DETECTION OF TRYPANOSOMA-CRUZI WITH THE POLYMERASE CHAIN-REACTION ANDIN-SITU HYBRIDIZATION IN INFECTED MURINE CARDIAC TISSUE

Chagas' disease is caused by the hemoflagellate protozoan Trypanosoma cruzi, which is predominantly found in South and Central America and M exico. Although the parasite is present in the United States, confirme d cases of human disease are rare. The most serious manifestation of c hronic Chagas' disease is a progressive inflammatory cardiomyopathy, H owever, T. cruzi has not been consistently demonstrated with histologi c techniques in inflammatory cardiac lesions. In this study, we used b oth polymerase chain reaction (PCR) amplification of extracted DNA fro m hematoxylin and eosin-stained tissue scrapings, and in situ hybridiz ation to detect the presence of T. cruzi in infected murine cardiac ti ssue sections. Three T, cruzi-specific DNA sequences were used: a 122- basepair (bp) sequence localized within the minicircle network (MCS), a 188-bp nuclear repetitive sequence (RS), and a 177-bp sequence withi n the open reading frame of a gene coding for a flagellar protein (FPS ). We found that ail three sequences are amplifiable from scrapings of murine cardiac tissue, The MCS and RS are detected at 0.167 and 0.24 amastigote DNA equivalents, while I;PS is barely detected at 0.24 amas tigote DNA equivalents, On the other hand, in situ hybridization with all three sequences allowed for the detection of T, cruzi amastigotes within the tissue. The MCS and FPS, however, consistently yielded a mo re intense signal, These results indicate that PCR and in situ hybridi zation may prove useful in establishing the prevalence of T. cruzi in human chagasic cardiomyopathy.