Contrasting vector competence of three main East African Anopheles malaria vector mosquitoes for Plasmodium falciparum

Contrasting vector competence of three main East African Anopheles malaria vector mosquitoes for Plasmodium falciparum | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Article Contrasting vector competence of three main East African Anopheles malaria vector mosquitoes for Plasmodium falciparum Prisca A. Kweyamba, Lorenz M. Hofer, Ummi A. Kibondo, Rehema Y. Mwanga, and 5 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-5038559/v1 This work is licensed under a CC BY 4.0 License Status: Under Review Version 1 posted 11 You are reading this latest preprint version Abstract There are three Anopheles mosquito species in East Africa that are responsible for the majority of malaria transmission, posing a significant public health concern. Understanding the vector competence of different mosquito species is crucial for targeted and cost-effective malaria control strategies. This study investigated the vector competence of laboratory reared strains of East African An. gambiae sensu stricto, An. funestus s.s. , and An. arabiensis mosquitoes towards local isolates of Plasmodium falciparum infection. Mosquito feeding assays using gametocytaemic blood from local donors revealed significant differences in both prevalence and intensity of oocyst and sporozoite infections among the three vectors. An. funestus mosquitoes presented the highest sporozoite prevalence 23.5% (95% confidence interval (CI): 17.5–29.6) and intensity of infection 6-58138 sporozoites. Relative to An. funestus , the odds ratio for sporozoites prevalence were 0.46 (95% CI: 0.25–0.85) in An. gambiae and 0.19 (95% CI: 0.07–0.51) in An. arabiensis , while the incidence rate ratio for sporozoite intensity was 0.31 (95% CI: 0.14–0.69) in An. gambiae and 0.66 (95% CI: 0.16–2.60) in An. arabiensis. Our findings indicate that all three malaria species contribute to malaria transmission in East Africa with An. funestus demonstrating superior vector competence. In conclusion, there is a need for comprehensive malaria control strategies targeting major malaria vector species, an update of malaria transmission models to consider vectoral competence and evaluation of malaria transmission blocking interventions in assays that include An. funestus mosquitoes. Biological sciences/Molecular biology Earth and environmental sciences/Ecology Figures Figure 1 Introduction Anopheles mosquitoes are the only arthropod vectors that transmit Plasmodium parasites that cause malaria in humans. The disease imposes a significant burden of mortality and morbidity, particularly in Sub-Saharan Africa (SSA) 1 where the most efficient mosquito vectors of malaria are found 2 . Human malaria is mediated only by female Anopheles , and of the estimated 460 species, only 40 species or species complexes are considered to be important vectors in the wild 3 , notably the Anopheles gambiae and Anopheles funestus species complexes that dominate malaria transmission throughout SSA 4 . In this paper, we focus on the three major East African vectors of human malaria: An. funestus sensu stricto , An. gambiae sensu stricto and An. arabiensis. The successful transmission of Plasmodium parasites between humans requires intricate transformations within the mosquito vector 5 , highlighting the key role of both vectorial capacity 6 and vector competence 7 in determining the local intensity of malaria transmission 7 . Vector competence refers to the ability of an arthropod vector to acquire, maintain and transmit a pathogen. This concept encompasses the inherent ability of a pathogen to effectively enter and reproduce within the vector and be released from the vector's salivary glands to initiate infection in another vertebrate host 7 , 8 . Vectorial capacity describes the potential intensity of transmission by mosquitoes. It is defined as the total number of infectious mosquito bites on humans that will arise from a single infected person on a single day 9 . This is influenced by a number of factors 6 , 10 , most notably the probability of mosquitoes to feed on humans 11 , daily vector survival 12 , environmental factors that affect the time it takes for parasites to develop in the mosquito host 13 , the availability of larval breeding sites 14 , presence of vector control tools 10 and vector competence 15 . The vectoral capacity of a particular vector is strongly influenced by its ecology. The larval stage of mosquitoes takes place in water where biological factors greatly influence the habitat suitability and carrying capacity. These factors influence vector presence 3 , fitness 16 , longevity 17 , which in turn affects the probability that a mosquito can acquire and maintain a parasite for long enough to become infectious 18 . In East Africa, the three major vectors have different regional distribution due to ecology 19 and hydrology 14 and varying contributions to malaria transmission across different seasons 20 – 23 . An. funestus mosquitoes have permanent breeding sites abundant with vegetation, making them likely to transmit malaria all year round 24 , 25 . An. arabiensis and An. gambiae s.s. typically dominate in temporary sunlit pools and their presence is strongly dependent on rainfall or irrigation 26 , 27 . An. gambiae s.s. requires high humidity to survive and occurs almost exclusively during humid and rainy periods 28 . An. arabiensis and An. funestus , are more resistant to desiccation, are commonly found in abundance during the peak of the wet season and continue into the dry season; sustaining malaria transmission for several months after the end of the rains 23 , 29 . Anopheles mosquitoes are dependent on vertebrate blood to provide proteins needed for egg development and undergo multiple cycles of feeding and egg development in their lifetime. Therefore, the preference of a vector for human blood has a direct impact on its efficiency as a vector by increasing its probability of acquiring and transmitting onward infection 30 . An. arabiensis have an opportunistic feeding behavior, targeting both human and animal hosts for its blood meals, so it may be a more or less important vector dependent on the relative proportion of cattle in an area 31 . An. gambiae 32 and An. funestus 33 are more specialized blood feeders, feeding almost entirely on humans, although this does depend on host availability. In addition, there is evidence that multiple blood meals increase the likelihood of Plasmodium developing in the mosquito 34 , 35 . As well as environmental factors that affect vectors' susceptibility to infection and the interactions between the vector, pathogen, and host that impact probabilities of onward transmission, vector competence is influenced by a variety of internal factors, including the genetics of both the vector and the pathogen 5 , 36 . Plasmodium takes resources from its definitive host that results in reduced fitness and reproductive output 37 . Therefore, the mosquito innate immune system either modulates or resists infection 38 , while the parasite counteracts mosquito defenses through host manipulation 39 and polyclonality 40 . Mosquito species show different levels of susceptibility to Plasmodium from refractory in the case of Anopheles quadriannulatus 41 , 42 to high susceptibility in Anopheles coluzzi 43 , 44 . In East Africa, there is evidence of higher proportions of infected An. funestus 45 , 46 relative to An. arabiensis and An. gambiae . This can be to an extent explained by the fact that An. funestus generally feeds almost exclusively on humans and has been shown to live longer than An. arabiensis 47 . However, any differences in degree of vector competence among the three primary malaria vectors has not been evaluated. Understanding vector competence is crucial in understanding the risk of malaria transmission, informing effective malaria control strategies 36 , 48 , 49 and parameterizing mathematical models, where mosquito-parasite interactions are rarely considered 15 , 50 . Therefore, this study investigated whether vector competence towards Plasmodium falciparum differs between local East African strains of An. gambiae s.s. , An. funestus and An. arabiensis mosquitoes. By experimentally infecting mosquitoes with field gametocytes using Direct Membrane Feeding Assays (DMFAs), we aim to compare the prevalence and intensity of P. falciparum infection between these Anopheles mosquito species. Results Prevalence and intensity of P. falciparum infection among local strains of An. gambiae s.s. , An. funestus and An. arabiensis mosquitoes Oocysts The prevalence of oocyst-infected mosquitoes varied among the three Anopheles mosquito species, with An. funestus presenting the highest oocyst infection rate of 13.5% (95% CI: 9.2–17.6), followed by An. gambiae s.s. at 10.7% (95% CI: 6.9–14.4), and An. arabiensis at 5.6% (95% CI: 2.5–8.7) (Table 1 ). The proportion of oocyst-infected An. arabiensis mosquitoes was significantly lower than An. funestus mosquitoes (OR = 0.40, 95% CI: 0.20–0.80, p = 0.010), but there was no difference between An. funestus and An. gambiae (Fig. 1 A). Additionally, there was no significant difference in the proportion of oocyst-infected mosquitoes between An. gambiae s.s. and An. arabiensis mosquitoes (OR = 0.52, 95% CI: 0.25–1.06, p = 0.072). There was no significant difference in oocyst intensity between the three species, although as with prevalence, oocyst intensity was similar between An. funestus , which ranged from 1–12 and An. gambiae s.s. , ranging from 1–14; and lower among An. arabiensis , which presented the lowest intensity of oocysts ranging from 1–3 (Fig. 1 C). Sporozoites The burden of sporozoite-infected mosquitoes varied among the three Anopheles mosquito species with An. funestus presenting the highest sporozoite infection rate of 23.5% (95% CI: 17.5–29.6), followed by An. gambiae s.s. at 11.4% (95% CI: 6.5–16.3), and An. arabiensis at 4.9% (95% CI: 0.6–9.1) (Table 1 ). An. funestus had a higher probability of being sporozoite-infected than An. gambiae s.s. (OR = 0.46, 95% CI: 0.25–0.85, p = 0.013) or An. arabiensis (OR = 0.19, 95% CI: 0.07–0.51, p = 0.001), (Fig. 1 B). Moreover, there was no statistically significant difference observed in the proportion of sporozoite infection between An. gambiae s.s. and An. arabiensis mosquitoes (OR = 0.41, 95% CI: 0.14–1.17 P = 0.098), although there were few infected An. arabiensis . Additionally, there were similar trends observed in sporozoite intensity among the three species, with highest intensity observed in An. funestus , ranging from 6–58,138 followed by An. gambiae s.s. , ranging from 21 − 7,976 and An. arabiensis ranging from 20–27,877. A significant difference was observed in sporozoite intensity between An. funestus and An. gambiae s.s. (IRR = 0.31, 95% CI: 0.14–0.69, p = 0.004) (Fig. 1 D), although not between An. funestus and An. arabiensis likely due to low sporozoite prevalence (5%) in An. arabiensis leading to uncertainty in the estimates. Table 1 Proportion and intensity of oocyst and sporozoite infected laboratory reared mosquitoes with gametocytaemic blood from participants in Bagamoyo, Tanzania Proportion infected Intensity Number infected (n/N) Prevalence (95%CI) OR (95%CI) Median (Min-Max) IRR (95%CI) Oocysts An. funestus s.s. 35/260 13.5 (9.2–17.6) 2 (1–12) 1 An.gambiae s.s. An. arabiensis 29/271 12/214 10.7 (6.9–14.4) 5.6 (2.5–8.7) 0.76 (0.44–1.31) * 0.40 (0.20–0.80) * 1 (1–14) 2 (1–3) 1.31 (0.83–2.06) 1.04 (0.55–1.95) Sporozoites An. funestus s.s. 46/195 23.5 (17.5–29.6) 1 3,983 (6–58,138) 1 An.gambiae s.s. An. arabiensis 19/166 05/102 11.4 (6.5–16.3) 4.9 (0.6–9.1) 0.46 (0.25–0.85) ** 0.19 (0.07–0.51) ** 2,447 (21 − 7,976) 714 (20–27,877) 0.31 (0.14–0.69) ** 0.66 (0.16–2.60) ** Note: OR were derived from mixed-effect logistic regression while IRR were derived from mixed effect negative binomial regression using study participant as a random effect * p-value < 0.05, * *p-value < 0.01 Discussion This study represents the first attempt to measure whether the degree of vector competence towards P. falciparum infection varies between East African An. funestus s.s. , An. arabiensis , and An. gambiae s.s. mosquitoes. Through experimental feeding of mosquitoes with gametocytaemic blood from donors in Tanzania, we observed significant differences in both prevalence and intensity of oocyst and sporozoite infections among the three major vector species. An. funestus s.s. mosquitoes presented with the highest burden and intensity of infection indicating that they are more competent than either An. gambiae s.s. or An. arabiensis . This is the first time, to our knowledge that the vectoral competence of An. funestus s.s. has been evaluated. The findings of this study largely agreed with the findings of a study in Burkina Faso, that found no difference in genetic susceptibility to P. falciparum measured by oocyst infection between three sympatric population groups of the An. gambiae s.l. complex including An. coluzzi, An. gambiae s.s. and An. arabiensis that had been reared from wild-caught larvae 44 . Anopheles mosquitoes are reported to have varying levels of vector competence, influenced by genetic factors, as well as environmental conditions, and host-parasite interactions 5 , 36 , 51 . An. gambiae s.s. has long been recognized as a highly polymorphic and efficient malaria vector, possibly due to the strong co-adaptation of P. falciparum to this specific mosquito species 52 . However, our study reveals that An. funestus were more competent. This finding is consistent with reports of increasing malaria cases caused by An. funestus in Tanzania and other parts of SSA 53 – 57 . The high prevalence and intensity of infection observed in An. funestus mosquitoes suggest its potential as a predominant malaria vector, particularly in areas with suitable breeding sites abundant in vegetation. Our results are consistent with the observation that the massive introgression event that lead to the evolution of Anopheles funestus 13,000 years ago that facilitated its adaptation to new environments resulting in its subsequent dramatic geographic range expansion across most of tropical Africa also enhanced vectorial capacity in Anopheles funestus mosquitoes 58 . Although An. arabiensis mosquitoes play a crucial role in malaria transmission, particularly in arid regions in the Horn of Africa 45 , our findings show lower prevalence and intensity of oocyst and sporozoite infections when compared to An. funestus and An. gambiae s.s. mosquito infections. While An. arabiensis is often abundant, and is widely discussed as a vector of residual and outdoor malaria, the more competent and endophilic malaria vectors An. funestus s.s. and An. gambiae s.s. should be targeted for control. While several studies have shown that insecticide resistant vectors are more competent to Plasmodium 59 , 60 , our study also found that