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Abstract Background Aedes albopictus is an important vector for pathogens such as dengue, Zika, and chikungunya viruses. While insecticides is the mainstay for mosquito control, their widespread and excessive use has led to the increased resistance in Ae. albopictus globally. Gut symbiotic bacteria are believed to play a potential role in insect physiology, potentially linking to mosquitoes’ metabolic resistance against insecticides. Methods We investigated the role of symbiotic bacteria in the development of resistance in Ae. albopictus by comparing gut symbiotic bacteria between deltamethrin-sensitive and deltamethrin-resistant populations. Adults were reared from field-collected larvae. Sensitive and resistant mosquitoes were screened using 0.03% and 0.09% deltamethrin, respectively, on the basis of the World Health Organization (WHO) tube bioassay. Sensitive and resistant field-collected larvae were screened using 5 × LC50 (lethal concentration at 50% mortality) and 20 × LC50 concentration of deltamethrin, respectively. Laboratory strain deltamethrin-sensitive adults and larvae were used as controls. The DNA of gut samples from these mosquitoes were extracted using the magnetic bead method. Bacterial 16S rDNA was sequenced using BGISEQ method. We isolated and cultured gut microorganisms from adult and larvae mosquitoes using four different media: Luria Bertani (LB), brain heart infusion (BHI), nutrient agar (NA), and salmonella shigella (SS). Results Sequencing revealed significantly higher gut microbial diversity in field-resistant larvae compared with field-sensitive and laboratory-sensitive larvae (P

Abstract Background To interrupt residual malaria transmission and achieve successful elimination of Plasmodium falciparum in low-transmission settings, the World Health Organization (WHO) recommends the administration of a single dose of 0.25 mg/kg (or 15 mg/kg for adults) primaquine (PQ) combined with artemisinin-based combination therapy (ACT), without glucose-6-phosphate dehydrogenase (G6PD) testing. However, due to the risk of haemolysis in patients with G6PD deficiency (G6PDd), PQ use is uncommon. Thus, this study aimed to assess the safety of a single low dose of PQ administered to patients with G6PD deficiency. Methods An observational cohort study was conducted with patients treated for uncomplicated P. falciparum malaria with either single-dose PQ (0.25 mg/kg) (SLD PQ) + ACT or ACT alone. Microscopy-confirmed uncomplicated P. falciparum malaria patients visiting public health facilities in Arjo Didessa, Southwest Ethiopia, were enrolled in the study from September 2019 to November 2022. Patients with uncomplicated P. falciparum malaria were followed up for 28 days through clinical and laboratory diagnosis, such as measurements of G6PD levels and haemoglobin (Hb) concentrations. G6PD levels were measured by a quantiative CareSTART™ POCT S1 biosensor machine. Patient interviews were also conducted, and the type and frequency of clinical complaints were recorded. Hb data were taken on days (D) 7, 14, 21, and 28 following treatment with SLD-PQ + ACT or ACT alone. Results A total of 249 patients with uncomplicated P. falciparum malaria were enrolled in this study. Of these, 83 (33.3%) patients received ACT alone, and 166 (66.7%) received ACT combined with SLD-PQ treatment. The median age of the patients was 20 (IQR 28–15) years. G6PD deficiency was found in 17 (6.8%) patients, 14 males and 3 females. There were 6 (7.2%) and 11 (6.6%) phenotypic G6PD-deficient patients in the ACT alone and ACT + SLD-PQ arms, respectively. The mean Hb levels in patients treated with ACT + SLD-PQ were reduced by an average of 0.45 g/dl (95% CI = 0.39 to 0.52) in the posttreatment phase (D7) compared to a reduction of 0.30 g/dl (95% CI = 0.14 to − 0.47) in patients treated with ACT alone (P = 0.157). A greater mean Hb reduction was observed on day 7 in the G6PDd ACT + SLD-PQ group (− 0.60 g/dL) than in the G6PDd ACT alone group (− 0.48 g/dL); however, there was no statistically significant difference (P = 0.465). Overall, D14 losses were 0.10 g/dl (95% CI = − 0.00 to 0.20) and 0.05 g/dl (95% CI = − 0.123 to 0.22) in patients with and without SLD-PQ, respectively (P = 0.412). Conclusions This study’s findings indicate that using SLD-PQ in combination with ACT is safe for uncomplicated P. falciparum malaria regardless of the patient's G6PD status in Ethiopian settings. Caution should be taken in extrapolating this finding in other settings with diverse G6DP phenotypes.

Abstract Background Understanding of malaria ecology is a prerequisite for designing locally adapted control strategies in resource-limited settings. The aim of this study was to utilize the spatial heterogeneity in malaria transmission for the designing of adaptive interventions. Methods Field collections of clinical malaria incidence, asymptomatic Plasmodium infection, and malaria vector data were conducted from 108 randomly selected clusters which covered different landscape settings including irrigated farming, seasonal flooding area, lowland dryland farming, and highlands in western Kenya. Spatial heterogeneity of malaria was analyzed and classified into different eco-epidemiological zones. Results There was strong heterogeneity and detected hot/cold spots in clinical malaria incidence, Plasmodium prevalence, and vector abundance. The study area was classified into four zones based on clinical malaria incidence, parasite prevalence, vector density, and altitude. The two irrigated zones have either the highest malaria incidence, parasite prevalence, or the highest malaria vector density; the highlands have the lowest vector density and parasite prevalence; and the dryland and flooding area have the average clinical malaria incidence, parasite prevalence and vector density. Different zones have different vector species, species compositions and predominant species. Both indoor and outdoor transmission may have contributed to the malaria transmission in the area. Anopheles gambiae sensu stricto (s.s.), Anopheles arabiensis, Anopheles funestus s.s., and Anopheles leesoni had similar human blood index and malaria parasite sporozoite rate. Conclusion The multi-transmission-indicator-based eco-epidemiological zone classifications will be helpful for making decisions on locally adapted malaria interventions.

Abstract Background Understanding the clustering of infections for persistent malaria transmission is critical to determining how and where to target specific interventions. This study aimed to determine the density, blood meal sources and malaria transmission risk of anopheline vectors by targeting malaria index cases, their neighboring households and control villages in Arjo-Didessa, southwestern Ethiopia. Methods An entomological study was conducted concurrently with a reactive case detection (RCD) study from November 2019 to October 2021 in Arjo Didessa and the surrounding vicinity, southwestern Ethiopia. Anopheline mosquitoes were collected indoors and outdoors in index case households and their surrounding households (neighboring households), as well as in control households, using pyrethrum spray cache (PSC) and U.S. Centers for Disease Control and Prevention (CDC) light traps. Adult mosquitoes were morphologically identified, and speciation in the Anopheles gambiae complex was done by PCR. Mosquito Plasmodium infections and host blood meal sources were detected by circumsporozoite protein enzyme-linked immunosorbent assay (CSP-ELISA) and cytochrome b-based blood meal PCR, respectively. Results Among the 770 anopheline mosquitoes collected, An. gambiae sensu lato (A. gambiae s.l.) was the predominant species, accounting for 87.1% (n = 671/770) of the catch, followed by the Anopheles coustani complex and Anopheles pharoensis, which accounted for 12.6% (n = 97/770) and 0.26% (n = 2/770) of the catch, respectively. From the sub-samples of An. gambiae s.l.analyzed with PCR, An. arabiensis and Anopheles amharicus were identified. The overall mean density of mosquitoes was 1.26 mosquitoes per trap per night using the CDC light traps. Outdoor mosquito density was significantly higher than indoor mosquito density in the index and neighboring households (P = 0.0001). The human blood index (HBI) and bovine blood index (BBI) of An. arabiensis were 20.8% (n = 34/168) and 24.0% (n = 41/168), respectively. The overall Plasmodium sporozoite infection rate of anophelines (An. arabiensis and An. coustani complex) was 4.4% (n = 34/770). Sporozoites were detected indoors and outdoors in captured anopheline mosquitoes. Of these CSP-positive species for Pv-210, Pv-247 and Pf, 41.1% (n = 14/34) were captured outdoors. A significantly higher proportion of sporozoite-infected mosquitoes were caught in index case households (5.6%, n = 8/141) compared to control households (1.1%, n = 2/181) (P = 0.02), and in neighboring households (5.3%, n = 24/448) compared to control households (P = 0.01). Conclusions The findings of this study indicated that malaria index cases and their neighboring households had higher outdoor mosquito densities and Plasmodium infection rates. The study also highlighted a relatively higher outdoor mosquito density, which could increase the potential risk of outdoor malaria transmission and may play a role in residual malaria transmission. Thus, it is important to strengthen the implementation of vector control interventions, such as targeted indoor residual spraying, long-lasting insecticidal nets and other supplementary vector control measures such as larval source management and community engagement approaches. Furthermore, in low transmission settings, such as the Arjo Didessa Sugarcane Plantation, providing health education to local communities, enhanced environmental management and entomological surveillance, along with case detection and management by targeting of malaria index cases and their immediate neighboring households, could be important measures to control residual malaria transmission and achieve the targeted elimination goals. Graphical Abstract

Abstract Background Water resource development projects, such as dams and irrigation schemes, have a positive impact on food security and poverty reduction. However, such projects could increase prevalence of vector borne disease, such as malaria. This study investigate the impact of different agroecosystems and prevalence of malaria infection in Southwest Ethiopia. Methods Two cross-sectional surveys were conducted in the dry and wet seasons in irrigated and non-irrigated clusters of Arjo sugarcane and Gambella rice development areas of Ethiopia in 2019. A total of 4464 and 2176 study participants from 1449 households in Arjo and 546 households in Gambella enrolled in the study and blood samples were collected, respectively. All blood samples were microscopically examined and a subset of microscopy negative blood samples (n = 2244) were analysed by qPCR. Mixed effect logistic regression and generalized estimating equation were used to determine microscopic and submicroscopic malaria infection and the associated risk factors, respectively. Results Prevalence by microscopy was 2.0% (88/4464) in Arjo and 6.1% (133/2176) in Gambella. In Gambella, prevalence was significantly higher in irrigated clusters (10.4% vs 3.6%) than in non-irrigated clusters (p

