Anopheles gambiae sensu lato mosquitoes are among the dominant malaria vectors in sub-Saharan Africa. However, not much is known about the oviposition behaviour of these species necessary for the development of malaria vector control strategies. With the aim of investigating cues associated with selected oviposition sites, artificial oviposition sites- ponds (soil mixed with water) were set-up in an open field at Mbita, Western Kenya in 2012 and 2013. Ponds were allowed to be colonized by wild An. gambiae s.l.. The numbers of Anopheles early instar larvae were counted and used as a proxy for oviposition preference. Water samples were then analysed for physicochemical, bacterial and chemical profiles. The bacterial profiles were analysed using denaturing gradient gel electrophoresis (DGGE) and the chemical profiles with gas chromatography-mass spectrometry (GC-MS). The detection of possible oviposition cues from oviposition substrates requires sensitive analytical methods. Volatiles detection was improved seven times. The detection of bacteria deoxyribonucleic acid (DNA) bands with DGGE was also improved to a minimum DNA concentration of 50 ng/µl. Results showed that ponds were colonized differently. Fresh ponds were preferred over slightly older ponds. Bacterial analysis revealed a low number of bacteria colony forming units (CFU) in preferred ponds. Some volatiles, including: 6,10-dimethyl-5,9-undecadien-2-one (geranylacetone) and 4-ethylbenzaldehyde, were associated with the oviposition preferred pond. In addition, low pH and high turbidity were associated with the ponds selected for oviposition. Finally, fungi isolated from the rhizomes of nut grass yielded a promising array of volatiles of which one is known to attract oviposition site seeking malaria mosquitoes. This finding opens the door for a cost effective and environmental friendly method of using fungi in an “attract and kill” strategy targeting malaria vectors. / Myggor i Anopheles gambiae sensu lato komplexet tillhör de myggor som är bäst på att sprida malaria parasiter i afrika söder om Sahara. Kunskapen om de här myggornas äggläggningsbeteende är begränsad. Den här kunskapen behövs för att kunna utveckla nya och förbättra tillgängliga malaria vektor kontroll metoder. Nya metoder som kan komplettera de som används idag (insecticides treated nets (ITNs) och indoor residual spraying (IRS)) behövs eftersom de metoderna har problem med resistensutveckling. Två studier utfördes på icipe fältstation i Mbita västra Kenya under 2012 och 2013 med målet att identifiera faktorer som påverkar myggornas äggläggningsbeteende. Baljor fyllda med en blandning av jord och vatten (äggläggningssubstrat) användes för att tillverka artificiella äggläggningsplatser som liknar de vattenpölar som de här myggarterna gärna lägger ägg i. Baljorna koloniserades av vilda myggor och antalet mygglarver som detekterades i baljorna jämfördes och användes som en proxy för äggläggningspreferens. Fysikaliska och kemiska parametrar mättes på jordvattenblandningarna i baljorna och prover togs för att analysera bakteriepopulationer med hjälp av denaturing gradient gel electrophoresis (DGGE) och flyktiga ämnen med hjälp av gas-kromatografi kopplat till mass-spektrometri (GC-MS). För att kunna detektera de låga halter av flyktiga ämnen och bakterier som fanns i de här proverna krävdes det känsliga metoder. Antalet flyktiga ämnen som kunde detekteras ökades sju gånger genom att tillsätta NaCl till vattenproverna innan doften insamlades och termisk desorption användes istället för lösningsmedels desorption. För att förbättra detektionsgränsen för bakterier amplifierades bakterie-DNA i två PCR reaktioner som sedan mixades och koncentrerades. Resultaten från fältstudierna med baljorna visade att de koloniserades olika av Anopheles myggorna. Baljor med nyblandat substrat innehöll dubbelt så många mygglarver som baljor med jord-vattensubstrat som åldrats under en längre tid. Lägre mängd bakterier, lägre pH och högre grumlighet var gemensamt för de baljor som myggorna föredrog. De flyktiga ämnen som detekterades i de olika baljor varierade mellan olika försök och inget ämne fanns med i alla upprepningar av ett försök. Trots det detekterades några ämnen oftare i de baljor som myggorna föredrog att lägga ägg jämfört med de med en mindre mängd mygglarver. De inkluderar geranylacetone och 4-ethylbenzaldehyde. Svampar isolerades från rotstockar av gräs som fanns i den jord som användes för att göra äggläggningssubstraten i fältstudierna. De flyktiga ämnen som avgavs från svampkulturerna analyserades. Bland annat så identifierades ett ämne som fungerar som en äggläggningsattrahent för An. gambiae s.l. myggor.Resultaten från den här avhandlingen kommer att kunna användas för att utveckla miljövänliga ”attract and kill” metoder för att kontrollera malaria myggor. / <p>QC 20160211</p>

