Genomic Diversity of NDM-Producing Klebsiella Species from Brazil, 2013–2022

Background: Since its first report in the country in 2013, NDM-producing Enterobacterales have been identified in all the Brazilian administrative regions. In this study, we characterized by antimicrobial susceptibility testing and by molecular typing a large collection of NDM-producing Klebsiella isolates from different hospitals in Brazil, mainly from the state of Sao Paulo, over the last decade. Methods: Bacterial isolates positive for bla(NDM)-genes were identified by MALDI-TOF MS and submitted to antimicrobial susceptibility testing by disk diffusion or broth microdilution (for polymyxin B). All isolates were submitted to pulsed-field gel electrophoresis, and isolates belonging to different clusters were submitted to whole genome sequencing by Illumina technology and downstream analysis. Mating out assays were performed by conjugation, plasmid sizes were determined by S1-PFGE, and plasmid content was investigated by hybrid assembly after MinIon long reads sequencing. Results: A total of 135 NDM-producing Klebsiella were identified, distributed into 107 different pulsotypes; polymyxin B was the only antimicrobial with high activity against 88.9% of the isolates. Fifty-four isolates presenting diversified pulsotypes were distributed in the species K. pneumoniae (70%), K. quasipneumoniae (20%), K. variicola (6%), K. michiganensis (a K. oxytoca Complex species, 2%), and K. aerogenes (2%); bla(NDM-1) was the most frequent allele (43/54, 80%). There was a predominance of Clonal Group 258 (ST11 and ST340) encompassing 35% of K. pneumoniae isolates, but another thirty-one different sequence types (ST) were identified, including three described in this study (ST6244 and ST6245 for K. pneumoniae, and ST418 for K. michiganensis). The bla(NDM-1) and bla(NDM-7) were found to be located into IncF and IncX3 type transferable plasmids, respectively. Conclusions: Both clonal (mainly driven by CG258) and non-clonal expansion of NDM-producing Klebsiella have been occurring in Brazil in different species and clones, associated with different plasmids, since 2013.