Vector borne-diseases make up a significant portion of morbidity and mortality worldwide, being responsible for around 700,000 deaths annually according to the World Health Organization. Neglected, tropical diseases such as Chagas disease have a significant impact on people in Latin America, affecting millions, and especially those residing in rural areas. Chagas disease is the number one cause for heart disease in Latin America, and is caused by the Trypanosoma cruzi parasite, carried by Triatominae insect vectors. The intricate life cycle of the parasite, ecology and behavior of the vector, and lack of disease treatment options, make Chagas disease challenging to control. Prevention measures are highly sought after, and implementation science approaches such as Ecohealth management engage affected communities in disease prevention. Knowing what insect vectors are feeding on sheds light on vector ecology and behavior, aiding in vector management which is pivotal in disease prevention. While DNA-based methods have traditionally been used to study vector blood meals, they come with limitations and challenges, such as the need for fresh, high abundance blood meals. Therefore, the goal of this research was to evaluate Chagas vector blood meal sources using an innovative protein mass spectrometry-based approach. We demonstrate first the ability to utilize liquid chromatography tandem mass spectrometry (LC-MS/MS) to correctly identify hemoglobin protein peptides from mouse blood and subsequently identify Chagas vector blood meal sources from field-collected insect vectors where blood meal identification is compared with traditional DNA-based methods as a control. An experimental feeding study allowed us to then demonstrate the longevity of hemoglobin protein peptides for blood meal detection, showing LC-MS/MS-based blood meal identification outperforms DNA-based polymerase chain reaction (PCR) at least 4 weeks post-feeding and 12 weeks post-molting. This allowed us to test the limits of our innovative detection method experimentally and comparatively. Finally, we evaluated blood meals in field-caught insect vectors collected as part of a large collaborative Ecohealth project in Central America. LC-MS/MS identified two times as many blood meals in insect vectors, including those that did not have blood meals detected with DNA-based PCR. As single vectors often feed on multiple sources, we also validated our ability to decipher multiple blood meals from an individual vector and showed the ability to quantify a blood meal using synthetic AQUA (Absolute QUAntification) peptides, a first step in using quantification data for identifying blood meals not currently in our underlying database. Furthermore, we show that lower resolution mass spectrometers are able to identify blood meals from taxa correctly, an important and strong attribute of our LC-MS/MS-based method, opening the door to using proteomics in countries where Chagas disease is endemic and resources are limited. Even though expertise and resources of research labs differ in locations across the globe, herein is described how LC-MS/MS is a valuable additional tool for fighting neglected tropical diseases. Ultimately, hemoglobin-based LC-MS/MS vector blood meal identification is a complementary technique to available molecular methods and can confidently identify Chagas vector blood meal sources to aid in understanding vector biology and ecology, and aid in developing appropriate Ecohealth vector control measures.

Trypanosoma cruzi is endemic in the U.S., infecting humans, dogs, and wildlife. This study identified a new geographic distribution for triatomine species within Texas based on 2,449 records obtained from published data and new field studies. Triatomine vectors of T. cruzi were reported from 97 counties covering all ecological zones. Triatoma gerstaeckeri was the most commonly collected species followed by T. sanguisuga. New field collections resulted in 233 specimens from 37 counties and a 52% T. cruzi infection rate. A second trypanosome, Blastocrithida triatomae was found in two specimens from different locations. A habitat suitability model for T. gerstaeckeri was developed using GIS and remote sensing applications. Forest and rangeland were the predominant land cover classes found within T. gerstaeckeri habitat, where as water and agriculture proved to have little influence on habitat suitability. Genetic variation of seven triatomine species from Texas was evaluated using cytochrome b DNA sequences from 61 new specimens. This is the first study of the taxonomic status of T. gerstaeckeri, T. indictiva, and T. neotomae using molecular markers. Intraspecific variation for T. sanguisuga and T. gerstaeckeri suggests significant gene flow across their ranges within Texas. Genetic variation of T. cruzi isolates from Texas was evaluated using SSU rRNA gene sequences. Included were 63 new sequences from five triatomine species, canine, baboon, and human isolates. Sequences partitioned into two groups in agreement with previous studies on U.S. isolates. Genetic variation of T. cruzi did not occur according to host, geographic location, or collection site. The extent of Chagas disease in domestic canines of Texas is described by geographic distribution, signalment, and clinical presentation and histopathology. Based on data from 553 cases, the geographic distribution in Texas is widespread (46 counties) and closely matches the distribution of the Triatomine vectors. Chagas disease was diagnosed in 33 breeds, primarily sporting/working dogs. This study represents the most comprehensive characterization of components of the Chagas disease transmission cycle in the U.S. to date. These findings should raise awareness among physicians, veterinarians, and public health practitioners regarding T. cruzi, its vectors, canine infection, and human risk for Chagas disease in Texas.

