title: Polymerase chain reaction-based method for the identification of Leishmania (Viannia) braziliensis and Leishmania (Viannia) guyanensis in mucosal tissues conserved in paraffin.
abstract: BACKGROUND
  In the Americas, mucosal leishmaniasis is primarily associated with infection by Leishmania (Viannia) braziliensis. However, Leishmania (Viannia) guyanensis is another important cause of this disease in the Brazilian Amazon. In this study, we aimed at detecting Leishmaniadeoxyribonucleic acid (DNA) within paraffin-embedded fragments of mucosal tissues, and characterizing the infecting parasite species.
  
  
  METHODS
  We evaluated samples collected from 114 patients treated at a reference center in the Brazilian Amazon by polymerase chain reaction (PCR) and restriction fragment length polymorphism (RFLP) analyses.
  
  
  RESULTS
  Direct examination of biopsy imprints detected parasites in 10 of the 114 samples, while evaluation of hematoxylin and eosin-stained slides detected amastigotes in an additional 17 samples. Meanwhile, 31/114 samples (27.2%) were positive for Leishmania spp. kinetoplast deoxyribonucleic acid (kDNA) by PCR analysis. Of these, 17 (54.8%) yielded amplification of the mini-exon PCR target, thereby allowing for PCR-RFLP-based identification. Six of the samples were identified as L. (V.) braziliensis, while the remaining 11 were identified as L. (V.) guyanensis.
  
  
  CONCLUSIONS
  The results of this study demonstrate the feasibility of applying molecular techniques for the diagnosis of human parasites within paraffin-embedded tissues. Moreover, our findings confirm that L. (V.) guyanensisis a relevant causative agent of mucosal leishmaniasis in the Brazilian Amazon.
authors.affiliation: Universidade do Estado do Amazonas, Manaus, Amazonas, Brazil.
authors.forename: Suzane Ribeiro
authors.initials: SR
authors.lastname: Prestes
authors.affiliation: Fundação de Medicina Tropical Dr. Heitor Vieira Dourado, Manaus, Amazonas, Brazil.
authors.forename: Jorge Augusto de Oliveira
authors.initials: JA
authors.lastname: Guerra
authors.affiliation: Núcleo de Medicina Tropical, Universidade de Brasília, Brasília, Distrito Federal, Brazil.
authors.forename: Gustavo Adolfo Sierra
authors.initials: GA
authors.lastname: Romero
authors.affiliation: Universidade do Estado do Amazonas, Manaus, Amazonas, Brazil.
authors.forename: Laylah Kelre Costa
authors.initials: LK
authors.lastname: Magalhaes
authors.affiliation: Universidade do Estado do Amazonas, Manaus, Amazonas, Brazil.
authors.forename: Rosa Amelia Gonçalves
authors.initials: RA
authors.lastname: Santana
authors.affiliation: Universidade do Estado do Amazonas, Manaus, Amazonas, Brazil.
authors.forename: Marcel Gonçalves
authors.initials: MG
authors.lastname: Maciel
authors.affiliation: Centro de Malária e outras Doenças Tropicais, Instituto de Higiene e Medicina Tropical, Lisboa, Portugal.
authors.forename: Ana
authors.initials: A
authors.lastname: Custódio
authors.affiliation: Universidade do Estado do Amazonas, Manaus, Amazonas, Brazil.
authors.forename: Maria das Graças Vale
authors.initials: Md
authors.lastname: Barbosa
authors.affiliation: Centro de Malária e outras Doenças Tropicais, Instituto de Higiene e Medicina Tropical, Lisboa, Portugal.
authors.forename: Henrique
authors.initials: H
authors.lastname: Silveira
mesh_terms: D016054:DNA, Protozoan / Q000032:analysis; D005260:Female; D006801:Humans; D007892:Leishmania braziliensis / Q000235:genetics* / Q000302:isolation & purification; D018334:Leishmania guyanensis / Q000235:genetics* / Q000302:isolation & purification; D007897:Leishmaniasis, Mucocutaneous / Q000469:parasitology*; D008297:Male; D009092:Mucous Membrane / Q000469:parasitology*; D010232:Paraffin; D016133:Polymerase Chain Reaction; D012150:Polymorphism, Restriction Fragment Length; D015800:Protozoan Proteins / Q000235:genetics
chemical_list: D016054:DNA, Protozoan; D015800:Protozoan Proteins; D010232:Paraffin