Caracterização fenotípica e genotípica de Klebsiella pneumoniae produtora de KPC: investigação da associação entre colonização e infecção

Abstract

Introduction: Nosocomial infections are often associated with KPC-producing Klebsiella pneumoniae (Kp-KPC). The gastrointestinal tract acts as an important reservoir for these strains, and previous colonization is an important risk factor for developing the disease. Despite this, the reasons why some patients with similar health status and Kp-KPC carriers progress to infection, while others do not, remain elusive. Objective: The aim of this study was to identify genetic determinants present in Kp-KPC isolates that may contribute to the transition from colonization to infection. Material and Methods: A retrospective study was carried out between May 2018 and June 2021, in which 374 Kp-KPC isolates obtained from surveillance cultures and infection samples were selected for analysis of the antibiotic susceptibility test and molecular typing by XbaI-PFGE. Results: The Kp-KPC population studied showed a wide genetic diversity and resistance to almost all the antibiotics tested. Most of the patients (69.8%) was in the ICU at the time the sample was taken for microbiological analysis, demonstrating that this is a cohort that requires special care. From the XbaI-PFGE typing; 51 isolates were selected for further characterization and subdivided into two groups: 24 Kp-KPC were categorized as colonizers only (ONLYCOL) and 27 as colonizers that transitioned to infection (COLINFEC). Kp-KPC from both groups were selected for whole genome sequencing (WGS) and additional phenotypic assays, such as biofilm production test, antibiotic susceptibility test and ability to infect Galleria mellonella. The antibiotic resistance and biofilm assays showed no significant differences between the ONLYCOL and COLINFEC groups. Multi Locus Sequence Typing (MLST) revealed eight ST-type sequences, with those of the 258 clonal complex (ST 11 and ST 340) predominating in both groups. However, ST 16 was associated with the COLINFEC group (p = 0.034). The blaKPC-2 allele was present in all 51 selected isolates. Yersiniabactin and colibactin toxin (clb-3) were widely representative in both groups. The IncF replicon was prevalent among the isolates and was more associated with the blaKPC-2 gene in the COLINFEC group. Comparative genomic analysis identified seven annotations associated with the COLINFEC group (p < 0.01), including two insertion sequences (IS5 family and ISNCY family), a protease (HtpX), a heat shock protein (IbpA), the Q antitermination protein, phospholipase D and a hypothetical protein. On the other hand, annotations related to the metabolism of metals, salts and minerals were associated with the ONLYCOL group. Kp-KPCs from the COLINFEC group showed greater virulence in G. mellonella larvae in the first 24 hours of infection, and the ST 11-KL64 strain showed high larval mortality, even at low concentrations. Conclusion: In conclusion, this study has reported elements specifically associated with infection, such as ST 16 and seven gene annotations. These findings contribute to the understanding of the molecular mechanisms underlying the pathogenicity of Kp-KPC and may have implications for the development of future interventions aimed at prevention.