Die Allele (*R1, *R2, *S1 und *S2) des A. gambiae complement-like thioester-containing Protein 1 (TEP1) bestimmen die Fitness der Mücken, welches die männlichen Fertilität und den Resistenzgrad der Mücke gegen Pathogene wie Bakterien und Malaria-Parasiten. Dieser Kompromiss zwischen Reproduktion und Immunnität hat Auswirkungen auf die Größe der Mückenpopulationen und die Rate der Malariaübertragung. Wie die genetische Diversität von TEP1 die genetische Struktur natürlicher Vektorpopulationen beeinflusst, ist noch unklar. Die Zielsetzung dieser Doktorarbeit waren: i) die biogeographische Kartographierung der TEP1 Allele und Genotypen in lokalen Malariavektorpopulationen in Mali, Burkina Faso, Kamerun, und Kenia, und ii) die Bemessung des Einflusses von TEP1 Polymorphismen auf die Entwicklung humaner P. falciparum Parasiten in der Mücke. Die Analysen der TEP1 Polymorphismen zeigten, dass die natürliche Selektion auf Exone, sowie Introne wirkt, was auf eine starke funktionale Beschränkung an diesem Lokus hindeutet. Außerdem zeigen unsere Daten die strukturierte Erhaltung natürlicher genetischer Variation im TEP1 Lokus, in welchem die Allele und Genotypen spezifische evolutionäre Wege verfolgen. Diese Ergebnisse weisen auf die Existenz von arten- und habitatspezifischen Selektionsdrücken hin, die auf den TEP1 Lokus wirken. Resultate haben gezeigt, dass TEP1*S1 und *S2 Mücken gleichermassen empfänglich für Plasmodium-Infektionen sind. Insgesamt tragen die Resultate der biogeographischen Kartographierung des TEP1 Lokus und der Züchtungs- und Infektionsexperimente zu einem besseren Verständnis über den Einfluss der verschiedenen Vektorarten und lokale Umwelteinflüsse auf die Vektorpopulationen und Malariaübertragung bei. Des weiteren kann die hier beschriebene hochdurchsatz-genotypisierungs Methode, zur Studie lokaler A. gambiae Mückenpopulationen, in der Feldforschungsarbeit eingesetzt werden. Dieser neue Ansatz wird die epidemiologisch relevante Überwachung und Vorhersage dynamischer Prozesse in lokalen Malariavektorpopulationen unterstützen, welche die Entwicklung neuer Strategien der Vektorkontrolle ermöglichen könnten. / The alleles (*R1, *R2, *S1 and *S2) and genotypes of A. gambiae complement-like thioester-containing protein 1 (TEP1) determine the fitness in male fertility and the degree of mosquito resistance to pathogens such as bacteria and malaria parasites. This trade-off between the reproduction and the immunity impacts directly on mosquito population abundance and malaria transmission respectively. How TEP1 genetic diversity influences the genetic structure of natural vector populations and development of human malaria parasites is unclear. The aims of this thesis were to: i) map distribution of TEP1 alleles and genotypes in local malaria vector populations in Mali, Burkina Faso, Cameroon and Kenya, and ii) assess the impact of TEP1 polymorphism on development of human P. falciparum parasites in mosquitoes. Analyses of TEP1 polymorphism revealed that natural selection acts in concert on both exons and introns, suggesting strong functional constrains acting at this locus. Moreover, our data demonstrate a structured maintenance of natural TEP1 genetic variation, where the alleles and the genotypes follow distinct evolutionary paths. These findings suggest the existence of species- and habitat-specific selection patterns that act on TEP1 locus. Results revealed that the TEP1*S1 and *S2 mosquitoes are equally susceptible to Plasmodium infections. Collectively, results of my thesis on the biogeographic TEP1 mapping, and on the breeding and infection experiments contribute to a better understanding of how the vector species and local environmental factors, shape vector population structures and malaria transmission. Furthermore, the high throughput TEP1 genotyping approach reported here could be used for field studies of local A. gambiae mosquito populations. This new approach will benefit surveilance and prediction of dynamics in local malaria vector populations that may have epidemiological significance, and therefore inform the development of novel vector control measures.

