Blood meal analysis of Anopheles vectors of simian malaria based on laboratory and field studies

Blood feeding and host-seeking behaviors of a mosquito play an imperative role in determining its vectorial capacity in transmitting pathogens. Unfortunately, limited information is available regarding blood feeding behavior of Anopheles species in Malaysia. Collection of resting Anopheles mosquitoes for blood meal analysis poses a great challenge especially for forest dwelling mosquitoes. Therefore, a laboratory-based study was conducted to evaluate the potential use of mosquitoes caught using human landing catch (HLC) for blood meal analysis, and subsequently to document blood feeding behavior of local Anopheles mosquitoes in Peninsular Malaysia. The laboratory-based experiment from this study revealed that mosquitoes caught using HLC had the potential to be used for blood meal analysis. Besides HLC, mosquitoes were also collected using manual aspirator and Mosquito Magnet. Overall, 47.4% of 321 field-caught Anopheles mosquitoes belonging to six species were positive for vertebrate host DNA in their blood meal. The most frequent blood meal source was human (45.9%) followed by wild boar (27.4%), dog (15.3%) and monkey (7.5%). Interestingly, only Anopheles cracens and Anopheles introlatus (Leucosphyrus Group) fed on monkey. This study further confirmed that members of the Leucosphyrus Group are the predominant vectors for knowlesi malaria transmission in Peninsular Malaysia mainly due to their simio-anthropophagic feeding behavior.

The Centres for Disease Control Light Trap (CDC LT) and The Human Decoy Trap (HDT) Compared to The Human Landing Catch (HLC) for Measuring Anopheles Biting in Rural Tanzania

BackgroundThe intensity of vector mosquito biting is an important measure for malaria epidemiology and control. The human landing catch (HLC) is an effective entomological surveillance tool, but is labour-intensive, expensive and raises safety issues. The Centres for Disease Control light trap (CDC LT) and the human decoy trap (HDT) are less costly and exposure-free alternatives. This study compared the CDC LT and HDT against the HLC for measuring Anopheles (An.) biting in rural Tanzania and assessed their suitability as HLC proxies.MethodsIndoor mosquito surveys using HLC and CDC LT and outdoor surveys using HLC and HDT were conducted in 2017 and in 2019 in Ulanga, Tanzania in 19 villages, with one trap per house per night. Species composition, sporozoite rates and the numbers of mosquitoes caught by different trap types were compared. Aggregating the data by village and month, the Bland-Altman approach was used to assess agreement. ResultsOverall, 66,807 Anopheles funestus and 14,606 An. arabiensis adult females were caught from 6,013 CDC LT, 339 indoor HLC, 136 HDT and 195 outdoor HLC collections. Overall, the CDC LT caught fewer malaria vectors than indoor HLC: An. arabiensis (Adjusted rate ratio (Adj.RR) =0.35 (95% confidence interval (CI):0.27-0.46)) and An. funestus (Adj.RR=0.63(95%CI:0.51-0.79)). HDT caught fewer malaria vectors than outdoor HLC: An. arabiensis (Adj.RR=0.04(95%CI:0.01-0.14)) and An. funestus (Adj.RR=0.10(95%CI:0.07-0.15)). The bias and variability of the ratios of geometric mean mosquitoes caught by CDC LT and HDT relative to HLC collections for the same village-month were dependent on mosquito densities. The relative efficacies of both CDC LT and HDT declined with mosquito abundance. The variability in the ratios was substantial for low HLC counts and decreased as mosquito abundance increased. CDCLT caught a higher proportion of infected An. arabiensis and An. funestus than HLC, and HDT caught no infected mosquitoes.ConclusionsIf caution is taken in appreciation of its limitations, the CDC LT is suitable for use in routine entomological surveys and may be preferable for measuring sporozoite rates for Afrotropical mosquitoes. Use of HLC is useful to estimate human exposure to mosquitoes for estimating Entomological Inoculation Rate (EIR). The present design of the HDT is not amenable for use to conduct large-scale entomological surveys.

