Thesis (M. Tech.) - Central University of Technology, Free State, 2007 / Malaria and the mosquito that induces the disease in humans have hounded the military for decades. Malaria represents one of the most important infectious disease threats to deployed military forces. Malaria in soldiers has a serious economic impact, both in terms of lost productivity and treatment cost for the state. A contingent of South African National Defence Force members has been deployed in Burundi since November 2001, as part of a peacekeeping mission. At the time of the study no information was available regarding the prevalence of malaria among military personnel during deployments in Burundi and East Africa. In Africa, the saying is that malaria is the disease of poverty and a cause of poverty. To combat malaria, it is of vital importance that the recommended medication be taken exactly as prescribed and that the course is completed. However, one of the greatest challenges facing the African continent in the present fight against malaria is drug resistance. The discovery of Mefloquine and the subsequent development of suitable drugs, have been intimately associated with military imperatives, contingencies and requirements. Since World War II, the development of Chloroquine-resistant falciparum malaria has driven the search for new drugs. Mefloquine, developed by the Walter Reed Army Institute of Research in the United States, was first shown effective as a prophylaxis and treatment of resistant falciparum malaria in the 1970’s. To obtain data, questionnaires were administered to SANDF soldiers deployed in Burundi, East Africa. The total size of the population under investigation was 336 with a final sample size of 111 respondents. The sample was selected by using simple random sampling. The questionnaire aimed to determine the perception of respondents regarding the malaria threat, their compliance with taking the medication, and their experiencing of possible side-effects which might occur due to the chemoprophylaxis and the prophylactic efficacy of Mefliam®. The fact that, of the 111 people who used Mefliam®, only four presented with any malaria symptoms, is an indicator that Mefliam® is an effective option as an antimalarial drug to be used in East Africa and Burundi. The results of this study will be used by the personnel of the South African National Defence Force (SANDF) and other military forces deployed in East Africa. It is envisaged that the results will be used by military policy- and decision-makers as a control programme and by others involved in the control of malaria. The findings and recommendations should also be of interest to anyone visiting the area.

Replication-deficient adenovirus (Ad) and modified vaccinia virus Ankara (MVA) vectors expressing single Plasmodium falciparum antigens can induce potent T cell and antibody responses and have entered clinical testing using a heterologous prime-boost immunisation approach (Ad_MVA). This thesis describes a number of pre-clinical approaches aimed at enhancing the efficacy of these viral vectored vaccines targeting the blood-stage of malaria. First, the development of a highly efficacious malaria vaccine is likely to require a multi-antigen and/or multi-stage subunit vaccine. The utility of an Ad_MVA immunisation regime combining vaccines expressing the 42kDa C-terminus of the blood- stage antigen merozoite surface protein 1 (MSP142) and the pre-erythrocytic antigen circumsporozoite protein (CSP) in the P. yoelii mouse model was investigated. It was found that vaccine co- administration leads to maintained antibody responses and efficacy against blood-stage infection, but reduced secondary CD8+ T cell responses and efficacy against liver-stage infection. CD8+ T cell interference can be minimised by co-administering the MVA vaccines at separate sites, resulting in enhanced liver-stage efficacy. The mechanisms of CD8+ T cell interference were explored. Second, Ad_MVA regimes expressing blood-stage antigens that can protect against P. chabaudi and P. yoelii blood-stage infection were tested against P. berghei, but did not confer protection. Similarly, IgG from rabbits immunised against P. falciparum MSP1 (PfMSP1) could not protect mice from a chimeric P. berghei parasite expressing PfMSP1. Third, two molecular adjuvants, the C4bp α-chain oligomerisation domain (IMX108/313) and the Fc fragment of murine IgG2a, were tested for their ability to enhance immunogenicity of recombinant adenoviruses when fused at the C-terminus of a blood-stage antigen. IMX108/313 was found to adjuvant T cell responses of small (< 80kDa) antigens and this was associated with antigen oligomerisation. However, the Fc fragment did not adjuvant responses. Finally, it was found that using a strong early promoter to drive antigen expression enhances the immunogenicity of single administration MVA vaccines, but that this did not enhance post-boost immunogenicity in an Ad_MVA regime.