Made available in DSpace on 2011-05-04T12:42:03Z (GMT). No. of bitstreams: 0 Previous issue date: 2009 / Até hoje não existem mecanismos eficientes de controle da malaria, uma das doenças infecciosas mais importantes do mundo. Nas últimas décadas, a transformação genética dos mosquitos transmissores tem sido proposta como uma alternativa para o controle de esta doença. Para isso, além do desenvolvimento de técnicas para a introdução de DNA em células germinais dos mosquitos e da identificação de genes capazes de bloquear ou reduzir a transmissão de parasitas aos humanos, é necessário o desenvolvimento de métodos eficientes para a introdução e fixação de genes refratários à malaria nas populações naturais de mosquitos. O uso de elementos de transposição tem sido sugerido para tais fins pelas características biológicas de invasão e propagação em genomas eucariotas que presentão estes elementos. Na presente tese, analisaram-se os elementos de transposição presentes no genoma do mosquito Anopheles gambiae, um dos principais vetores da malaria no mundo. Os resultados apresentados se encontram divididos em três partes, primeiramente a descrição de AnoTExcel, uma base de dados com informação detalhada dos elementos de transposição presentes neste genoma. Por outra, a partir desta base, foram identificados e caracterizados elementos novos, não detectados previamente em nenhum genoma. Por último, foram analisados elementos representativos pertencentes às diferentes classes de elementos de transposição desde uma perspectiva evolutiva e foi proposta uma análise de redes (Network analysis) para inferir as inter-relações entre elementos de transposição pertencentes à mesma família. / Up today, there are no efficient mechanisms for the control of Malaria, one of the most important diseases in the world. In the last decades, the genetic transformation of the malaria vectors have been proposed as an alternative. For achieving so, besides the development of techniques for the introduction of foreign DNA into the mosquitoes germinal cells and the identification of genes able to block or reduced the parasites transmission to human, the development of efficient methods for the introduction and fixation of the refractory genes into the mosquitoes natural populations is needed. In this regard, the use of transposable elements has been suggested as a driver system due to their biological characteristics of eukaryotic genomes invasion and propagations. In this thesis the analysis of the transposable elements in the genome of Anopheles gambiae, one of the most important vectors of malaria, has been proposed. The results are divided in three parts, first of all a description of AnoTExcel, a database with detailed information of the transposable elements present in the mosquito genome. The characterization of Novel elements that have not been described before is also described. Last, an evolutionary analysis of representative elements from different classes and families of elements has been performed. Finally, we have also proposed a network analysis for inferring the relationships between elements within the same family.

Une distribution nationale de moustiquaires imprégnées d'insecticide à longue durée d'action à destination des populations vulnérables du Niger a été effectuée fin 2005. Déjà montrée lors d'études pilotes à l'échelle du village, l'efficacité de cet outil dans le contrôle du paludisme restait à évaluer à l'occasion de vastes programmes opérationnels qui se multiplient en Afrique. Peu d'études des populations de vecteurs ont été publiées dans ce cadre. Nous avons mis en place un suivi entomologique au niveau de sites sentinelles répartis dans la zone Sahélienne du Niger, ayant couvert trois saisons de transmission, dont une avant intervention considérée comme période contrôle. Les paramètres entomologiques de la transmission ont été déterminés pour An. gambiae s.l., et la distribution spatiale des deux principaux vecteurs, An. gambiae et An. arabiensis, a été précisée. Le suivi temporel a mis en évidence une baisse globale du niveau de transmission de P. falciparum, probablement entrainée par la forte hausse d'utilisation de moustiquaires imprégnées. Cependant la hausse de la résistance des populations aux pyréthrinoïdes semble avoir été rapidement amorcée faisant craindre à moyen terme une perte d'efficacité de cet outil central des stratégies de lutte contre le paludisme. L'étude de la structure génétique des populations d'An. gambiae et d'An. arabiensis à l'aide de marqueurs microsatellites a montré une homogénéité génétique dans l'espace, entre les villages, même séparés par plusieurs centaines de kilomètres, ainsi que dans le temps, entre la saison de transmission 2005 contrôle et la saison 2006 après distribution. Ces résultats ont suggéré qu'au cours de la première année d'intervention, la couverture en moustiquaires imprégnées atteinte n'a pas eu d'effet de masse suffisant pour entrainer une baisse de la diversité génétique ou une modification des fréquences alléliques des populations. La faible différenciation spatiale observée pourrait être expliquée par des échanges de gènes importants à l'intérieur de la zone d'étude, hypothèse appuyée par l'expansion rapide de la mutation kdr dans l'ensemble des sites où An. gambiae est présent. L'évaluation rigoureuse de tels programmes de contrôle permettra d'améliorer les outils de contrôle et par exemple de préserver l'efficacité des pyréthrinoïdes, seule classe d'insecticides actuellement disponible pour l'imprégnation des moustiquaires.

