title: NEW PRIMERS FOR DETECTION OF Leishmania infantum USING POLYMERASE CHAIN REACTION.
abstract: Leishmania infantum causes visceral leishmaniasis (VL) in the New World. The diagnosis of VL is confirmed by parasitological and serological tests, which are not always sensitive or specific. Our aim was to design new primers to perform a Polymerase Chain Reaction (PCR) for detecting L. infantum. Sequences of the minicircle kinetoplast DNA (kDNA) were obtained from GenBank, and the FLC2/RLC2 primers were designed. Samples of DNA from L. infantum, Leishmania amazonensis, Leishmania braziliensis, Leishmania guyanensis, Leishmania naiffi, Leishmania lainsoni, Leishmania panamensis, Leishmania major and Trypanosoma cruzi were used to standardize the PCR. PCR with FLC2/RLC2 primers amplified a fragment of 230 bp and the detection limit was 0.2 fg of L. infantum DNA. Of the parasite species assayed, only L. infantum DNA was amplified. After sequencing, the fragment was aligned to GenBank sequences, and showed (99%) homology with L. infantum. In the analysis of blood samples and lesion biopsy from a dog clinically suspected to have VL, the PCR detected DNA from L. infantum. In biopsy lesions from humans and dogs with cutaneous leishmaniasis, the PCR was negative. The PCR with FLC2/RLC2 primers showed high sensitivity and specificity, and constitutes a promising technique for the diagnosis of VL.
authors.affiliation: Universidade Estadual de Maringá, Maringá, PR, Brasil.
authors.forename: Kézia Peres
authors.initials: KP
authors.lastname: Gualda
authors.affiliation: Universidade Estadual de Maringá, Maringá, PR, Brasil.
authors.forename: Lílian Mathias
authors.initials: LM
authors.lastname: Marcussi
authors.affiliation: Universidade Estadual de Maringá, Maringá, PR, Brasil.
authors.forename: Herintha Coeto
authors.initials: HC
authors.lastname: Neitzke-Abreu
authors.affiliation: Departamento de Análises Clínicas e Biomedicina, Universidade Estadual de Maringá, Maringá, PR, Brasil.
authors.forename: Sandra Mara Alessi
authors.initials: SM
authors.lastname: Aristides
authors.affiliation: Departamento de Análises Clínicas e Biomedicina, Universidade Estadual de Maringá, Maringá, PR, Brasil.
authors.forename: Maria Valdrinez Campana
authors.initials: MV
authors.lastname: Lonardoni
authors.affiliation: Departamento de Análises Clínicas e Biomedicina, Universidade Estadual de Maringá, Maringá, PR, Brasil.
authors.forename: Rosilene Fressatti
authors.initials: RF
authors.lastname: Cardoso
authors.affiliation: Departamento de Análises Clínicas e Biomedicina, Universidade Estadual de Maringá, Maringá, PR, Brasil.
authors.forename: Thaís Gomes Verzignassi
authors.initials: TG
authors.lastname: Silveira
mesh_terms: D000818:Animals; D017931:DNA Primers / Q000235:genetics*; D018105:DNA, Kinetoplast / Q000235:genetics*; D016054:DNA, Protozoan / Q000235:genetics*; D004283:Dog Diseases / Q000175:diagnosis*; D004285:Dogs; D006801:Humans; D018314:Leishmania infantum / Q000302:isolation & purification*; D016773:Leishmaniasis, Cutaneous / Q000175:diagnosis* / Q000662:veterinary; D007898:Leishmaniasis, Visceral / Q000175:diagnosis* / Q000662:veterinary; D016133:Polymerase Chain Reaction; D012680:Sensitivity and Specificity
chemical_list: D017931:DNA Primers; D018105:DNA, Kinetoplast; D016054:DNA, Protozoan