Plasmodium vivax malaria threatens nearly half the world’s population. This relapsing disease may be more severe than previously recognised and is proving refractory to current malaria control measures. This thesis aimed to describe the burden of anaemia and mortality attributable to vivax malaria in Southern Papua, Indonesia, an area endemic for multidrug-resistant P. vivax and P. falciparum, and to determine the potential of currently available antimalarial drugs to reduce transmission of P. vivax in co-endemic regions. Approximately 0.5 million uniquely identified clinical records from patients presenting to Mitra Masyarakat Hospital between April 2004 and May 2009 were matched with corresponding laboratory and pharmacy data in order to determine the burden of anaemia in the hospital setting and the effectiveness of primaquine prescription for preventing P. vivax relapses. Clinical information extracted from patient notes was used to clarify the contribution of P. vivax malaria to a series of deaths detected by an active hospital-based surveillance system. Additional secondary sources of data used in this thesis included a large house-to-house survey and multiple clinical trials of antimalarial therapy from both Southern Papua and Northwestern Thailand. In Southern Papua, P. vivax malaria is an important cause of haematological morbidity both in the hospital and community setting. This morbidity is most significant in the first year of life when P. vivax infection accounts for 23% of all severe anaemia (haemoglobin <5g/dL) in the hospital and approximately 28% of all moderate-to-severe anaemia (haemoglobin <7g/dL) in the community. In this region concomitant P. vivax infection accentuates haematological impairment associated with P. falciparum malaria. Plasmodium vivax in Southern Papua rarely causes death directly but rather indirectly contributes to mortality through exacerbation of comorbid conditions. In Northwestern Thailand, 53.8% of patients with falciparum malaria who were treated with a rapidly eliminated drug between 1991 and 2005 had a recurrence of vivax malaria within two months making P. vivax infection the most common cause of parasitological failure in these individuals. Slowly eliminated artemisinin combination therapies (ACT) provided the greatest protection against recurrent P. vivax parasitaemia during 63 days of follow-up. In three randomised controlled trials from Papua and Thailand, P. vivax gametocytaemia was shown to mirror asexual parasitaemia closely and to have the same characteristics in acute and recurrent infections. This emphasises that the most important chemotherapeutic means of blocking P. vivax transmission is prevention of future relapse. Primaquine is recommended for this purpose but analyses in this thesis suggest that in Southern Papua, unsupervised primaquine at a dose of 0.5mg/kg/day for 14 days, does not reduce the risk of subsequent relapse (Adjusted Hazard Ratio = 1.01 [95% confidence interval 0.95-1.07]). Plasmodium vivax malaria should not be neglected. High priority must be given to new hypnozoitocidal drug discovery. In the interim, optimising the safety and effectiveness of primaquine and adoption of a unified ACT-based blood schizontocidal treatment strategy for malaria of any parasitological cause in co-endemic regions will be crucial for controlling P. vivax malaria.