An. funestus mosquitoes which are resistant to pyrethroids showed increased susceptibility to Plasmodium infection. The susceptibility of mosquitoes to Plasmodium infection may result from either their increased survival and longevity 61 compared to susceptible mosquitoes, which are killed by insecticide 62 , or could be due to reduced immunity to parasites 63 . Therefore, we cannot rule out that the difference in insecticide susceptibility affected the results through a change in mosquito immunity to parasites. There is strong evidence suggesting that insecticide resistance mutations increase the vector competence of An. gambiae for Plasmodium , potentially sustaining malaria transmission 60 . However, the An. arabiensis used in the study were also pyrethroid resistant and were still relatively less susceptible to infection. An additional study has shown that insecticide resistance mechanisms have an effect on the activation of the mosquito immune system and its physiology, resulting in differences in parasite development and survival 64 . Variations in parasite burden may not significantly affect parasite transmission, however, the intensity of infection does influence the activation of the vector immune system 65 . Nonetheless, in our study, all three species showed susceptibility to Plasmodium infection, with An. funestus presenting a higher susceptibility compared to An. gambiae s.s. and An. arabiensis . It was suggested that the susceptibility of An. gambiae to Plasmodium infection is due to persistent immune suppression to prevent excessive activation of the immune response following blood meal ingestion 66 . Moreover, research has shown that genetic diversity within mosquito populations can also significantly influence their susceptibility to Plasmodium infection 67 . This study highlighted that, while An. gambiae mosquitoes are commonly used in DMFAs to assess different malaria transmission-blocking interventions 68 – 70 , the observed shift towards An. funestus as a major contributor to transmission in SSA 56 , with the highest infection burden, suggests the importance of incorporating An. funestus mosquitoes into assays for testing malaria transmission-blocking activity. A limitation of our study is the exclusive use of laboratory reared mosquito strains rather than actual field mosquitoes. In conclusion, we confirmed that between the three mosquito species, An. funestus was the most permissive to P. falciparum infection, which is coherent with consistently high sporozoite rates observed in this species across SSA, whereas An. arabiensis shows the greatest resistance coherent with its lower observed sporozoites rates. Nonetheless, our findings suggest that all the three vector species play an active role in malaria transmission. The observed differences in vector competence among the three Anopheles species highlight the complexity of malaria transmission dynamics and the need for comprehensive malaria control strategies that target key malaria vector species. Furthermore, malaria transmission models should be revised to account for vectoral competence, and efforts to support malaria transmission blocking interventions tested on multiple malaria vectors are essential for making sustainable progress towards malaria elimination. Methods Mosquito rearing An. gambiae s.s. mosquitoes were originally collected from the southern region of Tanzania (Njage-Mngeta villages, Ifakara district, Morogoro region) in 1996 and have been maintained at the Ifakara Health Institute (IHI) insectaries, Tanzania. This strain of Anopheles gambiae s.s. is susceptible to all classes of insecticides. Field-collected An. arabiensis mosquitoes were obtained from the southern region of Tanzania (Sakamaganga village, Ifakara district) in 2005 and have been maintained in IHI insectaries. This strain is resistant to pyrethroids at a 1x discriminating concentration and is susceptible to other classes. The An. funestus colony was established at IHI insectaries in 2018 and was originally derived from founder colony established in 2000 at the National Institute for Communicable Diseases (NICD) South Africa. This strain is resistant to pyrethroids at a 1x discriminating concentration and is susceptible to other classes. Mosquito larvae were maintained at a density of 200 larvae per litre of water and fed 0.3 g per larva on Tetramin fish food (Tetra Ltd., UK). For colony maintenance, the adult mosquitoes are provided with cow blood between 3 and 6 days after emergence for egg development using a Hemotek® membrane feeder (SP-6 System, Hemotek Ltd., Blackburn BB6 7FD, UK). Mosquitoes were provided with autoclaved 10% sucrose solution ad libitum. Temperature and humidity within the insectary are maintained between 27 ± 2 o C and 60%-85% relative humidity following the MR4 guidance 71 . Recruitment of asymptomatic gametocytaemic carriers Gametocytaemic carriers were selected by screening thick blood smears from participants aged 6–40 years located in the village of Wami-Mkoko, in Bagamoyo district located in the coastal region of Tanzania, between June 2023 and August 2023. Participants meeting the inclusion criteria (asymptomatic individuals aged 6–40 who consented and had microscopically detectable gametocytes) were enrolled for blood collection at IHI transmission facilities in Bagamoyo, Tanzania. Gametocytes were quantified by counting against 500 white blood cells in thick smears, and their density was calculated based on an estimated leukocyte density of 8000/ µL of whole blood. Five milliliters of blood were obtained from microscopically confirmed gametocyte carriers with gametocytes density exceeding three gametocytes/500 red blood cells, equivalent to 48 gametocytes/ µL of whole blood. Seven gametocytaemic individuals were recruited to donate blood for DMFAs. Autologous serum was replaced with pre-warmed malaria-naïve AB serum European donors. Experimental infection of P. falciparum in Anopheles mosquitoes through DMFAs Infectious gametocytaemic blood was administered to mosquitoes through water-jacketed glass feeders (14mm Ø, Chemglass, New Jersey, USA) covered with parafilm®, connected to a circulating water bath (39 o C, ELMI, Switzerland) via plastic tubing. On average, 200 mosquitoes from each mosquito strain were fed a blood meal from each participant for a duration of 15 minutes. After blood feeding, the cups containing mosquitoes were then transferred to Bugdorm plastic cages (30 cm x 30 cm x 30 cm, Megaview Science Co., LTD, Taiwan) and placed in a climatic chamber (S600PLH, AraLab, Lisbon, Portugal) maintained at 75 ± 2% humidity and 27 ± 1°C at 12:12 hours dark: light cycle. Mosquitoes were deprived of sugar for 48 hours to allow unfed mosquitoes to die. Dead mosquitoes were aspirated out after 48 hours and then cotton soaked with autoclaved 10% sucrose solution was provided and replaced daily. Oocyst and Sporozoite scoring Eight days post infection (dpi), one-third of mosquitoes from each mosquito strain was dissected and their midguts were stained with a 1% mercurochrome solution before examination for presence of oocysts microscopically. The remaining mosquitoes were kept up to day 16 dpi and the mosquito`s DNA was extracted using DNAzol® reagent 72 for molecular analysis and quantification of P. falciparum infection in mosquito stages from the mosquito heads and thoraces (sporozoites stages). Using quantitative reverse transcription polymerase chain reaction (RT-qPCR) 73 , absolute quantification of all sporozoites positive samples was performed using the standard curves generated on P. falciparum -specific 18S rDNA plasmid (GenBank: AF145334) from P. falciparum (BEI Resources, NIAID, MRA-177). Plasmid copy numbers per µl were calculated as described elsewhere 74 . The standard curves were generated on serial dilutions over eight magnitudes assuming an average of six copies of the 18S rDNA gene sequence per parasite genome 75 . Each concentration from the serial dilution was run in triplicates to determine qPCR efficiency, limit of detection, slope and y-intercept. Statistical analysis Data cleaning and analysis were conducted using STATA 17 software (StataCorp LLC, College Station TX, USA). Descriptive statistics were employed for data summarization, presenting the proportion of infected mosquitoes with a 95% confidence interval. For parasite intensity (oocysts or sporozoites), the median along with minimum and maximum values were reported. To evaluate vector competence towards P. falciparum infection among the three mosquito strains, mixed-effect regression was used, with mosquito strain as a fixed categorical effect and study participants included as a random effect. For oocyst and sporozoite prevalence logistic distribution was used. For oocyst and sporozoite intensity, negative binomial distribution was used and only infected mosquitoes were included in the intensity analysis. Declarations Acknowledgments The authors express their sincere gratitude to the village leaders and community in Bagamoyo for their unwavering cooperation and support throughout the study. A special thanks to Vector Control and Product testing Unit (VCPTU) management, administrators and colleagues who helped in organising logistics and materials, allowing smooth performance of the study. Authors ‘contribution SJM, PAK, MMT, LH conceived the study. PAK, LH and MMT developed the study protocol. RYM and PAK performed the DMFAs. RMS, PAK and FM dissected the mosquitoes. PAK, LH and RYM performed molecular analysis. PAK drafted the manuscript. SJM, MMT, DWL, and LH reviewed and edited drafts of the manuscript. Code, data, and materials availability Data is provided within the supplementary information files. Competing interests All authors declare no competing interest. Funding This study`s field work was funded by VCTPU and PAK, RYM, RMS and FM receive salary support from the Transmission Zero project (Bill and Melinda Gates Foundation (BMGF), grant no. OPP1158151). Ethics approval and consent to participate All adult participants provided written informed consent, while for children under 18 years old, consent was obtained from their parent or guardian, with the children's assent also sought for their participation. All study volunteers received artemisinin-lumefantrine treatment within 24 hours of diagnosis, following the Tanzania Guidelines for Diagnosis and Treatment of Malaria 76 , administered by a qualified nurse. No adverse effects were reported among the participants during the study period. The study activities were reviewed and approved by the Institutional Review Board of IHI (IHI/IRB/No: 44 – 2020) and the National Institute for Medical Research Tanzania (NIMR/HQ/R.8a/Vol.IX/3595). All research was performed in accordance to relevant guidelines and regulations. References WHO. The World Malaria Report 2023 (World Health Organization, 2023). Kiszewski, A. et al. A global index representing the stability of malaria transmission. Am. J. Trop. Med. Hyg. 70 , 486–498 (2004). Sinka, M. E. et al. A global map of dominant malaria vectors. Parasit. Vectors . 5 , 69. 10.1186/1756-3305-5-69 (2012). Sinka, M. E. et al. The dominant Anopheles vectors of human malaria in Africa, Europe and the Middle East: occurrence data, distribution maps and bionomic precis. Parasit. Vectors . 3 , 117. 10.1186/1756-3305-3-117 (2010). Lefevre, T., Vantaux, A., Dabire, K. R., Mouline, K. & Cohuet, A. Non-genetic determinants of mosquito competence for malaria parasites. PLoS Pathog. 9 , e1003365 (2013). Wu, S. L. et al. Vector bionomics and vectorial capacity as emergent properties of mosquito behaviors and ecology. PLoS computational biology 16, e1007446, doi: (2020). 10.1371/journal.pcbi.1007446 Cohuet, A., Harris, C., Robert, V. & Fontenille, D. Evolutionary forces on Anopheles : what makes a malaria vector? Trends Parasitol. 26 , 130–136 (2010). Reeves, W. C., Asman, S., Hardy, J., Milby, M. & Reisen, W. Epidemiology and control of mosquito-borne arboviruses in California, 1943–1987. (1990). Garrett-Jones, C. & Grab, B. The Assessment of Insecticidal Impact on the Malaria Mosquito’s Vectorial Capacity, from Data on the Proportion of Parous Females. Bull. WHO 31 (1964). Brady, O. J. et al. Vectorial capacity and vector control: reconsidering sensitivity to parameters for malaria elimination. Trans. R Soc. Trop. Med. Hyg. 110 , 107–117. 10.1093/trstmh/trv113 (2016). Garrett-Jones, C. The human blood index of malaria vectors in relation to epidemiological assessment. Bull. World Health Organ. 30 (1964). Matthews, J., Bethel, A. & Osei, G. An overview of malarial Anopheles mosquito survival estimates in relation to methodology. Parasit. Vectors . 13 , 233. 10.1186/s13071-020-04092-4 (2020). Ohm, J. R. et al. Rethinking the extrinsic incubation period of malaria parasites. Parasites Vectors . 11 , 178. 10.1186/s13071-018-2761-4 (2018). Smith, M. W. et al. Incorporating hydrology into climate suitability models changes projections of malaria transmission in Africa. Nat. Commun. 11 , 4353. 10.1038/s41467-020-18239-5 (2020). Smith, D. L. et al. Ross, Macdonald, and a Theory for the Dynamics and Control of Mosquito-Transmitted Pathogens. PLoS Pathog 8, e1002588. doi:1002510.1001371/journal.ppat.1002588 (2012). Okech, B. A., Gouagna, L. C., Yan, G., Githure, J. I. & Beier, J. C. Larval habitats of Anopheles gambiae s.s. (Diptera: Culicidae) influences vector competence to Plasmodium falciparum parasites. Malar. J. 6 , 50. 10.1186/1475-2875-6-50 (2007). Asmare, Y., Hopkins, R. J., Tekie, H., Hill, S. R. & Ignell, R. Grass Pollen Affects Survival and Development of Larval Anopheles arabiensis (Diptera: Culicidae). J. insect Sci. (Online) . 17 10.1093/jisesa/iex067 (2017). Shapiro, L. L., Murdock, C. C., Jacobs, G. R., Thomas, R. J. & Thomas, M. B. Larval food quantity affects the capacity of adult mosquitoes to transmit human malaria. Proc. Biol. Sci. 283 10.1098/rspb.2016.0298 (2016). Schapira, A. & Boutsika, K. Malaria Ecotypes and Stratification. Adv. Parasitol. 78 , 97–167. 10.1016/B978-0-12-394303-3.00001-3 (2012). Takken, W. & Lindsay, S. W. Factors affecting the vectorial competence of Anopheles gambiae : a question of scale. Ecol. Aspects Application Genetically Modified Mosquitoes Dordrecht: Kluwer Acad. Publishers , 75–90 (2003). Villena, O. C., Ryan, S. J., Murdock, C. C. & Johnson, L. R. Temperature impacts the environmental suitability for malaria transmission by Anopheles gambiae and Anopheles stephensi . Ecology . 103 , e3685 (2022). Moller-Jacobs, L. L., Murdock, C. C. & Thomas, M. B. Capacity of mosquitoes to transmit malaria depends on larval environment. Parasites vectors . 7 , 1–12 (2014). Charlwood, J. D. et al. The rise and fall of Anopheles arabiensis (Diptera: Culicidae) in a Tanzanian village. Bull. Entomol. Res. 85 , 37–44 (1995). Mendis, C. et al. Anopheles arabiensis and An. funestus are equally important vectors of malaria in Matola coastal suburb of Maputo, southern Mozambique. Med. Vet. Entomol. 