Abstract Background Water resource development projects are essential for increasing agricultural productivity and ensuring food security. However, these activities require the modification of pre-existing environmental settings, which may alter mosquito larval habitat availability and seasonality. The intensive utilization of current adult vector control tools results in insecticide resistance among the main vectors. When coupled with behavioural resistances, a shift in malaria vector feeding and resting behaviours could compromise the effectiveness of the current adult vector control strategies. Thus, it is important to look for new or alternative vector control interventions for immatures to complement adult control by focusing on different larval habitats and their seasonal availability. Thus, this study investigated larval habitat seasonality and seasonal larval abundance and distribution in irrigated sugar cane plantation settings in Ethiopia. Methods Anopheles mosquito larval habitats were surveyed and visited twice a month for a period of 14 months. Anopheline larvae and pupae were collected, reared, and fed finely ground fish food. Adults were provided with sucrose solution and kept under standard conditions. Female Anopheles mosquitoes were identified morphologically and using a species-specific PCR assay. Environmental parameters, which include habitats’ physico-chemical characteristics, were assessed. Larval habitat diversity and larval abundance and distribution were determined across different seasons. Results The study revealed that Anopheles gambiae sensu lato (s.l.) was the most predominant 4197(57%) vector species, followed by Anopheles coustani complex 2388 (32.8%). Molecular analysis of sub-samples of An. gambiae s.l. resulted in Anopheles arabiensis (77.9%) and Anopheles amharicus (21.5%), and the remaining 1.1% (n = 7) sub-samples were not amplified. Physico-chemical parameters such as temperature (t = 2.22, p = 0.028), conductivity (t = 3.21, p = 0.002), dissolved oxygen (t = 7.96, p = 0.001), nitrate ion (t = 2.51, p = 0.013), and ammonium ion (t = 2.26, p = 0.025) showed a significant and direct association with mosquito larval abundance. Furthermore, mosquito larval abundance was correlated with distance to the nearest houses (r = − 0.42, p = 0.001), exposure to sunlight (r = 0.34, p = 0.001), during long and short rainy season animal hoof prints, truck tires/road puddles and rain pools were negatively correlated (r = − 0.22, p = 0.01) and types of habitat (r = − 0.20, p = 0.01). Significant habitat type productivity were observed in man-made pools (t = 3.881, P = 0.01163), rain pools, animal hoof prints, (t = − 4.332, P = 0.00749 in both short and long rainy season, whereas, during dry seasons habitat type productivity almost similar and have no significance difference. Conclusion The study found that different larval habitats had variable productivity in different seasons, and that physical and physicochemical features like ammonium and nitrate, as well as the distance between larval habitats and households, are related to larval production. As a result, vector control should take into account the seasonality of Anopheles larval habitat as well as the impact of pesticide application on larval source management.

The invasion of Anopheles stephensi into Africa poses a potential threat to malaria control and elimination on the continent. However, it is not clear if the recent malaria resurgence in Ethiopia has linked to the expansion of An. stephensi. We obtained the clinical malaria case reports and malaria intervention data from the Ethiopian Ministry of Health (MoH) for the period 2001–2022. We analyzed clinical malaria hotspots and investigated the potential role of An. stephensi in the 2022 malaria outbreaks. Clinical malaria cases in Ethiopia decreased by 80%, from 5.2 million cases in 2004 to 1.0 million cases in 2018; however, cases increased steadily to 2.6 million confirmed cases in 2022. Plasmodium vivax cases and proportion have increased significantly in the past 5 years. Clinical malaria hotspots are concentrated along the western Ethiopian border areas and have grown significantly from 2017 to 2022. Major malaria outbreaks in 2022/2023 were detected in multiple sites across Ethiopia, and An. stephensi was the predominant vector in some of these sites, however, it was absence from many of the outbreak sites. The causes of recent upsurge in malaria in Ethiopia may be multi-factorial and it is a subject of further investigation.

Abstract Background The rise of insecticide resistance against malaria vectors in sub-Saharan Africa has resulted in the need to consider other methods of vector control. The potential use of biological methods, including larvivorous fish, Bacillus thuringiensis israelensis (Bti) and plant shading, is sustainable and environmentally friendly options. This study examined the survivorship of Anopheles arabiensis and Anopheles funestus larvae and habitat productivity in four permanent habitat types in Homa Bay county, western Kenya. Methods Predator densities were studied in a laboratory setup while habitat productivity and larval survivorship was studied in field setup. Results Fish were observed as the most efficient predator (75.8% larval reduction rate) followed by water boatman (69%), and dragonfly nymph (69.5%) in predation rates. Lower predation rates were observed in backswimmers (31%), water beetles (14.9%), water spiders (12.2%), mayflies (7.3%), and tadpoles (6.9%). Increase in predator density in the field setup resulted in decreased Culex larval density. Larval and pupa age–specific distribution was determined and their survivorship curves constructed. Combined larvae (Stage I–IV) to pupa mortality was over 97% for An. arabiensis and 100% for An. funestus. The highest larval stage survival rate was from larval stages I to II and the lowest from larval stage IV to pupa. Stage-specific life tables indicated high mortality rates at every developmental stage, especially at the larval stage II and III. Conclusion Determination of the efficiency of various larval predators and habitat productivity will help with the correct identification of productive habitats and selection of complementary vector control methods through environmental management and/or predator introduction (for instance fish) in the habitats.

Abstract Background Mosquitoes are vectors of many pathogens, such as malaria, dengue virus, yellow fever virus, filaria and Japanese encephalitis virus. Wolbachia are capable of inducing a wide range of reproductive abnormalities in their hosts, such as cytoplasmic incompatibility. Wolbachia has been proposed as a tool to modify mosquitoes that are resistant to pathogen infection as an alternative vector control strategy. This study aimed to determine natural Wolbachia infections in different mosquito species across Hainan Province, China. Methods Adult mosquitoes were collected using light traps, human landing catches and aspirators in five areas in Hainan Province from May 2020 to November 2021. Species were identified based on morphological characteristics, species-specific PCR and DNA barcoding of cox1 assays. Molecular classification of species and phylogenetic analyses of Wolbachia infections were conducted based on the sequences from PCR products of cox1, wsp, 16S rRNA and FtsZ gene segments. Results A total of 413 female adult mosquitoes representing 15 species were identified molecularly and analyzed. Four mosquito species (Aedes albopictus, Culex quinquefasciatus, Armigeres subalbatus and Culex gelidus) were positive for Wolbachia infection. The overall Wolbachia infection rate for all mosquitoes tested in this study was 36.1% but varied among species. Wolbachia types A, B and mixed infections of A × B were detected in Ae. albopictus mosquitoes. A total of five wsp haplotypes, six FtsZ haplotypes and six 16S rRNA haplotypes were detected from Wolbachia infections. Phylogenetic tree analysis of wsp sequences classified them into three groups (type A, B and C) of Wolbachia strains compared to two groups each for FtsZ and 16S rRNA sequences. A novel type C Wolbachia strain was detected in Cx. gelidus by both single locus wsp gene and the combination of three genes. Conclusion Our study revealed the prevalence and distribution of Wolbachia in mosquitoes from Hainan Province, China. Knowledge of the prevalence and diversity of Wolbachia strains in local mosquito populations will provide part of the baseline information required for current and future Wolbachia-based vector control approaches to be conducted in Hainan Province. Graphical Abstract

Abstract Background Future distribution of dengue risk is usually predicted based on predicted climate changes using general circulation models (GCMs). However, it is difficult to validate the GCM results and assess the uncertainty of the predictions. The observed changes in climate may be very different from the GCM results. We aim to utilize trends in observed climate dynamics to predict future risks of Aedes albopictus in China. Methods We collected Ae. albopictus surveillance data and observed climate records from 80 meteorological stations from 1970 to 2021. We analyzed the trends in climate change in China and made predictions on future climate for the years 2050 and 2080 based on trend analyses. We analyzed the relationship between climatic variables and the prevalence of Ae. albopictus in different months/seasons. We built a classification tree model (based on the average of 999 runs of classification and regression tree analyses) to predict the monthly/seasonal Ae. albopictus distribution based on the average climate from 1970 to 2000 and assessed the contributions of different climatic variables to the Ae. albopictus distribution. Using these models, we projected the future distributions of Ae. albopictus for 2050 and 2080. Results The study included Ae. albopictus surveillance from 259 sites in China found that winter to early spring (November–February) temperatures were strongly correlated with Ae. albopictus prevalence (prediction accuracy ranges 93.0–98.8%)—the higher the temperature the higher the prevalence, while precipitation in summer (June–September) was important predictor for Ae. albopictus prevalence. The machine learning tree models predicted the current prevalence of Ae. albopictus with high levels of agreement (accuracy > 90% and Kappa agreement > 80% for all 12 months). Overall, winter temperature contributed the most to Ae. albopictus distribution, followed by summer precipitation. An increase in temperature was observed from 1970 to 2021 in most places in China, and annual change rates varied substantially from -0.22 ºC/year to 0.58 ºC/year among sites, with the largest increase in temperature occurring from February to April (an annual increase of 1.4–4.7 ºC in monthly mean, 0.6–4.0 ºC in monthly minimum, and 1.3–4.3 ºC in monthly maximum temperature) and the smallest in November and December. Temperature increases were lower in the tropics/subtropics (1.5–2.3 ºC from February–April) compared to the high-latitude areas (2.6–4.6 ºC from February–April). The projected temperatures in 2050 and 2080 by this study were approximately 1–1.5 °C higher than those projected by GCMs. The estimated current Ae. albopictus risk distribution had a northern boundary of north-central China and the southern edge of northeastern China, with a risk period of June–September. The projected future Ae. albopictus risks in 2050 and 2080 cover nearly all of China, with an expanded risk period of April–October. The current at-risk population was estimated to be 960 million and the future at-risk population was projected to be 1.2 billion. Conclusions The magnitude of climate change in China is likely to surpass GCM predictions. Future dengue risks will expand to cover nearly all of China if current climate trends continue.