Philosophiae Doctor - PhD / The discovery of microRNAs (miRNAs) is one of the most exciting scientific breakthroughs in the last decade. miRNAs are short RNA molecules that do not encode proteins but instead, regulate gene expression. Over the past several years, thousands of miRNAs have been identified in various insect genomes through cloning and sequencing, and even by computational prediction. However, information concerning possible roles of miRNAs in mosquitoes is limited. Within this context, we report here the first systematic analysis of these tiny RNAs and their target mRNAs in one of the principal African malaria vectors, Anopheles funestus s.s. Firstly, to extend the known repertoire of miRNAs expressed in this insect, the small RNAs from the four developmental stages (egg, larvae, pupae and the adult females), were sequenced using next generation sequencing technology. A total of 98 miRNAs were identified, which included 65 known Anopheles miRNAs, 25 miRNAs conserved in other insects and 8 novel miRNAs that had not been reported in any species. We further characterized new variants for miR-2 and miR-927 and stem-loop precursors for miR-286 and miR-2944. The analysis showed that many miRNAs have stage-specific expression, and co-transcribed and co-regulated during development. Secondly, for a better understanding of the molecular details of the miRNAs function, we identified the target genes for the Anopheles miRNAs using a novel approach that identifies overlap genes among three target prediction tools followed by filtering genes based on functional enrichment of GO terms and KEGG pathways. We found that most of the miRNAs are metabolic regulators. Moreover, the results suggest implication of some miRNAs not only in the development but also in insect-parasite interaction. Finally, we developed the InsecTar database (http://insectar.sanbi.ac.za) for miRNA targets in the three mosquito species; Anopheles gambiae, Aedes aegypti, and Culex quinquefasciatus, which incorporates prediction and the functional analysis of these target genes. The proposed database will undoubtedly assist to explore the roles of these regulatory molecules in insects. This type of analysis is a key step towards improving our understanding of the complexity and regulationmode of miRNAs in mosquitoes. Moreover, this study opens the door for exploration of miRNA in regulation of critical physiological functions specific to vector arthropods which may lead to novel approaches to combat mosquito-borne infectious diseases.