14 , 171–180 (2000). Nambunga, I. H. et al. Aquatic habitats of the malaria vector Anopheles funestus in rural south-eastern Tanzania. Malar. J. 19 , 1–11 (2020). Lindsay, S. et al. Exposure of Gambian children to Anopheles gambiae malaria vectors in an irrigated rice production area. Med. Vet. Entomol. 9 , 50–58 (1995). Gillies, M. T. & Coetzee, M. A supplement to the Anophelinae of Africa South of the Sahara. Publ S Afr. Inst. Med. Res. 55 , 1–143 (1987). Mala, A. O. et al. Dry season ecology of Anopheles gambiae complex mosquitoes at larval habitats in two traditionally semi-arid villages in Baringo, Kenya. Parasites vectors . 4 , 1–11 (2011). Mwanziva, C. E. et al. Transmission intensity and malaria vector population structure in Magugu, Babati District in northern Tanzania. Tanzan. J. health Res. 13 , 54–61 (2011). Smith, D. L. & McKenzie, F. E. Statics and dynamics of malaria infection in Anopheles mosquitoes. Malar. J. 3 , 13. 10.1186/1475-2875-3-13 (2004). Kent, R. J., Thuma, P. E., Mharakurwa, S. & Norris, D. E. Seasonality, blood feeding behavior, and transmission of Plasmodium falciparum by Anopheles arabiensis after an extended drought in southern Zambia. Am. J. Trop. Med. Hyg. 76 , 267 (2007). Lyimo, I. N., Keegan, S. P., Ranford-Cartwright, L. C. & Ferguson, H. M. The impact of uniform and mixed species blood meals on the fitness of the mosquito vector Anopheles gambiae s.s : does a specialist pay for diversifying its host species diet? J. Evol. Biol. 25 , 452–460. 10.1111/j.1420-9101.2011.02442.x (2012). Kahamba, N. F. et al. Using ecological observations to improve malaria control in areas where Anopheles funestus is the dominant vector. Malar. J. 21 , 158. 10.1186/s12936-022-04198-3 (2022). Shaw, W. R. et al. Multiple blood feeding in mosquitoes shortens the Plasmodium falciparum incubation period and increases malaria transmission potential. PLoS Pathog . 16 , e1009131 (2020). Hofer, L. M. et al. Additional blood meals increase sporozoite infection in Anopheles mosquitoes but not Plasmodium falciparum genetic diversity. Sci. Rep. 14 , 17467 (2024). Beerntsen, B. T., James, A. A. & Christensen, B. M. Genetics of mosquito vector competence. Microbiol. Mol. Biol. Rev. 64 , 115–137 (2000). Hajkazemian, M., Bossé, C., Mozūraitis, R. & Emami, S. N. Battleground midgut: The cost to the mosquito for hosting the malaria parasite. Biol. Cell. 113 , 79–94. 10.1111/boc.202000039 (2021). Li, M. et al. Response of the mosquito immune system and symbiotic bacteria to pathogen infection. Parasit. Vectors . 17 , 69. 10.1186/s13071-024-06161-4 (2024). Emami, S. N. et al. A key malaria metabolite modulates vector blood seeking, feeding, and susceptibility to infection. Science . 355 , 1076–1080. 10.1126/science.aah4563 (2017). Nsango, S. E. et al. Genetic clonality of Plasmodium falciparum affects the outcome of infection in Anopheles gambiae . Int. J. Parasitol. 42 , 589–595. https://doi.org/10.1016/j.ijpara.2012.03.008 (2012). Takken, W. et al. Susceptibility of Anopheles quadriannulatus Theobald (Diptera: Culicidae) to Plasmodium falciparum . Trans. R Soc. Trop. Med. Hyg. 93 , 578–580. 10.1016/s0035-9203(99)90054-8 (1999). Habtewold, T., Povelones, M., Blagborough, A. M. & Christophides, G. K. Transmission blocking immunity in the malaria non-vector mosquito Anopheles quadriannulatus species A. PLoS Pathog . 4 , e1000070. 10.1371/journal.ppat.1000070 (2008). Habtewold, T., Groom, Z. & Christophides, G. K. Immune resistance and tolerance strategies in malaria vector and non-vector mosquitoes. Parasit. Vectors . 10 , 186. 10.1186/s13071-017-2109-5 (2017). Gnémé, A. et al. Equivalent susceptibility of Anopheles gambiae M and S molecular forms and Anopheles arabiensis to Plasmodium falciparum infection in Burkina Faso. Malar. J. 12 , 204. 10.1186/1475-2875-12-204 (2013). Aschale, Y. et al. Systematic review of sporozoite infection rate of Anopheles mosquitoes in Ethiopia, 2001–2021. Parasit. Vectors . 16 , 437. 10.1186/s13071-023-06054-y (2023). Kabula, B. et al. Malaria entomological profile in Tanzania from 1950 to 2010: a review of mosquito distribution, vectorial capacity and insecticide resistance. Tanzan. J. Health Res. 13 , 319–331 (2011). Ntabaliba, W. et al. Life expectancy of Anopheles funestus is double that of Anopheles arabiensis in southeast Tanzania based on mark-release-recapture method. Sci. Rep. 13 , 15775. 10.1038/s41598-023-42761-3 (2023). Brady, O. J. et al. Vectorial capacity and vector control: reconsidering sensitivity to parameters for malaria elimination. Trans. R. Soc. Trop. Med. Hyg. 110 , 107–117 (2016). Shaw, W. R. & Catteruccia, F. Vector biology meets disease control: using basic research to fight vector-borne diseases. Nat. Microbiol. 4 , 20–34 (2019). Reiner, R. C. Jr. et al. A systematic review of mathematical models of mosquito-borne pathogen transmission: 1970–2010. J. R Soc. Interface . 10 , 20120921. 10.1098/rsif.2012.0921 (2013). Barreaux, A. M., Barreaux, P., Thievent, K. & Koella, J. C. Larval environment influences vector competence of the malaria mosquito Anopheles gambiae . Malar. World J. 7 , 1–6 (2016). White, B. J., Collins, F. H. & Besansky, N. J. Evolution of Anopheles gambiae in relation to humans and malaria. Annu. Rev. Ecol. Evol. Syst. 42 , 111–132 (2011). Lwetoijera, D. W. et al. Increasing role of Anopheles funestus and Anopheles arabiensis in malaria transmission in the Kilombero Valley, Tanzania. Malar. J. 13 , 1–10 (2014). Matowo, N. S. et al. An increasing role of pyrethroid-resistant Anopheles funestus in malaria transmission in the Lake Zone, Tanzania. Sci. Rep. 11 , 13457 (2021). Ogola, E. O., Odero, J. O., Mwangangi, J. M. & Masiga, D. K. Tchouassi, D. P. Population genetics of Anopheles funestus , the African malaria vector, Kenya. Parasites vectors . 12 , 1–9 (2019). Msugupakulya, B. J. et al. Changes in contributions of different Anopheles vector species to malaria transmission in east and southern Africa from 2000 to 2022. Parasites Vectors . 16 , 408 (2023). Wangrawa, D. W., Odero, J. O., Baldini, F., Okumu, F. & Badolo, A. Distribution and insecticide resistance profile of the major malaria vector Anopheles funestus group across the African continent. Med. Vet. Entomol. 38 , 119–137 (2024). Small, S. T. et al. Radiation with reticulation marks the origin of a major malaria vector. Proc. Natl. Acad. Sci. U S A . 117 , 31583–31590. 10.1073/pnas.2018142117 (2020). Suh, P. F. et al. Impact of insecticide resistance on malaria vector competence: a literature review. Malar. J. 22 , 19 (2023). Alout, H. et al. Insecticide resistance alleles affect vector competence of Anopheles gambiae ss for Plasmodium falciparum field isolates. PLoS One . 8 , e63849 (2013). Barreaux, P., Koella, J. C., N’Guessan, R. & Thomas, M. B. Use of novel lab assays to examine the effect of pyrethroid-treated bed nets on blood-feeding success and longevity of highly insecticide-resistant Anopheles gambiae sl mosquitoes. Parasites Vectors . 15 , 111 (2022). Minetti, C., Ingham, V. A. & Ranson, H. Effects of insecticide resistance and exposure on Plasmodium development in Anopheles mosquitoes. Curr. Opin. insect Sci. 39 , 42–49 (2020). James, R. & Xu, J. Mechanisms by which pesticides affect insect immunity. J. Invertebr. Pathol. 109 , 175–182 (2012). Rivero, A., Vezilier, J., Weill, M., Read, A. F. & Gandon, S. Insecticide control of vector-borne diseases: when is insecticide resistance a problem? PLoS Pathog. 6 , e1001000 (2010). Mendes, A. M. et al. Infection intensity-dependent responses of Anopheles gambiae to the African malaria parasite Plasmodium falciparum. Infect. Immun. 79 , 4708–4715 (2011). Cirimotich, C. M., Ramirez, J. L. & Dimopoulos, G. Native microbiota shape insect vector competence for human pathogens. Cell. host microbe . 10 , 307–310 (2011). Harris, C. et al. Plasmodium falciparum produce lower infection intensities in local versus foreign Anopheles gambiae populations. PloS one . 7 , e30849 (2012). Henry, B. Assessment of the transmission blocking activity of antimalarial compounds by membrane feeding assays using natural Plasmodium falciparum gametocyte isolates from West-Africa. Plos one . 18 , e0284751 (2023). Kweyamba, P. A. et al. Sub-lethal exposure to chlorfenapyr reduces the probability of developing Plasmodium falciparum parasites in surviving Anopheles mosquitoes. Parasites Vectors . 16 , 342 (2023). Habtewold, T. et al. Streamlined SMFA and mosquito dark-feeding regime significantly improve malaria transmission-blocking assay robustness and sensitivity. Malar. J. 18 , 1–11 (2019). Benedict, M. (2018). Chomczynski, P., Mackey, K., Drews, R. & Wilfinger, W. DNAzol®: a reagent for the rapid isolation of genomic DNA. Biotechniques . 22 , 550–553 (1997). Schindler, T. et al. Molecular monitoring of the diversity of human pathogenic malaria species in blood donations on Bioko Island, Equatorial Guinea. Malar. J. 18 , 1–11 (2019). Whelan, J. A., Russell, N. B. & Whelan, M. A. A method for the absolute quantification of cDNA using real-time PCR. J. Immunol. Methods . 278 , 261–269 (2003). Mercereau-Puijalon, O., Barale, J. C. & Bischoff, E. Three multigene families in Plasmodium parasites: facts and questions. Int. J. Parasitol. 32 , 1323–1344 (2002). Health, M. & Welfare, S. (United Republic of Tanzania Dar es Salam, (2014). Additional Declarations No competing interests reported. Supplementary Files VCOocystsData.xlsx VCSporozoiteData.xlsx Cite Share Download PDF Status: Under Review Version 1 posted Editorial decision: Revision requested 11 Nov, 2024 Reviews received at journal 11 Nov, 2024 Reviewers agreed at journal 22 Oct, 2024 Reviews received at journal 23 Sep, 2024 Reviewers agreed at journal 16 Sep, 2024 Reviewers agreed at journal 15 Sep, 2024 Reviewers invited by journal 13 Sep, 2024 Editor assigned by journal 13 Sep, 2024 Editor invited by journal 13 Sep, 2024 Submission checks completed at journal 12 Sep, 2024 First submitted to journal 05 Sep, 2024 You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-5038559","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Article","associatedPublications":[],"authors":[{"id":368931511,"identity":"4aa81822-2b62-4a2d-be78-b130cc2681d5","order_by":0,"name":"Prisca A. Kweyamba","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAA2klEQVRIiWNgGAWjYDACHgiVwMDe2AYkSdLCcxCqhY1oLRIJULWEtJjznDF8zFOzLY9/5uO2Bw8YbPIZ5Nsf4NVi2dtjbMxz7HaxxO3EdoMEhjTLBjYCXjI4z7tNmoftdmLD7cQ2iQSGwwZAhx0gpGX7b55/txPn3zwI08LYgF/L2d5tzLxttxM33GCEaWHGq4PBsuf8Z8m5fbeLDc+AHGaQZsDGloZfizlPWuKHN99u58kdP/5M8keFjQE/8/EH+B0GxEw8yFyCMQnSwviDkKpRMApGwSgY2QAAPexFIO1t1cEAAAAASUVORK5CYII=","orcid":"","institution":"Ifakara Health Institute","correspondingAuthor":true,"prefix":"","firstName":"Prisca","middleName":"A.","lastName":"Kweyamba","suffix":""},{"id":368931512,"identity":"0c0f25f7-cbb8-4689-a304-6c8c653b9ef7","order_by":1,"name":"Lorenz M. Hofer","email":"","orcid":"","institution":"Swiss Tropical and Public Health Institute","correspondingAuthor":false,"prefix":"","firstName":"Lorenz","middleName":"M.","lastName":"Hofer","suffix":""},{"id":368931513,"identity":"63f269dd-9c94-4afc-a64b-2eeec879bc9b","order_by":2,"name":"Ummi A. Kibondo","email":"","orcid":"","institution":"Ifakara Health Institute","correspondingAuthor":false,"prefix":"","firstName":"Ummi","middleName":"A.","lastName":"Kibondo","suffix":""},{"id":368931514,"identity":"ec6b6dc5-c5db-4aea-97af-d5604206eb4f","order_by":3,"name":"Rehema Y. Mwanga","email":"","orcid":"","institution":"Ifakara Health Institute","correspondingAuthor":false,"prefix":"","firstName":"Rehema","middleName":"Y.","lastName":"Mwanga","suffix":""},{"id":368931515,"identity":"f4417202-943d-4abe-8ac2-8dfbaa38c7ff","order_by":4,"name":"Rajabu M. Sayi","email":"","orcid":"","institution":"Ifakara Health Institute","correspondingAuthor":false,"prefix":"","firstName":"Rajabu","middleName":"M.","lastName":"Sayi","suffix":""},{"id":368931516,"identity":"c89917db-72f4-4542-9253-7fd8ea79313c","order_by":5,"name":"Fatuma Matwewe","email":"","orcid":"","institution":"Ifakara Health Institute","correspondingAuthor":false,"prefix":"","firstName":"Fatuma","middleName":"","lastName":"Matwewe","suffix":""},{"id":368931517,"identity":"afc33d6d-b4ec-4c9b-a95c-83d8d5f4d7ec","order_by":6,"name":"Dickson W. Lwetoijera","email":"","orcid":"","institution":"Ifakara Health Institute","correspondingAuthor":false,"prefix":"","firstName":"Dickson","middleName":"W.","lastName":"Lwetoijera","suffix":""},{"id":368931518,"identity":"8ea891af-5013-45d4-a396-2a764148314b","order_by":7,"name":"Mgeni M. Tambwe","email":"","orcid":"","institution":"Ifakara Health Institute","correspondingAuthor":false,"prefix":"","firstName":"Mgeni","middleName":"M.","lastName":"Tambwe","suffix":""},{"id":368931519,"identity":"0e2efaaa-35da-48f0-a8f6-ee13893f6198","order_by":8,"name":"Sarah J. Moore","email":"","orcid":"","institution":"Ifakara Health Institute","correspondingAuthor":false,"prefix":"","firstName":"Sarah","middleName":"J.","lastName":"Moore","suffix":""}],"badges":[],"createdAt":"2024-09-05 13:36:14","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-5038559/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-5038559/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":67687589,"identity":"887ce0df-467f-4ff4-acb6-e991aeea2d8b","added_by":"auto","created_at":"2024-10-28 16:54:41","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":9713,"visible":true,"origin":"","legend":"\u003cp\u003eBurden and intensity of oocysts and sporozoites in Plasmodium infected \u003cem\u003eAnopheles \u003c/em\u003emosquitoes. A) Oocyst infected mosquitoes B) Sporozoite infected mosquitoes C) Number of oocysts D) Number of sporozoites\u003c/p\u003e","description":"","filename":"Onlinefloatimage1.png","url":"https://assets-eu.researchsquare.com/files/rs-5038559/v1/02c470eac1d76c81291a68d4.png"},{"id":67687590,"identity":"23895397-d628-412d-a183-07a27eda65a2","added_by":"auto","created_at":"2024-10-28 16:54:46","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":734297,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-5038559/v1/2b2e84c9-b84d-462a-9a97-0c67b49ad0b3.pdf"},{"id":67687587,"identity":"73905809-19e7-4c63-b9d5-b24ea3eb9492","added_by":"auto","created_at":"2024-10-28 16:54:41","extension":"xlsx","order_by":1,"title":"","display":"","copyAsset":false,"role":"supplement","size":22978,"visible":true,"origin":"","legend":"","description":"","filename":"VCOocystsData.xlsx","url":"https://assets-eu.researchsquare.com/files/rs-5038559/v1/142e4b345178f0e11ded8d3c.xlsx"},{"id":67687588,"identity":"49b1820e-5100-44e5-8529-4a16116bcff1","added_by":"auto","created_at":"2024-10-28 16:54:41","extension":"xlsx","order_by":2,"title":"","display":"","copyAsset":false,"role":"supplement","size":41908,"visible":true,"origin":"","legend":"","description":"","filename":"VCSporozoiteData.xlsx","url":"https://assets-eu.researchsquare.com/files/rs-5038559/v1/001d30881c887f2949c2bd3b.xlsx"}],"financialInterests":"No competing interests reported.","formattedTitle":"Contrasting vector competence of three main East African Anopheles malaria vector mosquitoes for Plasmodium falciparum","fulltext":[{"header":"Introduction","content":"\u003cp\u003e \u003cem\u003eAnopheles\u003c/em\u003e mosquitoes are the only arthropod vectors that transmit \u003cem\u003ePlasmodium\u003c/em\u003e parasites that cause malaria in humans. The disease imposes a significant burden of mortality and morbidity, particularly in Sub-Saharan Africa (SSA) \u003csup\u003e\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e\u003c/sup\u003e where the most efficient mosquito vectors of malaria are found \u003csup\u003e\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e\u003c/sup\u003e. Human malaria is mediated only by female \u003cem\u003eAnopheles\u003c/em\u003e, and of the estimated 460 species, only 40 species or species complexes are considered to be important vectors in the wild \u003csup\u003e\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e\u003c/sup\u003e, notably the \u003cem\u003eAnopheles gambiae\u003c/em\u003e and \u003cem\u003eAnopheles funestus\u003c/em\u003e species complexes that dominate malaria transmission throughout SSA\u003csup\u003e\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e\u003c/sup\u003e. In this paper, we focus on the three major East African vectors of human malaria: \u003cem\u003eAn. funestus sensu stricto\u003c/em\u003e, \u003cem\u003eAn. gambiae sensu stricto and An. arabiensis.