Caracterização molecular e estudo do genoma de Burkholderia mallei isoladas de equídeos nos estados de Pernambuco e Alagoas, Brasil

Abstract

Glanders is an infectious-contagious disease first reported between the 2nd and 4th centuries and that causes serious economic losses for equidae culture. The aim of this study was to genetically characterize Burkholderia mallei isolates obtained from equine glanders in the states of Pernambuco and Alagoas. During necropsies of seropositive animals, samples of pyogranulomatous lesions were collected. The material was subjected to bacterial isolation and bacterial cultures were analyzed by conventional and real time Polymerase Chain Reaction (PCR) for confirmation. In total, six isolates were identified on horse farms in both states, five from Pernambuco state and one from Alagoas. The amplicons were sequenced and compared to the database found at the National Center for Biotechnology Information (NCBI), where it was possible to observe similarity with the Turkey 10 isolate, first reported in Turkey. In order to confirm the result, the isolates were submitted to Polymerase Chain Reaction - High Resolution Melting (PCR-HRM) for genetic characterization through the variation of Single Nucleotide polymorphisms (SNPs) and subsequently to complete genome sequencing (WGS). The complete genome sequencing revealed a correlation between the circulating strains between the two states, but a genetic variation was observed among strains isolated from animals from Pernambuco. Two genotypes of Burkholderia mallei circulating in Brazil have been described, being the L3B2 genotype previously described for a Brazilian isolate and the L3B3Sb3Gp1SbGp1 genotype, never previously described for contemporary strains in Brazil. This is the first current description of an isolate with genetic similarity to other strains reported in Europe, suggesting more than one event of introduction of the disease in Brazil. The characterization of circulating B. mallei isolates is important for the development and improvement of serological diagnostic techniques and the development of specific molecular markers for the detection of Brazilian strains, in addition to being an additional means of controlling the disease, making it possible to establish links between the different cases and identify transmission chains.