Malaria continues to cause significant mortality in sub-Saharan Africa and elsewhere, and existing vector control measures are being threatened by growing resistance to pyrethroid insecticides. With the goal of developing new human-safe, resistance-breaking insecticides we have explored several classes of acetylcholinesterase inhibitors. In vitro assay studies demonstrate that tri- and difluoromethyl ketones can potentially inhibit An. gambiae AChE (AgAChE). These compounds inhibit the enzyme by making a covalent adduct with the catalytic serine of AChE. Trifluoromethyl ketones however are poor inhibitors of the G119S resistant mutant of AgAChE. However difluoromethyl ketones can inhibit G119S AgAChE and compound 3-10g showed an IC₅₀ value of 25.1 nM after 23h incubation time. Despite this potent inhibition of AgAChE, the tri-, di-, and (mono)fluoroketones showed very low toxicity to An. gambiae, perhaps due to hydration and rapid clearance. In an attempt to improve An. gambiae toxicity, oximes and oxime ethers of these compounds were prepared as potential prodrugs. These structures identified trifluoromethyl ketone oxime 3-2d as a potent toxin against both wild-type (G3-strain) and a multiply resistant (Akron) strain of An. gambiae. This compound is within 3-fold of the toxicity of propoxur to wild type An. gambiae (LC₅₀ values of 106 and 39 µg/mL, respectively). Most significantly, 3-2d was much more toxic than propoxur to multiply-resistant (Akron) strain An. gambiae (LC₅₀ = 112 and >5,000 µg/mL, respectively). However, thus far we have not been able to link the toxicity of these compounds to a cholinergic mechanism. Pre-incubation studies suggest that significant hydrolysis of these compounds to TFKs does not occur over 22 h at pH 7.7 or 5.5. The mechanism of action of 3-2d remains unknown. Our enzyme inhibition studies have demonstrated that 3-2d does not hydrolyze to the trifluoromethyl ketone 2-9d at pH 7.7. The high Akron toxicity of 3-2d and poor inhibition of G119S AgAChE by 2-9d argue against enzyme mediated conversion of 3-2d to 2-9d within the mosquito. Thus, we can rule out an AChE inhibition mechanism for toxicity. Additional experiments by our collaborator (Dr. Jeffrey Bloomquist, University of Florida) also rule out inhibition of mitochondrial respiration or agonism of the muscarinic acetylcholine receptor. Future work will address other potential insecticidal modes of action. / Ph. D.

Malaria is one of the deadliest diseases known to mankind. In 2010, 219 million cases were reported, and 666,000 deaths were attributed to this disease. In the past, pyrethroid-treated mosquito nets have shown efficacy in reducing malaria transmission in many malaria endemic regions. However, an upsurge in the mosquito population that is resistant to pyrethroids threatens to compromise the efficacy of pyrethroid-treated bed nets. In an effort to develop another class of insecticide with a different mode of action, we have explored three classes of five membered heterocyclic carbamates (isoxazol-3-yl, pyrazol-5-yl, and pyrazol-4-yl), and 3-oxoisoxazole- 2(3H)-carboxamide as acetylcholinesterase inhibitors (AChE) targeting wild type (G3) and resistant (Akron) malaria mosquito Anopheles gambiae (Ag). Isoxazole carboxamide and carbamates were obtained regioselectively through judicious use of two different protocols. The final products were characterized and identified using ¹H and ¹³C NMR, and mass spectroscopy. In addition, the carboxamide structure was confirmed using X-ray diffraction. Several of the novel carbamates and carboxamides evaluated exhibited excellent toxicity towards susceptible G3 and resistant Akron strain An. gambiae (48f LC₅₀ G3 = 41 μg/mL, LC₅₀ Akron = 58 μg/mL, and 47i LC₅₀ G3 = 38 μg/mL, LC₅₀ Akron = 40 μg/mL). Hence, achieving the resistance- breaking goal. On the contrary, the commercial aryl methylcarbamates currently approved for indoor residual sprays (IRS) showed no potency towards the resistant strain An. gambiae (LC₅₀ G3 = 16-42 μg/mL, and LC₅₀ Akron >5,000 μg/mL). Further, we observed low toxicological cross-resistance ratios (RR) for the toxic isoxazol-3-yl and pyrazol-4-yl carbamates, and 3- oxoisoxazole-2(3H)-carboxamides (RR = 0.5-2.0). Amongst the commercial AChE inhibitors approved for IRS, only aldicarb exhibited such low RR (RR = 0.5), whereas the RR for commercial aryl methylcarbamates exceed 130-fold. The low RR observed for these novel heterocyclic inhibitors would certainly be favorable for a new anticholinesterase-based mosquitocide targeting both the susceptible and resistant strain mosquitoes. Although the overall selectivity (Ag vs human) did not exceed 24-fold, the heterocyclic carbamates and carboxamides synthesized by the author showed appreciable inhibition of resistant AChE (G119S) in comparison to commercial aryl carbamates, which showed no inhibition at all. During the course of this project, the isoxazol-3-yl and pyrazol-5-yl methylcarbamates proved to be unstable, and thus could not be isolated. The synthesis of pyrazol-4-yl methylcarbamates using N-methylcarbamoyl chloride proved particularly challenging due to the formation of by-products called allophanates. The similar Rf of the by-product and the desired final product made the isolation laborious and time-consuming. We have successfully overcome this problem by employing a new protocol, where triphosgene served as the carbonylating agent and N-methylamine in THF was used as the amine source. In addition, we have also developed another one-pot protocol for a safer synthesis of pyrazol-4-yl methylcarbamates utilizing 1,1- carbonyldiimidazole (CDI), and N-methylamine hydrogen chloride salt. With the pyrazol-4-yl core, apart from achieving excellent toxicity towards both strains of An. gambiae, we have also achieved excellent AgAChE vs hAChE selectivity (Ag vs h >100-fold). Due to our continued interest in developing this core, we have devised a convenient, scalable, no-column approach for the synthesis an intermediate 103 that can be utilized to synthesize these compounds more efficiently. / Ph. D.