\u003c/em\u003e\u003c/p\u003e \u003cp\u003eThe successful transmission of \u003cem\u003ePlasmodium\u003c/em\u003e parasites between humans requires intricate transformations within the mosquito vector \u003csup\u003e\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e\u003c/sup\u003e, highlighting the key role of both vectorial capacity \u003csup\u003e\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e\u003c/sup\u003e and vector competence \u003csup\u003e\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e\u003c/sup\u003e in determining the local intensity of malaria transmission \u003csup\u003e\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e\u003c/sup\u003e.\u003c/p\u003e \u003cp\u003eVector competence refers to the ability of an arthropod vector to acquire, maintain and transmit a pathogen. This concept encompasses the inherent ability of a pathogen to effectively enter and reproduce within the vector and be released from the vector's salivary glands to initiate infection in another vertebrate host \u003csup\u003e\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e,\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e\u003c/sup\u003e. Vectorial capacity describes the potential intensity of transmission by mosquitoes. It is defined as the total number of infectious mosquito bites on humans that will arise from a single infected person on a single day \u003csup\u003e\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e\u003c/sup\u003e. This is influenced by a number of factors \u003csup\u003e\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e,\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e\u003c/sup\u003e, most notably the probability of mosquitoes to feed on humans \u003csup\u003e\u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e\u003c/sup\u003e, daily vector survival \u003csup\u003e\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e\u003c/sup\u003e, environmental factors that affect the time it takes for parasites to develop in the mosquito host \u003csup\u003e\u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e\u003c/sup\u003e, the availability of larval breeding sites \u003csup\u003e\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e\u003c/sup\u003e, presence of vector control tools\u003csup\u003e\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e\u003c/sup\u003e and vector competence\u003csup\u003e\u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e\u003c/sup\u003e.\u003c/p\u003e \u003cp\u003eThe vectoral capacity of a particular vector is strongly influenced by its ecology. The larval stage of mosquitoes takes place in water where biological factors greatly influence the habitat suitability and carrying capacity. These factors influence vector presence \u003csup\u003e\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e\u003c/sup\u003e, fitness \u003csup\u003e\u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e\u003c/sup\u003e, longevity \u003csup\u003e\u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e\u003c/sup\u003e, which in turn affects the probability that a mosquito can acquire and maintain a parasite for long enough to become infectious \u003csup\u003e\u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e18\u003c/span\u003e\u003c/sup\u003e. In East Africa, the three major vectors have different regional distribution due to ecology \u003csup\u003e\u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e\u003c/sup\u003e and hydrology \u003csup\u003e\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e\u003c/sup\u003e and varying contributions to malaria transmission across different seasons \u003csup\u003e\u003cspan additionalcitationids=\"CR21 CR22\" citationid=\"CR20\" class=\"CitationRef\"\u003e20\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e23\u003c/span\u003e\u003c/sup\u003e. \u003cem\u003eAn. funestus\u003c/em\u003e mosquitoes have permanent breeding sites abundant with vegetation, making them likely to transmit malaria all year round \u003csup\u003e\u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e24\u003c/span\u003e,\u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e25\u003c/span\u003e\u003c/sup\u003e. \u003cem\u003eAn. arabiensis\u003c/em\u003e and \u003cem\u003eAn. gambiae s.s.\u003c/em\u003e typically dominate in temporary sunlit pools and their presence is strongly dependent on rainfall or irrigation \u003csup\u003e\u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e26\u003c/span\u003e,\u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e27\u003c/span\u003e\u003c/sup\u003e. \u003cem\u003eAn. gambiae s.s.\u003c/em\u003e requires high humidity to survive and occurs almost exclusively during humid and rainy periods \u003csup\u003e\u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e28\u003c/span\u003e\u003c/sup\u003e. \u003cem\u003eAn. arabiensis\u003c/em\u003e and \u003cem\u003eAn. funestus\u003c/em\u003e, are more resistant to desiccation, are commonly found in abundance during the peak of the wet season and continue into the dry season; sustaining malaria transmission for several months after the end of the rains\u003csup\u003e\u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e23\u003c/span\u003e,\u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e29\u003c/span\u003e\u003c/sup\u003e.\u003c/p\u003e \u003cp\u003e \u003cem\u003eAnopheles\u003c/em\u003e mosquitoes are dependent on vertebrate blood to provide proteins needed for egg development and undergo multiple cycles of feeding and egg development in their lifetime. Therefore, the preference of a vector for human blood has a direct impact on its efficiency as a vector by increasing its probability of acquiring and transmitting onward infection\u003csup\u003e\u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e30\u003c/span\u003e\u003c/sup\u003e. \u003cem\u003eAn. arabiensis\u003c/em\u003e have an opportunistic feeding behavior, targeting both human and animal hosts for its blood meals, so it may be a more or less important vector dependent on the relative proportion of cattle in an area \u003csup\u003e\u003cspan citationid=\"CR31\" class=\"CitationRef\"\u003e31\u003c/span\u003e\u003c/sup\u003e. \u003cem\u003eAn. gambiae\u003c/em\u003e \u003csup\u003e\u003cspan citationid=\"CR32\" class=\"CitationRef\"\u003e32\u003c/span\u003e\u003c/sup\u003eand \u003cem\u003eAn. funestus\u003c/em\u003e \u003csup\u003e\u003cspan citationid=\"CR33\" class=\"CitationRef\"\u003e33\u003c/span\u003e\u003c/sup\u003eare more specialized blood feeders, feeding almost entirely on humans, although this does depend on host availability. In addition, there is evidence that multiple blood meals increase the likelihood of \u003cem\u003ePlasmodium\u003c/em\u003e developing in the mosquito \u003csup\u003e\u003cspan citationid=\"CR34\" class=\"CitationRef\"\u003e34\u003c/span\u003e,\u003cspan citationid=\"CR35\" class=\"CitationRef\"\u003e35\u003c/span\u003e\u003c/sup\u003e.\u003c/p\u003e \u003cp\u003eAs well as environmental factors that affect vectors' susceptibility to infection and the interactions between the vector, pathogen, and host that impact probabilities of onward transmission, vector competence is influenced by a variety of internal factors, including the genetics of both the vector and the pathogen \u003csup\u003e\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e,\u003cspan citationid=\"CR36\" class=\"CitationRef\"\u003e36\u003c/span\u003e\u003c/sup\u003e. \u003cem\u003ePlasmodium\u003c/em\u003e takes resources from its definitive host that results in reduced fitness and reproductive output \u003csup\u003e\u003cspan citationid=\"CR37\" class=\"CitationRef\"\u003e37\u003c/span\u003e\u003c/sup\u003e. Therefore, the mosquito innate immune system either modulates or resists infection \u003csup\u003e\u003cspan citationid=\"CR38\" class=\"CitationRef\"\u003e38\u003c/span\u003e\u003c/sup\u003e, while the parasite counteracts mosquito defenses through host manipulation \u003csup\u003e\u003cspan citationid=\"CR39\" class=\"CitationRef\"\u003e39\u003c/span\u003e\u003c/sup\u003e and polyclonality\u003csup\u003e\u003cspan citationid=\"CR40\" class=\"CitationRef\"\u003e40\u003c/span\u003e\u003c/sup\u003e. Mosquito species show different levels of susceptibility to \u003cem\u003ePlasmodium\u003c/em\u003e from refractory in the case of \u003cem\u003eAnopheles quadriannulatus\u003c/em\u003e \u003csup\u003e\u003cspan citationid=\"CR41\" class=\"CitationRef\"\u003e41\u003c/span\u003e,\u003cspan citationid=\"CR42\" class=\"CitationRef\"\u003e42\u003c/span\u003e\u003c/sup\u003e to high susceptibility in \u003cem\u003eAnopheles coluzzi\u003c/em\u003e \u003csup\u003e\u003cspan citationid=\"CR43\" class=\"CitationRef\"\u003e43\u003c/span\u003e,\u003cspan citationid=\"CR44\" class=\"CitationRef\"\u003e44\u003c/span\u003e\u003c/sup\u003e.\u003c/p\u003e \u003cp\u003eIn East Africa, there is evidence of higher proportions of infected \u003cem\u003eAn. funestus\u003c/em\u003e \u003csup\u003e\u003cspan citationid=\"CR45\" class=\"CitationRef\"\u003e45\u003c/span\u003e,\u003cspan citationid=\"CR46\" class=\"CitationRef\"\u003e46\u003c/span\u003e\u003c/sup\u003e relative to \u003cem\u003eAn. arabiensis and An. gambiae\u003c/em\u003e. This can be to an extent explained by the fact that \u003cem\u003eAn. funestus\u003c/em\u003e generally feeds almost exclusively on humans and has been shown to live longer than \u003cem\u003eAn. arabiensis\u003c/em\u003e \u003csup\u003e\u003cspan citationid=\"CR47\" class=\"CitationRef\"\u003e47\u003c/span\u003e\u003c/sup\u003e. However, any differences in degree of vector competence among the three primary malaria vectors has not been evaluated. Understanding vector competence is crucial in understanding the risk of malaria transmission, informing effective malaria control strategies \u003csup\u003e\u003cspan citationid=\"CR36\" class=\"CitationRef\"\u003e36\u003c/span\u003e,\u003cspan citationid=\"CR48\" class=\"CitationRef\"\u003e48\u003c/span\u003e,\u003cspan citationid=\"CR49\" class=\"CitationRef\"\u003e49\u003c/span\u003e\u003c/sup\u003e and parameterizing mathematical models, where mosquito-parasite interactions are rarely considered \u003csup\u003e\u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e,\u003cspan citationid=\"CR50\" class=\"CitationRef\"\u003e50\u003c/span\u003e\u003c/sup\u003e.\u003c/p\u003e \u003cp\u003eTherefore, this study investigated whether vector competence towards \u003cem\u003ePlasmodium falciparum\u003c/em\u003e differs between local East African strains of \u003cem\u003eAn. gambiae s.s.\u003c/em\u003e, \u003cem\u003eAn. funestus\u003c/em\u003e and \u003cem\u003eAn. arabiensis\u003c/em\u003e mosquitoes. By experimentally infecting mosquitoes with field gametocytes using Direct Membrane Feeding Assays (DMFAs), we aim to compare the prevalence and intensity of \u003cem\u003eP. falciparum\u003c/em\u003e infection between these \u003cem\u003eAnopheles\u003c/em\u003e mosquito species.\u003c/p\u003e"},{"header":"Results","content":"\u003cp\u003e \u003cb\u003ePrevalence and intensity of\u003c/b\u003e \u003cb\u003eP. falciparum\u003c/b\u003e \u003cb\u003einfection among local strains of\u003c/b\u003e \u003cb\u003eAn. gambiae s.s.\u003c/b\u003e, \u003cb\u003eAn. funestus\u003c/b\u003e \u003cb\u003eand\u003c/b\u003e \u003cb\u003eAn. arabiensis\u003c/b\u003e \u003cb\u003emosquitoes\u003c/b\u003e\u003c/p\u003e \u003cdiv id=\"Sec3\" class=\"Section2\"\u003e \u003ch2\u003eOocysts\u003c/h2\u003e \u003cp\u003eThe prevalence of oocyst-infected mosquitoes varied among the three \u003cem\u003eAnopheles\u003c/em\u003e mosquito species, with \u003cem\u003eAn. funestus\u003c/em\u003e presenting the highest oocyst infection rate of 13.5% (95% CI: 9.2\u0026ndash;17.6), followed by \u003cem\u003eAn. gambiae s.s.\u003c/em\u003e at 10.7% (95% CI: 6.9\u0026ndash;14.4), and \u003cem\u003eAn. arabiensis\u003c/em\u003e at 5.6% (95% CI: 2.5\u0026ndash;8.7) (Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e). The proportion of oocyst-infected \u003cem\u003eAn. arabiensis\u003c/em\u003e mosquitoes was significantly lower than \u003cem\u003eAn. funestus\u003c/em\u003e mosquitoes (OR\u0026thinsp;=\u0026thinsp;0.40, 95% CI: 0.20\u0026ndash;0.80, p\u0026thinsp;=\u0026thinsp;0.010), but there was no difference between \u003cem\u003eAn. funestus\u003c/em\u003e and \u003cem\u003eAn. gambiae\u003c/em\u003e (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eA). Additionally, there was no significant difference in the proportion of oocyst-infected mosquitoes between \u003cem\u003eAn. gambiae s.s.\u003c/em\u003e and \u003cem\u003eAn. arabiensis\u003c/em\u003e mosquitoes (OR\u0026thinsp;=\u0026thinsp;0.52, 95% CI: 0.25\u0026ndash;1.06, p\u0026thinsp;=\u0026thinsp;0.072). There was no significant difference in oocyst intensity between the three species, although as with prevalence, oocyst intensity was similar between \u003cem\u003eAn. funestus\u003c/em\u003e, which ranged from 1\u0026ndash;12 and \u003cem\u003eAn. gambiae s.s.\u003c/em\u003e, ranging from 1\u0026ndash;14; and lower among \u003cem\u003eAn. arabiensis\u003c/em\u003e, which presented the lowest intensity of oocysts ranging from 1\u0026ndash;3 (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eC).