Evaluation of the interaction between insecticide resistance-associated genes and malaria transmission in Anopheles gambiae sensu lato in central Côte d’Ivoire

Background There is evidence that the knockdown resistance gene (Kdr) L1014F and acetylcholinesterase-1 gene (Ace-1R) G119S mutations involved in pyrethroid and carbamate resistance in Anopheles gambiae influence malaria transmission in sub-Saharan Africa. This is likely due to changes in the behaviour, life history and vector competence and capacity of An. gambiae. In the present study, performed as part of a two-arm cluster randomized controlled trial evaluating the impact of household screening plus a novel insecticide delivery system (In2Care Eave Tubes), we investigated the distribution of insecticide target site mutations and their association with infection status in wild An. gambiae sensu lato (s.l.) populations. Methods Mosquitoes were captured in 40 villages around Bouaké by human landing catch from May 2017 to April 2019. Randomly selected samples of An. gambiae s.l. that were infected or not infected with Plasmodium sp. were identified to species and then genotyped for Kdr L1014F and Ace-1R G119S mutations using quantitative polymerase chain reaction assays. The frequencies of the two alleles were compared between Anopheles coluzzii and Anopheles gambiae and then between infected and uninfected groups for each species. Results The presence of An. gambiae (49%) and An. coluzzii (51%) was confirmed in Bouaké. Individuals of both species infected with Plasmodium parasites were found. Over the study period, the average frequency of the Kdr L1014F and Ace-1R G119S mutations did not vary significantly between study arms. However, the frequencies of the Kdr L1014F and Ace-1R G119S resistance alleles were significantly higher in An. gambiae than in An. coluzzii [odds ratio (95% confidence interval): 59.64 (30.81–131.63) for Kdr, and 2.79 (2.17–3.60) for Ace-1R]. For both species, there were no significant differences in Kdr L1014F or Ace-1R G119S genotypic and allelic frequency distributions between infected and uninfected specimens (P > 0.05). Conclusions Either alone or in combination, Kdr L1014F and Ace-1R G119S showed no significant association with Plasmodium infection in wild An. gambiae and An. coluzzii, demonstrating the similar competence of these species for Plasmodium transmission in Bouaké. Additional factors including behavioural and environmental ones that influence vector competence in natural populations, and those other than allele measurements (metabolic resistance factors) that contribute to resistance, should be considered when establishing the existence of a link between insecticide resistance and vector competence. Graphical Abstract

The effect of irrigation on malaria vector bionomics and transmission intensity in western Ethiopia

Background Irrigation schemes may result in subsequent changes in malaria disease dynamics. Understanding the mechanisms and effects of irrigation on malaria vector bionomics and transmission intensity is essential to develop new or alternative surveillance and control strategies to reduce or control malaria risk. This study was designed to assess the effect of rice irrigation on malaria vector bionomics and transmission intensity in the Gambella Region, Ethiopia. Methods Comparative cross-sectional study was conducted in Abobo District of the Gambella Region, Ethiopia. Accordingly, clusters (kebeles) were classified into nearby and faraway clusters depending on their proximity to the irrigation scheme. Adult mosquito survey was conducted in February, August and November 2018 from three nearby and three faraway clusters using Centers for Disease Control and Prevention (CDC) light traps (LTs). During the November survey, human landing catch (HLC) and pyrethrum spray catch (PSC) were also conducted. The collected mosquitoes were morphologically identified to species and tested for Plasmodium infection using circumsporozoite protein enzyme-linked immunosorbent assay (CSP-ELISA). Furthermore, species-specific polymerase chain reaction (PCR) was performed to identify member species of the Anopheles gambiae complex. Chi-square and t-tests were used to analyze the data using the SPSS version 20 software package. Results A total of 4319 female anopheline mosquitoes comprising An. gambiae sensu lato, An. funestus group, An. pharoensis, An. coustani complex and An. squamosus were collected. Overall, 84.5% and 15.5% of the anopheline mosquitoes were collected from the nearby and faraway clusters, respectively. Anopheles gambiae s.l. was the predominant (56.2%) anopheline species in the area followed by An. pharoensis (15.7%). The density of anopheline mosquitoes was significantly higher in the nearby clusters in both HLCs [t(3) = 5.14, P = 0.0143] and CDC LT catches [t(271.97) = 7.446, P < 0.0001). The overall sporozoite rate of anopheline species from the nearby clusters was 10-fold higher compared to the faraway clusters. Conclusions Significantly higher mosquito population density was observed in areas close to the irrigation sites. Sporozoite infection rate in the mosquito population was also markedly higher from the nearby clusters. Therefore, the irrigation scheme could increase the risk of malaria in the area. Graphical abstract