Thesis (Ph.D.)--Baylor University, 2008. / Includes bibliographical references (p. 361-367).

Thesis (Ph. D. in Chemistry)--Vanderbilt University, Dec. 2008. / Title from title screen. Includes bibliographical references.

Thesis (Ph. D.)--University of Texas at El Paso, 2008. / Title from title screen. Vita. CD-ROM. Includes bibliographical references. Also available online.

Thesis (Ph.D.)--Baylor University, 2008. / Includes bibliographical references (p. 296-303).

BACKGROUND: Although Chagas disease is not an endemic health concern in the United States, it is prevalent with estimates of approximately 300,000 infected individuals. Of this, an estimated 20 – 30% will develop severe, life-threatening consequences. While what is known about Chagas disease is extensive, there is limited knowledge of this disease, especially among medical providers and clinicians in the United States. In order to bridge the gap between what is known about Chagas disease and those who lack this knowledge, adequate and effective education interventions must be developed and delivered. Education that is tailored for medical providers and clinicians most likely to encounter individuals at greastest risk for having Chagas disease is essential. Once this knowledge is gained, accurate evaluation, diagnosis, and treatment of indivuals with Chagas disease may be pursued, ultimately decreasing and preventing disease-associated morbidity and mortality. PROPOSAL: Clinicians from six departments (infectious disease, cardiology, internal medicine, family medicine, pediatrics, and obstretrics and gynecology) will be recruited from Boston Medical Center where they will be given an educational module about Chagas disease. Their knowledge of Chagas disease and how to clinically apply it will be assessed prior to the educational module, immediately following the educational module and 1-month and 6-months following the educational module. CONCLUSION: Chagas disease is a burden to health systems in many countries worldwide including the United States, and awareness of Chagas disease is lacking among medical personnel of multiple specialities in the United States. Educational interventions have provided knowledge of various diseases leading to protocol development, ultimately influencing clinical practice to a degree that reduces morbidity and mortality. The same is needed with respect to Chagas disease. The goal of this educational intervention is to provide a knowledge base through teaching and resources for clinicians to learn, understand, and review the steps needed to clinically evaluate, diagnose, and treat patients with Chagas disease. Using the reseach identified in this study as well as the proposed educational intervention, it is hoped that this disease burden can be alleviated.