\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec4\" class=\"Section2\"\u003e \u003ch2\u003eSporozoites\u003c/h2\u003e \u003cp\u003eThe burden of sporozoite-infected mosquitoes varied among the three \u003cem\u003eAnopheles\u003c/em\u003e mosquito species with \u003cem\u003eAn. funestus\u003c/em\u003e presenting the highest sporozoite infection rate of 23.5% (95% CI: 17.5\u0026ndash;29.6), followed by \u003cem\u003eAn. gambiae s.s.\u003c/em\u003e at 11.4% (95% CI: 6.5\u0026ndash;16.3), and \u003cem\u003eAn. arabiensis\u003c/em\u003e at 4.9% (95% CI: 0.6\u0026ndash;9.1) (Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e). \u003cem\u003eAn. funestus\u003c/em\u003e had a higher probability of being sporozoite-infected than \u003cem\u003eAn. gambiae s.s.\u003c/em\u003e (OR\u0026thinsp;=\u0026thinsp;0.46, 95% CI: 0.25\u0026ndash;0.85, p\u0026thinsp;=\u0026thinsp;0.013) or \u003cem\u003eAn. arabiensis\u003c/em\u003e (OR\u0026thinsp;=\u0026thinsp;0.19, 95% CI: 0.07\u0026ndash;0.51, p\u0026thinsp;=\u0026thinsp;0.001), (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eB). Moreover, there was no statistically significant difference observed in the proportion of sporozoite infection between \u003cem\u003eAn. gambiae s.s.\u003c/em\u003e and \u003cem\u003eAn. arabiensis\u003c/em\u003e mosquitoes (OR\u0026thinsp;=\u0026thinsp;0.41, 95% CI: 0.14\u0026ndash;1.17 P\u0026thinsp;=\u0026thinsp;0.098), although there were few infected \u003cem\u003eAn. arabiensis\u003c/em\u003e. Additionally, there were similar trends observed in sporozoite intensity among the three species, with highest intensity observed in \u003cem\u003eAn. funestus\u003c/em\u003e, ranging from 6\u0026ndash;58,138 followed by \u003cem\u003eAn. gambiae s.s.\u003c/em\u003e, ranging from 21\u0026thinsp;\u0026minus;\u0026thinsp;7,976 and \u003cem\u003eAn. arabiensis\u003c/em\u003e ranging from 20\u0026ndash;27,877. A significant difference was observed in sporozoite intensity between \u003cem\u003eAn. funestus\u003c/em\u003e and \u003cem\u003eAn. gambiae s.s.\u003c/em\u003e (IRR\u0026thinsp;=\u0026thinsp;0.31, 95% CI: 0.14\u0026ndash;0.69, p\u0026thinsp;=\u0026thinsp;0.004) (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eD), although not between \u003cem\u003eAn. funestus\u003c/em\u003e and \u003cem\u003eAn. arabiensis\u003c/em\u003e likely due to low sporozoite prevalence (5%) in \u003cem\u003eAn. arabiensis\u003c/em\u003e leading to uncertainty in the estimates.\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab1\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 1\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eProportion and intensity of oocyst and sporozoite infected laboratory reared mosquitoes with gametocytaemic blood from participants in Bagamoyo, Tanzania\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"6\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c6\" colnum=\"6\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/th\u003e \u003cth align=\"left\" colspan=\"3\" nameend=\"c4\" namest=\"c2\"\u003e \u003cp\u003eProportion infected\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colspan=\"2\" nameend=\"c6\" namest=\"c5\"\u003e \u003cp\u003eIntensity\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u003cb\u003eNumber infected\u003c/b\u003e\u003c/p\u003e \u003cp\u003e\u003cb\u003e(n/N)\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u003cb\u003ePrevalence (95%CI)\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e\u003cb\u003eOR (95%CI)\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e\u003cb\u003eMedian (Min-Max)\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e\u003cb\u003eIRR (95%CI)\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eOocysts\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eAn. funestus s.s.\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e35/260\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e13.5 (9.2\u0026ndash;17.6)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e2 (1\u0026ndash;12)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e1\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eAn.gambiae s.s.\u003c/em\u003e\u003c/p\u003e \u003cp\u003e\u003cem\u003eAn. arabiensis\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e29/271\u003c/p\u003e \u003cp\u003e12/214\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e10.7 (6.9\u0026ndash;14.4)\u003c/p\u003e \u003cp\u003e5.6 (2.5\u0026ndash;8.7)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e0.76 (0.44\u0026ndash;1.31) *\u003c/p\u003e \u003cp\u003e0.40 (0.20\u0026ndash;0.80) *\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e1 (1\u0026ndash;14)\u003c/p\u003e \u003cp\u003e2 (1\u0026ndash;3)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e1.31 (0.83\u0026ndash;2.06)\u003c/p\u003e \u003cp\u003e1.04 (0.55\u0026ndash;1.95)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eSporozoites\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eAn. funestus s.s.\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e46/195\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e23.5 (17.5\u0026ndash;29.6)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e1\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e3,983 (6\u0026ndash;58,138)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e1\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cem\u003eAn.gambiae s.s.\u003c/em\u003e\u003c/p\u003e \u003cp\u003e\u003cem\u003eAn. arabiensis\u003c/em\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e19/166\u003c/p\u003e \u003cp\u003e05/102\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e11.4 (6.5\u0026ndash;16.3)\u003c/p\u003e \u003cp\u003e4.9 (0.6\u0026ndash;9.1)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e0.46 (0.25\u0026ndash;0.85) **\u003c/p\u003e \u003cp\u003e0.19 (0.07\u0026ndash;0.51) **\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e2,447 (21\u0026thinsp;\u0026minus;\u0026thinsp;7,976)\u003c/p\u003e \u003cp\u003e714 (20\u0026ndash;27,877)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e0.31 (0.14\u0026ndash;0.69) **\u003c/p\u003e \u003cp\u003e0.66 (0.16\u0026ndash;2.60) **\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003ctfoot\u003e \u003ctr\u003e\u003ctd colspan=\"6\"\u003e\u003cem\u003eNote: OR were derived from mixed-effect logistic regression while IRR were derived from mixed effect negative binomial regression using study participant as a random effect\u003c/em\u003e\u003c/td\u003e\u003c/tr\u003e \u003ctr\u003e\u003ctd colspan=\"6\"\u003e*\u003cem\u003ep-value\u0026thinsp;\u0026lt;\u0026thinsp;0.05, *\u003c/em\u003e*p-value\u0026thinsp;\u0026lt;\u0026thinsp;0.01\u003c/td\u003e\u003c/tr\u003e \u003c/tfoot\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003c/div\u003e"},{"header":"Discussion","content":"\u003cp\u003eThis study represents the first attempt to measure whether the degree of vector competence towards \u003cem\u003eP. falciparum\u003c/em\u003e infection varies between East African \u003cem\u003eAn. funestus s.s.\u003c/em\u003e, \u003cem\u003eAn. arabiensis\u003c/em\u003e, and \u003cem\u003eAn. gambiae s.s.\u003c/em\u003e mosquitoes. Through experimental feeding of mosquitoes with gametocytaemic blood from donors in Tanzania, we observed significant differences in both prevalence and intensity of oocyst and sporozoite infections among the three major vector species. \u003cem\u003eAn. funestus s.s. mosquitoes\u003c/em\u003e presented with the highest burden and intensity of infection indicating that they are more competent than either \u003cem\u003eAn. gambiae s.s.\u003c/em\u003e or \u003cem\u003eAn. arabiensis\u003c/em\u003e. This is the first time, to our knowledge that the vectoral competence of \u003cem\u003eAn. funestus s.s.\u003c/em\u003e has been evaluated. The findings of this study largely agreed with the findings of a study in Burkina Faso, that found no difference in genetic susceptibility to \u003cem\u003eP. falciparum\u003c/em\u003e measured by oocyst infection between three sympatric population groups of the \u003cem\u003eAn. gambiae s.l.\u003c/em\u003e complex including \u003cem\u003eAn. coluzzi, An. gambiae\u003c/em\u003e s.s. and \u003cem\u003eAn. arabiensis\u003c/em\u003e that had been reared from wild-caught larvae \u003csup\u003e\u003cspan citationid=\"CR44\" class=\"CitationRef\"\u003e44\u003c/span\u003e\u003c/sup\u003e.\u003c/p\u003e \u003cp\u003e \u003cem\u003eAnopheles\u003c/em\u003e mosquitoes are reported to have varying levels of vector competence, influenced by genetic factors, as well as environmental conditions, and host-parasite interactions \u003csup\u003e\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e,\u003cspan citationid=\"CR36\" class=\"CitationRef\"\u003e36\u003c/span\u003e,\u003cspan citationid=\"CR51\" class=\"CitationRef\"\u003e51\u003c/span\u003e\u003c/sup\u003e. \u003cem\u003eAn. gambiae s.s.\u003c/em\u003e has long been recognized as a highly polymorphic and efficient malaria vector, possibly due to the strong co-adaptation of \u003cem\u003eP. falciparum\u003c/em\u003e to this specific mosquito species \u003csup\u003e\u003cspan citationid=\"CR52\" class=\"CitationRef\"\u003e52\u003c/span\u003e\u003c/sup\u003e. However, our study reveals that \u003cem\u003eAn. funestus\u003c/em\u003e were more competent. This finding is consistent with reports of increasing malaria cases caused by \u003cem\u003eAn. funestus\u003c/em\u003e in Tanzania and other parts of SSA\u003csup\u003e\u003cspan additionalcitationids=\"CR54 CR55 CR56\" citationid=\"CR53\" class=\"CitationRef\"\u003e53\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR57\" class=\"CitationRef\"\u003e57\u003c/span\u003e\u003c/sup\u003e. The high prevalence and intensity of infection observed in \u003cem\u003eAn. funestus\u003c/em\u003e mosquitoes suggest its potential as a predominant malaria vector, particularly in areas with suitable breeding sites abundant in vegetation. Our results are consistent with the observation that the massive introgression event that lead to the evolution of \u003cem\u003eAnopheles funestus\u003c/em\u003e 13,000 years ago that facilitated its adaptation to new environments resulting in its subsequent dramatic geographic range expansion across most of tropical Africa also enhanced vectorial capacity in \u003cem\u003eAnopheles funestus\u003c/em\u003e mosquitoes \u003csup\u003e\u003cspan citationid=\"CR58\" class=\"CitationRef\"\u003e58\u003c/span\u003e\u003c/sup\u003e.\u003c/p\u003e \u003cp\u003eAlthough \u003cem\u003eAn. arabiensis\u003c/em\u003e mosquitoes play a crucial role in malaria transmission, particularly in arid regions in the Horn of Africa \u003csup\u003e\u003cspan citationid=\"CR45\" class=\"CitationRef\"\u003e45\u003c/span\u003e\u003c/sup\u003e, our findings show lower prevalence and intensity of oocyst and sporozoite infections when compared to \u003cem\u003eAn. funestus\u003c/em\u003e and \u003cem\u003eAn. gambiae s.s.\u003c/em\u003e mosquito infections. While \u003cem\u003eAn. arabiensis\u003c/em\u003e is often abundant, and is widely discussed as a vector of residual and outdoor malaria, the more competent and endophilic malaria vectors \u003cem\u003eAn. funestus s.s.\u003c/em\u003e and \u003cem\u003eAn. gambiae s.s.\u003c/em\u003e should be targeted for control.\u003c/p\u003e \u003cp\u003eWhile several studies have shown that insecticide resistant vectors are more competent to \u003cem\u003ePlasmodium\u003c/em\u003e \u003csup\u003e\u003cspan citationid=\"CR59\" class=\"CitationRef\"\u003e59\u003c/span\u003e,\u003cspan citationid=\"CR60\" class=\"CitationRef\"\u003e60\u003c/span\u003e\u003c/sup\u003e, our study also found that \u003cem\u003eAn. funestus\u003c/em\u003e mosquitoes which are resistant to pyrethroids showed increased susceptibility to \u003cem\u003ePlasmodium\u003c/em\u003e infection. The susceptibility of mosquitoes to \u003cem\u003ePlasmodium\u003c/em\u003e infection may result from either their increased survival and longevity\u003csup\u003e\u003cspan citationid=\"CR61\" class=\"CitationRef\"\u003e61\u003c/span\u003e\u003c/sup\u003e compared to susceptible mosquitoes, which are killed by insecticide\u003csup\u003e\u003cspan citationid=\"CR62\" class=\"CitationRef\"\u003e62\u003c/span\u003e\u003c/sup\u003e, or could be due to reduced immunity to parasites\u003csup\u003e\u003cspan citationid=\"CR63\" class=\"CitationRef\"\u003e63\u003c/span\u003e\u003c/sup\u003e. Therefore, we cannot rule out that the difference in insecticide susceptibility affected the results through a change in mosquito immunity to parasites. There is strong evidence suggesting that insecticide resistance mutations increase the vector competence of \u003cem\u003eAn. gambiae\u003c/em\u003e for \u003cem\u003ePlasmodium\u003c/em\u003e, potentially sustaining malaria transmission\u003csup\u003e\u003cspan citationid=\"CR60\" class=\"CitationRef\"\u003e60\u003c/span\u003e\u003c/sup\u003e. However, the An. arabiensis used in the study were also pyrethroid resistant and were still relatively less susceptible to infection.\u003c/p\u003e \u003cp\u003eAn additional study has shown that insecticide resistance mechanisms have an effect on the activation of the mosquito immune system and its physiology, resulting in differences in parasite development and survival \u003csup\u003e\u003cspan citationid=\"CR64\" class=\"CitationRef\"\u003e64\u003c/span\u003e\u003c/sup\u003e. Variations in parasite burden may not significantly affect parasite transmission, however, the intensity of infection does influence the activation of the vector immune system \u003csup\u003e\u003cspan citationid=\"CR65\" class=\"CitationRef\"\u003e65\u003c/span\u003e\u003c/sup\u003e. Nonetheless, in our study, all three species showed susceptibility to \u003cem\u003ePlasmodium\u003c/em\u003e infection, with \u003cem\u003eAn. funestus\u003c/em\u003e presenting a higher susceptibility compared to \u003cem\u003eAn. gambiae s.s.\u003c/em\u003e and \u003cem\u003eAn. arabiensis\u003c/em\u003e. It was suggested that the susceptibility of \u003cem\u003eAn. gambiae\u003c/em\u003e to \u003cem\u003ePlasmodium\u003c/em\u003e infection is due to persistent immune suppression to prevent excessive activation of the immune response following blood meal ingestion\u003csup\u003e\u003cspan citationid=\"CR66\" class=\"CitationRef\"\u003e66\u003c/span\u003e\u003c/sup\u003e. Moreover, research has shown that genetic diversity within mosquito populations can also significantly influence their susceptibility to \u003cem\u003ePlasmodium\u003c/em\u003e infection\u003csup\u003e\u003cspan citationid=\"CR67\" class=\"CitationRef\"\u003e67\u003c/span\u003e\u003c/sup\u003e.\u003c/p\u003e \u003cp\u003eThis study highlighted that, while \u003cem\u003eAn. gambiae\u003c/em\u003e mosquitoes are commonly used in DMFAs to assess different malaria transmission-blocking interventions\u003csup\u003e\u003cspan additionalcitationids=\"CR69\" citationid=\"CR68\" class=\"CitationRef\"\u003e68\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR70\" class=\"CitationRef\"\u003e70\u003c/span\u003e\u003c/sup\u003e, the observed shift towards \u003cem\u003eAn. funestus\u003c/em\u003e as a major contributor to transmission in SSA\u003csup\u003e\u003cspan citationid=\"CR56\" class=\"CitationRef\"\u003e56\u003c/span\u003e\u003c/sup\u003e, with the highest infection burden, suggests the importance of incorporating \u003cem\u003eAn. funestus\u003c/em\u003e mosquitoes into assays for testing malaria transmission-blocking activity. A limitation of our study is the exclusive use of laboratory reared mosquito strains rather than actual field mosquitoes.