Malaria Transmission, Vector Diversity, and Insecticide Resistance at a Peri-Urban Site in the Forest Zone of Ghana

IntroductionRecent surge of Anopheles resistance to major classes of World Health Organization (WHO)-approved insecticides globally necessitates the need for information about local malaria vector populations. It is believed that insecticide efficacy loss may lead to operational failure of control interventions and an increase in malaria infection transmission. We investigated the susceptibility levels of malaria vectors to all classes of WHO-approved vector control insecticides and described the dynamics of malaria transmission in a peri-urban setting.MethodsFit 3–5-day-old adults that emerged from Anopheles larvae collected from several different sites in the study area were subjected to the WHO bioassay for detecting insecticide resistance. The knockdown resistance gene (kdr) mutations within the vector populations were detected using PCR. Entomological inoculation rates were determined using the human landing catch technique and Plasmodium falciparum circumsporozoite ELISA.ResultsThe malaria vectors from the study area were resistant to all classes of insecticides tested. Out of the 284 Anopheles complex specimen assayed for the resistance study, 265 (93.30%) were identified as Anopheles gambiae s.s. The kdr gene was detected in 90% of the Anopheles gambiae s.s. assayed. In an area where Anopheles coluzzii resistance to insecticides had never been reported, the kdr gene was detected in 78% of the Anopheles coluzzii sampled. The entomological inoculation rate (EIR) for the dry season was 1.44 ib/m/n, whereas the EIR for the rainy season was 2.69 ib/m/n.ConclusionsThis study provides information on the high parasite inoculation rate and insecticide resistance of malaria vectors in a peri-urban community, which is critical in the development of an insecticide resistance management program for the community.

The human-baited host decoy trap (HDT) is an efficient sampling device for exophagic malaria mosquitoes within irrigated lands in southern Malawi

Irrigation schemes provide an ideal habitat for Anopheles mosquitoes particularly during the dry season. Reliable estimates of outdoor host-seeking behaviour are needed to assess the impact of vector control options and this is particularly the case for Anopheles arabiensis which displays a wide range of behaviours that circumvent traditional indoor-insecticide based control. In this study we compared the Host Decoy Trap (HDT) with the Human Landing Catch (HLC) and Suna trap in a repeated Latin square design in two villages on an irrigated sugar estate in southern Malawi. Over the course of 18 trapping nights we caught 379 female Anopheles, the majority of which were identified as An. arabiensis. Overall, the HDT and HLC caught a similar number of Anopheles per night with both methods catching significantly higher densities than the Suna trap across both villages. Regardless of the density of Anopheles mosquitoes in each village the HLC and HDT demonstrated broadly similar sampling efficacy. We conclude that the HDT is an effective sampling device for outdoor host seeking An. arabiensis in southern Malawi. The presence of An. arabiensis in irrigated lands during the dry season poses a challenge for ongoing indoor vector control efforts.

Contribution to the mosquito fauna (Diptera, Culicidae) of Lika, central Croatia, with special reference to invasive species

This study of mosquitoes in Lika, geographically, geologically and climatically a very specific region, was conducted to gain insight into the composition and relative abundance of the mosquito fauna, as well as to check for the possible presence of invasive species. Sampling took place from July to September 2020. Mosquito larvae were captured from medium and small breeding sites using a 25 cm diameter net and a plastic dipper. Adult specimens were sampled in both urban and rural areas using CO2 baited CDC traps, CO2 baited BG Sentinel traps with BG Lure and human landing catch. The study was conducted at 69 sites. We collected 5,126 specimens (100 adults and 5,026 larvae) and demonstrated the presence of 16 mosquito species. CDC traps were almost five times more effective (quantity and quality) than BG Sentinel traps. The most common mosquito taxa caught in Lika were Culex pipiens complex (78.36% of all larvae) and Ae. geniculatus (30.26% of all adults). Both invasive mosquito species present in Croatia, Aedes albopictus and Ae. japonicus, were recorded, with Ae. albopictus being detected in Lika for the first time. Aedes japonicus was widespread, recorded in 23 localities. According to molecular analysis, only An. maculipennis s.s. was confirmed in the An. maculipennis complex. Within the Cx. pipiens complex, both Cx. pipiens biotypes, pipiens and molestus were confirmed together with one single hybrid specimen. Analysis of mosquito occurrence with regard to altitude shows that most samples and species were collected between 601 and 700 m a.s.l., although the Shannon evenness index and Hill’s index show the highest value in the range of 901 to 1140 m a.s.l. In this systematic study on the mosquito fauna in Lika, a significant species diversity, including invasive species, was found.

