La dengue est une maladie virale des régions tropicales et subtropicales, transmise par les moustiques du genre Aedes. Le virus de la dengue (DENV) appartient à la famille des Flaviviridae, genre Flavivirus. Si la plupart des infections sont asymptomatiques ou se traduisent par un syndrome fébrile sans gravité, le virus peut aussi causer une maladie plus sévère caractérisée par une fuite plasmatique, avec ou sans hémorragie. Sans prise en charge adéquate, les formes les plus sévères peuvent évoluer vers un syndrome de choc, potentiellement mortel. Il n’existe pas de traitement spécifique de la dengue mais une réhydratation adaptée et débutée précocement permet de réduire la survenue de formes sévères de la maladie. Malheureusement, les symptômes initiaux de la dengue avant la survenue des éventuelles complications ne sont pas spécifiques et seul un diagnostic biologique basé sur la détection du génome viral, de l’antigène NS1 ou des anticorps anti-DENV dans le sang des patients permet de confirmer la nature exacte de l’infection. La dengue constitue à l’heure actuelle un problème majeur de santé publique du fait de son expansion mondiale et de l’augmentation annuelle du nombre de cas sévères. Pour assurer la surveillance épidémiologique et le contrôle de la maladie, il est indispensable de développer des outils diagnostiques performants et faciles à mettre en œuvre, à la fois utilisables par les médecins de toutes les structures médicales, des simples centres de soins de santé primaire aux centres de référence, et utilisables lors d’enquêtes épidémiologiques pour l’investigation de nouvelles épidémies. Le travail de cette thèse a porté sur plusieurs aspects de cette problématique. Dans une première partie, un test commercial de diagnostic rapide (TDR) permettant la détection simultanée de la NS1 et des IgG et IgM anti-DENV, a été évalué, en laboratoire spécialisé et sur le terrain, afin de comparer, à partir des mêmes échantillons, les performances du test dans deux situations différentes. La sensibilité s’est révélée plus faible lors de l’utilisation sur le terrain que lors de l’utilisation en laboratoire de référence. La majorité des discordances a été observées pour la détection des IgG et des IgM. L’impact de la mise à disposition du test sur la prise en charge des patients a également été évalué et il s’est avéré qu’au cours de cette étude les pédiatres cambodgiens ont ignorés les résultats du test rapide et ont préféré suivre leur instinct clinique.Un second volet a porté sur la faisabilité d’utiliser les urines et la salive en remplacement du sang veineux pour les tests employés en routine pour le diagnostic de la dengue. Les urines et la salive sont des fluides biologiques plus faciles à prélever que le sang veineux ce qui présente un avantage majeur pour les enquêtes épidémiologiques mais peut également secourir les médecins lorsqu’un prélèvement de sang veineux est difficile à obtenir, par exemple chez les enfants. Bien que les performances des différentes méthodes de diagnostic ne soient pas aussi bonnes avec de l’urine et la salive qu’avec du plasma, les résultats obtenus par PCR en temps réel et avec les ELISAs de détection des anticorps anti-DENV démontrent l’intérêt potentiel de ces deux fluides biologiques pour détecter les infections par le DENV lorsqu’il est difficile d’obtenir du sang veineux. Plusieurs TDR commerciaux développés pour permettre la détection de la NS1 et des anticorps anti-DENV (IgM, IgG et IgA) dans les urines et la salive ont été évalués mais les performances obtenues se sont révélées peu satisfaisantes.Une dernière partie du travail a été consacrée à l’étude de la protéinurie comme marqueur pronostic potentiel de sévérité de la dengue. Ce marqueur biologique ne s’est pas révélé être utile pour diagnostiquer précocement les formes sévères de la maladie. / Dengue is a viral disease transmitted by Aedes species mosquitoes, in tropical and subtropical regions. Dengue virus (DENV) belongs to the family Flaviviridae, genus Flavivirus. Although most DENV infections are asymptomatic or result in a self-limited febrile illness, severe diseases characterized by plasma leakage, with or without hemorrhage, can also occur. Patients with a severe dengue can rapidly progress into a life-threatening shock syndrome if no efficient clinical management is provided. There is no specific treatment available for dengue but an accurate and early fluid therapy substantially reduces the occurrence of severe forms of the disease. Dengue symptoms are typically non-specific until or unless complications develop. Only a biologic diagnosis based on DENV genome, NS1 antigen or anti-DENV antibodies detection enables to confirm dengue cases. Dengue is now a major public health problem due to both its geographical spread and the increase in the number of severe cases. New diagnostic tools are necessary to ensure epidemiological surveillance and control of the disease. These tools need to be effective and easy to use in every medical settings, from the smallest primary health centers to the biggest reference centers, and also usable for epidemiologic studies, e.g. for epidemic investigations. The work presented in this thesis was dedicated to this problematic.In a first part of the work, a rapid diagnostic test (RDT), designed to detect NS1 antigen, anti-DENV IgG and IgM, was