Background: Larval source management (LSM) was once central to malaria control before insecticide–treated nets and indoor residual spraying dominated. Renewed interest in LSM raises questions about its effectiveness in rural Africa, where habitats are dispersed, and vector species contribute unequally, and whether species–targeted larviciding could offer greater gains than broadcast approaches. Methods: This modelling study quantified the potential impact of larviciding in African settings where multiple vector species contribute unequally to malaria transmission. We modeled malaria transmission in southeastern Tanzania using agent–based simulations incorporating seasonal dynamics, insecticide resistance, and semi–field biolarvicide efficacy. Outcomes were entomological inoculation rate, malaria incidence in under–fives, and operational larviciding costs. Findings: Large–scale deployment of biolarvicides with >1–week residual activity substantially reduced malaria transmission, with disproportionately greater gains when control efforts were preferentially focused on the dominant vector species, Anopheles funestus , compared to broadcast approaches treating both An. funestus and An. arabiensis habitats. In the absence of ITNs, a four–month fortnightly larviciding campaign targeting An. funestus at 80% coverage reduced EIR by 58% and incidence by ~40%, versus ~55% incidence and ~70% EIR reductions under broadcast strategies; targeting An. arabiensis alone yielded ≤30% EIR and ≤13% incidence reductions. Starting with pre–existing 80% ITN coverage, funestus –targeted larviciding further reduced peak EIR by ~70% and incidence by ~77%, versus ~90% and ~85%, respectively, with broadcast strategies, suggesting broadcast larviciding provided limited additional reductions beyond those achieved by the funestus –targeted approach. At 40% ITN coverage, additional reductions were ~62% of EIR and ~46% in incidence ( funestus –targeted) versus ~76% and 63%, respectively (broadcast). The targeted campaigns preserved a 30–50% cost advantage while sustaining >50% dry–season transmission reductions. Finally, high–coverage (e.g., 80%) funestus –targeted larviciding campaigns achieved greater impacts than lower–coverage (e.g., 40–60%) targeting both species. Conclusions: In settings where multiple vector species contribute unequally to malaria transmission, preferentially targeting larviciding against the dominant vector species can deliver substantial epidemiological impact, with greater resource efficiency than broadcast approaches targeting multiple vectors. In Tanzania, where An. funestus drives most transmission; concentrating larviciding efforts on its characteristic aquatic habitats may offer a scalable, low–cost complement to established tools such as ITNs. ### Competing Interest Statement The authors have declared no competing interest. ### Funding Statement This work was supported in whole by the Bill & Melinda Gates Foundation [grant number OPP1214408 to FOO, Ifakara Health Institute]. Under the grant conditions of the Foundation, a Creative Commons Attribution 4.0 Generic License has already been assigned to the 783 Author Accepted Manuscript version that might arise from this submission. ### Author Declarations I confirm all relevant ethical guidelines have been followed, and any necessary IRB and/or ethics committee approvals have been obtained. Yes I confirm that all necessary patient/participant consent has been obtained and the appropriate institutional forms have been archived, and that any patient/participant/sample identifiers included were not known to anyone (e.g., hospital staff, patients or participants themselves) outside the research group so cannot be used to identify individuals. Yes I understand that all clinical trials and any other prospective interventional studies must be registered with an ICMJE-approved registry, such as ClinicalTrials.gov. I confirm that any such study reported in the manuscript has been registered and the trial registration ID is provided (note: if posting a prospective study registered retrospectively, please provide a statement in the trial ID field explaining why the study was not registered in advance). Yes I have followed all appropriate research reporting guidelines, such as any relevant EQUATOR Network research reporting checklist(s) and other pertinent material, if applicable. Yes All data supporting the main conclusions of this article are included within the article.