Made available in DSpace on 2014-06-11T19:33:24Z (GMT). No. of bitstreams: 0 Previous issue date: 2005-05-24Bitstream added on 2014-06-13T18:07:30Z : No. of bitstreams: 1 silva_ma_dr_arafcf.pdf: 976242 bytes, checksum: 452382843f6dbbd98e65fdd05a72d135 (MD5) / Universidade Estadual Paulista (UNESP) / Trypanosoma cruzi é um protozoário flagelado causador da doença de Chagas que, segundo estimativas da OMS afeta 16-18 milhões de pessoas na América Latina, causando danos sociais extremamente graves. A doença apresenta formas clínicas variadas que incluem a forma indeterminada, cardiomiopatia e alterações digestivas. Variações intraespecíficas nas diferentes cepas de Trypanosoma cruzi estudadas foram demonstradas em nível morfo-biológico, bioquímico e genético. Essa heterogeneidade poderia explicar a variabilidade nas manifestações clínicas da doença de Chagas e as diferenças regionais de sua morbidade. Com o objetivo de caracterizar quatro cepas de Trypanosoma cruzi isoladas de pacientes chagásicos crônicos, residentes na região de Araraquara - SP, parâmetros biológicos e moleculares foram avaliados. Para estudar o comportamento biológico das cepas, três grupos de camundongos Swiss, não isogênicos, pesando 10-12g foram infectados com formas sangüíneas das cepas em estudo. Foram avaliados os seguintes aspectos: período pré-patente, curvas de parasitemia, morfologia do parasita no sangue periférico, taxas de mortalidade e lesões histopatológicas. Três cepas apresentaram parasitemia patente com períodos pré-patentes variáveis, baixa parasitemia e formas tripomastigotas largas durante todo o curso da infecção. Uma cepa apresentou parasitemia sub-patente. Nenhum animal morreu durante o período do estudo. Exame histopatológico mostrou ninhos de formas amastigotas no coração de animais infectados para duas cepas e inflamação em vários órgãos para todas as cepas. Para a caracterização molecular o DNA do parasita foi extraído de formas epimastigotas mantidas em meio LIT. Parte do espaçador não transcrito do gene de mini-exon foi amplificado por PCR. Todas as cepas geraram produtos com 250 pb. Os dados biológicos e moleculares permitiram a classificação de todas as cepas como T.cruzi II. / Trypanosoma cruzi is a flagellate protozoan agent of Chagas disease that, according to the OMS estimation, affect 16-18 milion of people in Latin America, making serious social damage. The illness shows varied clinicals forms that include indeterminate form, cardiomiopathy and digestive alterations. Intra-specific variations among differents Trypanosoma cruzi strains studied have been demonstrated on morpho-biological, genetic and biochemical levels. Such heterogeneity can explain the variability of Chagas disease in clinical manifestations and in the regional differences of the morbidity rate. This research has the objective to characterize four strains of Trypanosoma cruzi isolated from humans patients, residents in Araraquara - SP region, by evaluating biological and molecular parameters. In order to study the biological behavior of Trypanosoma cruzi strains, three groups of albino Swiss mice, weighting 10-12g were intraperitoneally inoculated with bloodstreams forms of the strains in study. Pre-patent period, parasitaemia patterns, tripomastigote morphology, mortality rate and tissue distribution of the protozoan were analysed. Three strains showed patent parasitemy with variable pre-patent period, low parasitemy and broad forms during the whole infection period. One strain showed sub-patent parasitemy. Mortality rates were null. Histopathological analysis showed amastigotes forms in heart for two strains and inflamatory process in several organs for all the strains. For the molecular characterization, the parasite DNA was extracted from epimastigotes forms kept in LIT medium. Part of the non-transcribed spacer of the mini-exon gene was amplified by PCR. All the strains generated products with 250 pb. The molecular and biological informations allowed to classify all the strains as T.cruzi II.

