Aplicação de técnicas moleculares para identificação do complexo mycobacterium tuberculosis e genotipagem da linhagem beijing em cepas de m. tuberculosis isoladas em Manaus - AM

Abstract

The new technologies being developed have redefined the diagnosis of tuberculosis, providing a foundation for laboratory diagnostic techniques. The molecular diagnosis of tuberculosis by Polymerase Chain Reaction (PCR, polymerase chain reaction) that uses primers of high specificity (98%), with wide sensitivity variations (20-100%), has been used to identify genetic targets in the bacillus. The general objective of this work was to apply molecular techniques for the identification of bacteria of the Mycobacterium tuberculosis Complex and genotyping of the Beijing strain in strains of M. tuberculosis isolated in Manaus - AM. For this, the mycobacteria were preserved in Sauton's liquid medium or Middlebrook 7H9 medium containing Tween 80 and glycerol, kept at -20oC or -80oC in the mycobacteriology laboratory at INPA. With this, a DNA extraction was initially performed to use it in the PCR technique, followed by agarose gel electrophoresis and, at the end, the electrophoretic profile of each strain was set up based on genomic deletions for the primers 16SrRNA, Rv0577, IS1561 ', Rv1510, Rv1970, Rv3877/8, Rv3120 and Rv2073c. Finally, genotyping was carried out to verify the presence of the Beijing lineage among these strains, through the Multiplex PCR technique, where the primers ON-1002 (Fw), ON-1258 (R1) and ON-1127 (R2) were used. This work shows that among the CMtb species, only the Mycobacterium tuberculosis species was identified among the analyzed isolates (64.3%), as well as the presence of the Beijing genotype was not observed among the M. tuberculosis strains isolated in Manaus – AM and analyzed so far. Key words: M. tuberculosis; Identification; Molecular; Genotyping; Beijing.