Differentiation in a Single-Tube PCR between Leishmania mexicana and Leishmania braziliensis in Clinical Samples
Victor Monteon *
Biomedical Research Center, Campeche State University, Mexico.
Enrique Quen-Ramirez
Biomedical Research Center, Campeche State University, Mexico
Román Diaz-Arce
Biomedical Research Center, Campeche State University, Mexico
María Vargas
State Public Health Laboratory, Campeche, Mexico.
Ruth López
Biomedical Research Center, Campeche State University, Mexico.
Lidia Baylon
Infectomic and Molecular Pathogenesis Department, Research Center and Advanced Studies, Mexico.
José L. Rosales
Infectomic and Molecular Pathogenesis Department, Research Center and Advanced Studies, Mexico.
*Author to whom correspondence should be addressed.
Abstract
Aim: To assess the usefulness of a single-tube PCR end point for differentiating between Leishmania and Viannia subgenera in clinical samples of cutaneous leishmaniasis.
Methodology: Impression smears of 65 patients from Campeche State, Mexico with leishmaniasis-like lesions were analyzed by conventional Giemsa staining and a new simple PCR end point with primers named LU-5A designed by [Harris et al.1998] and LME (designed for this study and based on a mini-exon of L.(L) mexicana,GenBankX64317.1). This PCR generates an amplicon of 210 and 270 bp with L. (L) mexicana (MHOM/MX/84/SETGS) and 230 bp with L. (V) braziliensis (MHOM/BR/75/M2903).
Results: Out of 65 impression smears of clinical samples 50 were positive according to PCR (76.9%) and 29 were positive according to microscopic observation (44.6%). The primers did not hybridized with human, Trypanosoma cruzi, Staphylococcus aureus, Staphylococcus epidermidis or Pseudomona aeruginosa DNA. Of the positive smears, 84% (42/50) corresponded to L. (L) mexicana and 16% (8/50) to L. (V) braziliensis.
Conclusion: A one-step PCR with primers LU-5A and LME improves the diagnostic capacity to differentiate between Leishmania and Viannia subgenera in parallel with conventional tests.
Keywords: Leishmania mexicana, Leishmania braziliensis, diagnosis, PCR