Abstract Background In western Kenya, not all malaria cases are reported as stipulated in the community case management of malaria (CCMm) strategy. This underreporting affects the equity distribution of malaria commodities and the evaluation of interventions. The current study aimed to evaluate the effectiveness of community health volunteers’ active case detection and management of malaria in western Kenya. Methods Cross-sectional active case detection (ACD) of malaria survey was carried out between May and August 2021 in three eco-epidemiologically distinct zones in Kisumu, western Kenya: Kano Plains, Lowland lakeshore and Highland Plateau. The CHVs conducted biweekly ACD of malaria household visits to interview and examine residents for febrile illness. The Community Health Volunteers (CHVs) performance during the ACD of malaria was observed and interviews done using structured questionnaires. Results Of the total 28,800 surveyed, 2597 (9%) had fever and associated malaria symptoms. Eco-epidemiological zones, gender, age group, axillary body temperature, bed net use, travel history, and survey month all had a significant association with malaria febrile illness (p

Abstract Background Malaria in western Kenya is currently characterized by sustained high Plasmodial transmission and infection resurgence, despite positive responses in some areas following intensified malaria control interventions since 2006. This study aimed to evaluate long-term changes in malaria transmission profiles and to assess patterns of asymptomatic malaria infections in school children aged 5–15 years at three sites in western Kenya with heterogeneous malaria transmission and simultaneous malaria control interventions. Methods The study was conducted from 2018 to 2019 and is based on data taken every third year from 2005 to 2014 during a longitudinal parasitological and mosquito adult surveillance and malaria control programme that was initiated in 2002 in the villages of Kombewa, Iguhu, and Marani. Plasmodium spp. infections were determined using microscopy. Mosquito samples were identified to species and host blood meal source and sporozoite infections were assayed using polymerase chain reaction. Results Plasmodium falciparum was the only malaria parasite evaluated during this study (2018–2019). Asymptomatic malaria parasite prevalence in school children decreased in all sites from 2005 to 2008. However, since 2011, parasite prevalence has resurged by > 40% in Kombewa and Marani. Malaria vector densities showed similar reductions from 2005 to 2008 in all sites, rose steadily until 2014, and decreased again. Overall, Kombewa had a higher risk of infection compared to Iguhu (χ 2 = 552.52, df = 1, P

Abstract Background Malaria remains a public health problem in Kenya despite sustained interventions deployed by the government. One of the major impediments to effective malaria control is a lack of accurate diagnosis and effective treatment. This study was conducted to assess clinical malaria incidence and treatment seeking profiles of febrile cases in western Kenya. Methods Active case detection of malaria was carried out in three eco-epidemiologically distinct zones topologically characterized as lakeshore, hillside, and highland plateau in Kisumu County, western Kenya, from March 2020 to March 2021. Community Health Volunteers (CHVs) conducted biweekly visits to residents in their households to interview and examine for febrile illness. A febrile case was defined as an individual having fever (axillary temperature ≥ 37.5 °C) during examination or complaints of fever and other nonspecific malaria related symptoms 1–2 days before examination. Prior to the biweekly malaria testing by the CHVs, the participants' treatment seeking methods were based on their behaviors in response to febrile illness. In suspected malaria cases, finger-prick blood samples were taken and tested for malaria parasites with ultra-sensitive Alere® malaria rapid diagnostic tests (RDT) and subjected to real-time polymerase chain reaction (RT-PCR) for quality control examination. Results Of the total 5838 residents interviewed, 2205 residents had high temperature or reported febrile illness in the previous two days before the visit. Clinical malaria incidence (cases/1000people/month) was highest in the lakeshore zone (24.3), followed by the hillside (18.7) and the highland plateau zone (10.3). Clinical malaria incidence showed significant difference across gender (χ2 = 7.57; df = 2, p = 0.0227) and age group (χ2 = 58.34; df = 4, p

Abstract Background Identification and characterization of larval habitats, documentation of Anopheles spp. composition and abundance, and Plasmodium spp. infection burden are critical components of integrated vector management. The present study aimed to investigate the effect of landscape heterogeneity on entomological and parasitological indices of malaria in western Kenya. Methods A cross-sectional entomological and parasitological survey was conducted along an altitudinal transect in three eco-epidemiological zones: lakeshore along the lakeside, hillside, and highland plateau during the wet and dry seasons in 2020 in Kisumu County, Kenya. Larval habitats for Anopheles mosquitoes were identified and characterized. Adult mosquitoes were sampled using pyrethrum spray catches (PSC). Finger prick blood samples were taken from residents and examined for malaria parasites by real-time PCR (RT-PCR). Results Increased risk of Plasmodium falciparum infection was associated with residency in the lakeshore zone, school-age children, rainy season, and no ITNs (χ 2 = 41.201, df = 9, P

Abstract Background Evolutionary pressures lead to the selection of efficient malaria vectors either resistant or susceptible to Plasmodium parasites. These forces may favour the introduction of species genotypes that adapt to new breeding habitats, potentially having an impact on malaria transmission. Thioester-containing protein 1 (TEP1) of Anopheles gambiae complex plays an important role in innate immune defenses against parasites. This study aims to characterize the distribution pattern of TEP1 polymorphisms among populations of An. gambiae sensu lato (s.l.) in western Kenya. Methods Anopheles gambiae adult and larvae were collected using pyrethrum spray catches (PSC) and plastic dippers respectively from Homa Bay, Kakamega, Bungoma, and Kisumu counties between 2017 and 2020. Collected adults and larvae reared to the adult stage were morphologically identified and then identified to sibling species by PCR. TEP1 alleles were determined in 627 anopheles mosquitoes using restriction fragment length polymorphisms-polymerase chain reaction (RFLP-PCR) and to validate the TEP1 genotyping results, a representative sample of the alleles was sequenced. Results Two TEP1 alleles (TEP1*S1 and TEP1*R2) and three corresponding genotypes (*S1/S1, *R2/S1, and *R2/R2) were identified. TEP1*S1 and TEP1*R2 with their corresponding genotypes, homozygous *S1/S1 and heterozygous *R2/S1 were widely distributed across all sites with allele frequencies of approximately 80% and 20%, respectively both in Anopheles gambiae and Anopheles arabiensis. There was no significant difference detected among the populations and between the two mosquito species in TEP1 allele frequency and genotype frequency. The overall low levels in population structure (F ST = 0.019) across all sites corresponded to an effective migration index (Nm = 12.571) and low Nei’s genetic distance values (

Abstract Background The city of Guangzhou has been the epicenter of dengue fever in China since the 1990s, with Aedes albopictus being the primary vector. The main method used to control vectors and prevent dengue fever has been the application of chemical insecticides; however, this control strategy has resulted in the development of resistance to these insecticides in mosquitoes. Here we report our investigation of the patterns of knockdown resistance (kdr) mutations in 15 field populations of Ae. albopictus collected from 11 districts in Guangzhou. Results Four mutant alleles (V1016G, F1534S, F1534C, F1534L) were detected in domain II and III of the voltage-gated sodium channel (VGSC) gene. Various allele frequencies of kdr mutations were observed (3.1–25.9% for V1016G, 22.6–85.5% for F1534S, 0–29.0% for F1534L, 0.6–54.2% for F1534C). Seven kdr haplotypes (VF, VS, VL, VC, GF, GC, GS) were identified; the highest frequency of haplotypes was found for the single mutant haplotype VS (50.8%), followed by the wild-type VF haplotype (21.7%) and the single mutant haplotype VC (11.9%). Of the three double mutant haplotypes, GF was the most frequent (8.8%), followed by GC (1.2%) and GS (0.8%). Aedes albopictus showed spatial heterogeneity in deltamethrin resistance in populations collected in Guangzhou. We also observed significant differences in haplotype frequency. The frequency of the VC haplotype was significantly higher in high-risk dengue areas than in low-risk ones. Conclusions The kdr allele V1016G was discovered for the first time in Guangzhou. Genetic isolation in mosquito populations and long-term insecticide selection seem to be responsible for the persistent, patchy distribution of kdr mutant alleles. The small-scale spatial heterogeneity in the distribution and frequency of kdr mutations may have important implications for vector control operations and insecticide resistance management strategies. Graphical Abstract

Abstract Background Myanmar is one of the six countries in the Greater Mekong Subregion (GMS) of Southeast Asia. Malaria vectors comprise many Anopheles species, which vary in abundance and importance in malaria transmission among different geographical locations in the GMS. Information about the species composition, abundance, and insecticide resistance status of vectorial systems in Myanmar is scarce, hindering our efforts to effectively control malaria vectors in this region. Methods During October and November 2019, larvae and adult females of Anopheles mosquitoes were collected in three sentinel villages of Banmauk township in northern Myanmar. Adult female mosquitoes collected by cow-baited tent collection (CBTC) and adults reared from field-collected larvae (RFCL) were used to determine mortality rates and knockdown resistance (kdr) against deltamethrin using the standard WHO susceptibility test. Molecular species identification was performed by multiplex PCR and ITS2 PCR, followed by DNA sequencing. The kdr mutation at position 1014 of the voltage-gated sodium channel gene was genotyped by DNA sequencing for all Anopheles species tested. Results A total of 1596 Anopheles mosquitoes from seven morphologically identified species groups were bioassayed. Confirmed resistance to deltamethrin was detected in the populations of An. barbirostris (s.l.), An. hyrcanus (s.l.), and An. vagus, while possible resistance was detected in An. annularis (s.l.), An. minimus, and An. tessellatus. Anopheles kochi was found susceptible to deltamethrin. Compared to adults collected by CBTC, female adults from RFCL had significantly lower mortality rates in the four species complexes. A total of 1638 individuals from 22 Anopheles species were molecularly identified, with the four most common species being An. dissidens (20.5%) of the Barbirostris group, An. peditaeniatus (19.4%) of the Hyrcanus group, An. aconitus (13.4%) of the Funestus group, and An. nivipes (11.5%) of the Annularis group. The kdr mutation L1014F was only detected in the homozygous state in two An. subpictus (s.l.) specimens and in a heterozygous state in one An. culicifacies (s.l.) specimen. Conclusions This study provides updated information about malaria vector species composition and insecticide resistance status in northern Myanmar. The confirmed deltamethrin resistance in multiple species groups constitutes a significant threat to malaria vector control. The lack or low frequency of target-site resistance mutations suggests that other mechanisms are involved in resistance. Continual monitoring of the insecticide resistance of malaria vectors is required for effective vector control and insecticide resistance management. Graphical Abstract

Abstract Background Accurate malaria diagnosis and appropriate treatment at local health facilities are critical to reducing morbidity and human reservoir of infectious gametocytes. The current study assessed the accuracy of malaria diagnosis and treatment practices in three health care facilities in rural western Kenya. Methods The accuracy of malaria detection and treatment recommended compliance was monitored in two public and one private hospital from November 2019 through March 2020. Blood smears from febrile patients were examined by hospital laboratory technicians and re-examined by an expert microscopists thereafter subjected to real-time polymerase chain reaction (RT-PCR) for quality assurance. In addition, blood smears from patients diagnosed with malaria rapid diagnostic tests (RDT) and presumptively treated with anti-malarial were re-examined by an expert microscopist. Results A total of 1131 febrile outpatients were assessed for slide positivity (936), RDT (126) and presumptive diagnosis (69). The overall positivity rate for Plasmodium falciparum was 28% (257/936). The odds of slide positivity was higher in public hospitals, 30% (186/624, OR:1.44, 95% CI = 1.05–1.98, p