Doctor of Philosophy / Department of Entomology / Kun Yan Zhu / Chitin metabolism represents an attractive target site for combating insect pests as insect growth and development are strictly dependent on precisely toned chitin synthesis and degradation and this process is absent in humans and other vertebrates. However, current understanding on this process and the involved enzymes is rather limited in insects. In this study, two chitin synthase genes (AgCHS1 and AgCHS2 or AgCHSA and AgCHSB), and 20 chitinase and chitinase-like genes (groups I-VIII) presumably encoding the enzymes for chitin biosynthesis and degradation, respectively, were identified and characterized in African malaria mosquito, Anopheles gambiae. Immunohistochemistry analysis and developmental stage- and tissue-dependent transcript profiling by using reverse transcription PCR, real-time quantitative PCR, and in situ hybridization revealed new information on these genes. Current understanding on chitin synthases is extended by the expression profiles such as the localization of AgCHS1 and AgCHS2 transcripts in eggs, AgCHS2 transcripts in the posterior larval midgut, AgCHS1 and AgCHS2 proteins in the compound eyes, and AgCHS2 enzyme in pupal inter-segments. Chitinase and chitinase-like genes are highly diverse in their gene structure, domain organization, and stage- and tissue-specific expression patterns. Most of these genes were expressed in several stages. However, some genes are stage- and tissue-specific such as AgCht8 mainly in pupal and adult stages, AgCht2 and AgCht12 specifically in foregut, AgCht13 exclusively in midgut. Functional analysis of each chitin synthase gene was conducted by using the chitosan/dsRNA nanoparticle-based RNA interference (RNAi) through larval feeding. The repression of the AgCHS1 transcripts which are predominantly expressed in carcass initiated from the mosquito larval feeding of dsRNA suggests the systemic nature of RNAi in mosquito larvae. In addition, silencing of AgCHS1 increased larval susceptibilities to diflubenzuron, whereas silencing of AgCHS2 enhanced the peritrophic matrix disruption and thus increased larval susceptibilities to calcofluor white or dithiothreitol. Furthermore, a non-radioactive method was adapted and optimized to examine the chitin synthase activity in mosquitoes. By using this method, diflubenzuron and nikkomycin Z show limited in vitro inhibition on chitin synthase at high concentration in cell free system, whereas no in vivo inhibition was observed.

Le paludisme est la maladie tropicale la plus redoutable au monde. Un enfant africain meurt de paludisme toutes les 30 secondes. La complexité spécifique du paludisme serait liée à une forte interaction du Plasmodium et de son vecteur avec l’environnement. Les cartes de distribution géographique montrent que la présence et l’impact du paludisme en santé publique varient considérablement avec le climat. L’éco-épidémiologie du paludisme au Sahara est encore mal connu. L’absence de pluies pendant des années peut entraîner une rupture prolongée de la transmission, comme les années 1970 à 1973, où le paludisme s’était retiré du sahel du fait de la sécheresse. Lorsqu’il pleut abondamment, la transmission sporadique peut se transformer rapidement en épidémies de paludisme qui surprennent de temps en temps la population faiblement ou non immunisée. Le sahel est caractérisé par une variabilité accrue du climat dont les bases sont encore mal cernées. L’effet majeur prévisible du changement climatique serait la transformation de tout le cycle de l’eau dont les manifestations importantes en Afrique seront les sécheresses et les inondations. Les inondations provoquent d’importants déplacements de populations. Les pluies diluviennes devraient faciliter la propagation du paludisme et la colonisation de nouveaux territoires, voire une possibilité de migration retour de l’anophèle plus au Nord jusqu’au sud de l’Europe. Le manque d’immunité des populations nord-sahéliennes peut occasionner des phénomènes épidémiques de grande ampleur et à grande échelle. Notre étude visait à comprendre les mécanismes d’impact du climat sur le paludisme dans le Sahel malien. Nos ressources financières ne permettaient pas de mener une étude longue à l’échelle du climat. Néanmoins, nous