Dickson Wilson Lwetoijera

Background: Anopheles funestus , a major vector of malaria in Africa, has proven difficult to colonize in laboratory settings, impeding research on its biology and control. After several attempts, our team recently succeeded in colonizing a strain of An. funestus from Tanzania (FUTAZ). The objective of this study was to analyze the key fitness and genotypic characteristics of these mosquitoes during multiple filial generations of laboratory adaptation and compare them to wild An. funestus from Tanzania and a pre-existing colony of An. funestus from Mozambique (FUMOZ). Methods: Measures of mating success, body size, fecundity, and insecticide resistance were compared between the newly established FUTAZ colonies, the long-established FUMOZ colonies, and a colony of Anopheles arabiensis maintained in the same laboratory. The maternal lineages of the An. funestus mosquitoes were investigated through a hydrolysis probe analysis of their mitochondrial DNA to identify distinct clades. Other intragenomic variations were examined by D3-RFLP genotyping of 28S ribosomal DNA. These molecular markers were used to compare the FUTAZ colonies, FUMOZ colonies in Tanzania (TZ) and South African (SA) laboratories, and the wild-collected Tanzanian An. funestu s. Results: The mating success and body size of FUTAZ females declined significantly from filial generations F1 to F6 relative to the founder population (F0) then increased from F7 onwards, eventually matching FUMOZ by F9. Fecundity was similar across all colonies tested. However, it took significantly longer for 50% of the females in the FUTAZ and FUMOZ colonies (> 10 days) to mate compared to females in the An. arabiensis colony (~ 5 days). Resistance appeared to be lost during colonization, but this varied with insecticide classes. Majority of FUTAZ mosquitoes, and the wild-caught Tanzanian An. funestus belonged to Clade I (80.4–89.4%) and RFLP type “Y” (90.5–91.4%), while FUMOZ colonies in TZ and SA were mostly Clade II (65.5–88.5%) and RFLP type “MW” (90.5–91.5%). Conclusions: This study demonstrates that certain fitness traits of An. funestus , notably mating success and body size, declined during the initial phases of laboratory colonization but recovered as the population adapted to laboratory conditions. Having large founder populations is critical to survive these early-generation bottlenecks. Clade-specific and RFLP genotyping confirmed FUTAZ colony as genetically representative of wild-caught Tanzanian An. funestus . This newly established colony constitutes a valuable and reliable resource for malaria vector research.

Background: Residential window design is a primary determinant of natural ventilation in urban housing; however, the factors driving window design choices in rapidly urbanizing sub-Saharan African cities remain poorly understood. In Dar es Salaam Tanzania, sliding aluminum windows have become increasingly dominant in the residential housing market, yet evidence on how stakeholder perceptions of these designs relate to indoor ventilation adequacy, disease risk, and household energy use remains scarce. Method: This mixed-methods study was conducted in Dar es Salaam Tanzania, between July and October 2025. A structured questionnaire was administered to 611 participants (496 household respondents and 115 hardware store vendors), followed by two focus group discussions with housing stakeholders. Descriptive statistics (frequencies and proportions) summarized participant and household characteristics. Associations between window designs and self-reported diseases and use of artificial cooling systems were examined using chi-square tests and logistic regression, with statistical significance set at a p -value < 0.05. Focus group data were analyzed using thematic analysis, and a desktop review was done to assess the presence of multisectoral ventilation guidance. Results: Sliding aluminum windows dominated the market (57% of sales) and household installations (47%), driven primarily by aesthetic considerations (46%). Awareness of ventilation guidelines was low across all stakeholder groups (18%). Limited airflow was significantly associated with greater reliance on mechanical fans (χ ² (1) = 5.72, p = 0.017). Large household size was the strongest predictor of both malaria (OR = 2.65, 95% CI: 1.61–4.39, p < 0.001) and respiratory infections (OR = 3.74, 95% CI: 1.17–12.0, p = 0.026). Use of gas as cooking fuel was protective against respiratory infections (OR = 0.35, 95% CI: 0.14–0.85, p = 0.021). The desktop review revealed no enforceable benchmarks linking window design to ventilation performance, disease prevention, or energy efficiency in Tanzanian building regulations. Conclusion: Market dynamics and structural constraints shape window design choices in urban Dar es Salaam more than health-oriented performance standards. Bringing public health ideas into housing rules, creating better ventilation guidelines for specific areas, and raising community awareness could improve indoor air quality and reduce reliance on energy-intensive solutions in rapidly growing cities.