Abstract Background Land use change has increasingly been expanding throughout the world in the past decades. It can have profound effects on the spatial and temporal distribution of vector borne diseases like malaria through ecological and habitat change. Understanding malaria disease occurrence and the impact of prevention interventions under this intense environmental modification is important for effective and efficient malaria control strategy. Methods A descriptive ecological study was conducted by reviewing health service records at Abobo district health office. The records were reviewed to extract data on malaria morbidity, mortality, and prevention and control methods. Moreover, Meteorological data were obtained from Gambella region Meteorology Service Center and National Meteorology Authority head office. Univariate, bivariate and multivariate analysis techniques were used to analyze the data. Results For the twelve-year time period, the mean annual total malaria case count in the district was 7369.58. The peak monthly malaria incidence was about 57 cases per 1000 people. Only in 2009 and 2015 that zero death due to malaria was recorded over the past 12 years. Fluctuating pattern of impatient malaria cases occurrence was seen over the past twelve years with an average number of 225.5 inpatient cases. The data showed that there is a high burden of malaria in the district. Plasmodium falciparum (Pf) was a predominant parasite species in the district with the maximum percentage of about 90. There was no statistically significant association between season and total malaria case number (F3,8: 1.982, P:0.195). However, the inter-annual total case count difference was statistically significant (F11,132: 36.305, p

Abstract Background Aedes (Stegomyia) albopictus (Skuse, 1894) is the main vector of dengue virus in China. The resistance to insecticides is a huge obstacle for the control of this species, and determining its resistance status and mechanisms in China is essential for the implementation of vector management strategies. Methods We have investigated the larval and adult resistance status of Ae. albopictus to deltamethrin in eight field populations in China. Mutations at the voltage-gated sodium channel gene, related to the knockdown resistance (kdr) effect, were detected by sequencing of PCR products. The eight field populations were examined for pyrethroid resistance using the World Health Organization standard bioassays, and the association between the mutations and phenotypic resistance was tested. Results The eight field populations of larvae of Ae. albopictus in China exhibited high resistance to deltamethrin; the RR50 values ranged from 12 (ZJ) to 44 (GZ). Adult bioassay revealed that Ae. albopictus populations were resistant to deltamethrin (mortality rate

Abstract Background Environmental conditions affect the biology of mosquito vectors. Aedes albopictus is a major vector of many important diseases including dengue, Zika, and chikungunya in China. Understanding the development, fecundity, and survivorship of Ae. albopictus mosquitoes in different environmental conditions is beneficial for the implementation of effective vector control measures. Methods Aedes albopictus larval and adult life-table experiments were conducted under natural conditions in indoor, half-shaded, and fully shaded settings, simulating the three major habitat types in Hainan Province, a tropical island in the South China Sea. Temperature, humidity, and light intensity were recorded daily. Larval rearing used habitat water and tap water, with and without additional artificial food. Development time, survivorship, pupation rate, and adult emergence rates were monitored. Adult mosquito survivorship and fecundity were monitored daily and reproductive rates were determined, and age-dependent survivorship and reproduction were analyzed. Results The pupation time and male and female emergence times were significantly shorter in indoor conditions than in shaded and half-shaded conditions for both tap water with added food and habitat water with added food groups. For habitat water with added food, the shaded environment had the lowest pupation rate among the settings. For tap water with added food group, the shaded environment had the lowest pupation rate. The mean survival time of females was 27.3 ± 0.8 days in the indoor condition, which was significantly longer than that in the half-shaded (18.4 ± 0.6 days) and shaded (13.8 ± 1.2 days) conditions. Adult mortality was age-dependent, and the rate of change in mortality with age was not significantly different among different environmental conditions. The mean net replacement rate (R 0) of female mosquitoes showed no significant difference among the three conditions, whereas the per capita intrinsic growth rate (r) in the shaded condition was 42.0% and 20.4% higher than that in the indoor and half-shaded conditions, respectively. Female daily egg mass was also age-dependent in all the settings, decaying exponentially with age. Conclusions Our results imply that half-shaded conditions are likely the best natural condition for adult emergence and female reproduction, and food supply is crucial for larval development and pupation. The results provide new avenues for integrated mosquito management in indoor and outdoor areas, especially in half-shaded areas. Graphical Abstract

IntroductionSince the second half of the 20th century, Aedes albopictus, a vector for more than 20 arboviruses, has spread worldwide. Aedes albopictus is the main vector of infectious diseases transmitted by Aedes mosquitoes in China, and it has caused concerns regarding public health. A comprehensive understanding of the spatial genetic structure of this vector species at a genomic level is essential for effective vector control and the prevention of vector-borne diseases.MethodsDuring 2016–2018, adult female Ae. albopictus mosquitoes were collected from eight different geographical locations across China. Restriction site-associated DNA sequencing (RAD-seq) was used for high-throughput identification of single nucleotide polymorphisms (SNPs) and genotyping of the Ae. albopictus population. The spatial genetic structure was analyzed and compared to those exhibited by mitochondrial cytochrome c oxidase subunit 1 (cox1) and microsatellites in the Ae. albopictus population.ResultsA total of 9,103 genome-wide SNP loci in 101 specimens and 32 haplotypes of cox1 in 231 specimens were identified in the samples from eight locations in China. Principal component analysis revealed that samples from Lingshui and Zhanjiang were more genetically different than those from the other locations. The SNPs provided a better resolution and stronger signals for novel spatial population genetic structures than those from the cox1 data and a set of previously genotyped microsatellites. The fixation indexes from the SNP dataset showed shallow but significant genetic differentiation in the population. The Mantel test indicated a positive correlation between genetic distance and geographical distance. However, the asymmetric gene flow was detected among the populations, and it was higher from south to north and west to east than in the opposite directions.ConclusionsThe genome-wide SNPs revealed seven gene pools and fine spatial genetic structure of the Ae. albopictus population in China. The RAD-seq approach has great potential to increase our understanding of the spatial dynamics of population spread and establishment, which will help us to design new strategies for controlling vectors and mosquito-borne diseases.

Abstract 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

Abstract Background Malaria causes significant mortality and morbidity in sub-Saharan Africa, especially among children under five years of age and places a huge economic burden on individuals and health systems. While this burden has been assessed previously, few studies have explored how malaria comorbidities affect inpatient costs. This study in a malaria endemic area in Western Kenya, assessed the total treatment costs per malaria episode including comorbidities in children and adults. Methods Total economic costs of malaria hospitalizations were calculated from a health system and societal perspective. Patient-level data were collected from patients admitted with a malaria diagnosis to a county-level hospital between June 2016 and May 2017. All treatment documented in medical records were included as health system costs. Patient and household costs included direct medical and non-medical expenses, and indirect costs due to productivity losses. Results Of the 746 patients admitted with a malaria diagnosis, 64% were female and 36% were male. The mean age was 14 years (median 7 years). The mean length of stay was three days. The mean health system cost per patient was Kenyan Shilling (KSh) 4288 (USD 42.0) (95% confidence interval (CI) 95% CI KSh 4046–4531). The total household cost per patient was KSh 1676 (USD 16.4) (95% CI KSh 1488–1864) and consisted of: KSh 161 (USD1.6) medical costs; KSh 728 (USD 7.1) non-medical costs; and KSh 787 (USD 7.7) indirect costs. The total societal cost (health system and household costs) per patient was KSh 5964 (USD 58.4) (95% CI KSh 5534–6394). Almost a quarter of patients (24%) had a reported comorbidity. The most common malaria comorbidities were chest infections, diarrhoea, and anaemia. The inclusion of comorbidities compared to patients with-out comorbidities led to a 46% increase in societal costs (health system costs increased by 43% and patient and household costs increased by 54%). Conclusions The economic burden of malaria is increased by comorbidities which are associated with longer hospital stays and higher medical costs to patients and the health system. Understanding the full economic burden of malaria is critical if future malaria control interventions are to protect access to care, especially by the poor.

Cutaneous leishmaniasis (CL) is a notifiable disease in Sri Lanka with increasing case numbers reported from every part of the country. In addition to disease treatment and vector control measures, knowledge and perceptions in a community are key contributors to a successful intervention program. An island-wide survey was carried out to assess the knowledge and perceptions regarding CL across the island, with 252 confirmed CL cases and 2,608 controls. Data was collected by trained personnel, using a pre-tested Case Reporting Form (CRF). Although the percentage who referred to CL by its correct name was low (1.4%), majority stated that it is a fly induced skin disease (79.1%). Knowledge on the symptoms, curability and the name of the vector was high in these communities, but specific knowledge on vector breeding places, biting times and preventive methods were poor. The patients were more knowledgeable when compared to the controls. Differences in the level of knowledge could be identified according to the level of education of the participants as well as across the different areas of the country. The main source of information was through the healthcare system, but the involvement of media in educating the communities on the disease was minimal. While this study population was unaccustomed to the use of repellants or sprays, the use of bed nets was high (77.7% of the participants) in this study population. Although misconceptions and incorrect practices are rare in Sri Lankan communities, promoting health education programs which may improve disease awareness and knowledge on vector and its control will further strengthen the control and prevention strategies. Author summary Cutaneous leishmaniasis (CL) is a disease which causes skin ulcers and spread by the sand fly. In recent years, the number of CL patients reported from all parts Sri Lanka increased notably, indicating that the spread of the disease should be controlled. The general publics’ knowledge on the disease and its control methods, is very important to implement a successful control program. Thus, this study was conducted to assess the level of knowledge of the local community on CL, its vector and control methods. The required information was collected by interviewing CL patients as well as healthy persons. More than 50% of both groups gave correct answers to more general questions about the disease such as the disease name, symptoms and spread, indicating that the awareness of CL among the public is satisfactory. However, the study also indicated that the patients were more knowledgeable when compared to the healthy persons. Furthermore, it was evident that knowledge on more detailed information such as vector breeding places, vector biting times is poor in both groups and the healthcare system has been the primary source of information.

Aedes albopictus (Ae. albopictus), an important vector of dengue virus (DENV), is distributed worldwide. Identifying host proteins involved in flavivirus replication in Ae. albopictus and determining their natural antiviral mechanisms are critical to control virus transmission. Revealing the key proteins related to virus replication and exploring the host-pathogen interaction are of great significance in finding new pathways of the natural immune response in Ae. albopictus. Isobaric tags for relative and absolute quantification (iTRAQ) was used to perform a comparative proteomic analysis between the midgut of Ae. albopictus infected with DENV and the control. 3,419 proteins were detected, of which 162 were ≥ 1.2-fold differentially upregulated or ≤ 0.8-fold differentially downregulated (p < 0.05) during DENV infections. Differentially expressed proteins (DEPs) were mainly enriched in ubiquitin ligase complex, structural constituent of cuticle, carbohydrate metabolism, and lipid metabolism pathways. We found that one of the DEPs, a putative pupal cuticle (PC) protein could inhibit the replication of DENV and interact with the DENV-E protein. In addition, the result of immunofluorescence (IF) test showed that there was co-localization between ubiquitin carboxyl-terminal hydrolase (UCH) protein and the DENV-E protein, and virus infection reduced the level of this protein. iTRAQ-based proteomic analysis of the Ae. albopictus midgut identified dengue infection-induced upregulated and downregulated proteins. The interaction between the PC and UCH proteins in the midgut of Ae. albopictus might exert a natural antiviral mechanism in mosquito.

