outdoor-resting Anopheles gambiae and An. funestus mosquitoes collected from pit shelters, animal houses and granaries in two villages in Northern Ghana during rainy and dry seasons. Mosquitoes were marked with fluorescent dyes and released indoors. The experiments were controlled with indoor-resting mosquitoes, which were marked and released outdoors. Species of all recaptured mosquitoes were identified and assessed for consistency in their resting behaviour. Results A total of 4,460 outdoor-resting mosquitoes comprising An. gambiae sensu lato (s.l.) (2,630, 59%) and An. funestus complex (1,830, 41%) were marked and released. Overall, 31 (0.7%) mosquitoes were recaptured mostly from outdoor location comprising 25 (81%) An. gambiae s.l. and 6 (19%) An. funestus complex. Only 3 (10%) of the recaptured mosquitoes were found resting indoors where they were released. The majority of the outdoor-recaptured mosquitoes were An. arabiensis (11, 39%), followed by An. coluzzii (7, 25%); whereas all indoor-recaptured mosquitoes were An. coluzzii. For the control experiment, 324 indoor-resting mosquitoes constituting 313 (97%) An. gambiae s.l. and 11 (3%) An. funestus complex were marked and released. However, none of these was recaptured neither indoors nor outdoors. More mosquitoes were significantly captured and recaptured during rainy season ( Z = 6.579, P < 0.0001). Conclusions The results obtained suggested the tendency for the mosquitoes to retain their outdoor-resting behaviour. Further investigations are required to ascertain if emerging preference for outdoor resting behaviour in malaria vector populations is consistent or a random occurrence.

Impact of cattle on the abundance of indoor and outdoor resting malaria vectors in southern Malawi

Background Understanding the blood feeding preferences and resting habits of malaria vectors is important for assessing and designing effective malaria vector control tools. The presence of livestock, such as cattle, which are used as blood meal hosts by some malaria vectors, may impact malaria parasite transmission dynamics. The presence of livestock may provide sufficient blood meals for the vectors, thereby reducing the frequency of vectors biting humans. Alternatively, the presence of cattle may enhance the availability of blood meals such that infectious mosquitoes may survive longer, thereby increasing the risk of malaria transmission. This study assessed the effect of household-level cattle presence and distribution on the abundance of indoor and outdoor resting malaria vectors. Methods Houses with and without cattle were selected in Chikwawa district, southern Malawi for sampling resting malaria vectors. Prokopack aspirators and clay pots were used for indoor and outdoor sampling, respectively. Each house was sampled over two consecutive days. For houses with cattle nearby, the number of cattle and the distances from the house to where the cattle were corralled the previous night were recorded. All data were analysed using generalized linear models fitted with Poisson distribution. Results The malaria vectors caught resting indoors were Anopheles gambiae sensustricto (s.s.), Anopheles arabiensis and Anopheles funestuss.s. Outdoor collections consisted primarily of An. arabiensis. The catch sizes of indoor resting An. gambiae sensulato (s.l.) were not different in houses with and without cattle (P = 0.34). The presence of cattle near a house was associated with a reduction in the abundance of indoor resting An. funestuss.l. (P = 0.04). This effect was strongest when cattle were kept overnight ≤ 15 m away from the houses (P = 0.03). The blood meal hosts varied across the species. Conclusion These results highlight differences between malaria vector species and their interactions with potential blood meal hosts, which may have implications for malaria risk. Whereas An. arabiensis remained unaffected, the reduction of An. funestuss.s. in houses near cattle suggests a potential protective effect of cattle. However, the low abundance of mosquitoes reduced the power of some analyses and limited the generalizability of the results to other settings. Therefore, further studies incorporating the vectors’ host-seeking behaviour/human biting rates are recommended to fully support the primary finding.

