Detecção de DNA de Leishmania spp. no sangue e em esperma de indivíduos residentes na região Amazônica

Dissertação (mestrado)—Universidade de Brasília, Faculdade de Medicina, Pós-Graduação em Ciências Médicas, Laboratório Multidisciplinar de Pesquisa em Doença de Chagas, 2014A leishmaniose é uma zoonose transmitida por flebotomíneos e causada por várias espécies do gênero Leishmania. Na região Amazônica, em particular no estado do Pará, a doença de Chagas e as leishmanioses compartilham área de transmissão, o que leva frequentemente à presença de infecções mistas. A ocorrência dessas foi verificada em estudo prévio em população de chagásicos da Amazônia, identificando a presença de anticorpos anti-Leishmania spp. em 50% das amostras. No sêmen dos mesmos indivíduos, também foi detectado DNA do Trypanosoma cruzi, sugerindo a possibilidade de transmissão sexual. Com poucos relatos sobre a transmissão sexual das infecções causadas por tripanossomatídeos, a real importância epidemiológica dessa via de transmissão permanece desconhecida. Assim, o este estudo avaliou a presença de infecções mistas causadas por Leishmania spp. e T. cruzi, mediante a análise dos resultados obtidos no diagnóstico sorológico por ELISA e molecular por PCR ITS1, em uma população residente no estado do Pará. Além disso, foi realizada a pesquisa de DNA de Leishmania spp. em amostras de esperma dessa população. Os resultados sorológicos revelaram que 50/108 (46,29%) dos indivíduos apresentavam anticorpos específicos contra antígenos de Leishmania spp., enquanto que a reação de PCR foi positiva em somente 20/108 (18,51%) dos indivíduos testados, evidenciando uma clara discrepância de 27,78% entre os métodos de diagnóstico. O teste de PCR demonstrou-se mais eficiente para identificação dos indivíduos com infecção mista, já que o teste sorológico apresentou um elevado número de reações cruzadas. O método molecular também foi capaz de detectar DNA nuclear do parasito no esperma de 53,1% dos indivíduos. O sequenciamento dos produtos amplificados permitiu identificar as espécies L. amazonensis e L. infantum nas amostras, demonstrando que a possibilidade de transmissão sexual das leishmanioses necessita de estudos mais aprofundados. Nossos resultados reafirmam a necessidade de aprimorar as técnicas de diagnóstico utilizadas na rotina laboratorial e aponta a técnica de PCR como ferramenta promissora na identificação das infecções causadas por Leishmania spp. A pesquisa realizada apresenta informações relevantes sobre diagnóstico e vias de transmissão das leishmanioses na região Amazônica, que poderá favorecer o aprimoramento de medidas de controle, prevenção e assistência à população. __________________________________________________________________________ ABSTRACTLeishmanias is is a zoonosis transmitted by sandflies and caused by various species of the genus Leishmania. In Amazon, particularly at Pará State, Chagas disease and leishmaniasis share a common transmission area, which often leads to mixed infections. This scenario was observed in a previous study in a chagasic population of Amazon, which demonstrated the presence of anti-Leishmania spp. antibodies in almost 50% of the samples. Furthermore, itwas detected Trypanosoma cruzi DNA in semen from infected people, suggesting the possibility of sexual transmission of the parasite. There are few reports regarding the sexual transmission of infections caused by trypanosomatids and the actual epidemiologic alimportance of this transmission route remains unknown. Thus, the present study aimed to evaluate the presence of mixed infections caused by Leishmania spp. and T. cruzi byserological ELISA and molecular diagnosis PCR LITSR, in an endemic population at ParáState. Moreover, Leishmania spp. DNA search was carried out in sperm samples of this population. The results of serological tests revealed that 50/108 (46,29%) individuals hadspecific antibodies against Leishmania spp. antigens, whereas PCR was positive in only20/108 (18,51%) of tested samples, showing a clear discrepancy 27,78% between these methods. The study showed PCR was more efficient to identify individuals with mixedinfection, since serological assay showed a large number of cross-reactions. Molecularmethod was able to detect parasite nuclear DNA in semen from 53.1% individuals.Amplicons sequencing allowed us to identify the species L. amazonensis and L. infantum insamples, demonstrating the possibility of leishmaniasis sexual transmission needs to befurther investigated. Our results reaffirm the requirement of improve diagnostic techniquescurrently used in routine, and PCR arises as a promising tool for identification of infectionscaused by Leishmania spp. The study presented relevant information about diagnosis andleishmaniasis transmission routes at the Amazon region, which may favor the improvement of control measures, prevention and population care