avons voulu démontrer "ce qui peut se passer avec le paludisme au Sahara" en cas de pluies abondantes associées au changement climatique. Nous avons procédé à une analyse régionale des relations brutes entre facteurs environnementaux et transmission du paludisme, affinée par une étude locale sachant que la transmission est corrélée in fine aux variables climatiques locales. Contraints par les menaces terroristes au nord du Mali, l’étude locale à été conduite dans une localité du Sahel écologiquement proche du Sahara.Nous avons analysé l’évolution de la variabilité climatique par des modèles numériques en relation avec les données entomologiques intégrées aux données satellitaires du terrain, analysé comment la distribution spatiale des gîtes aquatiques et des habitats humains rythment la fréquence des pontes et des piqûres qui assure le transfert du Plasmodium entre les populations humaines et vectorielles. A large échelle, l’étude montre que la transmission du paludisme est globalement liée à la pluviométrie mais la durée de transmission relève des contraintes topographiques locales. L’abondance des anophèles est liée à la pluviométrie dans les zones exondées et au routage des eaux de pluies dans les zones inondées. L’étude a prouvé un divorce entre la densité des moustiques et le risque de transfert du Plasmodium. Anopheles gambiae infligeait à lui seule 16 fois plus de piqûres inondées qu’exondées. Le taux d’infection de gambiae était 3,5 fois plus élevé en zone exondée qu’inondée et son risque de transmission 2,68 fois plus élevé en zone exondée qu’inondée. A l’échelle réduite du village, gambiae était produit de façon continue bien qu’il pleuve seulement 2-3 mois sur 12 de l’année. Parallèlement, une transmission estivale de mare était relayée par une transmission hivernale. / Nous pensons que ces résultats aideront les acteurs de santé publique à mieux comprendre l’épidémiologie du paludisme au Sahel, en vue d’orienter d’avantage la recherche sur des outils additionnels de lutte pour freiner la propagation du paludisme grâce à la gestion environnementale. Ils fournissent une piste prédictive du comment le changement climatique impactera la transmission du paludisme en Afrique.

>Magister Scientiae - MSc / Tsetse flies (Glossina spp) are vectors of African trypanosome (Trypanosoma spp) parasites, causative agents of Human African trypanosomiasis (sleeping sickness) and Nagana in livestock. Research suggests that tsetse fly immunity factors are key determinants in the success and failure of infection and the maturation process of parasites. An analysis of tsetse fly immunity factors is limited by the paucity of genomic data for Glossina spp. Nevertheless, completely sequenced and assembled genomes of Drosophila melanogaster, Anopheles gambiae and Aedes aegypti provide an opportunity to characterize protein families in species such as Glossina by using a comparative genomics approach. In this study we characterize thioester-containing proteins (TEPs), a sub-family of immunity-related proteins, in Glossina by leveraging the EST data for G.morsitans and the genomic resources of D. melanogaster, A. gambiae as well as A.aegypti.A total of 17 TEPs corresponding to Drosophila (four TEPs), Anopheles (eleven TEPs) and Aedes aegypti (two TEPs) were collected from published data supplemented with Genbank searches. In the absence of genome data for G. morsitans, 124 000 G.morsitans ESTs were clustered and assembled into 18 413 transcripts (contigs and singletons). Five Glossina contigs (Gmcn1115, Gmcn1116, Gmcn2398, Gmcn2281 and Gmcn4297) were identified as putative TEPs by BLAST searches. Phylogenetic analyses were conducted to determine the relationship of collected TEP proteins.Gmcn1115 clustered with DmtepI and DmtepII while Gmcn2398 is placed in a separate branch, suggesting that it is specific to G. morsitans.The TEPs are highly conserved within D. melanogaster as reflected in the conservation of the thioester domain, while only two and one TEPs in A. gambiae and A. aegypti thioester domain show conservation of the thioester domain suggesting that these proteins are subjected to high levels of selection. Despite the absence of a sequenced genome for G. morsitans, at least two putative TEPs where identified from EST data.