Background: Micronutrient deficiencies and physical inactivity can adversely affect child growth and development. This study assessed the effects of school-based physical activity and multi-micronutrient supplementation on micronutrient status among schoolchildren in Kilombero district, Tanzania. Methods: In a cluster-randomized trial, children aged 6-12 years were allocated to physical activity, multi-micronutrient supplementation, combined physical activity plus supplementation, or placebo control. Anthropometric and biochemical assessments were conducted at baseline, 14 months, and 26 months. Dried blood spot samples were available for 923 children at baseline. Complete-case analyses used biomarker-specific subsamples with valid baseline and 26-month measurements. Results: The primary complete-case sample included 243 children with valid paired measurements for zinc and serum transferrin receptor; vitamin D analyses were restricted to 52 children because of missing or invalid samples. At baseline, iron and vitamin D deficiencies were common, affecting 42.8% and 39.9% of children, respectively, while zinc deficiency affected 11.9%. At 26 months, allocation to the physical activity intervention was associated with lower odds of zinc deficiency, both when delivered alone (OR = 0.16) and when combined with supplementation (OR = 0.57). Supplementation alone was not significantly associated with reduced zinc deficiency. Iron status did not differ between intervention groups. Vitamin D findings should be interpreted with caution because analyses were based on a very small biomarker-specific subsample. Conclusions: School-based physical activity, alone or combined with multi-micronutrient supplementation, was associated with lower odds of zinc deficiency among Tanzanian schoolchildren. Supplementation alone showed no clear benefit for zinc or iron status. Vitamin D findings remain inconclusive because of substantial biomarker-specific missingness. Future trials should strengthen adherence monitoring, biomarker follow-up, and repeated assessment of dietary and contextual factors.

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 analyse 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 (percentage of female mosquitoes inseminated), body size (wing length), fecundity (number of eggs laid per female), and insecticide susceptibility (percentage of 24-hour mortality after exposure to insecticides) were compared between the newly established colonies of Tanzanian An. funestus (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, I and II. Additionally, other intragenomic variations were examined through a PCR analysis of restriction fragment length polymorphisms (RFLP) on the third domain of 28S ribosomal DNA. These molecular markers were used to compare the FUTAZ colonies, FUMOZ colonies in Tanzania and South Africa, and the wild-collected An. funestus from Tanzania. Result: The mating success and body size of FUTAZ females declined significantly from filial generations F1 to F6 relative to the founder population (F0), but 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 (over 10 days) to mate compared to females in the An. arabiensis colony (approximately 5 days). Insecticide resistance appeared to be lost during colonization, but this varied with insecticide classes. Majority of mosquitoes in the FUTAZ colony, as well as the wild-caught Tanzanian An. funestus belonged to Clade I (80.4-89.4%) and RFLP type "Y" (90.5-91.4%), while the FUMOZ colonies were mostly Clade II (65.5-88.5%) and RFLP type "MW" (90.5-91.5%). Conclusion: This study suggests that the mating success and body size of An. funestus decreases significantly during the early stages of colonization, then increase as the mosquitoes adapt to laboratory conditions. It is therefore crucial to have a large enough founder population to persist through these early generations in order to achieve stable colonization of An. funestus. The Clade and RFLP genotyping demonstrated the genetic similarities between the FUTAZ mosquitoes and wild-caught Tanzanian An. funestus, but also showed that the new colony can be distinguished from the FUMOZ colony. ### Competing Interest Statement The authors have declared no competing interest.

Despite Anopheles funestus s.s. being a highly competent and widespread malaria vector in Africa, its population structure remains largely understudied in many countries, including Tanzania. Herein, we examine the genetic diversity, geographic isolation, and gene flow of An. funestus populations across ten administrative regions in mainland Tanzania. We employed 12 previously used microsatellite DNA markers to describe genetic diversity, isolation by distance, and gene flow patterns among ten An. funestus s.s. populations (n = 654) and one An. parensis population (n = 28), used as an outgroup, sampled across ten regions in mainland Tanzania. Overall, allelic richness (Na) and genetic diversity (HS) did not differ significantly among populations. Although some loci and populations showed significant departures from Hardy–Weinberg equilibrium, the patterns were not indicative of substructuring within locations. Pairwise genetic divergence (FST) values indicated clear separation between An. parensis and An. funestus s.s., with values exceeding 0.2, consistent with species-level differentiation. Among An. funestus s.s. populations, the highest divergence was observed between southeastern coastal populations (Mtwara, Ruvuma, Lindi) and inland populations, with FST values up to 0.288. There was no evidence of isolation by distance. Instead, patterns of genetic divergence suggested connectivity across the Rift Valley and heterogeneity among southeastern populations. Neighbor-joining analysis and Bayesian genotype clustering identified three distinct population groups: (i) An. parensis (Dodoma), (ii) a genetically distinct An. funestus s.s. population from Mtwara, and (iii) a more homogeneous cluster comprising the remaining An. funestus s.s. populations. Notably, the Mtwara population appeared highly differentiated, with divergence approaching that between An. funestus s.s. and An. parensis, supporting its distinctiveness but not undermining the role of An. parensis as an outgroup. Except for the Mtwara population, whose status will need to be clarified through whole-genome sequencing, moderate genetic divergence was found among An. funestus s.s. populations across Tanzania, despite geographical separation and the Rift Valley. The observed genetic structure suggests that anthropogenic gene flow may play a key role in shaping population divergence. Future studies should aim to delineate the effects of local adaptation from recent gene flow to further explore these dynamics