The role of gallery forests in maintaining Phlebotominae populations: potential Leishmania spp. vectors in the Brazilian savanna

Background: knowledge on synanthropic phlebotomines and their natural infection by Leishmania is necessary for the identification of potential areas for leishmaniasis occurrence. Objective: to analyse the occurrence of Phlebotominae in gallery forests and household units (HUs) in the city of Palmas and to determine the rate of natural infection by trypanosomatids. Methods: gallery forests and adjacent household areas were sampled on July (dry season) and November (rainy season) in 2014. The total sampling effort was 960 HP light traps and eight Shannon traps. Trypanosomatids were detected in Phlebotominae females through the amplification of the SSU rDNA region, and the positive samples were used in ITS1-PCR. Trypanosomatid species were identified using sequencing. Findings: a total of 1,527 sand flies representing 30 species were captured in which 949 (28 spp.) and 578 (22 spp.) were registered in July and November, respectively. In July, more specimens were captured in the gallery forests than in the HUs, and Nyssomyia whitmani was particularly frequent. In November, most of the specimens were found in the HUs, and again, Ny. whitmani was the predominant species. Lutzomyia longipalpis was commonly found in domestic areas, while Bichromomyia flaviscutellata was most frequent in gallery forests. Molecular analysis of 154 pools of females (752 specimens) identified Leishmania amazonensis, L. infantum, and Crithidia fasciculata in Ny. whitmani, as well as L. amazonensis in Lu. longipalpis, Trypanosoma sp. and L. amazonensis in Pintomyia christenseni, and L. amazonensis in both Psathyromyia hermanlenti and Evandromyia walkeri. Main conclusions: these results show the importance of gallery forests in maintaining Phlebotominae populations in the dry month, as well as their frequent occurrence in household units in the rainy month. This is the first study to identify Leishmania, Trypanosoma, and Crithidia species in Phlebotominae collected in Palmas, Tocantins, Brazil

Congenital Chagas disease: alert of research negligence

Abstract INTRODUCTION: Chagas disease (CD), a neglected endemic disease in Latin America, has acquired new epidemiological characteristics with an increase in the importance of alternative transmission routes such as congenital transmission. We evaluated the scientific research on this subject. METHODS: We searched the Scielo, BVS, and PubMed databases from 2006 to 2017. RESULTS: We identified a small number of published articles, mostly in journals with an impact factor less than 3.0. Studies on human congenital transmission of CD were carried out in only seven different countries. CONCLUSIONS: Our data highlight the lack of research on congenital CD

Toxocara spp. seroprevalence in pregnant women in Brasília, Brazil

Abstract INTRODUCTION: The impact of gestational toxocariasis is an understudied topic on female reproductive health. We estimated anti-Toxocara IgG prevalence among pregnant women in Brasília, Brazil, and investigated the association of the infection with history of abortion and contact with pets. METHODS: Infection was diagnosed using ELISA with excretory/secretory antigens. Participant information was obtained via questionnaires. RESULTS: Of 311 pregnant women, 23 were anti-Toxocara IgG positive. Twenty-two percent of anti-Toxocara IgG-positive participants and 26% had previously miscarried. Previous contact with pets was associated with higher toxocariasis prevalence. CONCLUSIONS: A direct relationship between toxocariasis and contact with pets was observed, but there was no relationship with the miscarriage prevalence

Congenital Chagas disease: alert of research negligence

INTRODUCTION: Chagas disease (CD), a neglected endemic disease in Latin America, has acquired new epidemiological characteristics with an increase in the importance of alternative transmission routes such as congenital transmission. We evaluated the scientific research on this subject. METHODS: We searched the Scielo, BVS, and PubMed databases from 2006 to 2017. RESULTS: We identified a small number of published articles, mostly in journals with an impact factor less than 3.0. Studies on human congenital transmission of CD were carried out in only seven different countries. CONCLUSIONS: Our data highlight the lack of research on congenital CD.Faculdade de Medicina (FMD