\u003c/p\u003e \u003cp\u003eIn conclusion, we confirmed that between the three mosquito species, \u003cem\u003eAn. funestus\u003c/em\u003e was the most permissive to \u003cem\u003eP. falciparum\u003c/em\u003e infection, which is coherent with consistently high sporozoite rates observed in this species across SSA, whereas \u003cem\u003eAn. arabiensis\u003c/em\u003e shows the greatest resistance coherent with its lower observed sporozoites rates. Nonetheless, our findings suggest that all the three vector species play an active role in malaria transmission. The observed differences in vector competence among the three \u003cem\u003eAnopheles\u003c/em\u003e species highlight the complexity of malaria transmission dynamics and the need for comprehensive malaria control strategies that target key malaria vector species. Furthermore, malaria transmission models should be revised to account for vectoral competence, and efforts to support malaria transmission blocking interventions tested on multiple malaria vectors are essential for making sustainable progress towards malaria elimination.\u003c/p\u003e"},{"header":"Methods","content":"\u003cdiv id=\"Sec7\" class=\"Section2\"\u003e \u003ch2\u003eMosquito rearing\u003c/h2\u003e \u003cp\u003e\u003cem\u003eAn. gambiae s.s.\u003c/em\u003e mosquitoes were originally collected from the southern region of Tanzania (Njage-Mngeta villages, Ifakara district, Morogoro region) in 1996 and have been maintained at the Ifakara Health Institute (IHI) insectaries, Tanzania. This strain of \u003cem\u003eAnopheles gambiae s.s.\u003c/em\u003e is susceptible to all classes of insecticides. Field-collected \u003cem\u003eAn. arabiensis\u003c/em\u003e mosquitoes were obtained from the southern region of Tanzania (Sakamaganga village, Ifakara district) in 2005 and have been maintained in IHI insectaries. This strain is resistant to pyrethroids at a 1x discriminating concentration and is susceptible to other classes. The \u003cem\u003eAn. funestus\u003c/em\u003e colony was established at IHI insectaries in 2018 and was originally derived from founder colony established in 2000 at the National Institute for Communicable Diseases (NICD) South Africa. This strain is resistant to pyrethroids at a 1x discriminating concentration and is susceptible to other classes. Mosquito larvae were maintained at a density of 200 larvae per litre of water and fed 0.3 g per larva on Tetramin fish food (Tetra Ltd., UK). For colony maintenance, the adult mosquitoes are provided with cow blood between 3 and 6 days after emergence for egg development using a Hemotek\u0026reg; membrane feeder (SP-6 System, Hemotek Ltd., Blackburn BB6 7FD, UK). Mosquitoes were provided with autoclaved 10% sucrose solution ad libitum. Temperature and humidity within the insectary are maintained between 27\u0026thinsp;\u0026plusmn;\u0026thinsp;2\u003csup\u003eo\u003c/sup\u003eC and 60%-85% relative humidity following the MR4 guidance\u003csup\u003e\u003cspan citationid=\"CR71\" class=\"CitationRef\"\u003e71\u003c/span\u003e\u003c/sup\u003e.\u003c/p\u003e \u003c/div\u003e\n\u003ch3\u003eRecruitment of asymptomatic gametocytaemic carriers\u003c/h3\u003e\n\u003cp\u003eGametocytaemic carriers were selected by screening thick blood smears from participants aged 6\u0026ndash;40 years located in the village of Wami-Mkoko, in Bagamoyo district located in the coastal region of Tanzania, between June 2023 and August 2023. Participants meeting the inclusion criteria (asymptomatic individuals aged 6\u0026ndash;40 who consented and had microscopically detectable gametocytes) were enrolled for blood collection at IHI transmission facilities in Bagamoyo, Tanzania.\u003c/p\u003e \u003cp\u003eGametocytes were quantified by counting against 500 white blood cells in thick smears, and their density was calculated based on an estimated leukocyte density of 8000/ \u0026micro;L of whole blood. Five milliliters of blood were obtained from microscopically confirmed gametocyte carriers with gametocytes density exceeding three gametocytes/500 red blood cells, equivalent to 48 gametocytes/ \u0026micro;L of whole blood. Seven gametocytaemic individuals were recruited to donate blood for DMFAs. Autologous serum was replaced with pre-warmed malaria-na\u0026iuml;ve AB serum European donors.\u003c/p\u003e \u003cp\u003e \u003cb\u003eExperimental infection of\u003c/b\u003e \u003cb\u003eP. falciparum\u003c/b\u003e \u003cb\u003ein\u003c/b\u003e \u003cb\u003eAnopheles\u003c/b\u003e \u003cb\u003emosquitoes through DMFAs\u003c/b\u003e\u003c/p\u003e \u003cp\u003eInfectious gametocytaemic blood was administered to mosquitoes through water-jacketed glass feeders (14mm \u0026Oslash;, Chemglass, New Jersey, USA) covered with parafilm\u0026reg;, connected to a circulating water bath (39\u003csup\u003eo\u003c/sup\u003eC, ELMI, Switzerland) via plastic tubing. On average, 200 mosquitoes from each mosquito strain were fed a blood meal from each participant for a duration of 15 minutes. After blood feeding, the cups containing mosquitoes were then transferred to Bugdorm plastic cages (30 cm x 30 cm x 30 cm, Megaview Science Co., LTD, Taiwan) and placed in a climatic chamber (S600PLH, AraLab, Lisbon, Portugal) maintained at 75\u0026thinsp;\u0026plusmn;\u0026thinsp;2% humidity and 27\u0026thinsp;\u0026plusmn;\u0026thinsp;1\u0026deg;C at 12:12 hours dark: light cycle. Mosquitoes were deprived of sugar for 48 hours to allow unfed mosquitoes to die. Dead mosquitoes were aspirated out after 48 hours and then cotton soaked with autoclaved 10% sucrose solution was provided and replaced daily.\u003c/p\u003e \u003cdiv id=\"Sec9\" class=\"Section2\"\u003e \u003ch2\u003eOocyst and Sporozoite scoring\u003c/h2\u003e \u003cp\u003eEight days post infection (dpi), one-third of mosquitoes from each mosquito strain was dissected and their midguts were stained with a 1% mercurochrome solution before examination for presence of oocysts microscopically. The remaining mosquitoes were kept up to day 16 dpi and the mosquito`s DNA was extracted using DNAzol\u0026reg; reagent \u003csup\u003e\u003cspan citationid=\"CR72\" class=\"CitationRef\"\u003e72\u003c/span\u003e\u003c/sup\u003e for molecular analysis and quantification of \u003cem\u003eP. falciparum\u003c/em\u003e infection in mosquito stages from the mosquito heads and thoraces (sporozoites stages). Using quantitative reverse transcription polymerase chain reaction (RT-qPCR)\u003csup\u003e\u003cspan citationid=\"CR73\" class=\"CitationRef\"\u003e73\u003c/span\u003e\u003c/sup\u003e, absolute quantification of all sporozoites positive samples was performed using the standard curves generated on \u003cem\u003eP. falciparum\u003c/em\u003e-specific 18S rDNA plasmid (GenBank: AF145334) from \u003cem\u003eP. falciparum\u003c/em\u003e (BEI Resources, NIAID, MRA-177). Plasmid copy numbers per \u0026micro;l were calculated as described elsewhere\u003csup\u003e\u003cspan citationid=\"CR74\" class=\"CitationRef\"\u003e74\u003c/span\u003e\u003c/sup\u003e. The standard curves were generated on serial dilutions over eight magnitudes assuming an average of six copies of the 18S rDNA gene sequence per parasite genome\u003csup\u003e\u003cspan citationid=\"CR75\" class=\"CitationRef\"\u003e75\u003c/span\u003e\u003c/sup\u003e. Each concentration from the serial dilution was run in triplicates to determine qPCR efficiency, limit of detection, slope and y-intercept.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec10\" class=\"Section2\"\u003e \u003ch2\u003eStatistical analysis\u003c/h2\u003e \u003cp\u003eData cleaning and analysis were conducted using STATA 17 software (StataCorp LLC, College Station TX, USA). Descriptive statistics were employed for data summarization, presenting the proportion of infected mosquitoes with a 95% confidence interval. For parasite intensity (oocysts or sporozoites), the median along with minimum and maximum values were reported.\u003c/p\u003e \u003cp\u003eTo evaluate vector competence towards \u003cem\u003eP. falciparum\u003c/em\u003e infection among the three mosquito strains, mixed-effect regression was used, with mosquito strain as a fixed categorical effect and study participants included as a random effect. For oocyst and sporozoite prevalence logistic distribution was used. For oocyst and sporozoite intensity, negative binomial distribution was used and only infected mosquitoes were included in the intensity analysis.\u003c/p\u003e \u003c/div\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eAcknowledgments\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors express their sincere gratitude to the village leaders and community in Bagamoyo for their unwavering cooperation and support throughout the study. A special thanks to\u0026nbsp;Vector Control and Product testing Unit (VCPTU) management, administrators and colleagues who helped in organising logistics and materials, allowing smooth performance of the study.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAuthors \u0026lsquo;contribution\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eSJM, PAK, MMT, LH conceived the study. PAK, LH and MMT developed the study protocol. RYM and PAK performed the DMFAs. RMS, PAK and FM dissected the mosquitoes. PAK, LH and RYM performed molecular analysis. PAK drafted the manuscript. SJM, MMT, DWL, and LH reviewed and edited drafts of the manuscript.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eCode, data, and materials availability\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eData is provided within the supplementary information files.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eCompeting interests\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eAll authors declare no competing interest.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eFunding\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThis study`s field work was funded by VCTPU and PAK, RYM, RMS and FM receive salary support from the Transmission Zero project (Bill and Melinda Gates Foundation (BMGF), grant no. OPP1158151).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eEthics approval and consent to participate\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eAll adult participants provided written informed consent, while for children under 18 years old, consent was obtained from their parent or guardian, with the children\u0026apos;s assent also sought for their participation. All study volunteers received artemisinin-lumefantrine treatment within 24 hours of diagnosis, following the Tanzania Guidelines for Diagnosis and Treatment of Malaria\u003csup\u003e76\u003c/sup\u003e, administered by a qualified nurse. No adverse effects were reported among the participants during the study period. The study activities were reviewed and approved by the Institutional Review Board of IHI (IHI/IRB/No: 44 \u0026ndash; 2020) and the National Institute for Medical Research Tanzania (NIMR/HQ/R.8a/Vol.IX/3595). All research was performed in accordance to relevant guidelines and regulations.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\u003cli\u003e\u003cspan\u003eWHO. \u003cem\u003eThe World Malaria Report 2023\u003c/em\u003e (World Health Organization, 2023).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eKiszewski, A. et al. A global index representing the stability of malaria transmission. \u003cem\u003eAm. J. Trop. Med. Hyg.\u003c/em\u003e \u003cb\u003e70\u003c/b\u003e, 486\u0026ndash;498 (2004).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eSinka, M. E. et al. A global map of dominant malaria vectors. \u003cem\u003eParasit. Vectors\u003c/em\u003e. \u003cb\u003e5\u003c/b\u003e, 69. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003e10.1186/1756-3305-5-69\u003c/span\u003e\u003cspan address=\"10.1186/1756-3305-5-69\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e (2012).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eSinka, M. E. et al. The dominant \u003cem\u003eAnopheles\u003c/em\u003e vectors of human malaria in Africa, Europe and the Middle East: occurrence data, distribution maps and bionomic precis. \u003cem\u003eParasit. Vectors\u003c/em\u003e. \u003cb\u003e3\u003c/b\u003e, 117. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003e10.1186/1756-3305-3-117\u003c/span\u003e\u003cspan address=\"10.1186/1756-3305-3-117\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e (2010).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eLefevre, T., Vantaux, A., Dabire, K. R., Mouline, K. \u0026amp; Cohuet, A. Non-genetic determinants of mosquito competence for malaria parasites. \u003cem\u003ePLoS Pathog.\u003c/em\u003e \u003cb\u003e9\u003c/b\u003e, e1003365 (2013).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eWu, S. L. et al. Vector bionomics and vectorial capacity as emergent properties of mosquito behaviors and ecology. \u003cem\u003ePLoS computational biology\u003c/em\u003e 16, e1007446, doi: (2020). \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003e10.1371/journal.pcbi.1007446\u003c/span\u003e\u003cspan address=\"10.1371/journal.pcbi.1007446\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eCohuet, A., Harris, C., Robert, V. \u0026amp; Fontenille, D. Evolutionary forces on \u003cem\u003eAnopheles\u003c/em\u003e: what makes a malaria vector? \u003cem\u003eTrends Parasitol.\u003c/em\u003e \u003cb\u003e26\u003c/b\u003e, 130\u0026ndash;136 (2010).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eReeves, W. C., Asman, S., Hardy, J., Milby, M. \u0026amp; Reisen, W. Epidemiology and control of mosquito-borne arboviruses in California, 1943\u0026ndash;1987. (1990).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eGarrett-Jones, C. \u0026amp; Grab, B. The Assessment of Insecticidal Impact on the Malaria Mosquito\u0026rsquo;s Vectorial Capacity, from Data on the Proportion of Parous Females. \u003cem\u003eBull. WHO\u003c/em\u003e \u003cb\u003e31\u003c/b\u003e (1964).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eBrady, O. J. et al. Vectorial capacity and vector control: reconsidering sensitivity to parameters for malaria elimination. \u003cem\u003eTrans. R Soc. Trop. Med. Hyg.\u003c/em\u003e \u003cb\u003e110\u003c/b\u003e, 107\u0026ndash;117. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003e10.1093/trstmh/trv113\u003c/span\u003e\u003cspan address=\"10.1093/trstmh/trv113\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e (2016).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eGarrett-Jones, C. The human blood index of malaria vectors in relation to epidemiological assessment. \u003cem\u003eBull. World Health Organ.