Most in-hospital deaths secondary to infections in under-five deaths within sub-Saharan Africa (SSA) occur in the initial 24 hours of admission and shock has been identified as a major risk factor for the early deaths. However, controversies exist on the appropriate clinical diagnosis of shock, choice of ideal fluid for resuscitation (crystalloid or colloid), and safety of fluid resuscitation in severe malnutrition or severe malaria. This thesis investigates these aspects and also reviews the evidence base of current paediatric fluid resuscitation guidelines for children (aged >60 days and ≤12 years) with severe febrile illnesses. Capillary refill time >2 seconds, weak pulse volume, or bradycardia, in the presence of abnormal temperature and severe disease are predictive of impaired perfusion (defined by lactic acidosis) and death. Tachycardia and temperature gradient are neither associated with increased risk of death nor predictive of hypoperfusion. Existing international definitions of shock have low sensitivities (FEAST=44%, WHO=2%, and ACCM=59%) and high specificities (FEAST=82%, WHO=100%, and ACCM=66%) for diagnosis of impaired perfusion. Clinical criteria derived (called derived shock) had a sensitivity of 30% and specificity of 93%. Shock in children with severe febrile illnesses in Kilifi has a complex presentation but mainly presents with hyperdynamic circulation (high cardiac index) and vasodilatation. Cases with low cardiac index (myocardial dysfunction) are relatively rare but increase the risk of mortality when present. Synthetic colloids (gelofusine, hydroxyethyl starch 130/0.4 (HES), and dextran 70) are safe for use in fluid resuscitation in children with severe malaria. However, HES is the most promising compared to other synthetic colloids concerns still remain about its renal safety. However, further evaluation of synthetic colloids for treatment of shock is not warranted due to the findings of FEAST trial. A Pilot trial shows that bolus isotonic fluids are safe, have better efficacy, and produce faster resolution of shock compared to low-sodium solutions at volumes and rates recommended by WHO in children with severe malnutrition. Evidence available from all ten the trials in children with sepsis show that fluid resuscitation using crystalloids and colloids result in similar survival. However, fluid bolus resuscitation results in increased mortality compared to no bolus (control) in children in SSA. This finding excludes children with gastroenteritis, trauma, burns, and malnutrition. Colloids are better than crystalloids for severe dengue shock but both have similar efficacy in moderate dengue shock.

Malaria caused by the protozan parasite Plasmodium is one of the most serious infectious diseases in the developing world. It is estimated that malaria causes an annual mortality rate of ~627,000. New drugs are urgently required, as the incidence of resistance is spreading rapidly. Glycosylphosphatidylinositol (GPI) anchored proteins decorate the merozoite surface and several of which, including merozoite surface proteins - 1 and -2 have previously been shown to be essential for erythrocyte invasion and parasite survival. Plasmodium GPI-anchors contain a glycan core consisting of four mannose residues. Therefore, the enzymes involved in the synthesis of activated mannose, guanidine diphosphomannose pyrophosphorylase (GDP-Man PP) and dolichol phosphate mannose synthase (DPMS), are thought to be crucial for GPI-anchor biosynthesis and as such potential drug targets. Double homologous recombination has been exploited to test whether PfGDP-Man PP and PfDPMS are essential during the erythrocytic portion of the parasite's life cycle. Additionally, overexpression parasite lines for both enzymes have been generated and have shown that the regulation of the two enzymes are intricately linked. Focused metabolomics by multi-reaction monitoring of the overexpression lines suggests that the fucosylation pathway may have a novel function within the parasite, possibly as a dynamic store for activated fucose/mannose. In order to determine the cellular concentration of key metabolites within the parasite, the volumes of the intra-erythrocytic stages have been determined and show that the concentration of metabolites in the ring stage parasites is substantially higher than previously thought. Furthermore, the sub-cellular localisation of GDP-Man PP and DPMS has been determined by immunofluorescence assay. The recombinant expression of DPMS in E. coli allowed its active site residues to be probed as well as establishing a platform for inhibitors to be screened against the enzyme. Finally, inhibitors of the T. brucei DPMS enzyme have been screened against the P. falciparum parasites in culture.

