title: [PCR-RFLP and RAPD for typing neotropical Leishmania].
abstract: BACKGROUND
  The analysis of the PCR-restriction fragment length polymorphism and random amplified polymorphic DNA have been useful tools for Leishmania identification.
  
  
  OBJECTIVE
  Molecular procedures were demonstrated for identification and typing of reference strains of New World Leishmania and their applicability was validated for clinical samples.
  
  
  METHODS
  DNA was extracted from 16 reference strains of Latin American Leishmania as well as from clinical samples of leishmaniasis patients. A sequence coding for cysteine proteinase B was amplified by PCR and subjected to restriction fragment length polymorphism analysis. The enzyme used was Taq1. For eight of the reference strains, the random amplified polymorphic desoxyribonucleic acid technique (RAPD) was applied. Band patterns for Leishmania species differentiation were established each each method. The sample size of the clinical sample was of 5.
  
  
  RESULTS
  PCR products of the cysteine proteinase B gene were obtained for L. braziliensis, L. peruviana, L. panamensis and L. guyanensis. For the other species, L. mexicana, L. amazonensis, L. garnhami, L. lainsoni, L. chagasi, L. naiffi, no amplification occurred. The patterns of restriction fragments revealed band patterns in common for L. peruviana, L. guyanensis and L. panamensis, whereas L. braziliensis had a distinctive pattern. When human samples were examined, amplification occurred for all cases, and the profiles corresponded to the common profile of L. peruviana, L. guyanensis and L. panamensis. The RAPD technique demonstrated reproducible and distinctive patterns for each of the 8 reference strains, L. mexicana, L. amazonensis, L. garnhami, L. lainsoni, L. chagasi, L. naiffi, making possible to differentiate all them. The advantages and limitations of each procedure are discussed.
  
  
  CONCLUSIONS
  The combination of RFP and RAPD methodologies provide useful tools to identify medical important species of Leishmania by recognizing DNA sequences characteristic of each species.
authors.affiliation: Departamento de Parasitología, Instituto Pedro Kourí, La Habana, Cuba. amontalvo@ipk.sld.cu
authors.forename: Ana Margarita
authors.initials: AM
authors.lastname: Montalvo
authors.forename: Lianet
authors.initials: L
authors.lastname: Monzote
authors.forename: Jorge
authors.initials: J
authors.lastname: Fraga
authors.forename: Ivón
authors.initials: I
authors.lastname: Montano
authors.forename: Carlos
authors.initials: C
authors.lastname: Muskus
authors.forename: Marcel
authors.initials: M
authors.lastname: Marín
authors.forename: Simonne
authors.initials: S
authors.lastname: de Doncker
authors.forename: Iván Darío
authors.initials: ID
authors.lastname: Vélez
authors.forename: Jean Claude
authors.initials: JC
authors.lastname: Dujardin
mesh_terms: D000818:Animals; D006801:Humans; D007891:Leishmania / Q000145:classification / Q000235:genetics; D007896:Leishmaniasis / Q000209:etiology / Q000503:physiopathology; D016133:Polymerase Chain Reaction / Q000379:methods*; D012150:Polymorphism, Restriction Fragment Length; D019105:Random Amplified Polymorphic DNA Technique; D014329:Tropical Climate