Leishmaniasis is a neglected tropical disease with an annual estimated 700,000 to 1 million new cases globally. The diseases affect the rural community and people living in poverty. It is transmitted by female phlebotomine sand-fly; a tiny 2–3 mm long insect vector. Along with the regional countries, Bhutan has embarked on the elimination goal. However, several challenges lies ahead on its path to elimination. There is no elimination strategy and actions. The reporting, surveillance system, control and prevention mechanisms are inadequate. Further, there is lack of knowledge among the health care providers that impedes elimination goals. Therefore, if Bhutan is serious about its elimination goal, all the gaps and current challenges needs to be addressed appropriately. Keywords: Bhutan; Elimination; Leishmaniasis.

Abstract Background Malaria control in Kenya is based on case management and vector control using long-lasting insecticidal nets (LLINs) and indoor residual spraying (IRS). However, the development of insecticide resistance compromises the effectiveness of insecticide-based vector control programs. The use of pesticides for agricultural purposes has been implicated as one of the sources driving the selection of resistance. The current study was undertaken to assess the status and mechanism of insecticide resistance in malaria vectors in irrigated and non-irrigated areas with varying agrochemical use in western Kenya. Methods The study was carried out in 2018–2019 in Homa Bay County, western Kenya. The bioassay was performed on adults reared from larvae collected from irrigated and non-irrigated fields in order to assess the susceptibility of malaria vectors to different classes of insecticides following the standard WHO guidelines. Characterization of knockdown resistance (kdr) and acetylcholinesterase-inhibiting enzyme/angiotensin-converting enzyme (Ace-1) mutations within Anopheles gambiae s.l. species was performed using the polymerase chain reaction (PCR) method. To determine the agricultural and public health insecticide usage pattern, a questionnaire was administered to farmers, households, and veterinary officers in the study area. Results Anopheles arabiensis was the predominant species in the irrigated (100%, n = 154) area and the dominant species in the non-irrigated areas (97.5%, n = 162), the rest being An. gambiae sensu stricto. In 2018, Anopheles arabiensis in the irrigated region were susceptible to all insecticides tested, while in the non-irrigated region reduced mortality was observed (84%) against deltamethrin. In 2019, phenotypic mortality was decreased (97.8–84% to 83.3–78.2%). In contrast, high mortality from malathion (100%), DDT (98.98%), and piperonyl butoxide (PBO)-deltamethrin (100%) was observed. Molecular analysis of the vectors from the irrigated and non-irrigated areas revealed low levels of leucine-serine/phenylalanine substitution at position 1014 (L1014S/L1014F), with mutation frequencies of 1–16%, and low-frequency mutation in the Ace-1R gene (0.7%). In addition to very high coverage of LLINs impregnated with pyrethroids and IRS with organophosphate insecticides, pyrethroids were the predominant chemical class of pesticides used for crop and animal protection. Conclusion Anopheles arabiensis from irrigated areas showed increased phenotypic resistance, and the intensive use of pesticides for crop protection in this region may have contributed to the selection of resistance genes observed. The susceptibility of these malaria vectors to organophosphates and PBO synergists in pyrethroids offers a promising future for IRS and insecticide-treated net-based vector control interventions. These findings emphasize the need for integrated vector control strategies, with particular attention to agricultural practices to mitigate mosquito resistance to insecticides. Graphic abstract

Abstract An extensive network of storm water conveyance systems in urban areas, often referred to as the “underground storm drain system” (USDS), serves as significant production habitats for mosquitoes. Knowledge of whether USDS habitats are suitable for newly introduced dengue vectors Aedes aegypti and Ae. albopictus will help guide surveillance and control efforts. To determine whether the USDS functions as a suitable larval habitat for Culex, Ae. aegypti and Ae. albopictus in southern California, we examined mosquito habitat utilization and larval survivorship using laboratory microcosm studies. The data showed that USDS constituted 4.1% of sampled larval habitats for Ae. aegypti and Ae. albopictus, and 22.0% for Cx. quinquefasciatus. Furthermore, USDS water collected in the summer completely inhibited Aedes larval development, but yielded a 15.0% pupation rate for Cx. quinquefasciatus. Food supplementation in the microcosms suggests that nutrient deficiency, toxins and other factors in the USDS water led to low success or complete failure of larval development. These results suggest that USDS habitats are currently not major productive larval habitats for Aedes mosquitoes in southern California. Our findings prompt inclusion of assessments of pupal productivity in USDS habitats and adult mosquito resting sites in the mosquito surveillance program.

Abstract Larval source management has gained renewed interest as a malaria control strategy in Africa but the widespread and transient nature of larval breeding sites poses a challenge to its implementation. To address this problem, we propose combining an integrated high resolution (50 m) distributed hydrological model and remotely sensed data to simulate potential malaria vector aquatic habitats. The novelty of our approach lies in its consideration of irrigation practices and its ability to resolve complex ponding processes that contribute to potential larval habitats. The simulation was performed for the year of 2018 using ParFlow-Common Land Model (CLM) in a sugarcane plantation in the Oromia region, Ethiopia to examine the effects of rainfall and irrigation. The model was calibrated using field observations of larval habitats to successfully predict ponding at all surveyed locations from the validation dataset. Results show that without irrigation, at least half of the area inside the farms had a 40% probability of potential larval habitat occurrence. With irrigation, the probability increased to 56%. Irrigation dampened the seasonality of the potential larval habitats such that the peak larval habitat occurrence window during the rainy season was extended into the dry season. Furthermore, the stability of the habitats was prolonged, with a significant shift from semi-permanent to permanent habitats. Our study provides a hydrological perspective on the impact of environmental modification on malaria vector ecology, which can potentially inform malaria control strategies through better water management.

Abstract The issues of pyrethroid resistance and outdoor malaria parasite transmission have prompted the WHO to call for the development and adoption of viable alternative vector control methods. Larval source management is one of the core malaria vector interventions recommended by the Ministry of Health in many African countries, but it is rarely implemented due to concerns on its cost-effectiveness. New long-lasting microbial larvicide can be a promising cost-effective supplement to current vector control and elimination methods because microbial larvicide uses killing mechanisms different from pyrethroids and other chemical insecticides. It has been shown to be effective in reducing the overall vector abundance and thus both indoor and outdoor transmission. In our opinion, the long-lasting formulation can potentially reduce the cost of larvicide field application, and should be evaluated for its cost-effectiveness, resistance development, and impact on non-target organisms when integrating with other malaria vector control measures. In this opinion, we highlight that long-lasting microbial larvicide can be a potential cost-effective product that complements current front-line long-lasting insecticidal nets (LLINs) and indoor residual spraying (IRS) programs for malaria control and elimination. Microbial larviciding targets immature mosquitoes, reduces both indoor and outdoor transmission and is not affected by vector resistance to synthetic insecticides. This control method is a shift from the conventional LLINs and IRS programs that mainly target indoor-biting and resting adult mosquitoes.

Abstract Background Mosquitoes are vectors of many tropical diseases. Understanding the ecology of local mosquito vectors, such as species composition, distributions, population dynamics, and species diversity is important for designing the optimal strategy to control the mosquito-borne diseases. Methods Entomological surveillance of adult mosquitoes was conducted in five sites representing different ecological settings across Hainan Island from January to December of 2018 using BG Sentinel (BGS) traps and Centers for Disease Prevention and Control (CDC) light traps. In each site, we selected three areas representing urban, suburban and rural settings. Eighteen trap-days were sampled in each setting at each site, and CDC light traps and BGS traps were setup simultaneously. Mosquito species composition, distribution, population dynamics, and species diversity were analyzed. Mosquito densities were compared between different study sites and between different settings. Results Nine species of mosquitoes belonging to four genera were identified. Culex quinquefasciatus (80.8%), Armigeres subalbatus (13.0%) and Anopheles sinensis (3.1%) were the top three species collected by CDC light traps; Cx. quinquefasciatus (91.9%), Ae. albopictus (5.1%), and Ar. subalbatus (2.8%) were the top three species collected by BGS traps. Predominant species varied among study sites. The population dynamics of Ae. albopictus, An. sinensis and Cx. quinquefasciatus showed clear seasonal variation regardless of study sites with a varied peak season for different species. Mosquito abundance of all species showed significant differences among different study sites and among urban, suburban and rural areas. Danzhou had the highest mosquito biodiversity, with an α, β, and Gini-Simpson biodiversity index of 8, 1.13 and 0.42, respectively. BGS traps captured Aedes mosquito at a higher efficiency than CDC light traps, whereas CDC light traps captured significantly more Anopheles and Armigeres mosquitoes than BGS traps. Conclusions Mosquitoes were abundant on Hainan Island with clear seasonality and spatial heterogeneity. Population density, species composition, distribution, and species diversity were strongly affected by the natural environment. Different tools are required for the surveillance of different mosquito species.

Anopheles sinensis is a major malaria vector in Southeast Asia. Resistance to pyrethroid insecticides in this species has impeded malaria control in the region. Previous studies found that An. sinensis populations from Yunnan Province, China were highly resistant to deltamethrin and did not carry mutations in the voltage-gated sodium channel gene that cause knockdown resistance. In this study, we tested the hypothesis that other genomic variants are associated with the resistance phenotype. Using paired-end whole genome sequencing (DNA-seq), we generated 108 Gb of DNA sequence from deltamethrin -resistant and -susceptible mosquito pools with an average coverage of 83.3× depth. Using a stringent filtering method, we identified a total of 916,926 single nucleotide variants (SNVs), including 32,240 non-synonymous mutations. A total of 958 SNVs differed significantly in allele frequency between deltamethrin -resistant and -susceptible mosquitoes. Of these, 43 SNVs were present within 37 genes that code for immunity, detoxification, cuticular, and odorant proteins. A subset of 12 SNVs were randomly selected for genotyping of individual mosquitoes by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) and showed consistent allele frequencies with the pooled DNA-seq derived allele frequencies. In addition, copy number variations (CNVs) were detected in 56 genes, including 33 that contained amplification alleles and 23 that contained deletion alleles in resistant mosquitoes compared to susceptible mosquitoes. The genomic variants described here provide a useful resource for future studies on the genetic mechanism of insecticide resistance in this important malaria vector species.