Influence of insecticide resistance on the biting and resting preferences of malaria vectors in the Gambia

Background The scale-up of indoor residual spraying and long-lasting insecticidal nets, together with other interventions have considerably reduced the malaria burden in The Gambia. This study examined the biting and resting preferences of the local insecticide-resistant vector populations few years following scale-up of anti-vector interventions. Method Indoor and outdoor-resting Anopheles gambiae mosquitoes were collected between July and October 2019 from ten villages in five regions in The Gambia using pyrethrum spray collection (indoor) and prokopack aspirator from pit traps (outdoor). Polymerase chain reaction assays were performed to identify molecular species, insecticide resistance mutations, Plasmodium infection rate and host blood meal. Results A total of 844 mosquitoes were collected both indoors (421, 49.9%) and outdoors (423, 50.1%). Four main vector species were identified, including An. arabiensis (indoor: 15%, outdoor: 26%); An. coluzzii (indoor: 19%, outdoor: 6%), An. gambiae s.s. (indoor: 11%, outdoor: 16%), An. melas (indoor: 2%, outdoor: 0.1%) and hybrids of An. coluzzii-An. gambiae s.s (indoors: 3%, outdoors: 2%). A significant preference for outdoor resting was observed in An. arabiensis (Pearson X2 = 22.7, df = 4, P<0.001) and for indoor resting in An. coluzzii (Pearson X2 = 55.0, df = 4, P<0.001). Prevalence of the voltage-gated sodium channel (Vgsc)-1014S was significantly higher in the indoor-resting (allele freq. = 0.96, 95%CI: 0.78–1, P = 0.03) than outdoor-resting (allele freq. = 0.82, 95%CI: 0.76–0.87) An. arabiensis population. For An. coluzzii, the prevalence of most mutation markers was higher in the outdoor (allele freq. = 0.92, 95%CI: 0.81–0.98) than indoor-resting (allele freq. = 0.78, 95%CI: 0.56–0.86) mosquitoes. However, in An. gambiae s.s., the prevalence of Vgsc-1014F, Vgsc-1575Y and GSTe2-114T was high (allele freq. = 0.96–1), but did not vary by resting location. The overall sporozoite positivity rate was 1.3% (95% CI: 0.5–2%) in mosquito populations. Indoor-resting An. coluzzii had mainly fed on human blood while indoor-resting An. arabiensis fed on animal blood. Conclusion In this study, high levels of resistance mutations were observed that could be influencing the mosquito populations to rest indoors or outdoors. The prevalent animal-biting behaviour demonstrated in the mosquito populations suggest that larval source management could be an intervention to complement vector control in this setting.

Is Outdoor-Resting Behaviour in Malaria Vectors Consistent? Short Report From Northern Ghana.

BackgroundRecent reports of a change in the resting behaviour of malaria vectors, from predominantly indoor resting to outdoor resting following blood feeding, have been attributed to selection pressure from use of vector control tools such as indoor residual spraying and long-lasting insecticide-treated nets. Recent studies have observed vectors resting predominantly outdoors in settings where anti-vector tools are extensively deployed. This present study examined if the outdoor resting behaviour in the vector population, is random or indicative of a consistent preference of one resting site over the other. MethodsMark-release-recapture (MRR) experiments were conducted with outdoor-resting Anopheles gambiae and An. funestus mosquitoes collected from pit shelters, animal houses and granaries in two villages in Northern Ghana. Mosquitoes were marked with fluorescent dyes and released indoors. The experiments were controlled with indoor-resting mosquitoes, which were marked and released outdoors. Species of all recaptured mosquitoes were identified and assessed for consistency in their resting behaviour. ResultsA total of 4,460 outdoor-resting mosquitoes comprising An. gambiae sensu lato (s.l.) (59%) and An. funestus complex (41%) were marked and released. Overall, 31 (0.7%) mosquitoes (25 An. gambiae s.l. and 6 An. funestus complex) were recaptured mostly from outdoor location. Only 3 of the recaptured mosquitoes were found resting indoors where they were released. The majority of the outdoor-recaptured mosquitoes were An. arabiensis (11, 39%), followed by An. coluzzii (7, 25%); whereas the indoor-recaptured mosquitoes were mainly (3) An. coluzzii. For the control experiment, 324 indoor-resting mosquitoes constituting 313 An. gambiae s.l. and 11 An. funestus complex were marked and released. However, none of these was recaptured neither indoors nor outdoors.ConclusionsThe mosquitoes demonstrated the tendency to retain their outdoor-resting behaviour. Further investigations are required to ascertain if emerging preference for outdoor resting behaviour in malaria vector populations is consistent or a random occurrence.

