Expression of Fluorescent Genes in Trypanosoma cruzi and Trypanosoma rangeli (Kinetoplastida: Trypanosomatidae): Its Application to Parasite-Vector Biology

Two Trypanosoma cruzi-derived cloning vectors, pTREX-n and pBs:CalB1/CUB01, were used to drive the expression of green fluorescent protein (GFP) and DsRed in Trypanosoma rangeli Tejera, 1920, and Trypanosoma cruzi Chagas, 1909, isolates, respectively. Regardless of the species, group, or strain, parasites harboring the transfected constructs as either episomes or stable chromosomal integrations showed high-level expression of fluorescent proteins. Tagged flagellates of both species were used to experimentally infect Rhodnius prolixus Stal, 1953. In infected bugs, single or mixed infections of T. cruzi and T. rangeli displayed the typical cycle of each species, with no apparent interspecies interactions. In addition, infection of kidney monkey cells (LLC-MK2) with GFP-T. cruzi showed that the parasite retained its fluorescent tag while carrying out its life cycle within cultured cells. The use of GFP-tagged parasites as a tool for biological studies in experimental hosts is discussed, as is the application of this method for copopulation studies of same-host parasitesFil: Guevara, Palmira. Universidad Central de Venezuela; VenezuelaFil: Dias, Manuel. Universidad de los Andes; ColombiaFil: Rojas, Agustina. Universidad de los Andes; ColombiaFil: Crisante, Gladys. Universidad de los Andes; ColombiaFil: Abreu Blanco, Maria Teresa. Universidad Central de Venezuela; VenezuelaFil: Umezawa, Eufrozina. Universidade de Sao Paulo; BrasilFil: Vazquez, Martin Pablo. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Investigaciones en Ingeniería Genética y Biología Molecular "Dr. Héctor N. Torres"; ArgentinaFil: Levin, Mariano Jorge. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Investigaciones en Ingeniería Genética y Biología Molecular "Dr. Héctor N. Torres"; ArgentinaFil: Añez, Nestor. Universidad de los Andes; ColombiaFil: Ramirez, Jose Luis. Instituto de Estudios Avanzados; Venezuel

Inconclusive results in conventional serological screening for Chagas` disease in blood banks: evaluation of cellular and humoral response

To find the most reliable screening method for Trypanosoma cruzi infection in blood banks. Epidemiological data, lymphoproliferation assay, parasitological, conventional serological tests: immunofluorescence, haemagglutination, ELISA with epimastigote and trypomastigote antigens and reference serological tests: trypomastigote excreted-secreted antigens (TESA) blot and chemiluminescent ELISA assay with mucine from trypomastigote forms were applied to individuals with inconclusive serology, non-chagasic individuals and chronic chagasic patients. TESA blot had the best performance when used as a single test in all the groups. In the inconclusive group 20.5% of individuals were positive for TESA blot, 23.3% for either lymphoproliferation or TESA blot, and 17.8% for lymphoproliferation only. Positive lymphoproliferation without detectable antibodies was observed in 5.47% of all inconclusive serology cases. Analysis of six parameters (three serological assays, at least one parasitological test, one lymphoproliferation assay and epidemiological data) in the inconclusive group showed that diagnosis of Chagas` disease was probable in 15 patients who were positive by two or more serological tests or for whom three of those six parameters were positive. TESA blot is a good confirmatory test for Chagas` disease in the inconclusive group. Although lymphoproliferation suggests the diagnosis of Chagas` disease in the absence of antibodies when associated with a high epidemiological risk of acquiring Chagas` disease, the data from this study and the characteristics of the lymphoproliferation assay (which is both laborious and time-consuming) do not support its use as a confirmatory test in blood-bank screening. However, our findings underscore the need to develop alternative methods that are not based on antibody detection to improve the diagnosis when serological tests are inconclusive

Inquérito sorológico para doença de Chagas em áreas rurais de Manaus, Coari e Tefé na Amazônia Ocidental