Quatre études épidémiologiques menées en Afrique ont montré que les enfants issus de mères ayant un placenta infecté par Plasmodium falciparum lors de l’accouchement font des infections palustres plus précocement que les autres enfants. Le fœtus serait sensibilisé in utero par les parasites infectants et développerait par la suite une tolérance aux infections palustres. Cette hypothèse nous conduit à supposer que i) la réponse anticorps spécifique de P. falciparum est différente chez les enfants en fonction du statut infectieux du placenta des mères à l’accouchement et ii) que cette sensibilité ne pourrait être induite que par les antigènes parasitaires porteurs des mêmes polymorphismes que ceux rencontrés in utero. Un troisième projet a consisté au développement d’une méthodologie permettant de distinguer les anticorps maternels de ceux néo-synthétisés par l’enfant dans le but de mesurer précisément l’acquisition de la réponse anticorps élaborée par l’enfant dès son plus jeune âge. Nous avons mis en place le suivi régulier et rapproché d’une cohorte de 620 nouveau-nés de la naissance à 18 mois au Bénin. Nous avons mesuré leurs réponses anticorps dirigées contre sept antigènes de P. falciparum candidats vaccins et constaté que le processus de maturation immunitaire commence à être mis en place à l’âge de 18 mois. L’infection palustre placentaire ne semble pas influer sur l’acquisition de la réponse anticorps spécifique jusqu’à 18 mois de vie. La méthodologie de distinction des anticorps maternels et néo-synthétisés a été validée. La caractérisation des polymorphismes des antigènes parasitaires présents à l’accouchement et pendant le suivi des enfants, mis en relation avec les données environnementales, a permis de valider en partie l’hypothèse de tolérance immunitaire. / Four epidemiological studies showed that infants born from mothers with Plasmodium falciparum placental malaria at delivery present a higher susceptibility to plasmodial infections than others. In connection with this observation, we hypothesized that i) the infants’ P. falciparum specific antibody responses are different according to presence or absence of placental malaria at delivery in their mothers and ii) susceptibility could only be induced by antigens that bring the same polymorphisms as those found in infected mothers. Another project consisted to develop a new methodology to distinguish maternal and neonatal antibodies in order to measure accurately neo-synthesized antibodies in the first months of life. A birth cohort of 620 newborns was established in an area endemic for malaria. Infants were followed-up until 18 months of age and their antibody responses specific for 7 P. falciparum antigens were quarterly measured. The emergence of the immune maturation process was observed in 18-months-infants. The acquisition of specific antibody responses was not impacted by placental malaria. The new methodological approach leading to distinguish maternal and neonatal antibodies was validated. The genetic characterization of the parasite antigen polymorphisms in mothers at delivery and their infants during the follow-up, in link to environmental data, led partially to the validation of the immune tolerance hypothesis.

Malariapigment Hemozoin wird üblicherweise als nicht-toxische, hochmolekulare, parasitäre Speicherform des nicht abgebauten, toxischen Häms aus dem Wirtszell-Hämoglobins betrachtet. Unaufgereinigtes Pigment, wie wir es im infizierten Erythroyzten finden und wie es nach Schizontenruptur freigesetzt wird, kann man als die "natürliche Diät" bezeichnen, die Makrophagen in Malaria-infizierten Wirten aufnehmen. Nach Aufnahme in den Makrophagen persistiert Hemozoin in den Lysosomen und wird nicht abgebaut. Das Häm-abbauende Enzym, die Häm-Oxygenase wird nicht induziert. Hemozoin ist eine potente Quelle für Radikale, woraus Lipoperoxide und davon abgeleitete Hydroxyaldehyde,wie 4-Hydroxynonenal resultieren . 