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La communauté Rurale de Mont-Rolland (région de Thiès, Sénégal) est un foyer endémique de leishmaniose canine décrit depuis 1970. Des études épidémiologiques récentes ont montré que l'agent étiologique est Leishmania (Leishmania) infantum et que le cycle de transmission est bien établi sur l'ensemble de la commune, avec plus de 30% des chiens et plus de 20% des hommes séropositifs. Néanmoins, malgré les différentes études épidémiologiques, le vecteur reste toujours inconnu. Jusqu'à présent, aucune espèce de Phlebotomus, connue pour être vecteur de L. infantum (sous genre Larroussius et secondairement Adlerius), n'a été identifiée au Sénégal. Dans ce contexte, l'objectif principal de cette thèse était d'identifier les vecteurs potentiels de la maladie et de caractériser le cycle de transmission de la leishmaniose canine par des études écologiques, parasitologiques et moléculaires des populations de phlébotomes. Ainsi, 7442 spécimens de phlébotomes ont été récoltés par piégeage adhésif, piégeage lumineux et par pulvérisations intradomiciliaires d'insecticides pyréthrinoïdes. Neuf espèces de phlébotomes ont été identifiées : deux appartiennent au genre Phlebotomus, P. duboscqi (vecteur de la leishmaniose cutanée au Sénégal) et P. rodhaini, les autres espèces appartiennent au genre Sergentomyia, S. adleri, S. clydei, S. antennata, S. buxtoni, S. dubia, S. schwetzi, et S. magna. La distribution spatiale des phlébotomes est hétérogène selon le village, selon l'écosystème et l'environnement de capture. Les études parasitologiques et les études moléculaires (PCR diagnostique de Leishmania) ont révélé trois espèces de Sergentomyia infectées par L. infantum, S. dubia (dissection et PCR), S. schwetzi (dissection et PCR) et S. magna (PCR). Il faut noter que plus de 2% des individus femelles à jeun testées étaient PCR positives, soulignant la survie des parasites chez ces espèces après la digestion. Toutes les autres espèces étaient négatives aussi bien en dissection qu'en PCR. Les analyses statistiques ont montré : que la distribution de S. dubia est significativement associée à la séroprévalence chez les chiens, que les individus de S. schwetzi positifs en PCR sont significativement associés à la séroprévalence chez le chien, qu'il y a un effet significatif du nombre de phlébotomes positifs en PCR (les 3 espèces confondues) sur la séroprévalence chez l'homme. Toutes ces données démontrent pour la première fois que 3 espèces du genre Sergentomyia, S. dubia, S. schwetzi et probablement S. magna, sont les vecteurs de la leishmaniose canine dans le foyer de Mont-Rolland. Ces résultats remettent en question le dogme qui stipule que seul le genre Phlebotomus serait capable de transmettre le parasite Leishmania dans l'Ancien-Monde. L'étude du comportement et de la distribution de ces trois espèces et de leurs spécimens infectés nous ont permis de proposer un modèle de transmission dans ce foyer. / AbstractThe rural community of Mont-Rolland (Thiès, Sénégal) is an endemic focus of canine leishmaniasis described since 1970. Recent epidemiological studies showed that the causative agent is Leishmania infantum with well established transmission cycle throughout the area. However, despite the several epidemiological studies, the vector species is still unknown. So far, no species of Phlebotomus belongs to Larroussius and Adlerius subgenera, known as vector of L. infantum, was identified in Senegal. In this context, the main objective of this thesis was to identify potential vectors of the disease and to characterize the cycle of canine leishmaniasis transmission by ecological, parasitological and molecular studies on sandfly populations. Thus, 7442 specimens were collected by sticky traps, light traps and indoor spraying with pyrethroid insecticide from different environments (intradomiciliary, peridomiciliary, culture area, isolated area) and different villages and ecosystems (sandy soils, lateritic gravel soils and sandy clay soils). Nine species of sand flies were identified; two belong to the Phlebotomus genus, P. duboscqi (vector of cutaneous leishmaniasis in Senegal) and P. rodhaini, the other species belong to the Sergentomyia genus, S. adleri, S. clydei, S. antennata, S. buxtoni, S. dubia, S. schwetzi, and S. magna. Two species of the Phlebotomus genus were significantly underrepresented (292 specimens) compared to species of the Sergentomyia genus (7150 specimens). The spatial distribution of sand flies is heterogeneous depending on the village, the ecosystem and environment, suggesting that several species are capable of transmitting the parasite. Parasitological and molecular studies (diagnostic PCR of Leishmania) revealed that three species of Sergentomyia were infected with L. infantum, S. dubia (dissection and PCR), S. schwetzi (dissection and PCR) and S. magna (PCR). It is worth noting that more than 2% of un-fed females of these species were PCR-positive, emphasizing the survival of parasites after digestion. All other species were found to be negative in both PCR and dissection. In addition, statistical analysis showed that 1) the distribution of S. dubia is significantly associated with seroprevalence in dogs, 2) PCR-positive individuals of S. schwetzi are significantly associated with seroprevalence in dogs, 3) there is a significant effect of PCR-positive sand flies (all species considered) on seroprevalence in humans. All these data demonstrated for the first time that three species of Sergentomyia, S. dubia, S. schwetzi and probably S. magna, are the vectors of the canine leishmaniasis in the Mont-Rolland commune. The results presented in this study challenge the dogma which states that only the Phlebotomus genus would be capable of transmitting Leishmania parasites in the Old World. The behaviour and distribution of these three species and their infected specimens allowed us to propose a model of transmission in the focus.

Sand flies (Diptera, Phlebotominae) are small biting insects and vectors of Leishmania spp. which cause medically and veterinary important disease - leishmaniasis. During the piercing of the host skin, sand fly females inject saliva to facilitate the blood feeding. The sand fly saliva is composed of many bioactive molecules which were shown to possess anti-inflammatory and anti-haemostatic functions. The saliva affects host's immunity in the bite site and consequently enhances the survival and development of transmitted pathogens. Most of the studies focus on salivary proteins and enzymes of sand flies belonging to Phlebotomus and Lutzomyia genera, while salivary proteins from sand flies of the third genus Sergentomyia were neglected so far. In this thesis we focused on comparison of salivary proteins from two Phlebotomus species, namely Phlebotomus perniciosus and Phlebotomus orientalis, and Sergentomyia schwetzi. These sand fly species differ not only by the ecology and geographical distribution but also by host preferences. Both Phlebotomus species prefer large or medium-size mammals as the bloodmeal source, particularly rabbits, hares and dogs for P. perniciosus and cattle, goats, sheep and humans for P. orientalis. Contrarily, Sergentomyia sand flies are known for preferred feeding on reptiles...

Introdução. A identificação dos flebotomíneos baseia-se principalmente na morfologia do adulto, o que pode ser problemático quando as espécies são morfologicamente muito semelhantes. Psychodopygus é um gênero de flebotomíneos de grande interesse em saúde pública devido ao papel de algumas espécies na veiculação de Leishmania spp. no Brasil. No entanto, este gênero inclui espécies com fêmeas morfologicamente indistinguíveis que pertencem à Série Chagasi, sendo elas: P. chagasi, P. complexus, P. squamiventris maripaensis, P. squamiventris squamiventris e P. wellcomei. Objetivos. Investigar a possibilidade de distinguir essas espécies por meio de análises morfométrica e molecular, além de produzir uma distribuição geográfica atualizada para o grupo analisando a probabilidade de ocorrência das espécies através da análise de modelagem de nicho ecológico. Material e Métodos. Foi realizada a análise discriminante na morfometria geométrica (cabeça e asa) e linear, morfologia (usando microscopia óptica e eletrônica de varredura) e a análise do citocromo c oxidase subunidade 1 (COI), avaliando-se um total de 752 espécimes (460 fêmeas e 292 machos) dos seguintes estados Amapá, Amazonas, Ceará, Mato Grosso, Pará, Rondônia, Roraima e Tocantins. Mapas de distribuição foram produzidos através de dados obtidos do material analisado e de revisão bibliográfica. Resultados. A análise discriminante usando caracteres morfométricos lineares mostrou-se capaz de diferenciar todas as espécies, exceto P. complexus, que apresentou 2,2% de erro de identificação. A morfometria geométrica das asas foi incapaz de separar completamente as espécies através da conformação, mas o tamanho do centróide dos espécimes fêmeas falhou apenas em distinguir P. complexus de P. s. maripaensis. Por outro lado, a morfometria geométrica das cabeças foi capaz de distinguir todas as espécies com grande eficiência ao usar tanto a forma como o tamanho do centróide. A análise morfológica revelou que a coloração torácica, principalmente do pronoto e do pós-noto, pode ser usada para separar as cinco espécies em três grupos: P. chagasi, P. wellcomei / P. complexus e P. s. mariapaensis / P. s. squamiventris. Os resultados da análise de DNA Barcoding, mostraram um agrupamento semelhante ao observado na morfologia; embora os espécimes de P. wellcomei do estado do Ceará mostrem uma grande distância genética da população do estado do Pará, evidenciando que essa espécie possa representar um complexo. Quanto à microscopia eletrônica de varredura, foram avaliadas detalhadamente as estruturas das antenas, tórax e genitália masculina. Salientamos que no anepímero (tórax) foi observada uma escama tipo \"raquete\" modificada apenas em Psychodopygus s. squamiventris. A revisão da distribuição geográfica mostrou que as espécies possuem uma distribuição cis-andina, ocorrendo principalmente no bioma Amazônico. A nítida separação de algumas espécies pelo rio Amazonas, sugere que o surgimento do grupo ocorreu no período que se estende da orogênese dos Andes até a formação deste rio. Conclusões. O estudo possibilitou diferenciar completamente as fêmeas das cinco espécies da Série Chagasi utilizando o conjunto de dados obtidos por morfometria linear e geométrica e análises morfológicas e também apresentar novos caracteres morfológicos e padrões distribucionais que facilitarão a identificação de machos e fêmeas dessas espécies. / Introduction. The identification of sand flies is mainly based on adult morphology, which can be problematic when species are morphologically very similar. Psychodopygus is one of the sand fly genera of great interest in public health, due to the role of some species in the transmission of Leishmania spp. in Brazil. However, this genus includes species with morphologically indistinguishable females that belong to the Chagasi series, which includes: P. chagasi, P. complexus, P. squamiventris maripaensis, P. squamiventris squamiventris and P. wellcomei. Objectives. To investigate the possibility of distinguishing among these species by means of morphometric and molecular analyses in addition to producing an updated geographical distribution for the group, analyzing the probability of the occurrence of the species by the analysis of ecological niche modeling. Material and methods. The analyses of the cytochrome c oxidase subunit 1 (COI), geometrical (head and wing) and of linear morphometry and morphology (using optical microscopy and scanning electron microscopy) were carried out using a total of 752 specimens (460 females and 292 males) from the following states: Amapá, Amazonas, Ceará, Mato Grosso, Pará, Rondônia, Roraima e Tocantins. Distribution maps were produced on the basis of data obtained from the material analyzed and a bibliographical review. Results. The discriminant analysis using linear morphometric characters was able to differentiate among all the species, except for P. complexus, which presented a 2.2% error of identification. The geometric morphometry of the wings was unable to completely separate the species by means of the shape analyses, but the centroid size of the female specimens only failed to distinguish P. complexus from P. s. maripaensis. Otherwise, the geometric morphometry of the heads was sufficient to distinguish all the species with great efficiency, when using both the head-shape and the centroid size. The morphological analysis revealed that the thoracic coloration, mainly of the pronotum and the post-notum, can be used to separate the five species into three groups: P. chagasi, P. wellcomei / P. complexus, P. s. mariapaensis / P. s. squamiventris. The results of the Barcoding DNA analyses showed a cluster similar to that observed in the morphology; however, P. wellcomei specimens from the Ceará population showed a great genetic distance from the population of Pará, evidencing that this species may represent a complex. As for the scanning electron microscopy, the structures of the antennae, thorax and male genitalia were evaluated in detail. In the anepimerum (thorax) a modified \"racket\"-type scale was observed only in Psychodopygus s. squamiventris. The review of the geographical distribution showed that the species have a cis-Andean distribution, occurring mainly in the Amazonian biome. The separation of some species from the others by the Amazon river suggests that the appearance of the Chagasi series occurred in the period from the orogenesis of the Andes to the formation of this river. Conclusions. The results clearly differentiate the females of the five species of the Chagasi series using the data set of linear and geometric morphometry and morphological analyses, providing new morphological and distributional data that will facilitate the identification of the males and females of this group.