Abstract Background In the past two decades, the massive scale-up of long-lasting insecticidal nets (LLINs) and indoor residual spraying (IRS) has led to significant reductions in malaria mortality and morbidity. Nonetheless, the malaria burden remains high, and a dozen countries in Africa show a trend of increasing malaria incidence over the past several years. This underscores the need to improve the effectiveness of interventions by optimizing first-line intervention tools and integrating newly approved products into control programs. Because transmission settings and vector ecologies vary from place to place, malaria interventions should be adapted and readapted over time in response to evolving malaria risks. An adaptive approach based on local malaria epidemiology and vector ecology may lead to significant reductions in malaria incidence and transmission risk. Methods/design This study will use a longitudinal block-cluster sequential multiple assignment randomized trial (SMART) design with longitudinal outcome measures for a period of 3 years to develop an adaptive intervention for malaria control in western Kenya, the first adaptive trial for malaria control. The primary outcome is clinical malaria incidence rate. This will be a two-stage trial with 36 clusters for the initial trial. At the beginning of stage 1, all clusters will be randomized with equal probability to either LLIN, piperonyl butoxide-treated LLIN (PBO Nets), or LLIN + IRS by block randomization based on their respective malaria risks. Intervention effectiveness will be evaluated with 12 months of follow-up monitoring. At the end of the 12-month follow-up, clusters will be assessed for “response” versus “non-response” to PBO Nets or LLIN + IRS based on the change in clinical malaria incidence rate and a pre-defined threshold value of cost-effectiveness set by the Ministry of Health. At the beginning of stage 2, if an intervention was effective in stage 1, then the intervention will be continued. Non-responders to stage 1 PBO Net treatment will be randomized equally to either PBO Nets + LSM (larval source management) or an intervention determined by an enhanced reinforcement learning method. Similarly, non-responders to stage 1 LLIN + IRS treatment will be randomized equally to either LLIN + IRS + LSM or PBO Nets + IRS. There will be an 18-month evaluation follow-up period for stage 2 interventions. We will monitor indoor and outdoor vector abundance using light traps. Clinical malaria will be monitored through active case surveillance. Cost-effectiveness of the interventions will be assessed using Q-learning. Discussion This novel adaptive intervention strategy will optimize existing malaria vector control tools while allowing for the integration of new control products and approaches in the future to find the most cost-effective malaria control strategies in different settings. Given the urgent global need for optimization of malaria control tools, this study can have far-reaching implications for malaria control and elimination. Trial registration US National Institutes of Health, study ID NCT04182126 . Registered on 26 November 2019.

Abstract Background Surveillance of outdoor host-seeking malaria vectors is crucial to monitor changes in vector biting behaviour and evaluate the impact of vector control interventions. Human landing catch (HLC) has been considered the most reliable and gold standard surveillance method to estimate human-biting rates. However, it is labour-intensive, and its use is facing an increasing ethical concern due to potential risk of exposure to infectious mosquito bites. Thus, alternative methods are required. This study was conducted to evaluate the performance of human-odour-baited CDC light trap (HBLT) and human-baited double net trap (HDNT) for outdoor host-seeking malaria vector surveillance in Kenya and Ethiopia. Methods The sampling efficiency of HBLT and HDNT was compared with CDC light trap and HLC using Latin Square Design in Ahero and Iguhu sites, western Kenya and Bulbul site, southwestern Ethiopia between November 2015 and December 2018. The differences in Anopheles mosquito density among the trapping methods were compared using generalized linear model. Results Overall, 16,963 female Anopheles mosquitoes comprising Anopheles gambiae sensu lato (s.l.), Anopheles funestus s.l., Anopheles pharoensis, Anopheles coustani and Anopheles squamosus were collected. PCR results (n = 552) showed that Anopheles arabiensis was the only member of An. gambiae s.l. in Ahero and Bulbul, while 15.7% An. arabiensis and 84.3% An. gambiae sensu stricto (s.s.) constituted An. gambiae s.l. in Iguhu. In Ahero, HBLT captured 2.23 times as many An. arabiensis and 2.11 times as many An. funestus as CDC light trap. In the same site, HDNT yielded 3.43 times more An. arabiensis and 3.24 times more An. funestus than HBLT. In Iguhu, the density of Anopheles mosquitoes did not vary between the traps (p > 0.05). In Bulbul, HBLT caught 2.19 times as many An. arabiensis as CDC light trap, while HDNT caught 6.53 times as many An. arabiensis as CDC light trap. The mean density of An. arabiensis did not vary between HDNT and HLC (p = 0.098), whereas the HLC yielded significantly higher density of An. arabiensis compared to HBLT and CDC light trap. There was a significant density-independent positive correlation between HDNT and HLC (r = 0.69). Conclusion This study revealed that both HBLT and HDNT caught higher density of malaria vectors than conventional CDC light trap. Moreover, HDNT yielded a similar vector density as HLC, suggesting that it could be an alternative tool to HLC for outdoor host-seeking malaria vector surveillance.

Leishmaniasis, a neglected tropical disease, is on the decline in South Asia. However, cases of cutaneous leishmaniasis have risen in Sri Lanka since 2001, and the lack of in-depth research on its epidemiologic characteristics hampers control efforts. We analyzed data collected from patients with cutaneous leishmaniasis in Sri Lanka during 2001–2018 to study temporal and geographic trends and identify and monitor disease hotspots. We noted a progression in case rates, including a sharp rise in 2018, showing temporal expansion of disease-prevalent areas and 2 persistent hotspots. The northern hotspot shifted and shrank over time, but the southern hotspot progressively expanded and remained spatially static. In addition, we noted regional incidence differences for age and sex. We provide evidence of temporally progressive and spatially expanding incidence of leishmaniasis in Sri Lanka with distinct geographic patterns and disease hotspots, signaling an urgent need for effective disease control interventions.

BackgroundPersons with submicroscopic malaria infection are a major reservoir of gametocytes that sustain malaria transmission in sub-Saharan Africa. Despite recent decreases in the national malaria burden in Kenya due to vector control interventions, malaria transmission continues to be high in western regions of the country bordering Lake Victoria. The objective of this study was to advance knowledge of the topographical, demographic and behavioral risk factors associated with submicroscopic malaria infection in the Lake Victoria basin in Kisumu County.MethodsCross-sectional community surveys for malaria infection were undertaken in three eco-epidemiologically distinct zones in Nyakach sub-County, Kisumu. Adjacent regions were topologically characterized as lakeshore, hillside and highland plateau. Surveys were conducted during the 2019 and 2020 wet and dry seasons. Finger prick blood smears and dry blood spots (DBS) on filter paper were collected from 1,777 healthy volunteers for microscopic inspection and real time-PCR (RT-PCR) diagnosis of Plasmodium infection. Persons who were PCR positive but blood smear negative were considered to harbor submicroscopic infections. Topographical, demographic and behavioral risk factors were correlated with community prevalence of submicroscopic infections.ResultsOut of a total of 1,777 blood samples collected, 14.2% (253/1,777) were diagnosed as submicroscopic infections. Blood smear microscopy and RT-PCR, respectively, detected 3.7% (66/1,777) and 18% (319/1,777) infections. Blood smears results were exclusively positive for P. falciparum, whereas RT-PCR also detected P. malariae and P. ovale mono- and co-infections. Submicroscopic infection prevalence was associated with topographical variation (χ2 = 39.344, df = 2, pConclusionTopographic features of the local landscape and seasonality are major correlates of submicroscopic malaria infection in the Lake Victoria area of western Kenya. Diagnostic tests more sensitive than blood smear microscopy will allow for monitoring and targeting geographic sites where additional vector interventions are needed to reduce malaria transmission.

The susceptibility of Asian tiger mosquitoes to DENV-2 in different seasons was observed in simulated field environments as a reference to design dengue fever control strategies in Guangzhou. The life table experiments of mosquitoes in four seasons were carried out in the field. The susceptibility of Ae. albopictus to dengue virus was observed in both environments in Guangzhou in summer and winter. Ae. albopictus was infected with dengue virus by oral feeding. On day 7 and 14 after infection, the viral load in the head, ovary, and midgut of the mosquito was detected using real-time fluorescent quantitative PCR. Immune-associated gene expression in infected mosquitoes was performed using quantitative real-time reverse transcriptase PCR. The hatching rate and pupation rate of Ae. albopictus larvae in different seasons differed significantly. The winter hatching rate of larvae was lower than that in summer, and the incubation time was longer than in summer. In the winter field environment, Ae. albopictus still underwent basic growth and development processes. Mosquitoes in the simulated field environment were more susceptible to DENV-2 than those in the simulated laboratory environment. In the midgut, viral RNA levels on day 7 in summer were higher than those on day 7 in winter (F = 14.459, P = 0.01); ovarian viral RNA levels on day 7 in summer were higher than those on day 7 in winter (F = 8.656, P < 0.001), but there was no significant difference in the viral load at other time points (P > 0.05). Dicer-2 mRNA expression on day 7 in winter was 4.071 times than that on day 7 in summer: the viral load and Dicer-2 expression correlated moderately. Ae. albopictus could still develop and transmit dengue virus in winter in Guangzhou. Mosquitoes under simulated field conditions were more susceptible to DENV-2 than those under simulated laboratory conditions.