4- Hydroxynonenal in Konzentrationen, wie sie in Hemozoin-beladenen Monozyten nachgewiesen wurden, hemmen die Proteinkinase C. In immunopräzipitierter Proteinkinase C aus Hemozoin- haltigen Makrophagen wurden ProteinkinaseC-Hydroxynonenal-Komplexe nachgewiesen. Die Hydroxynonenal-bedingte Hemmung der Proteinkinase C (und anderer bisher nicht untersuchter Enzyme und Prozesse) könnte die Hemozoineffekte auf den oxydativen burst und die Phagozytose erklären. Der Phorbolester-induzierte oxydative burst ist irreversibel gehemmt in Monozyten, die entweder Hemozoin oder aber Hemozoin-haltige infizierte Erythrozyten phagozytiert haben. Die Hemmung der NADPH-Oxydase, das für den oxydativen burst verantwortliche Enzym, durch intrazelluläres Hemozoin, sollte beträchtlich zur burst -Hemmung beitragen. Monozyten phagozytieren Hemozoin-haltige, infizierte Erythrozyten oder isoliertes Hemozoin , sind danach jedoch unfähig, erneut zu phagozytieren, wie es Monozyten nach Phagozytose und Verdau von nicht-infizierten Erythrozyten physiologischerweise tun. Schließlich ist die Expression von Membranantigenen, die für die Immunantwort von Bedeutung sind, in Hemozoin-haltigen Monozyten vermindert. Die Induktion des für die Präsentation externer Antigene verantwortlichen Histokompatibilitätskoplexes (MHC) Klasse II durch Interferon-gamma ist in Hemozoin-beladenen Monozyten aufgehoben. Sowohl das Interzelluläre Adhäsionsprotein 1 (CD54) als auch p150,95 Integrin (CD11c) sind in Hemozoin-haltigen Monozyten vermindert Oberflächen-exprimiert. Trotz der verschiedenen funktionellen Einschränkungen sind Hemozoin-beladene Phagozyten vital. Bei Plasmodium-falciparum-Malaria enthält ein hoher Anteil von Gewebsmakrophagen und zirkulierender Monozyten und Leukozyten große Mengen an Hemozoin. Wichtige Funktionen wie oxydativer burst , Phagozytose und die Expression von MHC Klasse II sind in Hemozoin- beladenen Phagozyten gestört. Es scheint deshalb gerechtfertigt, die Hemozoin-Beladung als wichtigen Faktor in der gestörten Immunantwort bei der P.falciparum-Malaria zu betrachten. / Malaria pigment hemozoin is generally considered to be a non-toxic, high-molecular-weight, parasitic storage form of undigested,toxic, host-hemoglobin-heme.Crude pigment, as present in infected erythrocytes and shed after schizont rupture, may be considered the 'natural diet' ingested by macrophages in malaria-infected hosts. After ingestion by macrophages hemozoin persists in the lysosomes without being degraded. The heme-degrading enzyme, the heme-oxygenase, is not induced. Hemozoin is a powerfull source of radicals that generates lipoperoxides and derived, toxic hydroxyaldehydes such as 4-hydroxynonenal. High concentrations of 4-hydroxynonenal, which have been detected in hemozoin-fed macrophages, inhibit protein kinase C. Complexes between hydroxynonenal and protein kinase C have been detected in immunoprecipitated protein kinase C from hemozoin-fed macrophages. Hydroxynonenal-mediated inhibition of protein kinase C (and of other as yet unidentified enzymes and processes) may explain hemozoin-mediated effects on oxidative burst and phagocytosis. The phorbol ester-eliceted oxidative burst is irreversibly suppressed in monocytes fed with hemozoin or hemozoin-containing, infected erythrocytes. The inhibition of NADPH-oxidase, the enzyme responsible for oxidative burst, by ingested hemozoin should considerably contribute to burst inhibition. Monocytes avidly ingest infected hemozoin-containing erythrocytes or isolated hemozoin but are unable to repeat the phagocytic cycle as monocytes do after phagocytosis and digestion of non-infected erythrocytes. Finally , the expression of membrane antigens involved in the immune response is decreased in hemozoin-loaded monocytes. The induction of the major histocompatibility complex (MHC) class II by interferon-gamma, that is responsible for presentation of external antigens, is abrogated in hemozoin-loaded monocyte. The intercellular adhesion molecule 1 (CD54) as well as the p150,95 integrin (CD11c) are decreased on the surface of monocytes containing hemozoin. Despite multiple functional impairments, hemozoin-loaded phagocytes remain alive. In Plasmodium-falciparum malaria large portions of resident macrophages and circulating monocytes and leukocytes contain massive amounts of hemozoin. Important functions like oxidative burst, phagocytosis and the expression of MHC class II are severely impaired in hemozoin-fed phagocytes. It seems therefore likly that hemozoin loading may play an important role in the impairment of the immune response seen in P.falciparum malaria.