The integration of larviciding as a supplementary tool for malaria vector control requires effective larvicide formulations that are specifically suited to the target vector species, environmentally friendly, and cost-effective to reflect the needs of resource-limited settings in sub-Saharan Africa. This study evaluated the sublethal effect and its residual activity of two microencapsulated pyriproxyfen formulations containing 33% and 50% active ingredient (AI) against Anopheles arabiensis larvae exposed to sublethal doses. Sublethal effects on fitness parameters, including fecundity and wing length, were evaluated at concentrations that resulted in 20%, 50%, and 70% emergence inhibition, obtained from a dose-response curve. In three replicates across all three concentrations, fecundity was assessed in 25 adult females, while wing length was measured in 33 adult males and 33 adult females. Residual activity was assessed by exposing 200 3^rd-instar larvae of An. arabiensis to each formulation at 0.06 mg/L 33% AI and 0.09 mg/L 50% AI in 5 replicates of artificial habitats. Once all mosquitoes had either emerged or died, a new batch of larvae was introduced every 2 weeks for 6 months. Females emerged from concentrations inhibiting 70% of larval emergence exhibited significantly reduced fecundity; 88% for 33% AI [RR = 0.12, 95% CI: 0.11, 0.13, p < 0.001] and 85% for 50% AI [RR = 0.14, 95% CI: 0.07, 0.32, p < 0.001]. Adults emerged from sublethal concentrations had reduced body size, decreasing as concentration increased. Residual activity on emergence inhibition was 14.4% [13.2%, 15.6%], and 23.1% [18.7%, 27.5%], relative to 23% and 14% for 33% and 50% AI, respectively in the 6^th month. Control mortality was consistently below 10% during the entire evaluation period. These findings demonstrate the efficaciousness and long-lasting potential of microencapsulated pyriproxyfen formulations to target An. arabiensis, and highlight its consideration for application in larviciding programmes.

Background: Pyriproxyfen (PPF), a juvenile hormone analog, is a promising chemical for autodissemination strategies, where mosquitoes aid in transferring insecticides to their breeding sites. This study evaluated the effects of sublethal PPF doses on Anopheles arabiensis fitness (fecundity, fertility, and body size) and pyrethroids susceptibility across three generations. Methods: Laboratory-reared, pyrethroid-resistant mosquito larvae were exposed once to sublethal PPF concentrations of (0.0003, 0.0006, 0.0008, and 0.001 mg a.i./L), which caused emergence inhibition rates of 5%, 10%, 15%, and 20%, respectively, alongside a control group. Emerged adults were tested for susceptibility to 0.75% permethrin, 3.75% permethrin, and 0.05% deltamethrin using WHO tube bioassays. Knockdown was recorded at 1 h, mortality at 24 h post-exposure, while fecundity, fertility, and body size were measured across three generations. Results: First-generation Anopheles arabiensis emerging from PPF-treated larvae showed reduced susceptibility to 0.75% permethrin and 0.05% deltamethrin compared to controls. Mortality from permethrin dropped from 22.7% in the controls to 11% and then 3% at 0.0003 and 0.001 mg a.i./L. For deltamethrin, mortality fell from 62.7% to 36% and then 23.3% at the same concentrations. Knockdown at 60 min was also significantly reduced, with permethrin KDT₆₀ dropping from 41.3% in controls to 9.3% and 3.3%, and deltamethrin KDT₆₀ from 79.7% to 66.7% and 65%. No significant differences were observed in subsequent generations (p > 0.05). PPF exposure also induced notable fitness costs in the first generation: mean wing length decreased from 3.07 mm in controls to 2.88–2.66 mm (6–13% reduction), mean egg production dropped from 30.1 to 13.9–18.8 eggs per female (37–54% reduction), and egg hatching rate declined from 87% to 79–82% (6–9% reduction). Conclusions: These findings suggest that sublethal PPF doses can temporarily enhance insecticide resistance without leading to heritable resistance and negatively impact key mosquito fitness traits. PPF may thus be a valuable addition to integrated vector management strategies.