Insecticide-based strategies for mosquito control are increasingly constrained by insecticide resistance, sparking interest in eco-friendly alternatives, one of which is the use of aquatic predators to reduce mosquito densities. Evidence from laboratory, semi-field, and localized field studies indicates that aquatic predators can significantly reduce mosquito larval and adult densities. However, direct evidence linking these ecological effects to sustained reductions in mosquito-borne disease transmission remains limited. This review synthesizes current studies on the use of aquatic predators for mosquito control and examines their potential as a complementary biological strategy within integrated vector management programs. While aquatic predators show promise for suppressing mosquito populations through both consumptive and non-consumptive effects, further research is needed to evaluate their epidemiological impact, operational feasibility, cost-effectiveness, scalability, and long-term sustainability. Addressing these knowledge gaps will be essential for determining how predator-based approaches can be effectively integrated into existing vector control strategies.

Gene-drive mosquitoes could transform malaria control in Africa, but their rapid, autonomous spread requires rigorous post-release monitoring. Most malaria-endemic countries already conduct some entomological surveillance, although it is often limited, fragmented, and externally funded. Molecular diagnostics are also expanding but remain mostly research focused and ad hoc. These imperfect systems offer workable foundations for strategic upgrades to support essential gene-drive monitoring. Priority investments should strengthen field-entomology, high-throughput genotyping for drive alleles and resistance, technical expertise, and integrated data for decision-making. Fortunately, first-generation gene drives already align with common phenotyping and genotyping workflows, avoiding major infrastructure overhauls, and permit simpler evaluation metrics than conventional interventions. This feature review examines key technical and operational considerations for monitoring gene drives and recommends how countries can adapt their vector surveillance systems to effectively monitor gene-drive mosquito releases.

Accurate estimation of mosquito age is critical for malaria surveillance, as only older female Anopheles mosquitoes can transmit the parasite. Traditional age-grading methods, include the Polovodova technique in which dilatations on the ovarian pedicel to tally completed gonotrophic cycles (GC) are counted. Despite their widespread use, these techniques remain poorly validated under realistic operational conditions, with limited evidence on their precision and accuracy across different parity rates, GC, and species. In this study, the accuracy and precision of the Polovodova method was evaluated across multiple age-graders and species under both laboratory and field conditions. A blinded validation study was conducted using laboratory-reared Anopheles arabiensis and Anopheles funestus (N > 3,600) spanning four gonotrophic cycles, and field-collected mosquitoes from Ulanga district, Tanzania (N = 600). Three blinded researchers performed dissections and readings, while a fourth unblinded researcher served as the reference reader (R0) to obtain ground truth (for parity) or assumed truth (for GC) for laboratory mosquitoes. Accuracy and precision for parity and gonotrophic cycle classification were assessed using pairwise inter-rater comparisons, percent agreement, Fleiss' kappa and Cohen's kappa (κ) statistics. For field mosquitoes, which had no age-referenced group, only precision was assessed. The Polovodova method accurately classified parity in both species, correctly identifying 98-100% of nulliparous and 94-96% of parous mosquitoes compared with the reference dataset. Inter-rater reliability for parity was almost perfect in An. arabiensis and An. funestus (κ = 0.81-0.83), with 82-96% pairwise agreement. GC classification was highly accurate for early stages (GC0-GC2: 85-100%) but declined at later stages GC4(34-46%), reflecting systematic underestimation of mosquito age. Overall GC reliability was moderate (κ = 0.56-0.59). Field classifications showed almost perfect agreement (κ = 0.87-0.88), supporting operational applicability. The Polovodova method provides robust estimates of parity and early gonotrophic-cycle status in Anopheles mosquitoes, but late-cycle classifications are more prone to reader disagreement and systematic underestimation. Operational use should therefore rely on trained multiple-reader workflows, including adjudication of discordant or late-cycle specimens, rather than single-reader classification. These findings provide empirical benchmarks for quality-assured Polovodova age-grading in malaria vector surveillance and intervention evaluation.