A malária continua sendo um problema em saúde global. Nas Américas, em 2017, o Brasil e Venezuela foram os países que mais contribuíram no número de casos. No Brasil, 99% dos casos ocorreram na Amazônia Legal. Apesar dos grandes progressos do Brasil desde 2005, nos últimos anos tem se encontrado um aumento dos casos e a persistência de áreas de alta transmissão. São poucos os estudos epidemiológicos recentes que abordam este problema para a Amazônia Legal. Existem associações entre a transmissão da malária e fatores de risco ambientais e socioeconômicos, mas, existe uma falta de consenso nestes fatores, influenciados pela escala de análise. Objetivo: Caracterizar e analisar os fatores de risco da transmissão da malária nos municípios da Amazônia Legal entre 2010 e 2015. Métodos: Foram utilizados os dados de malária autóctone por município entre 2010 e 2015 da base de dados SIVEP-malária da Secretaria de Vigilância em Saúde. Os fatores de risco analisados foram: índice Gini, taxa de analfabetismo, presença de minas, área de reservas indígenas no município, taxa de floresta no município e duração da estação seca. Para a seleção dos fatores de risco ambientais foi realizada uma revisão sistematizada e os fatores socioeconômicos foram embasados na literatura. A unidade espacial foram os 310 municípios dos estados do Acre, Amazonas, Amapá, Pará, Rondônia e Roraima. Os casos de malária foram analisados mediante o Índice Parasitário Anual (IPA) e a incidência mensal através da estratificação local dos dados. Para os fatores de risco uma regressão logística foi executada para um modelo ambiental, socioeconômico e completo, sendo o IPA a variável dependente e estratificando o risco de transmissão em baixo, médio e alto. Resultados: No período 2010 - 2015 houve redução de 61% no IPA, cumprindo a meta proposta pelo Plano Nacional de Controle e Prevenção da Malária no Brasil. No entanto, essa redução não aconteceu de igual forma em todos os municípios, e um conjunto de 13 municípios apresentam um mínimo de 40% dos casos ao longo dos 5 anos. É relevante destacar que tem estados que reduziram o efeito da sazonalidade enquanto outros não. A revisão sistematizada deixou claro que existem divergências na seleção dos fatores de risco ambientais e da influência na transmissão dependendo do espaço tempo e da escala. A análise dos fatores de risco apresentou que os modelos que incluíam fatores ambientais e socioeconômicos tinham um melhor desempenho ao longo dos anos e nos estratos de transmissão. Positivamente o índice Gini e negativamente a duração da estação seca foram os fatores de risco mais importantes para a transmissão. Conclusão: A malária é multifatorial e deve ser abordada tendo em conta o espaço tempo e a escala de atuação, para implementar intervenções eficientemente. As inequidades na população se apresentam como o grande empecilho para obter melhores resultados no seu controle e eliminação, e embora o Brasil tinha feito progressos muito meritórios, não conseguirá acabar com o problema da malária enquanto não aborda-lo como um problema à saúde global que vai além de programas específicos contra a doença. / Introduction: Malaria is still a global health problem. In the Americas in 2017, Brazil and Venezuela were the countries that most contributed to the number of cases. In Brazil, 99% of the cases are in the Legal Amazon. In the last years, there has been an increase of cases and areas of high transmission despite the significant progress in Brazil since 2005. Few recent epidemiological studies approached this issue for the Legal Amazon. There is a relationship between malaria transmission and environmental and socioeconomic risk factors. However, there is a lack of consensus on the influence of them due to the scale of analysis. Objective: Characterize and analyze the malaria transmission risk factors in the Legal Amazon municipalities from 2010 to 2015. Methodology: We used data of autochthonous malaria by the municipality from 2010 to 2015, extracted from the SIVEP-malária database from the Bureau of Health Surveillance. We analyzed the following risk factors: Gini index, illiteracy rate, mines presence, areas of indigenous reserve by municipality, forest rate by municipality and length of dry season. A systematized literature review was performed to select the environmental risk factors. Socioeconomic risk factors were selected based on the literature. The 310 municipalities in the states of Acre, Amazonas, Amapá, Pará, Rondônia and Roraima were our unit of analysis. Local stratification of Annual Parasite Index (API) and monthly incidence were used to analyze the data. Logistic regression was used to perform an environmental, socioeconomic and full model, stratifying by low, moderate and high-risk transmission; the dependent variable API. Results: From 2010 to 2015, there was a 61% API reduction, achieving the Brazilian National Malaria Control and Prevention Plan goal. However, the decline was heterogeneous among municipalities. There are 13 municipalities that at least had the 40% of all cases during the 5 years period. It is relevant to note that some states minimized the seasonality while others do not. The systematized review made clear the divergences to select the risk factors and the influence of them into the transmission due to the space-time and the scale. The full model showed a better performance to analyze the risk factors by strata and transmission intensity. Gini index positively and length of dry season negatively were the most important risk factors. Conclusion: Malaria is a multifactorial disease that has to be approached taking into account the space-time and the scale of analysis to implement effective interventions. Population inequities are the biggest obstacle to obtain better results in malaria control and elimination. Brazil advanced significantly but to end with the problem malaria has to be understood as a global health problem that goes beyond specific disease programs.

