Evaluación de las pruebas de PCR TcH2AF-R y S35-S36 para la detección de Trypanosoma cruzi en tejido cardiaco de ratón

Abstract

Introducción. El trasplante es una opción terapéutica en la cardiomiopatía chagásica. Para la detección temprana de una posible reactivación de la infección, se propone el uso de pruebas de reacción en cadena de la polimerasa (PCR) a partir de biopsias endomiocárdicas; el modelo de ratón es una aproximación preliminar para evaluar la aplicación de éstas. Objetivo. Evaluar la aplicación de las pruebas de PCR basadas en los iniciadores TcH2AF-R y S35-S36 para la detección de T. cruzi en tejido cardiaco de ratones infectados con el parásito. Materiales y métodos. Se infectaron por vía intraperitoneal dos grupos de ratones ICR de 15 y 10 individuos con 0,3 ml de PBS que contenían 1x10 6 tripomastigotes de la cepa MHOM/CO/ 2001/D.A. (T. cruzi I) o 1x10 4 tripomastigotes de la cepa MHOM/BR/00/Y (T. cruzi II), respectivamente. El seguimiento de la parasitemia se realizó mediante microhematocrito y presencia de parásitos en el corazón a los 30, 60 (modelo agudo), 100 y 150 (modelo crónico) días por medio de histopatología y de las PCR TcH2AF-R y S35-S36. Resultados. La histopatología mostró alteraciones en el miocardio y presencia de amastigotes en los modelos agudo y crónico. En contraste al microhematocrito y al análisis histopatológico, la PCR S35-S36 permitió la detección de ambas cepas del parásito. La PCR TcH2AF-R detectó la cepa T. cruzi I con un desempeño superior al microhematocrito y al análisis histopatológico. Conclusiones. El uso de ambas pruebas de PCR puede ser útil en la confirmación de la reactivación de la infección postrasplante.Palabras clave: Trypanosoma cruzi, enfermedad de Chagas, cardiomiopatía chagásica, reacción en cadena de la polimerasa. Evaluation of TcH2AF-R and S35-S36 primers in PCR tests for the detection of Trypanosoma cruzi in mouse cardiac tissueIntroduction. Heart transplant is a therapeutic option in the treatment of chagasic cardiomyopathy. For early detection of Chagas reactivation cases, the use of PCR tests using endomyocardial biopsies has been proposed. Development of an animal model will be the first step in evaluating the applicability of this approach. Objective. PCR tests based on the TcH2AF-R and S35-S36 primers were evaluated for the detection of T. cruzi in heart tissue of mice experimentally infected with the parasite. Parasitemia and cardiac parasitic infection were determined at 30, 60 (acute model), 100 and 150 (chronic model) days by means of histopathological examination and by PCR, using the TcH2AF-R and S35-S36 primers. Results. The histopathological findings revealed alterations in the heart and the presence of intracellular amastigotes in acute and chronic models. In contrast to parasitemia levels and

Histopathological findings of infections caused by canine distemper virus, Trypanosoma cruzi, and other parasites in two free-ranging White-nosed Coatis Nasua narica (Carnivora: Procyonidae) from Costa Rica

“…This is, however, a non-pathogenic parasite and it is not observed in mammalian tissue (Vallejo et al 2015). PCR reaction conditions were carried out as previously described with the S35-S36 set with some modifications (Barrera et al 2008). Briefly, PCR was carried out using a DNA thermal cycler (Gene Amp PCR System.…”

CD8+ T Cell Response Quality Is Related to Parasite Control in an Animal Model of Single and Mixed Chronic Trypanosoma cruzi Infections

“…Trypomastigotes of the Y strain were passaged in female inbred BALB/cAnNCr mice at least three times to maintain virulence. Both T. cruzi strains represent the most predominant genetic groups in Latin America and have shown different inflammatory profiles and infection outcomes in animal models (Barrera et al, 2008;Cruz et al, 2015;Sales-Campos et al, 2015;Santi-Rocca et al, 2017;Zingales, 2018;Ledezma et al, 2020). For instance, an infection with the T. cruzi DA strain has shown a prolonged infection with preferential migration to cardiac tissue and reappearance of parasitemia in chronic stages of infection (Barrera et al, 2008;Cruz et al, 2015).…”

“…Both T. cruzi strains represent the most predominant genetic groups in Latin America and have shown different inflammatory profiles and infection outcomes in animal models (Barrera et al, 2008;Cruz et al, 2015;Sales-Campos et al, 2015;Santi-Rocca et al, 2017;Zingales, 2018;Ledezma et al, 2020). For instance, an infection with the T. cruzi DA strain has shown a prolonged infection with preferential migration to cardiac tissue and reappearance of parasitemia in chronic stages of infection (Barrera et al, 2008;Cruz et al, 2015). In contrast, strain Y shows an aggressive illness with high parasitemia levels in the acute phase and tropisms towards the liver and colon (Mateus et al, 2019).…”

“…Because several studies have documented that the genetics and phenotypic heterogeneity of T. cruzi strains may be associated with distinct infection outcomes (Santi-Rocca et al, 2017;Zingales, 2018), we first analyzed the parasitological and histological outcomes during T. cruzi infection in experimentally infected mice with T. cruzi trypomastigotes belonging to two strains of different genotypes (Y or DA) as described in the Materials and Methods section. The Y and DA strains were isolated from infected patients and belong to the TcII and TcI genotypes of T. cruzi, respectively (Pinto et al, 1999;Barrera et al, 2008;Amato Neto, 2010;Cruz et al, 2015). We pooled acutely and chronically infected mice from days 10 and 30 and days 100 and 260, respectively, as described previously (Mateus et al, 2019).…”

“…A comprehensive study that included several genetic T. cruzi groups found strains from different genotypes induced a high degree of heterogeneity in inflammation degree or immune response in acutely and chronically infected mice (Santi-Rocca et al, 2017). For example, TcI and TcII strains, which are the most predominant T. cruzi genotypes in Latin America, have shown high diversity in inflammation outcomes ranging from mild to severe both chagasic patients and animal models (Barrera et al, 2008;Cruz et al, 2015;Sales-Campos et al, 2015;Hernandez et al, 2016;Zingales, 2018;Ledezma et al, 2020). Indeed, epidemiological studies of individuals with positive Chagas serological tests have shown that continuous exposure to infections with distinct T. cruzi genotypes increases the risk of progression to chronic Chagas cardiomyopathy (Zicker et al, 1990;Basquiera et al, 2003;Sabino et al, 2013;Perez et al, 2014).…”