Abstract Background Larval source management (LSM) can be highly effective for controlling malaria vectors such as Anopheles funestus s.s., which typically exploit large and permanent aquatic habitats. While these habitats can persist year-round in endemic regions of Africa, their availability and use shift between wet and dry seasons. Understanding these seasonal changes is essential for identifying the habitats that sustain vector populations and for determining when and where LSM would be most effective. Methods We investigated the availability and use of An. funestus larval habitats across wet and dry seasons in south-eastern Tanzania, and the environmental factors that influence these patterns. Cross-sectional surveys were conducted in five villages during the dry season (September–November 2021) and rainy season (February–May 2022) to map and characterize aquatic habitats and identify those colonized by An. funestus. Results In total, 2824 aquatic habitats were identified, of which 27% were positive for An. funestus. Remotely sensed land cover data and directly measured habitat characteristics were incorporated into generalized linear mixed models to evaluate seasonal and environmental predictors of larval presence and abundance. Larval occurrence and density were significantly influenced by habitat type, village, season, and their interactions, as well as by key physicochemical factors including water depth, vegetation type, algae, water clarity, and water source. An. funestus was commonly found in river streams, ground pools, and ditches across both seasons. During the wet season, however, it also occupied spring-fed wells, rice fields, and dug pits, indicating broader habitat use. Conclusion These findings demonstrate a clear seasonal shift in larval habitat use by An. funestus, reflecting its ecological adaptability. While the species generally favors permanent habitats, its expanded use of diverse sites in the wet season has important implications for LSM. Targeting persistent habitats during the dry season may offer a more efficient and feasible window for implementing this intervention.

BackgroundMalaria transmission is highly sensitive to climatic variability, as changes in temperatures and rainfall, directly influence mosquito breeding, survival, and parasite development. Extreme climatic events, such as flooding, further exacerbate malaria risk by disrupting access to preventive, diagnostic and treatment services. However, there is limited evidence on how communities in malaria-endemic settings perceive and respond to the health impacts of climate variability and change. This study explored community knowledge, perceptions, and practices related to the relationship between climate variability and malaria transmission in south-eastern Tanzania.MethodsAn explanatory mixed-methods cross-sectional study was conducted in malaria-endemic villages in south-eastern Tanzania. Quantitative data were collected through structured questionnaires administered to 384 community members, while qualitative data were obtained through 11 key informant interviews and 12 focus group discussions involving 72 participants. Survey data were analysed descriptively, and qualitative data were analysed thematically.ResultsAmong survey respondents, 86% reported experiencing climate-related changes, including altered cropping seasons, increased flooding, and a perceived rise in vector-borne diseases. Approximately two-thirds (67.5%) recognized a link between climate change and malaria transmission. Perceived vulnerability was high, with 59.5% reporting increased risk of vector-borne diseases and 70% indicating higher malaria occurrence during the rainy season compared to the dry season. Access to timely climate and health information was limited, as only 26.6% regularly received updates, despite 96.6% expressing a desire for such information. Findings from focus group discussions and key informant interviews corroborated these perceptions and highlighted the need for targeted community awareness and education on climate-related malaria risks.ConclusionsCommunity members demonstrated awareness of climate change and its perceived impacts on malaria and livelihoods. These findings highlight the importance of integrating community perspectives and local knowledge into climate-adaptation and malaria-control strategies to enhance locally relevant and community-centered resilience.