Gene drive technology presents a transformative approach to combatting malaria by introducing genetic modifications into wild mosquito populations to reduce their vectorial capacity. Although effective modifications have been developed, these efforts have been confined to laboratories in the global north. We previously demonstrated that modifying Anopheles gambiae to express two exogenous antimicrobial peptides inhibits the sporogonic development of laboratory-cultured Plasmodium falciparum , with models predicting substantial contributions to malaria elimination in Africa when integrated with gene drive ^1–3 . However, the effectiveness of this modification against genetically diverse, naturally circulating parasite isolates remained unknown. To address this critical gap, we adapted our technology for an African context by establishing infrastructural and research capacity in Tanzania, enabling the engineering of local A. gambiae under containment. Here we report the generation of a transgenic strain equipped with non-autonomous gene drive capabilities that robustly inhibits genetically diverse P. falciparum isolates obtained from naturally infected children. These genetic modifications were efficiently inherited by progeny when supplemented with Cas9 endonuclease provided by another locally engineered strain. Our work brings gene drive technology a critical step closer to application, providing a locally tailored and powerful tool for malaria eradication through the targeted dissemination of beneficial genetic traits in wild mosquito populations.

Background: Pyriproxyfen (PPF), a juvenile hormone analogue is a promising chemistry for autodissemination strategies, where mosquitoes aid in transferring insecticides to their breeding sites. This study evaluated the effects of sublethal PPF exposure on Anopheles arabiensis fitness (fecundity, fertility, and body size) and pyrethroids susceptibility across three generations. Methods: Laboratory-reared, pyrethroid-resistant mosquito larvae were exposed once to sublethal PPF concentrations of (0.0003, 0.0006, 0.0008, and 0.001 mg a.i./L), which caused emergence inhibition rates of 5%, 10%, 15%, and 20%, respectively, alongside a control group. Emerged adults were tested for susceptibility to 0.75% permethrin, 3.75% permethrin and 0.05% deltamethrin using WHO tube bioassays. Knockdown was recorded at 1 h, mortality at 24 h post-exposure, while fecundity, fertility, and body size were measured across three generations. Results: First-generation Anopheles arabiensis emerging from PPF-treated larvae showed reduced susceptibility to 0.75% permethrin and 0.05% deltamethrin compared to controls. Mortality from permethrin dropped from 22.7% in the controls to 11% and then 3% at 0.0003 and 0.001 mg a.i./L. For deltamethrin, mortality fell from 62.7% to 36% and then 23.3% at the same concentrations. Knockdown at 60 min was also significantly reduced, with permethrin KDT₆₀ dropping from 41.3% in controls to 9.3% and 3.3%, and deltamethrin KDT₆₀ from 79.7% to 66.7% and 65%. No significant differences were observed in subsequent generations (p > 0.05). PPF exposure also induced notable fitness costs in the first generation: mean wing length decreased from 3.07 mm in controls to 2.88–2.66 mm (6–13% reduction), mean egg production dropped from 30.1 to 13.9–18.8 eggs per female (37–54% reduction), and egg hatching rate declined from 87% to 79–82% (6–9% reduction). Conclusion: These findings suggest that sublethal PPF doses can temporarily enhance insecticide resistance without leading to heritable resistance and negatively impact key mosquito fitness traits. PPF may thus be a valuable addition to integrated vector management strategies.