\u003c/em\u003e \u003cb\u003e30\u003c/b\u003e (1964).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eMatthews, J., Bethel, A. \u0026amp; Osei, G. An overview of malarial \u003cem\u003eAnopheles\u003c/em\u003e mosquito survival estimates in relation to methodology. \u003cem\u003eParasit. Vectors\u003c/em\u003e. \u003cb\u003e13\u003c/b\u003e, 233. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003e10.1186/s13071-020-04092-4\u003c/span\u003e\u003cspan address=\"10.1186/s13071-020-04092-4\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e (2020).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eOhm, J. R. et al. Rethinking the extrinsic incubation period of malaria parasites. \u003cem\u003eParasites Vectors\u003c/em\u003e. \u003cb\u003e11\u003c/b\u003e, 178. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003e10.1186/s13071-018-2761-4\u003c/span\u003e\u003cspan address=\"10.1186/s13071-018-2761-4\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e (2018).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eSmith, M. W. et al. Incorporating hydrology into climate suitability models changes projections of malaria transmission in Africa. \u003cem\u003eNat. Commun.\u003c/em\u003e \u003cb\u003e11\u003c/b\u003e, 4353. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003e10.1038/s41467-020-18239-5\u003c/span\u003e\u003cspan address=\"10.1038/s41467-020-18239-5\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e (2020).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eSmith, D. L. et al. Ross, Macdonald, and a Theory for the Dynamics and Control of Mosquito-Transmitted Pathogens. \u003cem\u003ePLoS Pathog\u003c/em\u003e 8, e1002588. doi:1002510.1001371/journal.ppat.1002588 (2012).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eOkech, B. A., Gouagna, L. C., Yan, G., Githure, J. I. \u0026amp; Beier, J. C. Larval habitats of \u003cem\u003eAnopheles gambiae s.s.\u003c/em\u003e (Diptera: Culicidae) influences vector competence to Plasmodium falciparum parasites. \u003cem\u003eMalar. J.\u003c/em\u003e \u003cb\u003e6\u003c/b\u003e, 50. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003e10.1186/1475-2875-6-50\u003c/span\u003e\u003cspan address=\"10.1186/1475-2875-6-50\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e (2007).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eAsmare, Y., Hopkins, R. J., Tekie, H., Hill, S. R. \u0026amp; Ignell, R. Grass Pollen Affects Survival and Development of Larval \u003cem\u003eAnopheles arabiensis\u003c/em\u003e (Diptera: Culicidae). \u003cem\u003eJ. insect Sci. (Online)\u003c/em\u003e. \u003cb\u003e17\u003c/b\u003e \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003e10.1093/jisesa/iex067\u003c/span\u003e\u003cspan address=\"10.1093/jisesa/iex067\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e (2017).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eShapiro, L. L., Murdock, C. C., Jacobs, G. R., Thomas, R. J. \u0026amp; Thomas, M. B. Larval food quantity affects the capacity of adult mosquitoes to transmit human malaria. \u003cem\u003eProc. Biol. Sci.\u003c/em\u003e \u003cb\u003e283\u003c/b\u003e \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003e10.1098/rspb.2016.0298\u003c/span\u003e\u003cspan address=\"10.1098/rspb.2016.0298\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e (2016).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eSchapira, A. \u0026amp; Boutsika, K. Malaria Ecotypes and Stratification. \u003cem\u003eAdv. Parasitol.\u003c/em\u003e \u003cb\u003e78\u003c/b\u003e, 97\u0026ndash;167. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003e10.1016/B978-0-12-394303-3.00001-3\u003c/span\u003e\u003cspan address=\"10.1016/B978-0-12-394303-3.00001-3\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e (2012).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eTakken, W. \u0026amp; Lindsay, S. W. Factors affecting the vectorial competence of \u003cem\u003eAnopheles gambiae\u003c/em\u003e: a question of scale. \u003cem\u003eEcol. Aspects Application Genetically Modified Mosquitoes Dordrecht: Kluwer Acad. Publishers\u003c/em\u003e, 75\u0026ndash;90 (2003).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eVillena, O. C., Ryan, S. J., Murdock, C. C. \u0026amp; Johnson, L. R. Temperature impacts the environmental suitability for malaria transmission by \u003cem\u003eAnopheles gambiae\u003c/em\u003e and \u003cem\u003eAnopheles stephensi\u003c/em\u003e. \u003cem\u003eEcology\u003c/em\u003e. \u003cb\u003e103\u003c/b\u003e, e3685 (2022).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eMoller-Jacobs, L. L., Murdock, C. C. \u0026amp; Thomas, M. B. Capacity of mosquitoes to transmit malaria depends on larval environment. \u003cem\u003eParasites vectors\u003c/em\u003e. \u003cb\u003e7\u003c/b\u003e, 1\u0026ndash;12 (2014).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eCharlwood, J. D. et al. The rise and fall of \u003cem\u003eAnopheles arabiensis\u003c/em\u003e (Diptera: Culicidae) in a Tanzanian village. \u003cem\u003eBull. Entomol. Res.\u003c/em\u003e \u003cb\u003e85\u003c/b\u003e, 37\u0026ndash;44 (1995).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eMendis, C. et al. \u003cem\u003eAnopheles arabiensis\u003c/em\u003e and \u003cem\u003eAn. funestus\u003c/em\u003e are equally important vectors of malaria in Matola coastal suburb of Maputo, southern Mozambique. \u003cem\u003eMed. Vet. Entomol.\u003c/em\u003e \u003cb\u003e14\u003c/b\u003e, 171\u0026ndash;180 (2000).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eNambunga, I. H. et al. Aquatic habitats of the malaria vector \u003cem\u003eAnopheles funestus\u003c/em\u003e in rural south-eastern Tanzania. \u003cem\u003eMalar. J.\u003c/em\u003e \u003cb\u003e19\u003c/b\u003e, 1\u0026ndash;11 (2020).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eLindsay, S. et al. Exposure of Gambian children to \u003cem\u003eAnopheles gambiae\u003c/em\u003e malaria vectors in an irrigated rice production area. \u003cem\u003eMed. Vet. Entomol.\u003c/em\u003e \u003cb\u003e9\u003c/b\u003e, 50\u0026ndash;58 (1995).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eGillies, M. T. \u0026amp; Coetzee, M. A supplement to the Anophelinae of Africa South of the Sahara. \u003cem\u003ePubl S Afr. Inst. Med. Res.\u003c/em\u003e \u003cb\u003e55\u003c/b\u003e, 1\u0026ndash;143 (1987).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eMala, A. O. et al. Dry season ecology of \u003cem\u003eAnopheles gambiae\u003c/em\u003e complex mosquitoes at larval habitats in two traditionally semi-arid villages in Baringo, Kenya. \u003cem\u003eParasites vectors\u003c/em\u003e. \u003cb\u003e4\u003c/b\u003e, 1\u0026ndash;11 (2011).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eMwanziva, C. E. et al. Transmission intensity and malaria vector population structure in Magugu, Babati District in northern Tanzania. \u003cem\u003eTanzan. J. health Res.\u003c/em\u003e \u003cb\u003e13\u003c/b\u003e, 54\u0026ndash;61 (2011).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eSmith, D. L. \u0026amp; McKenzie, F. E. Statics and dynamics of malaria infection in \u003cem\u003eAnopheles\u003c/em\u003e mosquitoes. \u003cem\u003eMalar. J.\u003c/em\u003e \u003cb\u003e3\u003c/b\u003e, 13. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003e10.1186/1475-2875-3-13\u003c/span\u003e\u003cspan address=\"10.1186/1475-2875-3-13\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e (2004).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eKent, R. J., Thuma, P. E., Mharakurwa, S. \u0026amp; Norris, D. E. Seasonality, blood feeding behavior, and transmission of \u003cem\u003ePlasmodium falciparum\u003c/em\u003e by \u003cem\u003eAnopheles arabiensis\u003c/em\u003e after an extended drought in southern Zambia. \u003cem\u003eAm. J. Trop. Med. Hyg.\u003c/em\u003e \u003cb\u003e76\u003c/b\u003e, 267 (2007).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eLyimo, I. N., Keegan, S. P., Ranford-Cartwright, L. C. \u0026amp; Ferguson, H. M. The impact of uniform and mixed species blood meals on the fitness of the mosquito vector \u003cem\u003eAnopheles gambiae s.s\u003c/em\u003e: does a specialist pay for diversifying its host species diet? \u003cem\u003eJ. Evol. Biol.\u003c/em\u003e \u003cb\u003e25\u003c/b\u003e, 452\u0026ndash;460. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003e10.1111/j.1420-9101.2011.02442.x\u003c/span\u003e\u003cspan address=\"10.1111/j.1420-9101.2011.02442.x\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e (2012).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eKahamba, N. F. et al. Using ecological observations to improve malaria control in areas where \u003cem\u003eAnopheles funestus\u003c/em\u003e is the dominant vector. \u003cem\u003eMalar. J.\u003c/em\u003e \u003cb\u003e21\u003c/b\u003e, 158. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003e10.1186/s12936-022-04198-3\u003c/span\u003e\u003cspan address=\"10.1186/s12936-022-04198-3\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e (2022).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eShaw, W. R. et al. Multiple blood feeding in mosquitoes shortens the \u003cem\u003ePlasmodium falciparum\u003c/em\u003e incubation period and increases malaria transmission potential. \u003cem\u003ePLoS Pathog\u003c/em\u003e. \u003cb\u003e16\u003c/b\u003e, e1009131 (2020).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eHofer, L. M. et al. Additional blood meals increase sporozoite infection in \u003cem\u003eAnopheles\u003c/em\u003e mosquitoes but not Plasmodium falciparum genetic diversity. \u003cem\u003eSci. Rep.\u003c/em\u003e \u003cb\u003e14\u003c/b\u003e, 17467 (2024).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eBeerntsen, B. T., James, A. A. \u0026amp; Christensen, B. M. Genetics of mosquito vector competence. \u003cem\u003eMicrobiol. Mol. Biol. Rev.\u003c/em\u003e \u003cb\u003e64\u003c/b\u003e, 115\u0026ndash;137 (2000).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eHajkazemian, M., Boss\u0026eacute;, C., Mozūraitis, R. \u0026amp; Emami, S. N. Battleground midgut: The cost to the mosquito for hosting the malaria parasite. \u003cem\u003eBiol. Cell.\u003c/em\u003e \u003cb\u003e113\u003c/b\u003e, 79\u0026ndash;94. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003e10.1111/boc.202000039\u003c/span\u003e\u003cspan address=\"10.1111/boc.202000039\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e (2021).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eLi, M. et al. Response of the mosquito immune system and symbiotic bacteria to pathogen infection. \u003cem\u003eParasit. Vectors\u003c/em\u003e. \u003cb\u003e17\u003c/b\u003e, 69. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003e10.1186/s13071-024-06161-4\u003c/span\u003e\u003cspan address=\"10.1186/s13071-024-06161-4\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e (2024).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eEmami, S. N. et al. A key malaria metabolite modulates vector blood seeking, feeding, and susceptibility to infection. \u003cem\u003eScience\u003c/em\u003e. \u003cb\u003e355\u003c/b\u003e, 1076\u0026ndash;1080. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003e10.1126/science.aah4563\u003c/span\u003e\u003cspan address=\"10.1126/science.aah4563\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e (2017).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eNsango, S. E. et al. Genetic clonality of \u003cem\u003ePlasmodium falciparum\u003c/em\u003e affects the outcome of infection in \u003cem\u003eAnopheles gambiae\u003c/em\u003e. \u003cem\u003eInt. J. Parasitol.\u003c/em\u003e \u003cb\u003e42\u003c/b\u003e, 589\u0026ndash;595. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.1016/j.ijpara.2012.03.008\u003c/span\u003e\u003cspan address=\"10.1016/j.ijpara.2012.03.008\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e (2012).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eTakken, W. et al. Susceptibility of \u003cem\u003eAnopheles quadriannulatus Theobald\u003c/em\u003e (Diptera: Culicidae) to \u003cem\u003ePlasmodium falciparum\u003c/em\u003e. \u003cem\u003eTrans. R Soc. Trop. Med. Hyg.\u003c/em\u003e \u003cb\u003e93\u003c/b\u003e, 578\u0026ndash;580. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003e10.1016/s0035-9203(99)90054-8\u003c/span\u003e\u003cspan address=\"10.1016/s0035-9203(99)90054-8\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e (1999).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eHabtewold, T., Povelones, M., Blagborough, A. M. \u0026amp; Christophides, G. K. Transmission blocking immunity in the malaria non-vector mosquito \u003cem\u003eAnopheles quadriannulatus\u003c/em\u003e species A. \u003cem\u003ePLoS Pathog\u003c/em\u003e. \u003cb\u003e4\u003c/b\u003e, e1000070. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003e10.1371/journal.ppat.1000070\u003c/span\u003e\u003cspan address=\"10.1371/journal.ppat.1000070\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e (2008).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eHabtewold, T., Groom, Z. \u0026amp; Christophides, G. K. Immune resistance and tolerance strategies in malaria vector and non-vector mosquitoes. \u003cem\u003eParasit. Vectors\u003c/em\u003e. \u003cb\u003e10\u003c/b\u003e, 186. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003e10.1186/s13071-017-2109-5\u003c/span\u003e\u003cspan address=\"10.1186/s13071-017-2109-5\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e (2017).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eGn\u0026eacute;m\u0026eacute;, A. et al. Equivalent susceptibility of \u003cem\u003eAnopheles gambiae\u003c/em\u003e M and S molecular forms and Anopheles arabiensis to \u003cem\u003ePlasmodium falciparum\u003c/em\u003e infection in Burkina Faso. \u003cem\u003eMalar. J.\u003c/em\u003e \u003cb\u003e12\u003c/b\u003e, 204. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003e10.1186/1475-2875-12-204\u003c/span\u003e\u003cspan address=\"10.1186/1475-2875-12-204\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e (2013).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eAschale, Y. et al. Systematic review of sporozoite infection rate of \u003cem\u003eAnopheles\u003c/em\u003e mosquitoes in Ethiopia, 2001\u0026ndash;2021. \u003cem\u003eParasit. Vectors\u003c/em\u003e. \u003cb\u003e16\u003c/b\u003e, 437. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003e10.1186/s13071-023-06054-y\u003c/span\u003e\u003cspan address=\"10.1186/s13071-023-06054-y\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e (2023).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eKabula, B. et al. Malaria entomological profile in Tanzania from 1950 to 2010: a review of mosquito distribution, vectorial capacity and insecticide resistance. \u003cem\u003eTanzan. J. Health Res.\u003c/em\u003e \u003cb\u003e13\u003c/b\u003e, 319\u0026ndash;331 (2011).