Widespread insecticide resistance increasingly threatens malaria elimination, prompting a reassessment of vector control strategies. As Tanzania transitions from standard pyrethroid-only insecticide-treated nets (ITNs) to new-generation nets, evaluating the impact of this shift on malaria transmission and resistance is critical. Using the agent-based malaria model, EMOD, we assessed the impact of three ITN types, standard pyrethroid-only nets, pyrethroid-PBO nets (Olyset® Plus/PermaNet® 3.0), and the dual-active Interceptor® G2 nets (IG2) on malaria transmission and on modelled changes in resistance genotype frequencies under a simplified multi-locus representation of insecticide resistance. We also evaluated different sequences for introducing the new-generation nets, and the impact of combining ITNs with indoor residual spraying (IRS). The model was calibrated using incidence and prevalence data from two regions in northwestern Tanzania, incorporating transmission seasonality and heterogeneity, insecticide resistance, and behaviours of dominant vectors Anopheles funestus (highly anthropophilic, endophilic) and Anopheles arabiensis (more opportunistic readily biting non-human hosts outdoors). Changing from standard pyrethroid-only ITNs to pyrethroid-PBO and thereafter to IG2 ITNs reduced homozygous-resistant An. funestus and An. arabiensis by 71.9

The spread of Anopheles stephensi into the Horn of Africa represents one of the main challenges for malaria control, given the species’ ecological plasticity and resistance to multiple insecticides. In response to the World Health Organization’s 2022 vector alert, an adaptive, model-based spatial surveillance framework was developed and evaluated to improve detection, mapping accuracy, and operational responsiveness during invasion. Adaptive surveillance utilises initial observations to guide subsequent surveillance, linking the surveillance design to the underlying geographical characteristics of Anopheles stephensi distribution through observed data. This dynamic approach targets areas of high uncertainty and/or abundance, making the design responsive rather than predetermined. Focusing on Djibouti and selected regions of Ethiopia and Kenya, the adaptive surveillance was designed on previous in-country Anopheles stephensi surveillance data integrated with assembled open-source environmental, epidemiological, and demographic covariates. Key driver factors of the average monthly Anopheles stephensi catches varied geographically, although seasonality was universally important. Adaptive site allocation was optimised using a multicriteria target function which combines the trapping probability and uncertainty from previous surveys, with a simulation based on peaks-over-threshold (generalized Pareto) modelling of exceedances and Bayes factor–guided prioritisation. The selected adaptive surveillance design is the one that minimise the uncertainty in Anopheles stephensi trapping probability in hotspot areas. Optimal adaptive designs required between 50 to 59 sites per country, with uncertainty reductions in the probability of trapping projected up to 36% in Djibouti and more than 60% in Ethiopia and Kenya, with more than 60% site implementation halving uncertainty in Djibouti and Kenya and reducing it by up to 75% in Ethiopia. The proposed adaptive surveillance framework operationalises WHO guidance, accelerates hotspot identification, and inform targeted ecological studies and control interventions. It is extensible to other urban vectors (e.g., Aedes aegypti), enabling integrated, cross-border surveillance essential to contain Anopheles stephensi during ongoing invasion. ### Competing Interest Statement The authors have declared no competing interest. ### Funding Statement Yes ### Author Declarations I confirm all relevant ethical guidelines have been followed, and any necessary IRB and/or ethics committee approvals have been obtained. Yes I confirm that all necessary patient/participant consent has been obtained and the appropriate institutional forms have been archived, and that any patient/participant/sample identifiers included were not known to anyone (e.g., hospital staff, patients or participants themselves) outside the research group so cannot be used to identify individuals. Yes I understand that all clinical trials and any other prospective interventional studies must be registered with an ICMJE-approved registry, such as ClinicalTrials.gov. I confirm that any such study reported in the manuscript has been registered and the trial registration ID is provided (note: if posting a prospective study registered retrospectively, please provide a statement in the trial ID field explaining why the study was not registered in advance). Yes I have followed all appropriate research reporting guidelines, such as any relevant EQUATOR Network research reporting checklist(s) and other pertinent material, if applicable. Yes The entomological data used in this study are available in the Zenodo open research repository at the following link: https://doi.org/10.5281/zenodo.18642425

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). 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. funestus. 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