Abstract Background Aedes albopictus is a highly invasive mosquito species and a major vector of numerous viral pathogens. Many recent dengue fever outbreaks in China have been caused solely by the vector. Mapping of the potential distribution ranges of Ae. albopictus is crucial for epidemic preparedness and the monitoring of vector populations for disease control. Climate is a key factor influencing the distribution of the species. Despite field studies indicating seasonal population variations, very little modeling work has been done to analyze how environmental conditions influence the seasonality of Ae. albopictus. The aim of the present study was to develop a model based on available observations, climatic and environmental data, and machine learning methods for the prediction of the potential seasonal ranges of Ae. albopictus in China. Methods We collected comprehensive up-to-date surveillance data in China, particularly records from the northern distribution margin of Ae. albopictus. All records were assigned long-term (1970–2000) climatic data averages based on the WorldClim 2.0 data set. Machine learning regression tree models were developed using a 10-fold cross-validation method to predict the potential seasonal (or monthly) distribution ranges of Ae. albopictus in China at high resolution based on environmental conditions. The models were assessed based on sensitivity, specificity, and accuracy, using area under curve (AUC). WorldClim 2.0 and climatic and environmental data were used to produce environmental conduciveness (probability) prediction surfaces. Predicted probabilities were generated based on the averages of the 10 models. Results During 1998–2017, Ae. albopictus was observed at 200 out of the 242 localities surveyed. In addition, at least 15 new Ae. albopictus occurrence sites lay outside the potential ranges that have been predicted using models previously. The average accuracy was 98.4% (97.1–99.5%), and the average AUC was 99.1% (95.6–99.9%). The predicted Ae. albopictus distribution in winter (December–February) was limited to a small subtropical-tropical area of China, and Ae. albopictus was predicted to occur in northern China only during the short summer season (usually June–September). The predicted distribution areas in summer could reach northeastern China bordering Russia and the eastern part of the Qinghai-Tibet Plateau in southwestern China. Ae. albopictus could remain active in expansive areas from central to southern China in October and November. Conclusions Climate and environmental conditions are key factors influencing the seasonal distribution of Ae. albopictus in China. The areas predicted to potentially host Ae. albopictus seasonally in the present study could reach northeastern China and the eastern slope of the Qinghai-Tibet Plateau. Our results present new evidence and suggest the expansion of systematic vector population monitoring activities and regular re-assessment of epidemic risk potential.

Abstract Background The Asian tiger mosquito, Aedes albopictus, is one of the 100 worst invasive species in the world and the vector for several arboviruses including dengue, Zika and chikungunya viruses. Understanding the population spatial genetic structure, migration, and gene flow of vector species is critical to effectively preventing and controlling vector-borne diseases. Little is known about the population structure and genetic differentiation of native Ae. albopictus in China. The aim of this study was to examine the patterns of the spatial genetic structures of native Ae. albopictus populations, and their relationship to dengue incidence, on a large geographical scale. Methods During 2016–2018, adult female Ae. albopictus mosquitoes were collected by human landing catch (HLC) or human-bait sweep-net collections in 34 localities across China. Thirteen microsatellite markers were used to examine the patterns of genetic diversity, population structure, and gene flow among native Ae. albopictus populations. The correlation between population genetic indices and dengue incidence was also examined. Results A total of 153 distinct alleles were identified at the 13 microsatellite loci in the tested populations. All loci were polymorphic, with the number of distinct alleles ranging from eight to sixteen. Genetic parameters such as PIC, heterozygosity, allelic richness and fixation index (F ST) revealed highly polymorphic markers, high genetic diversity, and low population genetic differentiation. In addition, Bayesian analysis of population structure showed two distinct genetic groups in southern-western and eastern-central-northern China. The Mantel test indicated a positive correlation between genetic distance and geographical distance (R 2 = 0.245, P = 0.01). STRUCTURE analysis, PCoA and GLS interpolation analysis indicated that Ae. albopictus populations in China were regionally clustered. Gene flow and relatedness estimates were generally high between populations. We observed no correlation between population genetic indices of microsatellite loci in Ae. albopictus populations and dengue incidence. Conclusion Strong gene flow probably assisted by human activities inhibited population differentiation and promoted genetic diversity among populations of Ae. albopictus. This may represent a potential risk of rapid spread of mosquito-borne diseases. The spatial genetic structure, coupled with the association between genetic indices and dengue incidence, may have important implications for understanding the epidemiology, prevention, and control of vector-borne diseases.

Abstract Background The Asian tiger mosquito, Aedes albopictus, an increasingly relevant arboviral vector, has spread worldwide. However, currently available tools are limited in terms of effective monitoring of vector populations and accurate determination of the extent of viral transmission, especially before and during outbreaks. Therefore, it is essential to develop novel monitoring and surveillance tools, particularly those that target adult mosquitoes and enhance the trapping efficiency for Ae. albopictus. Methods A variety of human body odorants associated with different types of mosquito olfactory receptors were selected, and their attractiveness to Ae. albopictus was tested by a four-arm olfactometer. The optimal compatibility and proportion of the odorants, Mix-5, was observed via orthogonal design analyses. The attractiveness of Mix-5 to Ae. albopictus in the laboratory was assessed using Mosq-ovitraps and Electric Mosquito Killers. In the field, the effectiveness of generic BG-Lure, Mix-5 and a control treatment was compared with a baited Biogents Sentinel trap (BGS-trap) using a Latin square design. Results In the olfactometer experiments, the attractiveness of the selected candidate compounds at varying dilutions was poor when the individual compounds were used alone. The optimal combination, Mix-5, was generated based on orthogonal design analyses. In the laboratory, the average numbers of female Ae. albopictus mosquitoes attracted by the synthetic odorant blend Mix-5 were 27.00 and 27.50, compared with 12.00 and 14.83 for the control, when using Mosq-ovitraps and Electric Mosquito Killers, respectively. In the field, the average number of Ae. albopictus female mosquitoes trapped by Mix-5 was 9.67 females/trap, whereas the average numbers for BG-Lure and the control were 7.78 and 4.47, respectively. The lure also played an important role in attracting Culex quinquefasciatus mosquitoes, and the average numbers of Cx. quinquefasciatus female mosquitoes attracted by Mix-5, BG-Lure and the control were 18.78, 25.11 and 12.22, respectively. Conclusions A human odor-based bait blend was developed and exhibited enhanced effectiveness at attracting Ae. albopictus This blend can be used to monitor and trap dengue vector mosquitoes in Chinese cities.

Abstract Background Zika virus (ZIKV) is an emerging mosquito-borne arbovirus that can produce serious public health consequences. In 2016, ZIKV caused an epidemic in many countries around the world, including the United States. ZIKV surveillance and vector control is essential to combating future epidemics. However, challenges relating to the timely publication of case reports significantly limit the effectiveness of current surveillance methods. In many countries with poor infrastructure, established systems for case reporting often do not exist. Previous studies investigating the H1N1 pandemic, general influenza and the recent Ebola outbreak have demonstrated that time- and geo-tagged Twitter data, which is immediately available, can be utilized to overcome these limitations. Methods In this study, we employed a recently developed system called Cloudberry to filter a random sample of Twitter data to investigate the feasibility of using such data for ZIKV epidemic tracking on a national and state (Florida) level. Two auto-regressive models were calibrated using weekly ZIKV case counts and zika tweets in order to estimate weekly ZIKV cases 1 week in advance. Results While models tended to over-predict at low case counts and under-predict at extreme high counts, a comparison of predicted versus observed weekly ZIKV case counts following model calibration demonstrated overall reasonable predictive accuracy, with an R2 of 0.74 for the Florida model and 0.70 for the U.S. model. Time-series analysis of predicted and observed ZIKV cases following internal cross-validation exhibited very similar patterns, demonstrating reasonable model performance. Spatially, the distribution of cumulative ZIKV case counts (local- & travel-related) and zika tweets across all 50 U.S. states showed a high correlation (r = 0.73) after adjusting for population. Conclusions This study demonstrates the value of utilizing Twitter data for the purposes of disease surveillance. This is of high value to epidemiologist and public health officials charged with protecting the public during future outbreaks.

Abstract Background Understanding the complex heterogeneity of risk factors that can contribute to an increased risk of malaria at the individual and household level will enable more effective use of control measures. The objective of this study was to understand individual and household factors that influence clinical malaria infection among individuals in the highlands of Western Kenya. Methods This was a matched case–control study undertaken in the Western Kenya highlands. Clinical malaria cases were recruited from health facilities and matched to asymptomatic individuals from the community who served as controls. Each participant was screened for malaria using microscopy. Follow-up surveys were conducted with individual households to collect socio-economic data. The houses were also checked using pyrethrum spray catches to collect mosquitoes. Results A total of 302 malaria cases were matched to 604 controls during the surveillance period. Mosquito densities were similar in the houses of both groups. A greater percentage of people in the control group (64.6%) used insecticide-treated bed nets (ITNs) compared to the families of malaria cases (48.3%). Use of ITNs was associated with lower level of clinical malaria episodes (odds ratio 0.51; 95% CI 0.39–0.68; P

Abstract Background Surveillance of outdoor resting malaria vector populations is crucial to monitor possible changes in vector resting and feeding behaviour following the widespread use of indoor-based vector control interventions. However, it is seldom included in the routine vector surveillance system in Africa due to lack of well standardized and efficient traps. This study was conducted to evaluate the performance of sticky pots for outdoor resting malaria vector surveillance in western Kenya. Methods Mosquito collections were conducted from September 2015 to April 2016 in Ahero and Iguhu sites, western Kenya using sticky pots, pit shelters, clay pots, exit traps, Prokopack aspirator and CDC light traps (outdoor and indoor). Species within Anopheles gambiae (s.l.) were identified using polymerase chain reaction (PCR). Enzyme-linked immunosorbent assay (ELISA) was used to determine blood meal sources of malaria vectors. Results A total of 23,772 mosquitoes were collected, of which 13,054 were female anophelines comprising An. gambiae (s.l.) (72.9%), An. funestus (13.2%), An. coustani (8.0%) and An. pharoensis (5.9%). Based on PCR assay (n = 672), 98.6% An. arabiensis and 1.4% An. gambiae (s.s.) constituted An. gambiae (s.l.) in Ahero, while this was 87.2% An. gambiae (s.s.) and 12.8% An. arabiensis in Iguhu. The sticky pots and pit shelters showed similar performance with regard to the relative abundance and host blood meal indices of An. gambiae (s.l.) and An. funestus. In terms of density per trap, a pit shelter caught on average 4.02 (95% CI: 3.06–5.27) times as many An. gambiae (s.l.) as a sticky pot, while a sticky pot captured 1.60 (95% CI: 1.19–2.12) times as many An. gambiae (s.l.) as a clay pot. Exit traps yielded a significantly lower number of An. gambiae (s.l.) than all other traps in Ahero, but a higher number of An. gambiae (s.l.) compared to the other outdoor traps in Iguhu. Indoor CDC light traps captured a significantly higher number of An. funestus than other traps. Conclusions Sticky pots could be a useful and complementary tool for outdoor resting malaria vector surveillance, in settings where using pit shelters is not feasible and less productive. The lower vector density in the sticky pots compared to pit shelters suggests that batches of sticky pots (i.e. four per compound) need to be deployed in order to make a direct comparison. This study also highlighted the need to concurrently undertake indoor and outdoor vector surveillance to better understand residual malaria transmission.