Evaluation of Mosquito Magnet and other collection tools for Anopheles mosquito vectors of simian malaria

Background Vector surveillance is essential in determining the geographical distribution of mosquito vectors and understanding the dynamics of malaria transmission. With the elimination of human malaria cases, knowlesi malaria cases in humans are increasing in Malaysia. This necessitates intensive vector studies using safer trapping methods which are both field efficient and able to attract the local vector populations. Thus, this study evaluated the potential of Mosquito Magnet as a collection tool for Anopheles mosquito vectors of simian malaria along with other known collection methods. Methods A randomized 4 × 4 Latin square designed experiment was conducted to compare the efficiency of the Mosquito Magnet against three other common trapping methods: human landing catch (HLC), CDC light trap and human baited trap (HBT). The experiment was conducted over six replicates where sampling within each replicate was carried out for 4 consecutive nights. An additional 4 nights of sampling was used to further evaluate the Mosquito Magnet against the “gold standard” HLC. The abundance of Anopheles sampled by different methods was compared and evaluated with focus on the Anopheles from the Leucosphyrus group, the vectors of knowlesi malaria. Results The Latin square designed experiment showed HLC caught the greatest number of Anopheles mosquitoes (n = 321) compared to the HBT (n = 87), Mosquito Magnet (n = 58) and CDC light trap (n = 13). The GLMM analysis showed that the HLC method caught significantly more Anopheles mosquitoes compared to Mosquito Magnet (P = 0.049). However, there was no significant difference in mean nightly catch of Anopheles mosquitoes between Mosquito Magnet and the other two trapping methods, HBT (P = 0.646) and CDC light traps (P = 0.197). The mean nightly catch for both An. introlatus (9.33 ± 4.341) and An. cracens (4.00 ± 2.273) caught using HLC was higher than that of Mosquito Magnet, though the differences were not statistically significant (P > 0.05). This is in contrast to the mean nightly catch of An. sinensis (15.75 ± 5.640) and An. maculatus (15.78 ± 3.479) where HLC showed significantly more mosquito catches compared to Mosquito Magnet (P < 0.05). Conclusions Mosquito Magnet has a promising ability to catch An. introlatus and An. cracens, the important vectors of knowlesi and other simian malarias in Peninsular Malaysia. The ability of Mosquito Magnet to catch some of the Anopheles mosquito species is comparable to HLC and makes it an ethical and safer alternative. Graphic Abstract

Comparison between the human landing catch (HLC), an exposure-free mosquito-electrocuting trap (MET) and a BG-Sentinel trap (BGS) for evaluation of transfluthrin emanator against Aedes aegypti using choice and no-choice tests in a semi-field system

