Identificação e validação in silico de potenciais marcadores moleculares para o gênero Leishmania

Abstract

Leishmaniasis encompasses a wide spectrum of diseases caused by the parasite of the genus Leishmania. Molecular methods are being increasingly used for diagnosis, taxonomic, phylogenetic and epidemiological studies involving this parasite. The development of molecular markers has become essential for more accurate diagnoses in the identification of pathogens of interest to man. The objective of this work was to develop molecular markers in silico for Leishmania and evaluate its applicability which was developed using free bioinformatics programs available on the internet. Eleven NCBI sequences were selected for Leishmania infantum and Leishmania donovani, in which a multiple alignment was performed with the achievement of a consensus sequence. Once the conserved regions were identified, the region of the primer to be developed was defined. Three primers were developed, taking into account the number of base pairs, the percentage of guanine and cytosine, which ranged from 37,5% to 47,62% and Melting temperature that has a range of 56,48°C to 59,08°C. The developed primers have a validation rate of 85% to 100%. The PCR-RFLP in silico, proved to be adequate for the identification of Leishmania at the species level. The genetic variation in sequences in mini kDNA circles between L. donovani and L. infantum isolates, can be detected through the SNPs present in the respective genomes. The restriction enzyme RsaI was considered ideal to determine the band variation between the two species.