Orientador: Chung Man Chin / Banca: Maria do Carmo Longo / Banca: Maria Luiz Cruz / Resumo: A doença de Chagas representa um dos mais sérios problemas médico-sanitários da América Latina. Nos 21 países endêmicos, estima-se que 18 a 20 milhões de pessoas estejam infectadas e cem milhões estão sob risco. No Brasil apenas o benznidazol está disponível para o tratamento apresentando alta incidência de efeitos adversos e ineficácia na fase crônica da doença. O desenvolvimento de fármacos mais eficazes e com menor toxicidade contra a doença de Chagas é necessário, e para tal é importante o conhecimento dos alvos terapêuticos para o planejamento racional por novos compostos. Estudos demostraram que a tripanotiona redutase (TR) é a enzima chave do metabolismo anti-oxidativo do T. cruzi. Os nitrofuranos atuam como substratos da TR e são também efetivos inibidores enzimáticos da redução do tripanotiona dissulfeto. Entre os compostos nitrofurânicos, o nitrofural apresentou atividade contra T. cruzi, mas alta toxicidade por via oral e severos efeitos adversos. A latenciação de fármacos é um dos métodos de modificação molecular que tem como objetivo melhorar as características de fármacos protótipos. Sabendo disto, Chung (1996) sintetizou uma série de pró-fármacos recíprocos seletivos de nitrofural e primaquina, culminando na obtenção do derivado hidroximetilnitrofural (NFOH). Este se mostrou mais ativo de todos os derivados sintetizados, tanto em formas amastigotas quanto em tripomastigotas nos ensaios in vitro contra T. cruzi. Os compostos que possuem grupos nitro na sua molécula geralmente apresentam atividade mutagênica, estudos indicaram que a latenciação do nitrofural em NFOH foi capaz de diminuir a atividade mutagênica em aproximadamente 4 vezes. Diante do exposto, o objetivo deste trabalho foi iniciar os estudos de toxicidade do derivado... (Resumo completo, clicar acesso eletrônico abaixo). / Abstract: The Chagas' disease represents one of the most serious sanitary medical problems of Latin America. In the 21 endemic countries, it is estimated that 18 to 20 million people are infected and 100 million are under risk. Only one drug is available in Brazilian market: benznidazole, which shows high side effects and is ineffective in the chronic phase of the disease. The development of more efficient drugs with lower toxicity against the Chagas disease is necessary and for it, it is important the knowledge of therapeutic targets for the design of new compounds. Studies showed that trypanothione reductase (TR) is a key enzyme of the antioxidant metabolism of T. cruzi. Nitrofurans act as substrates of TR and are also effective enzymatic inhibitors of the reduction of trypanothione disulfide. Among the nitrofurans conpounds, nitrofurazone presented activity against T cruzi, but showed high toxicity by oral route and severe side effects. Latentiation is one process of molecular modification, which has the purpose to improve pharmaceutical and physical-chemical properties and decrease drug toxicity. In this way, Chung (1996) has synthesized the prodrug hydroxymethylnitrofurazone (NFOH), showing high activity in amastigotes forms in vitro assay against T. cruzi. Nitrocompounds generally presents mutagenic activitys. Studies suggested that the latentiantion of nitrofurazone giving NFOH was able to reduce toxicity activity in four times. According to this, the purpose of this work was to accomplish toxicity studies of the derivative compound. The results obtained in the tests acute toxicity in mice and rate suggested a lower toxicity of NFOH, obtaining a median lethal dose in relation to NF...(Complete abstract, click electronic address below). / Mestre

