Polymerase chain reaction amplification of three different Trypanosoma cruzi DNA sequences from human chagasic cardiac tissue.

Abstract

Abstract. Chagas' disease is caused by the hemoflagellate protozoan Trypanosoma cruzi. The most common, serious manifestation of Chagas' disease is a progressive inflammatory cardiomyopathy, which occurs decades after primary infection. The inability to consistently demonstrate T. cruzi by histologic techniques in inflammatory cardiac lesions has suggested that the parasites' persistence may not be required for the pathology of the chronic phase. In this report we further analyze the persistence and localization of T. cruzi DNA in the hearts of seven patients with chronic chagasic cardiomyopathy, along with four indeterminate patients and seven control patients seronegative for T. cruzi infection. In the seven patients with chronic chagasic cardiomyopathy, we extracted DNA from selected inflammatory foci-positive (IFP) and inflammatory foci-negative (IFN) areas of' hematoxylin and eosin-stained cardiac tissue. We then used polymerase chain reaction methodology to amplify three different T. cruzi sequences (a minicircle sequence [MCS], a satellite repetitive sequence [RS], and, a low copy number sequence within the gene coding for a flagellar protein [FPS]). The MCS was detected in ϳ100% of both the IFP and IFN areas analyzed. The RS was detected in 37.5% and 23% of the IFP and IFN areas, respectively (difference not statistically significant; P Ͼ 0.10, degrees of freedom ϭ 1, G test of independence ϭ 1.9522). The FPS was rarely detected (2%), and was only present in DNA extracted from IFP areas. The MCS was also detected in most indeterminate cases (none of whom had inflammatory lesions) although with a markedly diminished amplification signal relative to cardiomyopathy cases. The MCS was not amplified from the cardiac tissues from seronegative controls. These results suggest that the quantity of T. cruzi DNA persisting in hearts of patients with Chagas' disease correlates with cardiomyopathy, but may not be preferentially associated with inflammatory foci.Chagas' disease is caused by the flagellate protozoan Trypanosoma cruzi. Ten to twenty million people in South and Central America are infected with the parasite, and it is estimated that 35 million people are at risk of infection.

Trypanosoma cruzi persistence at oral inflammatory foci in chronic chagasic patients

“…This statement is supported by previous reports from experimental hosts or natural infected individuals (Tarleton et al, 1997;Olivares-Villagomez et al, 1998). The finding of persistent myocardial T. cruzi forms in CCP has been correlated with the presence and degree of inflammatory lesions which tend to be diffuse and progressive (Almeida et al, 1984;Tavares-Neto, 1990;Jones et al, 1993;Añez et al, 1999).…”

Detection of Trypanosoma cruzi DNA within murine cardiac tissue sections by in situ polymerase chain reaction

“…In addition, topological constraints involved in the physical organization of minicircles within the kinetoplast may inhibit efficient amplification (Chen et al 1995). Our results confirm previous data demonstrating that MCS is consistently found in myocardial tissue from chagasic patients with cardiomyopathy using either fluid phase or in situ PCR followed by hybridization (Lane et al 1997, Olivares-Villagomez et al 1998.…”

“…However, Tarleton et al (1997) reported a lack of autoimmune-type rejection for inflammatory response in neonatal hearts transplanted into mice chronically infected with T. cruzi. Olivares-Villagomez et al (1998) analyzed T. cruzi specific DNA sequences using fluid phase polymerase chain reaction (PCR) and demonstrated that the quantity of T. cruzi DNA in the hearts of chagasic patients correlates with cardiomyopathy but may not be associated with inflammatory foci. Palomino et al (2000) attempted to associate the presence of parasite antigens with histopathologic lesions found in affected tissues from 12 chronic chagasic cardiomyopathies.…”

The pathogenesis of Chagas' disease: when autoimmune and parasite-specific immune responses meet

“…Several studies have demonstrated that T. cruzi parasites or antigens can be found, although rarely, in individuals with chronic infection (Barbosa Jr andAndrade 1984, Jones et al 1993, Palomino et al 2000). However, the analyses of human heart fragments have failed to show a correlation between intensity of inflammation and of parasitism, even with the use of highly sensitive techniques, such as PCR or immunohistochemistry (Palomino et al 2000, Olivares-Villagomez et al 1998. In fact, this remarkable feature of the disease was immediately pointed out 90 years ago by Vianna (1911), the excellent pathologist working in collaboration with Carlos Chagas.…”