A malária continua sendo um problema em saúde global. Nas Américas, em 2017, o Brasil e Venezuela foram os países que mais contribuíram no número de casos. No Brasil, 99% dos casos ocorreram na Amazônia Legal. Apesar dos grandes progressos do Brasil desde 2005, nos últimos anos tem se encontrado um aumento dos casos e a persistência de áreas de alta transmissão. São poucos os estudos epidemiológicos recentes que abordam este problema para a Amazônia Legal. Existem associações entre a transmissão da malária e fatores de risco ambientais e socioeconômicos, mas, existe uma falta de consenso nestes fatores, influenciados pela escala de análise. Objetivo: Caracterizar e analisar os fatores de risco da transmissão da malária nos municípios da Amazônia Legal entre 2010 e 2015. Métodos: Foram utilizados os dados de malária autóctone por município entre 2010 e 2015 da base de dados SIVEP-malária da Secretaria de Vigilância em Saúde. Os fatores de risco analisados foram: índice Gini, taxa de analfabetismo, presença de minas, área de reservas indígenas no município, taxa de floresta no município e duração da estação seca. Para a seleção dos fatores de risco ambientais foi realizada uma revisão sistematizada e os fatores socioeconômicos foram embasados na literatura. A unidade espacial foram os 310 municípios dos estados do Acre, Amazonas, Amapá, Pará, Rondônia e Roraima. Os casos de malária foram analisados mediante o Índice Parasitário Anual (IPA) e a incidência mensal através da estratificação local dos dados. Para os fatores de risco uma regressão logística foi executada para um modelo ambiental, socioeconômico e completo, sendo o IPA a variável dependente e estratificando o risco de transmissão em baixo, médio e alto. Resultados: No período 2010 - 2015 houve redução de 61% no IPA, cumprindo a meta proposta pelo Plano Nacional de Controle e Prevenção da Malária no Brasil. No entanto, essa redução não aconteceu de igual forma em todos os municípios, e um conjunto de 13 municípios apresentam um mínimo de 40% dos casos ao longo dos 5 anos. É relevante destacar que tem estados que reduziram o efeito da sazonalidade enquanto outros não. A revisão sistematizada deixou claro que existem divergências na seleção dos fatores de risco ambientais e da influência na transmissão dependendo do espaço tempo e da escala. A análise dos fatores de risco apresentou que os modelos que incluíam fatores ambientais e socioeconômicos tinham um melhor desempenho ao longo dos anos e nos estratos de transmissão. Positivamente o índice Gini e negativamente a duração da estação seca foram os fatores de risco mais importantes para a transmissão. Conclusão: A malária é multifatorial e deve ser abordada tendo em conta o espaço tempo e a escala de atuação, para implementar intervenções eficientemente. As inequidades na população se apresentam como o grande empecilho para obter melhores resultados no seu controle e eliminação, e embora o Brasil tinha feito progressos muito meritórios, não conseguirá acabar com o problema da malária enquanto não aborda-lo como um problema à saúde global que vai além de programas específicos contra a doença. / Introduction: Malaria is still a global health problem. In the Americas in 2017, Brazil and Venezuela were the countries that most contributed to the number of cases. In Brazil, 99% of the cases are in the Legal Amazon. In the last years, there has been an increase of cases and areas of high transmission despite the significant progress in Brazil since 2005. Few recent epidemiological studies approached this issue for the Legal Amazon. There is a relationship between malaria transmission and environmental and socioeconomic risk factors. However, there is a lack of consensus on the influence of them due to the scale of analysis. Objective: Characterize and analyze the malaria transmission risk factors in the Legal Amazon municipalities from 2010 to 2015. Methodology: We used data of autochthonous malaria by the municipality from 2010 to 2015, extracted from the SIVEP-malária