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eNtabaliba, W. et al. Life expectancy of \u003cem\u003eAnopheles funestus\u003c/em\u003e is double that of \u003cem\u003eAnopheles arabiensis\u003c/em\u003e in southeast Tanzania based on mark-release-recapture method. \u003cem\u003eSci. Rep.\u003c/em\u003e \u003cb\u003e13\u003c/b\u003e, 15775. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003e10.1038/s41598-023-42761-3\u003c/span\u003e\u003cspan address=\"10.1038/s41598-023-42761-3\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e (2023).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eBrady, O. J. et al. Vectorial capacity and vector control: reconsidering sensitivity to parameters for malaria elimination. \u003cem\u003eTrans. R. Soc. Trop. Med. Hyg.\u003c/em\u003e \u003cb\u003e110\u003c/b\u003e, 107\u0026ndash;117 (2016).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eShaw, W. R. \u0026amp; Catteruccia, F. Vector biology meets disease control: using basic research to fight vector-borne diseases. \u003cem\u003eNat. Microbiol.\u003c/em\u003e \u003cb\u003e4\u003c/b\u003e, 20\u0026ndash;34 (2019).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eReiner, R. C. Jr. et al. A systematic review of mathematical models of mosquito-borne pathogen transmission: 1970\u0026ndash;2010. \u003cem\u003eJ. R Soc. Interface\u003c/em\u003e. \u003cb\u003e10\u003c/b\u003e, 20120921. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003e10.1098/rsif.2012.0921\u003c/span\u003e\u003cspan address=\"10.1098/rsif.2012.0921\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e (2013).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eBarreaux, A. M., Barreaux, P., Thievent, K. \u0026amp; Koella, J. C. Larval environment influences vector competence of the malaria mosquito \u003cem\u003eAnopheles gambiae\u003c/em\u003e. \u003cem\u003eMalar. World J.\u003c/em\u003e \u003cb\u003e7\u003c/b\u003e, 1\u0026ndash;6 (2016).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eWhite, B. J., Collins, F. H. \u0026amp; Besansky, N. J. Evolution of \u003cem\u003eAnopheles gambiae\u003c/em\u003e in relation to humans and malaria. \u003cem\u003eAnnu. Rev. Ecol. Evol. Syst.\u003c/em\u003e \u003cb\u003e42\u003c/b\u003e, 111\u0026ndash;132 (2011).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eLwetoijera, D. W. et al. Increasing role of \u003cem\u003eAnopheles funestus\u003c/em\u003e and \u003cem\u003eAnopheles arabiensis\u003c/em\u003e in malaria transmission in the Kilombero Valley, Tanzania. \u003cem\u003eMalar. J.\u003c/em\u003e \u003cb\u003e13\u003c/b\u003e, 1\u0026ndash;10 (2014).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eMatowo, N. S. et al. An increasing role of pyrethroid-resistant \u003cem\u003eAnopheles funestus\u003c/em\u003e in malaria transmission in the Lake Zone, Tanzania. \u003cem\u003eSci. Rep.\u003c/em\u003e \u003cb\u003e11\u003c/b\u003e, 13457 (2021).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eOgola, E. O., Odero, J. O., Mwangangi, J. M. \u0026amp; Masiga, D. K. Tchouassi, D. P. Population genetics of \u003cem\u003eAnopheles funestus\u003c/em\u003e, the African malaria vector, Kenya. \u003cem\u003eParasites vectors\u003c/em\u003e. \u003cb\u003e12\u003c/b\u003e, 1\u0026ndash;9 (2019).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eMsugupakulya, B. J. et al. Changes in contributions of different \u003cem\u003eAnopheles\u003c/em\u003e vector species to malaria transmission in east and southern Africa from 2000 to 2022. \u003cem\u003eParasites Vectors\u003c/em\u003e. \u003cb\u003e16\u003c/b\u003e, 408 (2023).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eWangrawa, D. W., Odero, J. O., Baldini, F., Okumu, F. \u0026amp; Badolo, A. Distribution and insecticide resistance profile of the major malaria vector \u003cem\u003eAnopheles funestus\u003c/em\u003e group across the African continent. \u003cem\u003eMed. Vet. Entomol.\u003c/em\u003e \u003cb\u003e38\u003c/b\u003e, 119\u0026ndash;137 (2024).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eSmall, S. T. et al. Radiation with reticulation marks the origin of a major malaria vector. \u003cem\u003eProc. Natl. Acad. Sci. U S A\u003c/em\u003e. \u003cb\u003e117\u003c/b\u003e, 31583\u0026ndash;31590. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003e10.1073/pnas.2018142117\u003c/span\u003e\u003cspan address=\"10.1073/pnas.2018142117\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e (2020).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eSuh, P. F. et al. Impact of insecticide resistance on malaria vector competence: a literature review. \u003cem\u003eMalar. J.\u003c/em\u003e \u003cb\u003e22\u003c/b\u003e, 19 (2023).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eAlout, H. et al. Insecticide resistance alleles affect vector competence of \u003cem\u003eAnopheles gambiae\u003c/em\u003e ss for \u003cem\u003ePlasmodium falciparum\u003c/em\u003e field isolates. \u003cem\u003ePLoS One\u003c/em\u003e. \u003cb\u003e8\u003c/b\u003e, e63849 (2013).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eBarreaux, P., Koella, J. C., N\u0026rsquo;Guessan, R. \u0026amp; Thomas, M. B. Use of novel lab assays to examine the effect of pyrethroid-treated bed nets on blood-feeding success and longevity of highly insecticide-resistant \u003cem\u003eAnopheles gambiae sl\u003c/em\u003e mosquitoes. \u003cem\u003eParasites Vectors\u003c/em\u003e. \u003cb\u003e15\u003c/b\u003e, 111 (2022).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eMinetti, C., Ingham, V. A. \u0026amp; Ranson, H. Effects of insecticide resistance and exposure on \u003cem\u003ePlasmodium\u003c/em\u003e development in Anopheles mosquitoes. \u003cem\u003eCurr. Opin. insect Sci.\u003c/em\u003e \u003cb\u003e39\u003c/b\u003e, 42\u0026ndash;49 (2020).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eJames, R. \u0026amp; Xu, J. Mechanisms by which pesticides affect insect immunity. \u003cem\u003eJ. Invertebr. Pathol.\u003c/em\u003e \u003cb\u003e109\u003c/b\u003e, 175\u0026ndash;182 (2012).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eRivero, A., Vezilier, J., Weill, M., Read, A. F. \u0026amp; Gandon, S. Insecticide control of vector-borne diseases: when is insecticide resistance a problem? \u003cem\u003ePLoS Pathog.\u003c/em\u003e \u003cb\u003e6\u003c/b\u003e, e1001000 (2010).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eMendes, A. M. et al. Infection intensity-dependent responses of \u003cem\u003eAnopheles gambiae\u003c/em\u003e to the African malaria parasite Plasmodium falciparum. \u003cem\u003eInfect. Immun.\u003c/em\u003e \u003cb\u003e79\u003c/b\u003e, 4708\u0026ndash;4715 (2011).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eCirimotich, C. M., Ramirez, J. L. \u0026amp; Dimopoulos, G. Native microbiota shape insect vector competence for human pathogens. \u003cem\u003eCell. host microbe\u003c/em\u003e. \u003cb\u003e10\u003c/b\u003e, 307\u0026ndash;310 (2011).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eHarris, C. et al. Plasmodium falciparum produce lower infection intensities in local versus foreign Anopheles gambiae populations. \u003cem\u003ePloS one\u003c/em\u003e. \u003cb\u003e7\u003c/b\u003e, e30849 (2012).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eHenry, B. Assessment of the transmission blocking activity of antimalarial compounds by membrane feeding assays using natural \u003cem\u003ePlasmodium falciparum\u003c/em\u003e gametocyte isolates from West-Africa. \u003cem\u003ePlos one\u003c/em\u003e. \u003cb\u003e18\u003c/b\u003e, e0284751 (2023).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eKweyamba, P. A. et al. Sub-lethal exposure to chlorfenapyr reduces the probability of developing \u003cem\u003ePlasmodium falciparum\u003c/em\u003e parasites in surviving \u003cem\u003eAnopheles\u003c/em\u003e mosquitoes. \u003cem\u003eParasites Vectors\u003c/em\u003e. \u003cb\u003e16\u003c/b\u003e, 342 (2023).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eHabtewold, T. et al. Streamlined SMFA and mosquito dark-feeding regime significantly improve malaria transmission-blocking assay robustness and sensitivity. \u003cem\u003eMalar. J.\u003c/em\u003e \u003cb\u003e18\u003c/b\u003e, 1\u0026ndash;11 (2019).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eBenedict, M. (2018).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eChomczynski, P., Mackey, K., Drews, R. \u0026amp; Wilfinger, W. DNAzol\u0026reg;: a reagent for the rapid isolation of genomic DNA. \u003cem\u003eBiotechniques\u003c/em\u003e. \u003cb\u003e22\u003c/b\u003e, 550\u0026ndash;553 (1997).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eSchindler, T. et al. Molecular monitoring of the diversity of human pathogenic malaria species in blood donations on Bioko Island, Equatorial Guinea. \u003cem\u003eMalar. J.\u003c/em\u003e \u003cb\u003e18\u003c/b\u003e, 1\u0026ndash;11 (2019).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eWhelan, J. A., Russell, N. B. \u0026amp; Whelan, M. A. A method for the absolute quantification of cDNA using real-time PCR. \u003cem\u003eJ. Immunol. Methods\u003c/em\u003e. \u003cb\u003e278\u003c/b\u003e, 261\u0026ndash;269 (2003).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eMercereau-Puijalon, O., Barale, J. C. \u0026amp; Bischoff, E. Three multigene families in Plasmodium parasites: facts and questions. \u003cem\u003eInt. J. Parasitol.\u003c/em\u003e \u003cb\u003e32\u003c/b\u003e, 1323\u0026ndash;1344 (2002).\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eHealth, M. \u0026amp; Welfare, S. (United Republic of Tanzania Dar es Salam, (2014).\u003c/span\u003e\u003c/li\u003e\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":false,"highlight":"","institution":"","isAcceptedByJournal":true,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"scientific-reports","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"scirep","sideBox":"Learn more about [Scientific Reports](http://www.nature.com/srep/)","snPcode":"","submissionUrl":"","title":"Scientific Reports","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"stoa","reportingPortfolio":"Scientific Reports","inReviewEnabled":true,"inReviewRevisionsEnabled":true},"keywords":"","lastPublishedDoi":"10.21203/rs.3.rs-5038559/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-5038559/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eThere are three \u003cem\u003eAnopheles\u003c/em\u003e mosquito species in East Africa that are responsible for the majority of malaria transmission, posing a significant public health concern. Understanding the vector competence of different mosquito species is crucial for targeted and cost-effective malaria control strategies. This study investigated the vector competence of laboratory reared strains of East African \u003cem\u003eAn. gambiae sensu stricto, An. funestus s.s.\u003c/em\u003e, and \u003cem\u003eAn. arabiensis\u003c/em\u003e mosquitoes towards local isolates of \u003cem\u003ePlasmodium falciparum\u003c/em\u003e infection. Mosquito feeding assays using gametocytaemic blood from local donors revealed significant differences in both prevalence and intensity of oocyst and sporozoite infections among the three vectors. \u003cem\u003eAn. funestus\u003c/em\u003e mosquitoes presented the highest sporozoite prevalence 23.5% (95% confidence interval (CI): 17.5\u0026ndash;29.6) and intensity of infection 6-58138 sporozoites. Relative to \u003cem\u003eAn. funestus\u003c/em\u003e, the odds ratio for sporozoites prevalence were 0.46 (95% CI: 0.25\u0026ndash;0.85) in \u003cem\u003eAn. gambiae\u003c/em\u003e and 0.19 (95% CI: 0.07\u0026ndash;0.51) in \u003cem\u003eAn. arabiensis\u003c/em\u003e, while the incidence rate ratio for sporozoite intensity was 0.31 (95% CI: 0.14\u0026ndash;0.69) in \u003cem\u003eAn. gambiae\u003c/em\u003e and 0.66 (95% CI: 0.16\u0026ndash;2.60) in \u003cem\u003eAn. arabiensis.\u003c/em\u003e Our findings indicate that all three malaria species contribute to malaria transmission in East Africa with \u003cem\u003eAn. funestus\u003c/em\u003e demonstrating superior vector competence. In conclusion, there is a need for comprehensive malaria control strategies targeting major malaria vector species, an update of malaria transmission models to consider vectoral competence and evaluation of malaria transmission blocking interventions in assays that include \u003cem\u003eAn. funestus\u003c/em\u003e mosquitoes.\u003c/p\u003e","manuscriptTitle":"Contrasting vector competence of three main East African Anopheles malaria vector mosquitoes for Plasmodium falciparum","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2024-10-28 16:54:36","doi":"10.21203/rs.3.rs-5038559/v1","editorialEvents":[{"type":"communityComments","content":0},{"type":"decision","content":"Revision requested","date":"2024-11-12T03:16:00+00:00","index":"","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2024-11-11T10:41:41+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"207527577628726492618723903564383188669","date":"2024-10-22T07:59:04+00:00","index":"hide","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2024-09-23T08:12:44+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"248303870113067086102197252791958114058","date":"2024-09-16T11:45:45+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"236111602084710045072745144815052293920","date":"2024-09-15T17:36:25+00:00","index":"hide","fulltext":""},{"type":"reviewersInvited","content":"","date":"2024-09-13T12:58:34+00:00","index":"","fulltext":""},{"type":"editorAssigned","content":"","date":"2024-09-13T12:55:49+00:00","index":"","fulltext":""},{"type":"editorInvited","content":"","date":"2024-09-13T12:36:19+00:00","index":"","fulltext":""},{"type":"checksComplete","content":"","date":"2024-09-12T13:00:54+00:00","index":"","fulltext":""},{"type":"submitted","content":"Scientific Reports","date":"2024-09-05T13:33:10+00:00","index":"","fulltext":""}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"scientific-reports","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"scirep","sideBox":"Learn more about [Scientific Reports](http://www.nature.com/srep/)","snPcode":"","submissionUrl":"","title":"Scientific Reports","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"stoa","reportingPortfolio":"Scientific Reports","inReviewEnabled":true,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"3b27baa6-1d85-46e8-838d-ce9228b02ca9","owner":[],"postedDate":"October 28th, 2024","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"under-review","subjectAreas":[{"id":39256154,"name":"Biological sciences/Molecular biology"},{"id":39256155,"name":"Earth and environmental sciences/Ecology"}],"tags":[],"updatedAt":"2025-01-10T06:08:10+00:00","versionOfRecord":[],"versionCreatedAt":"2024-10-28 16:54:36","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-5038559","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-5038559","identity":"rs-5038559","version":["v1"]},"buildId":"8U1c8b4HqxoKbykW_rLl7","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}