Most malaria vector control strategies target female Anopheles mosquitoes that transmit parasites, but emerging approaches such as gene drive, sterile insect technique, or Wolbachia require the release of males. Successful deployment depends on reliable knowledge of the species composition and age structure of male populations to overcome potential barriers to mating. Although approaches including near-infrared spectroscopy have been explored for male species identification and age grading, these methods remain constrained by limited validation, scalability, and accuracy under operational field conditions. Here we tested mid-infrared spectroscopy (MIRS) coupled with machine learning (ML) as a rapid and cost-effective approach for classifying species and age of male mosquitoes within the Anopheles gambiae complex. We used male mosquitoes from two laboratory-reared colonies in Burkina Faso (Anopheles coluzzii and An. gambiae) to develop a MIRS-ML model for species and age group (1–4, 5–10, 11–17 days) prediction under controlled conditions. This model was tested against a genetic and environmentally variable dataset consisting of male offspring obtained from gravid or blood fed Anopheles coluzzii and An. gambiae females collected from houses from two villages, Vallée du Kou and Soumousso and reared to adulthood in a semi-field system. The MIRS spectra from 2,120 males, representing both species, all age groups and both laboratory and semi-field backgrounds, were analysed using an extreme gradient boosting (XGBoost) algorithm to assess the ability to correctly predict age group and species. The XGBoost model classified mosquito species (86%) and age groups (85%) accurately in laboratory data, but performance declined under semi-field conditions (64% for species, 50% for age), reflecting environmental variability. Incorporating semi-field samples through transfer learning improved accuracy to 73% for species and 70% for age, underscoring both the limits of laboratory-only models and the value of transfer learning for enhancing generalisability in field settings. Our results demonstrate that mid-infrared spectroscopy with supervised machine learning (MIRS-ML) holds potential as a rapid tool for identifying the species and age group of cryptic male malaria vectors and represent one of the first applications of this approach to male Anopheles gambiae s.l. evaluated under semi-field conditions. However, before the approach can be used, larger datasets are needed to improve the classification algorithms and validate them for prediction in field populations.

Gene drive-modified mosquitoes (GDMMs) are gaining attention as sustainable tools to complement existing malaria control strategies. Their ability to self-propagate and spread through wild mosquito populations offers the promise of low-cost, long-lasting impact, but also raises ecological, ethical, and governance concerns. In this evolving debate, civil society organizations (CSOs) are pivotal actors in shaping dialogue, representing community concerns, and influencing policy decisions. This study examined the perspectives and recommendations of biodiversity-oriented CSOs on the governance, testing, and potential application of GDMMs for malaria control in Tanzania. An exploratory qualitative design was employed, involving eight in-depth interviews, one focus group discussion, and three large group discussions with representatives from ten biodiversity-focused CSOs in Tanzania. Participants were selected purposively based on prior involvement in national or regional dialogues related to biotechnology; and the discussions focused on concerns, uncertainties and needs associated with testing and potential use of GDMMs for malaria control, as well as the balance of prospective benefits against long-term environmental risks. Transcripts were analyzed thematically using NVivo 12 Plus. Participants expressed cautious support for research on GDMMs for malaria control but raised concerns about scientific uncertainty, limited local expertise, inadequate transparency, potential transboundary effects and technological dependency. They emphasized the importance of generating robust, context-specific evidence before considering any environmental releases of gene drives; and highlighted concerns over inadequate accountability, particularly the lack of clarity on who would assume responsibility if adverse outcomes arise. They also advocated for early, inclusive, transparent, and continuous engagement with both target communities and the broader public. Lastly, to ensure objective and impartial oversight, they recommended development of local expertise that is independent of technology developers and sponsors. The CSOs’ perspectives were diverse but broadly aligned with the precautionary principle, calling for preventive action amid uncertainty, clear accountability, and the pursuit of safer alternatives. Although many expressed serious reservations about gene drive mosquitoes, there was a shared recognition that research on the technology is necessary, provided it is conducted under controlled, transparent, and auditable conditions. Overall, these exploratory discussions underscored the need for: (i) balanced dialogue between advocates and skeptics, (ii) robust ethical and regulatory frameworks covering the full life cycle of the technology, (iii) sustained community and stakeholder engagement from the early stages of research and development, (iv) enhancements of in-country capacity, and (v) national sovereignty in decision-making regarding GDMMs. Demonstrating and effectively communicating these elements will be as critical as ensuring their existence.