Insecticide resistance status of indoor and outdoor resting malaria vectors in a highland and lowland site in Western Kenya

Background Long Lasting Insecticidal Nets (LLINs) and indoor residual spraying (IRS) represent powerful tools for controlling malaria vectors in sub-Saharan Africa. The success of these interventions relies on their capability to inhibit indoor feeding and resting of malaria mosquitoes. This study sought to understand the interaction of insecticide resistance with indoor and outdoor resting behavioral responses of malaria vectors from Western Kenya. Methods The status of insecticide resistance among indoor and outdoor resting anopheline mosquitoes was compared in Anopheles mosquitoes collected from Kisumu and Bungoma counties in Western Kenya. The level and intensity of resistance were measured using WHO-tube and CDC-bottle bioassays, respectively. The synergist piperonyl butoxide (PBO) was used to determine if metabolic activity (monooxygenase enzymes) explained the resistance observed. The mutations at the voltage-gated sodium channel (Vgsc) gene and Ace 1 gene were characterized using PCR methods. Microplate assays were used to measure levels of detoxification enzymes if present. Results A total of 1094 samples were discriminated within Anopheles gambiae s.l. and 289 within An. funestus s.l. In Kisian (Kisumu county), the dominant species was Anopheles arabiensis 75.2% (391/520) while in Kimaeti (Bungoma county) collections the dominant sibling species was Anopheles gambiae s.s 96.5% (554/574). The An. funestus s.l samples analysed were all An. funestus s.s from both sites. Pyrethroid resistance of An.gambiae s.l F1 progeny was observed in all sites. Lower mortality was observed against deltamethrin for the progeny of indoor resting mosquitoes compared to outdoor resting mosquitoes (Mortality rate: 37% vs 51%, P = 0.044). The intensity assays showed moderate-intensity resistance to deltamethrin in the progeny of mosquitoes collected from indoors and outdoors in both study sites. In Kisian, the frequency of vgsc-L1014S and vgsc-L1014F mutation was 0.14 and 0.19 respectively in indoor resting malaria mosquitoes while those of the outdoor resting mosquitoes were 0.12 and 0.12 respectively. The ace 1 mutation was present in higher frequency in the F1 of mosquitoes resting indoors (0.23) compared to those of mosquitoes resting outdoors (0.12). In Kimaeti, the frequencies of vgsc-L1014S and vgsc-L1014F were 0.75 and 0.05 respectively for the F1 of mosquitoes collected indoors whereas those of outdoor resting ones were 0.67 and 0.03 respectively. The ace 1 G119S mutation was present in progeny of mosquitoes from Kimaeti resting indoors (0.05) whereas it was absent in those resting outdoors. Monooxygenase activity was elevated by 1.83 folds in Kisian and by 1.33 folds in Kimaeti for mosquitoes resting indoors than those resting outdoors respectively. Conclusion The study recorded high phenotypic, metabolic and genotypic insecticide resistance in indoor resting populations of malaria vectors compared to their outdoor resting counterparts. The indication of moderate resistance intensity for the indoor resting mosquitoes is alarming as it could have an operational impact on the efficacy of the existing pyrethroid based vector control tools. The use of synergist (PBO) in LLINs may be a better alternative for widespread use in these regions recording high insecticide resistance.

Biting and resting preferences of malaria vectors in The Gambia

BackgroundThe scale-up of indoor residual spraying and long-lasting insecticidal nets, together with other interventions have considerably reduced the malaria burden in The Gambia. This study examined the biting and resting preferences of the local insecticide-resistant vector populations few years following scale-up of anti-vector interventions.MethodIndoor and outdoor-resting Anopheles gambiae mosquitoes were collected between July and October 2019 from ten villages in five regions in The Gambia using pyrethrum spray collection (indoor) and prokopack aspirator from pit traps (outdoor). Polymerase chain reaction assays were performed to identify molecular species, insecticide resistance mutations, Plasmodium infection rate and host blood meal. ResultsA total of 844 mosquitoes were collected both indoors (421, 49.9%) and outdoors (423, 50.1%). Four main vector species were identified, including An. arabiensis (indoor: 15%, outdoor: 26%); An. coluzzii (indoor: 19%, outdoor: 6%), An. gambiae s.s. (indoor: 11%, outdoor: 16%), An. melas (indoor: 2%, outdoor: 0.1%) and hybrids of An. coluzzii-An. gambiae (indoors: 3%, outdoors: 2%). A significant preference for outdoor resting was observed in An. arabiensis (Pearson X2=22.7, df=4, P<0.001) and for indoor resting in An. coluzzii (Pearson X2=55.0, df=4, P<0.001). Prevalence of the voltage-gated sodium channel (Vgsc)-1014S was higher in the indoor-resting (allele freq. = 0.96, 95%CI: 0.78–1) than outdoor-resting (allele freq. = 0.82, 95%CI: 0.76–0.87) An. arabiensis population. For An. coluzzii, the prevalence of most mutation markers were higher in the outdoor (allele freq. = 0.92, 95%CI: 0.81–0.98) than indoor-resting (allele freq. = 0.78, 95%CI: 0.56–0.86) mosquitoes. Sporozoite positivity rate was 1.3% (95% CI: 0.5–2%). Indoor-resting An. coluzzii had mainly fed on human blood while indoor-resting An. arabiensis, animal blood.ConclusionThe indoor-resting behavior of An. arabiensis that preferred animal blood and had low sporozoite rates, may be determined by the Vgsc-1014S mutation. Control interventions may include complementary vector control approaches such as zooprophylaxis.