Abstract Background Physiological characteristics (age and blood feeding status) of malaria vectors can influence their susceptibility to the current vector control tools that target their feeding and resting behaviour. To ensure the sustainability of the current and future vector control tools an understanding of how physiological characteristics may contribute to insecticide tolerance in the field is fundamental for shaping resistance management strategies and vector control tools. The aim of this study was to determine whether blood meal and mosquito age affect pyrethroid tolerance in field-collected Anopheles gambiae from western Kenya. Methods Wild mosquito larvae were reared to adulthood alongside the pyrethroid-susceptible Kisumu strain. Adult females from the two populations were monitored for deltamethrin resistance when they were young at 2–5 days old and older 14–16 days old and whether fed or unfed for each age group. Metabolic assays were also performed to determine the level of detoxification enzymes. Mosquito specimens were further identified to species level using the polymerase chain reaction (PCR) method. Results Anopheles gambiae sensu stricto was the predominant species comprising 96% of specimens and 2.75% Anopheles arabiensis. Bioassay results showed reduced pyrethroid induced mortality with younger mosquitoes compared to older ones (mortality rates 83% vs. 98%), independently of their feeding status. Reduced mortality was recorded with younger females of which were fed compared to their unfed counterparts of the same age with a mortality rate of 35.5% vs. 83%. Older blood-fed females showed reduced susceptibility after exposure when compared to unfed females of the same age (mortality rates 86% vs. 98%). For the Kisumu susceptible population, mortality was straight 100% regardless of age and blood feeding status. Blood feeding status and mosquito age had an effect on enzyme levels in both populations, with blood fed individuals showing higher enzyme elevations compared to their unfed counterparts (P

BackgroundLeishmaniasis is a neglected tropical vector-borne disease, which is on the rise in Sri Lanka. Spatiotemporal and risk factor analyses are useful for understanding transmission dynamics, spatial clustering and predicting future disease distribution and trends to facilitate effective infection control.MethodsThe nationwide clinically confirmed cutaneous leishmaniasis and climatic data were collected from 2001 to 2019. Hierarchical clustering and spatiotemporal cross-correlation analysis were used to measure the region-wide and local (between neighboring districts) synchrony of transmission. A mixed spatiotemporal regression-autoregression model was built to study the effects of climatic, neighboring-district dispersal, and infection carryover variables on leishmaniasis dynamics and spatial distribution. Same model without climatic variables was used to predict the future distribution and trends of leishmaniasis cases in Sri Lanka.ResultsA total of 19,361 clinically confirmed leishmaniasis cases have been reported in Sri Lanka from 2001-2019. There were three phases identified: low-transmission phase (2001-2010), parasite population buildup phase (2011-2017), and outbreak phase (2018-2019). Spatially, the districts were divided into three groups based on similarity in temporal dynamics. The global mean correlation among district incidence dynamics was 0.30 (95% CI 0.25-0.35), and the localized mean correlation between neighboring districts was 0.58 (95% CI 0.42-0.73). Risk analysis for the seven districts with the highest incidence rates indicated that precipitation, neighboring-district effect, and infection carryover effect exhibited significant correlation with district-level incidence dynamics. Model-predicted incidence dynamics and case distribution matched well with observed results, except for the outbreak in 2018. The model-predicted 2020 case number is about 5,400 cases, with intensified transmission and expansion of high-transmission area. The predicted case number will be 9115 in 2022 and 19212 in 2025.ConclusionsThe drastic upsurge in leishmaniasis cases in Sri Lanka in the last few year was unprecedented and it was strongly linked to precipitation, high burden of localized infections and inter-district dispersal. Targeted interventions are urgently needed to arrest an uncontrollable disease spread.

Aedes albopictus is a vector of over 20 arboviruses that has spread throughout the world, mainly in the second half of the twentieth century. Approximately 50–100 million people are infected with dengue virus (DENV) transmitted by Aedes mosquitoes each year, leading to heavy economic burdens for both governments and individuals, among countless other negative consequences. Understanding the vector competence of vector species is critical for effectively preventing and controlling vector-borne diseases. Accordingly, in this study, vector competence was evaluated by quantitative analysis of DENV-2 loads in mosquito tissues (midguts, heads, and salivary glands) and whole mosquitoes through real-time quantitative polymerase chain reaction (RT-qPCR) analysis. Wolbachia and the expression of immune-associated genes (Rel1, Rel2, Dicer2, and STAT) in mosquitoes were also detected by RT-qPCR to explore their impact on vector competence. The amount of DENV-2 in the mosquito midguts, heads, and salivary glands from southern-western China were found to be lower than those from eastern-central-northern China. The DENV-2 loads in whole mosquitoes showed a negative correlation with Rel1 gene (r = -0.285, P = 0.011) and STAT gene expression levels (r = -0.289, P = 0.009). In terms of Wolbachia strains, the density of the wAlbB strain was found to be significantly higher than that of the wAlbA strain in the eight Ae. albopictus populations, and the relative density of the wAlbB strain in mosquitoes from southern-western China was higher than those from eastern-central-northern China. The relative density of the wAlbB strain showed a negative correlation with the mean loads of DENV-2 in the heads (r = -0.729, P = 0.040), salivary glands (r = -0.785, P = 0.021), and whole mosquitoes (r = -0.909, P = 0.002). Thus, there are lower DENV-2 loads in the mosquitoes from southern-western China, which may be related to the innate immunity of mosquitoes as affected by Rel1 in the Toll pathway, STAT in the JAK-STAT pathway, and the relative density of the wAlbB strain.

BackgroundAn. funestus is a major Afrotropical vector of human malaria. This study sought to investigate the larval ecology, sporozoite infection rates and blood meal sources of An. funestus in western Kenya.MethodsLarval surveys were carried out in Bungoma (Highland) and Kombewa (lowland) of western Kenya. Aquatic habitats were identified, characterized, georeferenced and carefully examined for mosquito larvae and predators. Indoor resting mosquitoes were sampled using pyrethrum spray catches. Adults and larvae were morphologically and molecularly identified to species. Sporozoite infections and blood meal sources were detected using real-time PCR and ELISA respectively.ResultsOf the 151 aquatic habitats assessed, 62/80 (78%) in Bungoma and 58/71(82%) in Kombewa were positive for mosquito larvae. Of the 3,193 larvae sampled, An. funestus larvae constitute 38% (1224/3193). Bungoma recorded a higher number of An. funestus larvae (85%, 95%, CI, 8.722-17.15) than Kombewa (15%, 95%, CI, 1.33-3.91). Molecular identification of larvae showed that 89% (n = 80) were An. funestus. Approximately 59%, 35% and 5% of An. funestus larvae co-existed with An. gambiae s.l, Culex spp and An. coustani in the same habitats respectively. Of 1,221 An. funestus s.l adults sampled, molecular identifications revealed that An. funestus constituted 87% (n = 201) and 88% (n = 179) in Bungoma and Kombewa, respectively. The Plasmodium falciparum sporozoite rate of An. funestus in Bungoma and Kombewa was 2% (3/174) and 1% (2/157), respectively, and the human blood index of An. funestus was 84% (48/57) and 89% (39/44) and for Bungoma and Kombewa, respectively.ConclusionMan-made ponds had the highest abundance of An. funestus larvae. Multiple regression and principal component analyses identified the distance to the nearest house as the key environmental factor associated with the abundance of An. funestus larvae in aquatic habitats. This study serves as a guide for the control of An. funestus and other mosquito species to complement existing vector control strategies.

Plasmodium falciparum parasites have evolved genetic adaptations to overcome immune responses mounted by diverse Anopheles vectors hindering malaria control efforts. Plasmodium falciparum surface protein Pfs47 is critical in the parasite's survival by manipulating the vector's immune system hence a promising target for blocking transmission in the mosquito. This study aimed to examine the genetic diversity, haplotype distribution, and population structure of Pfs47 and its implications on malaria infections in endemic lowlands in Western Kenya. Cross-sectional mass blood screening was conducted in malaria endemic regions in the lowlands of Western Kenya: Homa Bay, Kombewa, and Chulaimbo. Dried blood spots and slide smears were simultaneously collected in 2018 and 2019. DNA was extracted using Chelex method from microscopic Plasmodium falciparum positive samples and used to genotype Pfs47 using polymerase chain reaction (PCR) and DNA sequencing. Thirteen observed haplotypes of the Pfs47 gene were circulating in Western Kenya. Population-wise, haplotype diversity ranged from 0.69 to 0.77 and the nucleotide diversity 0.10 to 0.12 across all sites. All the study sites displayed negative Tajima's D values although not significant. However, the negative and significant Fu's Fs statistical values were observed across all the study sites, suggesting population expansion or positive selection. Overall genetic differentiation index was not significant (FST = -0.00891, P > 0.05) among parasite populations. All Nm values revealed a considerable gene flow in these populations. These results could have important implications for the persistence of high levels of malaria transmission and should be considered when designing potential targeted control interventions.

BACKGROUND:Aedes albopictus is a major vector for several tropical infectious diseases. Characterization of Ae. albopictus development under natural conditions is crucial for monitoring vector population expansion, dengue virus transmission, and disease outbreak preparedness. METHODS:This study employed mosquito traits as a proxy to understanding life-table traits in mosquitoes using a semi-field study. Ae. albopictus larval and adult life-table experiments were conducted using microcosms under semi-field conditions in Guangzhou. Stage-specific development times and survivorship rates were determined and compared under semi-field conditions in different seasons from early summer (June) to winter (January), to determine the lower temperature limit for larval development and adult survivorship and reproductivity. RESULTS:The average egg- hatching rate was 60.1%, with the highest recorded in October (77.1%; mid-autumn). The larval development time was on average 13.2 days (range, 8.5-24.1 days), with the shortest time observed in September(8.7 days; early autumn) and longest in November (22.8 days). The pupation rates of Ae. albopictus larvae were on average 88.9% (range, 81.6-93.4%); they were stable from June to September but decreased from October to November. The adult emergence rates were on average 82.5% (range, 76.8-87.9%) and decreased from July to November. The median survival time of Ae. albopictus adults was on average 7.4 (range, 4.5-9.8), with the shortest time recorded in September. The average lifetime egg mass under semi-field conditions was 37.84 eggs/female. The larvae could develop into adults at temperatures as low as 12.3°C, and the adults could survive for 30.0 days at 16.3°C and still produce eggs. Overall, correlation analysis found that mean temperature and relative humidity were variables significantly affecting larval development and adult survivorship. CONCLUSION:Ae. albopictus larvae could develop and emerge and the adults could survive and produce eggs in early winter in Guangzhou. The major impact of changes in ambient temperature, relative humidity, and light intensity was on the egg hatching rates, adult survival time, and egg mass production, rather than on pupation or adult emergence rates.