FundaÃÃo Cearense de Apoio ao Desenvolvimento Cientifico e TecnolÃgico / Chagas disease is one of the most important parasitic diseases in Latin America, transmitted by triatomine, has reached more than 10 million people. This illness is typical of rural environments and inadequate housing provides shelter to the vector the disease transmitter. The aim of this study was to investigate the presence of Trypanosoma cruzi in triatomines captured in intradomiciliary and surroundings in cities in the southeast region of CearÃin. This is a retrospective descriptive study, by consulting the archives of Control Program of Chagas Disease in Regional Laboratory of Endemic Diseases of Limoeiro do Norte â CE, which attends the cities that were studied from 2009 to 2011. During the study were captured a total of 18.408 specimens of insects, including nymphs and adults inside the home and outside homes. Of this total, 17.910 specimens were examined, representing 97,29%. The number of nymphs was taken about three times greater than the number of adults. The year of 2010 was the one with the highest number of captures, with a total of 8.548 triatomines, distributed among nymphs (6.115) and adults (2.433), and 637 inside of the houses and 7.911 around the houses. This year presented an infection rate of 1.30%, with 107 positive triatomines, being Quixerà the city with the highest infection rate in that year. The infection rate in adults triatomine (1.92%) was higher than in nymphs (1.21%). The species captured during the study period were Triatoma pseudomaculta, Triatoma brasiliensis, Panstrongylus megistus, Panstrongylus lutzi and Rhodnius nasutus. Of these species, T. pseudomaculta was the most captured throughout the study period, with 12.643 specimes. During this study, were 11 cities studied, and the most infested was Tabuleiro do Norte with 3.976 specimens, followed by Ãrere with 3.289. The city with the highest infection rate during the study period was Limoeiro do Norte (5,00%) with a total of 125 triatomines positive, followed by Quixerà (2.39%). At the end of the study, we can conclude that it is still necessary to intensify Control Program of Chagas disease, in order to avoid the maximum transmission of this disease. Finally we highlight the importance of conducting educational programs to the population in order to provide guidance to the public on disease prevention, such as towing houses, fix cracks and maintain clean environments at home and around the homes to prevent colonization of the approach and vectors. / A doenÃa de Chagas à uma das doenÃas parasitÃrias mais importante da AmÃrica Latina, transmitida por triatomÃneos jà atingiu mais de 10 milhÃes de pessoas. Essa enfermidade à tÃpica de ambientes rurais e habitaÃÃes inadequadas que oferecem abrigo ao vector transmissor da doenÃa. O presente estudo tem como objetivo investigar a presenÃa de Trypanosoma cruzi em triatomÃneos capturados nos intra e peridomicÃlios, em municÃpios da regiÃo sudestedo estado do CearÃ. Trata-se de um estudo retrospectivo descritivo, de consulta aos arquivos do Programa de Controle da DoenÃa de Chagas do LaboratÃrio Regional de Endemias do Limoeiro do Norte â CE, que atende os municÃpios que foram estudados no perÃodo de 2009 a 2011. Durante o estudo foi capturado um total de 18.408 exemplares de triatomÃneos, entre ninfas e adultos no intradomicÃlio e peridomicÃlio. Desse total, 17.910 exemplares foram examinados, representando 97,29 % dos capturados. O nÃmero de ninfas capturadas foi cerca de trÃs vezes maior que o nÃmero de adultos. O ano de 2010 foi o ano com maior nÃmero de capturas, com um total de 8.548 triatomÃneos capturados, distribuÃdos entre ninfas (6.115) e adultos (2.433), sendo 637 no intradomicÃlio e 7.911 no peridomicÃlio. O Ãndice de infecÃÃo nesse ano foi de 1,30%, com 107 triatomÃneos positivos, sendo Quixerà o municÃpio com maior Ãndice de infecÃÃo. O Ãndice de infecÃÃo em adultos (1,92%) foi maior do que em ninfas (1,21%). As espÃcies capturadas durante o perÃodo de estudo foram Triatoma pseudomaculta, Triatoma brasiliensis, Panstrongylus megistus, Panstrongylus lutzi e Rhodnius nasutus. Dessas espÃcies, T. pseudomaculta foi a mais capturada durante todo o perÃodo de estudo, com 12.643 exemplares. Durante o perÃodo de estudo foram estudados 11 municÃpios, sendo o mais infestado Tabuleiro do Norte com 3.976 exemplares, seguido de Ãrere com 3.289 exemplares. O municÃpio que apresentou maior Ãndice de infecÃÃo durante o perÃodo de estudo foi Limoeiro do Norte (5,00%) com um total de 125 triatomÃneos positivos, seguido de Quixerà (2,39%). Ao fim do estudo, pode-se concluir que ainda faz-se necessÃria a intensificaÃÃo do Programa de Controle da DoenÃa de Chagas, para poder evitar ao mÃximo a transmissÃo da doenÃa. Finalmente destaca-se a importÃncia de realizar programas educativos à populaÃÃo com a finalidade de dar orientaÃÃes à populaÃÃo na prevenÃÃo da doenÃa, tais como rebocar as casas, corrigir frestas e manter limpos os ambientes no peridomicÃlio e no domicÃlio para evitar a aproximaÃÃo e colonizaÃÃo dos vetores.