Background: Semi-field experiments are an efficient way of assessing the impacts of potential new vector control tools (VCTs) before field trials. However, their design is critically important to ensure their results are unbiased and informative. An essential element of the design of semi-field experiments is power analysis, which empowers researchers to ensure that only designs with adequate statistical power are adopted. In this study, a methodology was developed, and its use was demonstrated in a tutorial, to determine the required number of semi-field chambers, sampling frequency and the number of mosquitoes required to achieve sufficient power for evaluating the impact of a single VCT or two in combination. Methods: By analysing data simulated from a generalized linear mixed-effects model, power was estimated for various experimental designs, including short- (24 h) vs. long-term (3 months) experiments and single vs. combined application of interventions (e.g., insecticide-treated nets combined with pyriproxyfen autodissemination). Results: Although power increased with increasing number of chambers, sampling frequency and the number of mosquitoes, the number of chambers and variance between chambers were the dominant factors determining power relative to all other design choices. High variance between chambers decreased power, highlighting the importance of making conditions similar among chambers, by reducing variation if possible and by rotating variables if not. As compared to a single intervention, an additional intervention required an increase in the number of chambers, while short and long experiments were similar in terms of key aspects such as the number of chambers per treatment. Conclusion: Determining the most efficient experimental design for a semi-field experiment will depend on a balance of design choices and resource constraints. The power analysis framework and tutorial provided here can aid in the robust design of these widely used experiments and ultimately facilitate the development of new vector control tools (VCTs).

Interspecific competition between mosquito larvae may affects adult vectorial capacity, potentially reducing disease transmission. It also influences population dynamics, and cannibalistic and predatory behaviors. However, knowledge of interspecific competition between Ae. aegypti and Anopheles species is limited. The study examined interspecific competition between Ae. aegypti larvae and either An.arabiensis, An. gambiae, or An. funestus on individual fitness in semi-field settings. The experiments involved density combinations of 100:100, 200:0, and 0:200 (Ae. aegypti: Anopheles), reared with and without food, in small habitats (8.5 cm height × 15 cm diameter) with 0.5 liters of water and large habitats (15 cm height × 35 cm diameter) with 1 liter of water. The first group received Tetramin^® fish food (0.02 g), while the second group was unfed to assess cannibalism and predation. While interspecific competition affected both genera, Anopheles species experienced greater effects, with reduced survival and delayed development, compared to Ae. aegypti. The mean wing lengths of all species were significantly small in small habitats with a mixed population (p < 0.001). The presence of food reduced cannibalism and predation compared to its absence. These interactions have implications for diseases transmission dynamics and can serve as biological indicators to signal the impacts of vector control interventions.

Interspecific competition between mosquito larvae may affects adult vectorial capacity, potentially reducing disease transmission. It also influences population dynamics, cannibalistic and predatory behaviors. However, knowledge of interspecific competition between Ae. aegypti and Anopheles species is limited. The study examined interspecific competition between Ae. aegypti larvae and either An. arabiensis, An. gambiae, or An. funestus on individual fitness in semi-field settings. The experiments involved density combinations of 100:100, 200:0, and 0:200 (Ae. aegypti: Anopheles), reared with and without food, in small habitat (8.5 cm height × 15 cm diameter) with 0.5 litre and medium habitats (15 cm height × 35 cm diameter) with 1 litre of water. The first group received Tetramin® fish food (0.02 g), while the second group was unfed to assess cannibalism and predation. While, interspecific competition affected both genera, Anopheles species experienced greater effect, with reduced survival and delayed development, compared to Ae. aegypti. The mean wing lengths of all species were significantly small in small habitats in mixed population (p < 0.001). The presence of food reduced cannibalism and predation compared to its absence. These interactions have implications for diseases transmission dynamics and can serve as biological indicators to signal the impacts of vector control interventions.