LIMITATION OF PRIMERS USED IN PCR FOR THE CHARACTERIZATION OF LEISHMANIA INFANTUM.

de Araújo, Helton Krisman; de Oliveira Castro, Silvana; da Silva Valejo, Maria Joelma Alves; da Costa Lima Junior, Manoel Sebastião; Neitzke-Abreu, Herintha Coeto

de Araújo, Helton Krisman; de Oliveira Castro, Silvana; da Silva Valejo, Maria Joelma Alves; da Costa Lima Junior, Manoel Sebastião; Neitzke-Abreu, Herintha Coeto.

Afiliação

de Araújo HK; Faculty of Health Sciences, Universidade Federal da Grande Dourados (UFGD), 79804-070, Dourados, Mato Grosso do Sul State, Brazil.
de Oliveira Castro S; Faculty of Health Sciences, Universidade Federal da Grande Dourados (UFGD), 79804-070, Dourados, Mato Grosso do Sul State, Brazil.
da Silva Valejo MJA; Postgraduate Program in Health Sciences, Universidade Federal da Grande Dourados (UFGD), 79804-070, Dourados, Mato Grosso do Sul State, Brazil.
da Costa Lima Junior MS; Oswaldo Cruz Foundation (FIOCRUZ), Instituto Aggeu Magalhães, Recife, Pernambuco State, Brazil.
Neitzke-Abreu HC; Faculty of Health Sciences, Universidade Federal da Grande Dourados (UFGD), 79804-070, Dourados, Mato Grosso do Sul State, Brazil.

J Parasitol ; 109(5): 445-449, 2023 10 01.

Article em En | MEDLINE | ID: mdl-37668295

RESUMO

Conventional PCR provides Leishmania species characterization with even a small amount of biological material. Species-specific primers have been a widely used alternative; however, nonspecific amplifications are a reality, interfering with PCR efficiency. In endemic areas with multiple etiological agents for leishmaniasis, there is a requirement for higher specificity of primers. This study evaluates 3 pairs of primers described for the identification and characterization of Leishmania infantum. Primers RV1/RV2, LEISH1/LEISH2, and FLC2/RLC2 were used with the DNA of L. infantum, Leishmania amazonensis, and Leishmania braziliensis. An initial temperature curve was performed (52-62 C) to determine the optimal annealing temperature, followed by a dilution curve of Leishmania DNA (500 pg/µl, 50 pg/µl, 5 pg/µl, 500 fg/µl, 50 fg/µl, 5 fg/µl, and 0.5 fg/µl) to be used for analytical sensitivity. RV1/RV2 PCR amplified L. infantum and L. amazonensis at all analyzed temperatures; LEISH1/LEISH2 PCR amplified all 3 species of Leishmania, although at some temperatures L. infantum was specifically amplified, and, finally, FLC2/RLC2 PCR amplified only L. infantum at all temperatures analyzed. In terms of sensitivity, RV1/RV2 PCR detected 1 fg of L. infantum DNA and 100 pg of L. amazonensis DNA; LEISH1/LEISH2 PCR detected 1 fg of L. infantum DNA, 100 fg of L. amazonensis DNA, and 10 fg of L. braziliensis DNA; and FLC2/RLC2 PCR detected 10 fg of L. infantum DNA. Thus, PCR with FLC2/RLC2 primers is best suited for the molecular characterization of L. infantum, especially in areas where there is an incidence of more than 1 Leishmania species, such as South America.

Assuntos

Leishmania infantum; Leishmania mexicana; Leishmania infantum/genética; Reação em Cadeia da Polimerase; América do Sul; Especificidade da Espécie

Palavras-chave

Leishmania; PCR; Primer

Texto completo

Adicionar na Minha BVS

Imprimir

XML

PubMed Links

Buscar no Google

Texto completo: 1 Coleções: 01-internacional Base de dados: MEDLINE Assunto principal: Leishmania mexicana / Leishmania infantum País/Região como assunto: America do sul Idioma: En Revista: J Parasitol Ano de publicação: 2023 Tipo de documento: Article País de afiliação: Brasil País de publicação: Estados Unidos

Texto completo

Adicionar na Minha BVS

Imprimir

XML

PubMed Links

Buscar no Google