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The polymerase chain reaction (PCR) is a potentially interesting diagnostic tool for detecting congenital Trypanosoma cruzi infection at birth. We have compared the sensitivity and capacity of a group of T. cruzi PCR primers in detecting the complete spectrum of known T. cruzi lineages, and to improve and simplify the detection of infection in neonatal blood. We found that the two primers, Tcz1/Tcz2 and Diaz1/Diaz2, which target the 195-basepair satellite repeat, detected all parasitic lineages with the same sensitivity. However, the intensity of the amplicon was somewhat higher with Tcz1/Tcz2. For other tested primers (nuclear DNA primers BP1/BP2, O1/O2, Pon1/Pon2, and Tca1/Tca2 and kinetoplast DNA primers S35'/S36' and 121/122), either the intensity of amplicons varied according to T. cruzi lineages or the PCR assay was less sensitive. The use of the Tcz1/Tcz2 primers, which target a tandem repetitive sequence, requires a careful determination of the appropriate amount of Taq polymerase to avoid the formation of smears and multiple amplicon bands. The Tcz1/Tcz2 primers resulted in an intense 200-basepair amplicon with DNA extracted from blood equivalent to 0.02 parasites per assay when used with a simple DNA extraction method and of a low amount of Taq polymerase from a standard PCR kit. To better assess such PCR protocol, we assayed 311 samples of neonatal blood previously tested by parasitologic methods. The reliability of our PCR test was demonstrated, since all the 18 blood samples from newborns with congenital T. cruzi infection were positive, whereas the remaining samples (30 from control newborns of uninfected mothers and 262 of 263 from babies born to infected mothers) were negative. Since our PCR method is simple, reliable, robust, and inexpensive, it appears suitable for the detection of T. cruzi infection in neonatal blood, even in laboratories that are not equipped for performing the PCR.
Received November 20, 2002. Accepted for publication February 6, 2003.
Acknowledgments: Myrna Virreira was a fellow of the Coopération Technique Belge. Cristina Alonso-Vega is a fellow of the Association pour la Promotion de l’Éducation et la Formation à l’Étranger (Association pour la Promotion de l’Education et de la Formation a l’Etranger, Communauté Française de Belgique). We thank Eduardo Suarez, Marisol Cordova, and the staff of the maternity German Urquidi (Cochabamba, Bolivia) for the management of patients; Patricia Rodriguez, Rudy Parrado, and Myrian Huanca (Centro Universitario de Medicina Tropical/Laboratorio de Medicine, Universidad Mayor de San Simon, Cochabamba, Bolivia) for the serologic and parasitologic diagnosis of patients; Gaelle Leloux and Pascale Deblandre for technical assistance; and Michel Tibayrenc and Christian Barnabé (Génétique des Maladies Infectieuses, Unité Mixte, Centre National de Recherche Scientifique, Institut de Recherche pour le Developpement no. 9926, Montpellier, France) for providing the DNA of T. cruzi lineages and T. rangeli.
Financial support: This study was supported by the Conseil Interuniversitaire de la Communauté Française de Belgique, the Centre de Recherche Interuniversitaire en Vaccinologie sponsored by the Région Wallonne and GlaxoSmithKline (Rixensart, Belgium), and the Fonds National de la Recherche Scientifique Médicale (Convention 3.4595.99).
Authors’ addresses: Myrna Virreira, Laboratoire de Chimie Biologique et Laboratoire de Parasitologie, Faculté de Médecine, Université Libre de Bruxelles, Brussels, Belgium. Faustino Torrico, Cristina Alonso-Vega, and Marco Solano, Centro Universitario de Medicina Tropical/Laboratorio de Medicina, Faculdad de Medecina, Universidad Mayor de San Simon, Cochabamba, Bolivia. Carine Truyens and Yves Carlier, Laboratoire de Parasitologie, Faculté de Médecine, Université Libre de Bruxelles, Brussels, Belgium. Michal Svoboda, Laboratoire de Chimie Biologique, Faculté de Médecine, Université Libre de Bruxelles, 808 Route de Lennik, CP 611, B-1070 Bruxelles, Belgium, Telephone: 32-2-555-6225, Fax: 32-2-555-6230, E-mail: msvobod{at}ulb.ac.be
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