Larval source management (LSM) was once central to malaria control before insecticide-treated nets and indoor residual spraying dominated. Renewed interest in LSM raises questions about its effectiveness in rural Africa, where habitats are dispersed, and vector species contribute unequally, and whether species-targeted larviciding could offer greater gains than broadcast approaches. This modelling study quantified the potential impact of larviciding in African settings where multiple vector species contribute unequally to malaria transmission. We modeled malaria transmission in southeastern Tanzania using the EMOD v2.20, an individual-based malaria transmission model incorporating seasonal dynamics, insecticide resistance, and semi-field biolarvicide efficacy. Outcomes were entomological inoculation rate, malaria incidence in under-fives, and operational larviciding costs. Large-scale deployment of biolarvicides with > 1-week residual activity substantially reduced malaria transmission, with disproportionately greater gains when control efforts were preferentially focused on the dominant vector species, Anopheles funestus, compared to broadcast approaches treating both An. funestus and An. arabiensis habitats. In the absence of ITNs, a four-month fortnightly larviciding campaign targeting An. funestus at 80

Although more than half of the urban population in sub-Saharan Africa reside in informal housing, knowledge about the material and spatial characteristics of such dwellings remains limited. This study examines informal housing practices in the Mabibo neighborhood of Dar es Salaam, Tanzania, using detailed surveys of the built environment and household data collection to investigate how dwellings are adapted to contextual conditions. Sociocultural adaptation strategies include spatial configurations to accommodate complex household structures, where extended families and tenants often reside in separate sub-units within the same plot, driving emergence of new courtyard housing typologies typified by built environment densification and increasing rental accommodation. Economic adaptation strategies include integration of income-generating functions in dwellings such as shops and rental units, incremental dwelling expansion based on available resources, and subdivision of plots to accommodate additional households. The study documents evolving construction practices with traditional housing being replaced by dwellings built from industrial materials and repurposed waste. Environmental adaptation strategies include raised floors to mitigate flooding, window screening to reduce exposure to mosquito-borne diseases, and suspended ceilings reported by residents as improving indoor thermal comfort. These findings show how peri-urban informal housing practices in rapidly urbanizing African cities evolve through adaptation to contextual conditions, such as land scarcity, constrained household economies, and availability of industrial construction materials linked to global supply chains.

Abstract Background Evidence of natural infection with Wolbachia and its negative correlation with Plasmodium falciparum among wild malaria vectors has opened new avenues for utilization of Wolbachia in malaria vector control. However, the interaction between Wolbachia and Plasmodium parasites in mosquitoes tends to be species-specific and may show ecological variations. Among the primary malaria vectors in Tanzania, natural Wolbachia infection has only been observed in Anopheles arabiensis, while there is still limited information on Wolbachia natural infection in Anopheles funestus sensu lato, and its interaction with P. falciparum in the mosquito species. Therefore, this study investigated the prevalence of natural infection and co-infection of Wolbachia and P. falciparum in the An. funestus s.l. in southeastern Tanzania, and characterized the Wolbachia strains detected. Methods The study was conducted in five villages in southeastern Tanzania between March and June 2024. Mosquitoes were collected from 52 households using Centers for Disease Control and Prevention (CDC) light traps and Prokopack aspirators, followed by morphological identification. Detection of An. funestus sibling species and Wolbachia was performed using conventional polymerase chain reaction (PCR) and nested PCR (Wolbachia only). Sanger sequencing was performed as a confirmatory test followed by phylogenetic analysis of the detected Wolbachia strains. P. falciparum sporozoites were detected using enzyme-linked immunosorbent assay (ELISA). Results Wolbachia was detected in almost half of all wild An. funestus s.l. tested using the primary PCR (prevalence = 46.5%, N = 400); and more than half when nested PCR approach was used (prevalence = 70.8%, N = 400). Only three mosquitoes carried P. falciparum sporozoites (prevalence = 0.8%, N = 400) and only one showed co-infection with Wolbachia (prevalence = 0.3%, n = 400). Sequencing and phylogenetic analysis involving both the 16S rDNA, coxA, and wsp Wolbachia genes showed that the detected strains clustered with Wolbachia supergroup B, specific for Dipterans. Conclusions Unlike findings from the previous study, this study demonstrates that An. funestus s.l. in southeastern Tanzania are infected with Wolbachia, at a surprisingly high prevalence. This study also provides the first report on Wolbachia–P. falciparum co-infection status in An. funestus s.l. in Tanzania. Further studies with larger sample sizes are needed to confirm the association between native Wolbachia and P. falciparum in wild An. funestus s.l. in southeastern Tanzania. Graphical Abstract