database from the Bureau of Health Surveillance. We analyzed the following risk factors: Gini index, illiteracy rate, mines presence, areas of indigenous reserve by municipality, forest rate by municipality and length of dry season. A systematized literature review was performed to select the environmental risk factors. Socioeconomic risk factors were selected based on the literature. The 310 municipalities in the states of Acre, Amazonas, Amapá, Pará, Rondônia and Roraima were our unit of analysis. Local stratification of Annual Parasite Index (API) and monthly incidence were used to analyze the data. Logistic regression was used to perform an environmental, socioeconomic and full model, stratifying by low, moderate and high-risk transmission; the dependent variable API. Results: From 2010 to 2015, there was a 61% API reduction, achieving the Brazilian National Malaria Control and Prevention Plan goal. However, the decline was heterogeneous among municipalities. There are 13 municipalities that at least had the 40% of all cases during the 5 years period. It is relevant to note that some states minimized the seasonality while others do not. The systematized review made clear the divergences to select the risk factors and the influence of them into the transmission due to the space-time and the scale. The full model showed a better performance to analyze the risk factors by strata and transmission intensity. Gini index positively and length of dry season negatively were the most important risk factors. Conclusion: Malaria is a multifactorial disease that has to be approached taking into account the space-time and the scale of analysis to implement effective interventions. Population inequities are the biggest obstacle to obtain better results in malaria control and elimination. Brazil advanced significantly but to end with the problem malaria has to be understood as a global health problem that goes beyond specific disease programs.

Os genes var de Plasmodium falciparum codificam a proteína PfEMP1 expressa na superfície de eritrócitos infectados e que medeia os fenômenos de citoaderência e \"rosetting\". Ambos os fenômenos estão diretamente associados à malária grave, e seu domínio mais N-terminal, DBL1alfa, media especificamente \"rosetting\". Análise de seqüências DBL1alfa de isolados brasileiros e de outros países revelou que a similaridade entre elas não pode predizer origem geográfica. Com o objetivo de determinar se existem seqüências DBL1alfa associadas à malária grave, analisamos as seqüências DBL1alfa expressas em parasitas obtidos de pacientes brasileiros com esta manifestação clínica e encontramos que as seqüências predominantemente expressas apresentavam uma ou duas deleções de cisteínas. Significativamente, apesar de freqüentes no genoma de parasitas de pacientes com malária não grave, essas seqüências foram raramente expressas. Esses dados demonstram a primeira associação de seqüências PfEMP1 expressas e malária grave em pacientes da Amazônia Brasileira. / Plasmodium falciparum var genes code for PfEMP1, a protein expressed on the surface of infected erythrocytes, and which mediates cytoadherence and rosetting. Both phenomena are directly associated with severe malaria and the most N-terminal domain, DBL1alfa, specifically mediates rosetting. DBL1alfa sequence analysis from Brazilian and worldwide isolates revealed that sequence similarities cannot predict geographical origin. To determine whether there are DBL1alfa sequences associated with severe malaria, we examined expressed var DBL1alfa sequences in patients with severe malaria from the Brazilian Amazon and found that the predominantly expressed DBL1alfa sequences from these parasites lacked 1-2 cysteine residues. Significantly, these sequences were amply found on the genomic repertoire of parasites from patients with mild malaria and yet they were rarely expressed. These data demonstrate the first association of particular PfEMP1 expressed sequences and severe malaria in patients from the Brazilian Amazon.