evaluated, both in a specialized laboratory and in the field, in order to compare the test performances in two different settings, with the same samples. Interestingly, sensitivity was lower when the test was used in the field compared to the sensitivity of the test when performed in the specialized laboratory. Discordances were mainly observed for IgM and IgG detection. Impact of the use of the RDT on clinical management was also assessed during the field study and it revealed that Cambodian pediatricians ignored the results of the RDT and followed their clinical instinct.A second part of the work was dedicated to the assessment of the usefulness of urine and saliva for dengue diagnostic. Dengue diagnostic normally requires a venous blood sample that can be difficult to obtain in certain conditions such as in children or during epidemiological studies. Urine and saliva are easier to collect as the procedure is non-invasive. We showed that, although the performances of the different diagnostic methods were not as good in saliva and urine as in plasma specimens, the results obtained by qRT-PCR and by anti-DENV antibody ELISA could well justify the use of these two body fluids to detect dengue infection in situations when the collection of blood samples is difficult. Performances of commercial RDTs developed for NS1 and anti-DENV antibodies (IgM, IgG and IgA) detection in urine and saliva specimens were not satisfactory.In the last part of the thesis, the potential use of proteinuria as a prognostic marker of severity was assessed but it didn’t prove to be a useful marker for risk prediction.

Introduction: Ensuring universal access to primary health care (PHC) is a key component of the Ethiopian nationalhealth policy. The policy also emphasises promoting and enhancing national self-reliance in health development bymobilizing and efficiently utilizing resources including community participation. To this end, the government introducedthe accelerated expansion of the PHC strategy through a comprehensive health extension programme (HEP). HEP is afamily and community-based health care delivery system institutionalised at health post level which combines carefullyselected high impact promotive, preventive and basic curative interventions. All HEP interventions are promotive and preventive except the malaria intervention which, in addition, incorporates a curative service. In the country, malaria is a leading disease. Unlike most Sub-Saharan African countries where P. falciparum accounts for almost all malaria infections, in Ethiopia both P. falciparum and P. vivax are co-dominant. Considering this peculiar epidemiological nature, the national guideline recommends alternative diagnosis and treatment strategies. Rationale: The lack of adequate resources and the efficiency with which available resources are being utilised are the main challenges in any health care setting. Therefore, if the HEP which consumes consideral amount of resource desires to reach its intended goal, monitoring and improving its efficiency is of great public heath importance. HEP has been successful in improving access to PHC including the malaria diagnosis and treatment service. Though this is a crucial measure, its quality ought to be considered. For the malaria curative service, studying the cost-effectiveness of the available strategy and patients’ adherence to the treatment regimen can be considered as proxy measures of quality for which local evidence is lacking. However, none of the existing studies in this field of research has addressed the Ethiopian malaria epidemiological context and its diagnosis and treatment guideline. In Tigray, for more than two decades, access to malaria early diagnosis and prompt treatment was facilitated by volunteer community healthworkers (CHWs). However, with the introduction of artemether-lumefantrine (AL) the service was compromised mainly for reasons of cost, safety and logistic. Therefore, it was important to explore the feasibility and the impact of community deployment of AL with rapid diagnostic tests (RDTs). The aim: to explore the overall performance of HEP and particularly the access to and quality of malaria early diagnosis and prompt treatment in the Tigray region of Ethiopia. Methods: Different study designs and populations were used for each of the four specific objectives. Data envelop analysis (DEA) was applied to assess the HEP efficiency. For this, register data for the output variables and primary data for the input and the environmental factors were collected. A health provider perspective cost-effectiveness analysis was used to determine which among the currently available diagnostic and treatment strategies is best for the country. Effectiveness data were generated from a stratified cross-sectional survey and secondary data were used to calculate the cost. For measuring adherence to the six-dose AL regimen, an assessment questionnaire and pill count was employed at patients´ home. To determine whether deploying AL with RDT at community level was feasible and effective, a number of designs were used: longitudinal follow-up, cross-sectional surveys, cost analysis, verbal autopsyquestionnaires and focal group discussions. Main findings: More than three-quarters of the health posts were found to be technically inefficient with an average score of 42%, which implies