Epidemiologia molecular, resistência aos antimicrobianos e perfil de virulência de isolados clínicos de Klebsiella pneumoniae resistentes aos carbapenêmicos

Abstract

Carbapenem-resistant Klebsiella pneumoniae (CRKP) represents a global public health threat as there are limited therapeutic options available to control this pathogen. This study aimed to analyze the antimicrobial resistance, virulence profile and molecular epidemiology of CRKP strains isolated from patients in hospitals from the Metropolitan Region of Greater Vitória, Espírito Santo. The susceptibility profile to antimicrobials was evaluated by agar diffusion and broth microdilution tests. Detection of genes that confer resistance to beta-lactams was performed by the polymerase chain reaction (PCR) technique. The blaKPC insertion region was evaluated using whole genome sequencing (WGS). The virulence genes were detected by PCR and in vivo virulence was analyzed using the Galleria mellonella model. Genetic polymorphism was analyzed using the pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST) techniques. The blaKPC and different genes that encoding extended-spectrum beta-lactamases (ESBL) were detected in all strains, which were also resistant to multiple drugs (MDR), whilst some were also resistant to last-resort antimicrobials, such as colistin and tigecycline. It was identified through the analysis of the blaKPC insertion region that this gene is mostly transported by the Tn4401 transposon, emphasizing its importance in the dissemination of blaKPC. The Galleria mellonella model showed the presence of CRKP virulent strains in health-related environments and suggests that strains resistant to colistin were associated with greater virulence. The profile of virulence genes was conserved, not correlating to the G. mellonela results, showing that these pathogens' virulence is complex and multifaceted. MLST analysis revealed eight different sequence types (ST11, ST37, ST147, ST340, ST384, ST394, ST437 e ST628), two (ST628 e ST394) being reported for the first time in Brazil. The 2B strain presented a different genetic context compared to the other strains, where Tn4401 was modified with the insertion of the Tn5403 element, providing evidence of a new insertion region of blaKPC-2 in an ST11 strain belonging to the CC258, considered a high-risk clone. In conclusion, this study shows the dissemination of virulent CRKP-MDR strains in hospitals at Greater Vitória and notably the presence of highly-virulent CRKP-MDR ST628 strains, demonstrating that virulent and resistant clones can rapidly arise. Furthermore, it reinforces the importance of the use of genomic tools on the epidemiological vigilance of CRKP, as they can contribute to the understanding of the molecular evolution of these strains, assisting in tracing and controlling its dissemination.