Insecticide resistance status of indoor and outdoor resting malaria vectors in a highland and lowland site in Western Kenya

BackgroundLong Lasting Insecticidal Nets (LLINs) and indoor residual spraying (IRS) represent powerful tools for controlling malaria vectors in sub-Saharan Africa. The success of these interventions relies on their capability to inhibit indoor feeding and resting of malaria mosquitoes. This study sought to understand the interaction of insecticide resistance with indoor and outdoor resting behavioral responses of malaria vectors from Western Kenya.MethodsThe status of insecticide resistance among indoor and outdoor resting anopheline mosquitoes was compared in Anopheles mosquitoes collected from Kisumu and Bungoma counties in Western Kenya. The level and intensity of resistance were measured using WHO-tube and CDC-bottle bioassays, respectively. The synergist piperonyl butoxide (PBO) was used to determine if metabolic activity (monooxygenase enzymes) explained the resistance observed. The mutations at the voltage-gated sodium channel (Vgsc) gene and Ace 1 gene were characterized using PCR methods. Microplate assays were used to measure levels of detoxification enzymes if present.ResultsA total of 1094 samples were discriminated within Anopheles gambiae s.l. and 289 within An. funestus s.l. In Kisian (Kisumu county), the dominant species was Anopheles arabiensis 75.2% (391/520) while in Kimaeti (Bungoma county) collections the dominant sibling species was Anopheles gambiae s.s 96.5% (554/574). The An. funestus s.l samples analysed were all An. funestus s.s from both sites. Pyrethroid resistance of An.gambiae s.l F1 progeny was observed in all sites. Lower mortality was observed against deltamethrin for the progeny of indoor resting mosquitoes compared to outdoor resting mosquitoes (Mortality rate: 37% vs 51%, P=0.044). The intensity assays showed moderate-intensity resistance to deltamethrin in the progeny of mosquitoes collected from indoors and outdoors in both study sites. In Kisian, the frequency of vgsc-L1014S and vgsc-L1014F mutation was 0.14 and 0.19 respectively in indoor resting malaria mosquitoes while those of the outdoor resting mosquitoes were 0.12 and 0.12 respectively. The ace 1 mutation was present in higher frequency in the F1 of mosquitoes resting indoors (0.23) compared to those of mosquitoes resting outdoors (0.12). In Kimaeti, the frequencies of vgsc-L1014S and vgsc-L1014F were 0.75 and 0.05 respectively for the F1 of mosquitoes collected indoors whereas those of outdoor resting ones were 0.67 and 0.03 respectively. The ace 1 G119S mutation was present in progeny of mosquitoes from Kimaeti resting indoors (0.05) whereas it was absent in those resting outdoors. Monooxygenase activity was elevated by 1.83 folds in Kisian and by 1.33 folds in Kimaeti for mosquitoes resting indoors than those resting outdoors respectively.ConclusionThe study recorded high phenotypic, metabolic and genotypic insecticide resistance in indoor resting populations of malaria vectors compared to their outdoor resting counterparts. The indication of moderate resistance intensity for the indoor resting mosquitoes is alarming as it could have an operational impact on the efficacy of the existing pyrethroid based vector control tools. The use of synergist (PBO) in LLINs may be a better alternative for widespread use in these regions recording high insecticide resistance.