Background The human landing catch (HLC) measures human exposure to mosquito bites and evaluates the efficacy of vector control tools. However, it may expose volunteers to potentially infected mosquitoes. The mosquito electrocuting trap (MET) and BG-Sentinel traps (BGS) represent alternative, exposure-free methods for sampling host-seeking mosquitoes. This study investigates whether these methods can be effectively used as alternatives to HLC for measuring the efficacy transfluthrin against Aedes aegypti. Methods The protective efficacy (PE) of freestanding passive transfluthrin emanators (FTPEs), measured by HLC, MET and BGS were compared in no-choice and choice tests. The methods were located 2 m from an experimental hut with FTPEs positioned 3 m on either side of them. For the choice experiment, a competitor HLC was included 10 m from the first collection point. One hundred laboratory-reared Ae. aegypti mosquitoes were released and collected for three consecutive hours. Results In the no-choice test, each method measured similar PE; HLC: 66% (95% confidence interval [CI]: 50–82), MET: 55% (95% CI: 48–63) and BGS: 64% (95% CI: 54–73). The proportion of mosquitoes recaptured was consistent between methods (20–24%) in treatment and varied (47–71%) in the control. However, in choice tests, the PE measured by each method varied; HLC: 37% (95% CI: 25–50%), MET: 76% (95% CI: 61–92) and BGS trap: 0% (95% CI: 0–100). Recaptured mosquitoes were no longer consistent between methods in treatment (2–26%) and remained variable in the control (7–42%). FTPE provided 50% PE to the second HLC 10 m away. In the control, the MET and the BGS were less efficacious in collecting mosquitoes in the presence of a second HLC. Conclusions Measurement of the PE in isolation was fairly consistent for HLC, MET and BGS. Because HLC is not advisable, it is reasonable to use MET / BGS as a proxy for HLC for testing VP in areas of active arboviruses endemic areas. The presence of a human host in close proximity invalidated the PE estimates from BGS and METs. Findings also indicated that transfluthrin can protect multiple people in the peridomestic area and that at short-range mosquitoes select humans over the BGS.

Indoor and outdoor malaria transmission in two ecological settings in rural Mali: implications for vector control

Background Implementation and upscale of effective malaria vector control strategies necessitates understanding the multi-factorial aspects of transmission patterns. The primary aims of this study are to determine the vector composition, biting rates, trophic preference, and the overall importance of distinguishing outdoor versus indoor malaria transmission through a study at two communities in rural Mali. Methods Mosquito collection was carried out between July 2012 and June 2016 at two rural Mali communities (Dangassa and Koïla Bamanan) using pyrethrum spray-catch and human landing catch approaches at both indoor and outdoor locations. Species of Anopheles gambiae complex were identified by polymerase chain reaction (PCR). Enzyme-Linked -Immuno-Sorbent Assay (ELISA) were used to determine the origin of mosquito blood meals and presence of Plasmodium falciparum sporozoite infections. Results A total of 11,237 An. gambiae sensu lato (s.l.) were collected during the study period (5239 and 5998 from the Dangassa and Koïla Bamanan sites, respectively). Of the 679 identified by PCR in Dangassa, Anopheles coluzzii was the predominant species with 91.4% of the catch followed by An. gambiae (8.0%) and Anopheles arabiensis (0.6%). At the same time in Koïla Bamanan, of the 623 An. gambiae s.l., An. coluzzii accounted for 99% of the catch, An. arabiensis 0.8% and An. gambiae 0.2%. Human Blood Index (HBI) measures were significantly higher in Dangassa (79.4%; 95% Bayesian credible interval (BCI) [77.4, 81.4]) than in Koïla Bamanan (15.9%; 95% BCI [14.7, 17.1]). The human biting rates were higher during the second half of the night at both sites. In Dangassa, the sporozoite rate was comparable between outdoor and indoor mosquito collections. For outdoor collections, the sporozoite positive rate was 3.6% (95% BCI [2.1–4.3]) and indoor collections were 3.1% (95% BCI [2.4–5.0]). In Koïla Bamanan, the sporozoite rate was higher indoors at 4.3% (95% BCI [2.7–6.3]) compared with outdoors at 2.4% (95% BCI [1.1–4.2]). In Dangassa, corrected entomological inoculation rates (cEIRs) using HBI were 13.74 [95% BCI 9.21–19.14] infective bites/person/month (ib/p/m) at indoor, and 18.66 [95% BCI 12.55–25.81] ib/p/m at outdoor. For Koïla Bamanan, cEIRs were 1.57 [95% BCI 2.34–2.72] ib/p/m and 0.94 [95% BCI 0.43–1.64] ib/p/m for indoor and outdoor, respectively. EIRs were significantly higher at the Dangassa site than the Koïla Bamanan site. Conclusion The findings in this work may indicate the occurrence of active, outdoor residual malaria transmission is comparable to indoor transmission in some geographic settings. The high outdoor transmission patterns observed here highlight the need for additional strategies to combat outdoor malaria transmission to complement traditional indoor preventive approaches such as long-lasting insecticidal nets (LLINs) and indoor residual spraying (IRS) which typically focus on resting mosquitoes.