potentially they could improve their efficiency by 58%. Scale of operation was not a cause of inefficiency. None of the considered environmental factors was associated with efficiency. The Parascreen-based strategy (multispecies RDT-BS) was found to be the most cost-effective strategy, which allowed treating correctly an additional 65% of patients with less cost than the paracheck-BS. Presumptive-BS was highly dominated. Among P.falciparum positive patients to whom AL was prescribed, more than a quarter did not finish their treatment. The main reasons for interrupting the dose were ‘too many tablets’ and ‘felt better before finishing the dose’. The ownership of aradio, the belief that malaria cannot be treated traditionally and a delay of more than one day in seeking treatment after the onset of fever were significantly associated with being adherent. Deploying AL with RDT at community level was demonstrated to be effective and feasible. In the intervention district, almost 60% of suspected cases were managed by CHWs. Malaria transmission was lower at least threefold and malaria mortality risk by around 40% compared to the control district. The use of RDTs reduced cost and possibly the risk of drug resistance development. Conclusion: Though improving access to health care is important, it should be considered a means, not an end. Themore accessible a system is the more people could utilise it to improve their health. Thus, ensuring the access obtainedthrough HEP is maintained, its quality is improved and efficiently utilised to its optimal productivity level is a necessarytask. The DEA study revealed a high level of inefficiency where majority of the health posts needed improvement.This thesis also found parascreen-BS to be the most cost-effective strategy and that there is no epidemiological andeconomical contextual justification to keep both, the presumptive-BS and the RDT-BS specific only to P.falciparum.The high poor adherence levels raises great concern as it leads to recurrent malaria attacks of the patient, speed upthe development and spread of drug resistance strains and reduces the effect of the drug on the transmission. Therefore,providing effective drug alone is not sufficient; assessing and monitoring adherence to the treatment is by faressential. Deployment of AL with RDT through a community-based service has shown an enormous impact in termsof cost, transmission, morbidity and mortality. However, it is worth noting that this results came from an area wherea community-based service has been involved in the PHC system for more than three decades.

Le paludisme est un problème de santé public dans de nombreux pays. Cinq espèces infectent l'homme : P. falciparum, responsable de la grande majorité des décès, et P. vivax, P. ovale, P. malariae et P. knowlesi qui provoquent des formes bénignes de la maladie. Le diagnostic qui fait partie des moyens de lutte, est une urgence médicale. Les tests de diagnostic rapides (TDRs) dont l'usage est recommandés par l'OMS, sont donc de plus en plus employés. Cependant, la détection et l'identification des espèces non P. falciparum par ces tests est insuffisante. Le besoin en nouveaux couples « antigènes-anticorps » est une nécessité pour améliorer les TDRs. Au cours de ce travail, de nouveaux anticorps anti LDH de P.malariae ont été produits.Une recherche de nouveaux antigènes a également été entreprise. Pour cela, certaines enzymes de la voie de la glycolyse ont été étudiées. Pour la première fois des séquences des enzymes de cette voie ont été obtenues pour P. ovale et P. malariae. Elles ont permis de déterminer de nombreux épitopes cibles potentiels spécifiques et ceux communs à toutes les espèces. Dans un deuxième temps, une recherche en protéomique a été menée pour identifier des biomarqueurs parasitaires. L'étude du culot globulaire et du plasma de patients infectés a permis la sélection de 8 protéines cibles originales. Ces travaux préparent la fabrication et la commercialisation par la société Whidiag d'une nouvelle génération de TDRs pour le paludisme. / Malaria is a public health problem in many countries. Five species infect humans: P. falciparum, responsible for the vast majority of deaths, and P. vivax, P. ovale, P. malariae and P. knowlesi causing mild forms of the disease. The diagnostic is a means of control and a medical emergency. The rapid diagnostic tests (RDT) whose are recommended by WHO, are increasingly used. However, the detection and identification of not P. falciparum species is insufficient. New "antigen-antibody" couples are a need to improve the RDTs performance. In this work, new anti LDH antibodies from P. malariae were produced. A search for new antigens was also undertaken. For this purpose, some enzyme of glycolysis pathway were studied. For the first time the sequences of the enzymes from this pathway were obtained for P. ovale and P. malariae. We identified many potential target epitopes specific and common to all those species. In a second step, a proteomics approches has been conducted to identify parasites biomarkers. The study of red blood cells and plasma of infected patients has led to the selection of 8 original target proteins. This work prepares the manufacturing and marketing of a new generation of RDTs for malaria by the company Whidiag

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