Submitted by Sandra Infurna ([email protected]) on 2017-06-13T16:28:06Z No. of bitstreams: 1 josecoura_etal_IOC_2011.pdf: 713583 bytes, checksum: 4b85288d95ee4f1ee962b5f383123892 (MD5)Approved for entry into archive by Sandra Infurna ([email protected]) on 2017-06-13T16:43:07Z (GMT) No. of bitstreams: 1 josecoura_etal_IOC_2011.pdf: 713583 bytes, checksum: 4b85288d95ee4f1ee962b5f383123892 (MD5)Made available in DSpace on 2017-06-13T16:43:07Z (GMT). No. of bitstreams: 1 josecoura_etal_IOC_2011.pdf: 713583 bytes, checksum: 4b85288d95ee4f1ee962b5f383123892 (MD5) Previous issue date: 2011Universidade do Estado do Amazonas. Programa de Pós-graduação em Medicina Tropical. Escola Superior de Ciências da Saúde. Manaus, AM, Brasil.Fundação de Medicina Tropical Heitor Vieira Dourado. Gerência de Entolomogia e Leishmaniose. Manaus, AM, Brasil / Universidade Federal do Amazonas. Faculdade de Farmácia. Manaus, AM, Brasil / Universidade Federal do Amazonas. Faculdade de Medicina. Manaus, AM, BrasilUniversidade do Estado do Amazonas. Programa de Pós-graduação em Medicina Tropical. Escola Superior de Ciências da Saúde. Manaus, AM, Brasil.Universidade Federal do Amazonas. Faculdade de Farmácia. Manaus, AM, Brasil / Universidade Federal do Amazonas. Faculdade de Medicina. Manaus, AM, BrasilInstituto de Medicina Tropical de São Paulo. Laboratório de Doenças Parasitárias. São Paulo, SP, Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Doenças Parasitárias. Rio de Janeiro, RJ. Brasil.Universidade do Estado do Amazonas. Programa de Pós-graduação em Medicina Tropical. Escola Superior de Ciências da Saúde. Manaus, AM, Brasil./ Fundação de Medicina Tropical Heitor Vieira Dourado. Gerência de Entolomogia e Leishmaniose. Manaus, AM, Brasil.Universidade do Estado do Amazonas. Programa de Pós-graduação em Medicina Tropical. Escola Superior de Ciências da Saúde. Manaus, AM, Brasil./ Fundação de Medicina Tropical Heitor Vieira Dourado. Gerência de Entolomogia e Leishmaniose. Manaus, AM, Brasil / Universidade Nilton Lins. Departamento de Biologia. Manaus, AM, Brasil.Introduction: Deforestation, uncontrolled forest, human population migration from endemic areas, and the large number of reservoirs and wild vectors naturally infected by Trypanosoma cruzi promote the endemicity of Chagas disease in the Amazon region. Methods: We conducted an initial serological survey (ELISA) in a sample of 1,263 persons; 1,095 (86.7%) were natives of the state of Amazonas, 666 (52.7%) were male, and 948 (75.1%) were over 20 years old. Serum samples that were found to be reactive, indeterminate, or inconclusive by indirect immunofluorescence (IFI) or positive with low titer by IFA were tested by Western blot (WB). Serologically confirmed patients (WB) were evaluated in terms of epidemiological, clinical, ECG, and echocardiography characteristics. Results: Fifteen patients had serologically confirmed T. cruzi infection, and 12 of them were autochthonous to the state of Amazonas, for an overall seroprevalence of 1.2% and 0.9% for the state of Amazonas. Five of the 15 cases were males, and the average age was 47 years old; most were farmers with low education. One patient who was not autochthonous, having originated from Alagoas, showed right bundle branch block, bundle branch block, and anterosuperior left ventricular systolic dysfunction with an ejection fraction of 54%. Conclusions: The results of this study ratify the importance of monitoring CD cases in Amazonia, particularly in the state of Amazonas

Serological survey for Chagas disease in the rural areas of Manaus, Coari, and Tefé in the Western Brazilian Amazon

INTRODUCTION: Deforestation, uncontrolled forest, human population migration from endemic areas, and the large number of reservoirs and wild vectors naturally infected by Trypanosoma cruzi promote the endemicity of Chagas disease in the Amazon region. METHODS: We conducted an initial serological survey (ELISA) in a sample of 1,263 persons; 1,095 (86.7%) were natives of the State of Amazonas, 666 (52.7%) were male, and 948 (75.1%) were over 20 years old. Serum samples that were found to be reactive, indeterminate, or inconclusive by indirect immunofluorescence (IFI) or positive with low titer by IFA were tested by Western blot (WB). Serologically confirmed patients (WB) were evaluated in terms of epidemiological, clinical, ECG, and echocardiography characteristics. RESULTS: Fifteen patients had serologically confirmed T. cruzi infection, and 12 of them were autochthonous to the state of Amazonas, for an overall seroprevalence of 1.2% and 0.9% for the state of Amazonas. Five of the 15 cases were males, and the average age was 47 years old; most were farmers with low education. One patient who was not autochthonous, having originated from Alagoas, showed right bundle branch block, bundle branch block, and anterosuperior left ventricular systolic dysfunction with an ejection fraction of 54%. CONCLUSIONS: The results of this study ratify the importance of monitoring CD cases in Amazonia, particularly in the state of Amazonas