Temperature is a key environmental factor influencing the development, survival, and transmission potential of malaria vectors. While most laboratory studies use constant temperature (CT) regimes, mosquitoes in natural habitats experience fluctuating temperatures (FTs), which may affect their life-history traits. We investigated the effects of CT (27 °C) and FT (27 ± 3 °C) on larval and adult traits of 2 major malaria vectors, Anopheles gambiae and An. coluzzii, under laboratory conditions. We measured larval survival, development time, adult body size, and adult survival, using survival and mixed-effects models. Species-specific and stage-specific responses to temperature regimes were observed. An. gambiae larvae exhibited higher survival under FT, while An. coluzzii larvae survived better under CTs. However, this pattern reversed in adulthood: An. coluzzii adults showed increased survival and larger body size under FT, whereas An. gambiae adults performed better under CT. Development time was slightly longer under FT for both species, with An. coluzzii pupating faster overall. These opposing patterns suggest that differential larval survival under FTs may influence adult fitness in a species-specific manner. The contrasting and reversed responses of An. gambiae and An. coluzzii across life stages might reflect their ecological adaptations: An. gambiae, found in small and thermally variable habitats, performed better under FT during larval stages, while An. coluzzii, associated with larger, more thermally stable habitats, showed improved adult performance under FT. These findings underscore the importance of incorporating species-specific, stage-dependent thermal responses into models of vector dynamics and control strategies under climate change.

Abstract Background Malaria control gains are stagnating or reversing, in part due to widespread insecticide and drug resistance, as well as increasing financial and implementation challenges. This study assessed the national strategies for insecticide resistance monitoring and management in Tanzania to identify key barriers and misalignments between policy and actual practice. Methods We initially reviewed technical documents from Tanzania’s National Malaria Control Programme (NMCP) produced between 2014 and 2024, along with World Health Organization (WHO) guidelines and reports on insecticide resistance monitoring and management. This was followed by in-depth interviews with key informants in Tanzania, including policymakers, representatives from funding agencies, technical experts, scientists, and vector control implementers at district and regional levels. The qualitative data was analyzed thematically using NVivo software. Findings The document review revealed strong policy alignments of the NMCP strategies for monitoring and management of insecticide resistance with international practice as recommended by WHO. However, implementation of the policy intentions remained limited, with resistance monitoring being conducted in only 22 of Tanzania’s 184 district councils. Interviews with 24 stakeholders highlighted significant gaps between the stated policies and guidelines and the actual practice. These gaps were driven by inadequate financing, donor dependence, insufficient coordination and dissemination of guidelines, limited technical capacity, and weak engagement of communities and district-level operators. Participants emphasized that without strengthened surveillance systems, sustainable local financing, and greater community and intersectoral collaboration, achieving the current strategic goal of malaria elimination by 2030 remains unlikely. Conclusion Despite strong policy alignment and strategic planning, Tanzania’s capacity to implement insecticide-resistance management initiatives remains constrained by several policy-practice gaps, particularly in funding, staffing and coordination. Addressing these persistent challenges will be essential for more effective malaria vector control and will require sustained financing, stronger coordination, improved vector monitoring, and greater community and intersectoral engagement.

Establishing and maintaining laboratory colonies of the malaria vector, Anopheles funestus using newly collected material has proven challenging, in part because of their low propensity to mate in captivity. In this study we assessed how cage conditions influence the mating success of two An. funestus strains originating from different geographic areas, Angola (FANG) and Mozambique (FUMOZ). The visual environment in adult mosquito-rearing cages was manipulated either by covering the cages in different planes with black opaque cloth (referred to as black horizons) or by placing black visual markers at various positions inside the cages. Mating success was assessed by dissecting the spermathecae capsule of the females after the standard 10-day mating period. Insemination rates were consistently higher in the An. funestus FANG strain than in the FUMOZ strain in both the black horizon (odds ratio [OR] 0.31, 95