Adaptation to pandrug resistance of clinical strain E. coli M19736 ST615

CARRERA PÁEZ LC; KNECHT C; GONZALES MACHUCA A; CARPIO DÍAZ E; VARGAS CV; ÁLVAREZ VE; PIEKAR M; DONIS, N; QUIROGA MP; CENTRÓN D

Congreso; VII Congreso de Bioquímica y Biología Molecular de Bacterias; 2023

Escherichia coli could acquire and harbor multiple mobile genetic elements (MGE) associated to antimicrobial genes (ARG) that contribute to the phenotypes of multidrug, extreme and pandrug resistance (MDR, XDR, PDR). These genetic elements can be transferred by conjugation, transformation, transduction, and by the most recent mechanism discovered named outer membrane vesicles (OMV) within bacterial communities. In the present study, we used as biological model one of the first clinical MDR colistin resistant isolate from Argentina, E. coli M19736 which harbors a plasmid with a mcr-1 gene. We studied its ability to receive crucial ARG by transformation and conjugation. We used as donors several plasmids from different genera from clinical isolates sequenced by Illumina technology: i) pDCAG-1, (>112.000 pb, IncFII) that harbors blaCTX-M-15, ii) pDCCK1-KPC (>77.218 pb, IncM1) that harbors blaKPC-2, iii) pDCVA3 (IncFII) that harbors blaNDM-5, iv) pDCASG6-NDM (137.269 pb, IncC) that harbors blaNDM-1, and v) paadB (5877 bp). Gene acquisition was verified by PCR, minimum inhibitory concentration, and phenotypic detection of beta-lactamases. Then, each transconjugant and transformant was evaluated for its ability to maintain the ARG on the first and 10th days after to being subcultured without antibiotic pressure. E. coli M19736 acquired blaCTX-M-15, blaKPC-2, blaNDM-5, blaNDM-1, aadB and maintained these ARG 100%, 73,3%, 82,20, 100% and 100%, at the 10th day of subculture, respectively. When evolved PDR E. coli M19736 acquired progressively blaCTX-M-15, blaNDM-1 and aadB genes, a different pattern of maintenance was found with 6,6%, 98,8% and 100%, respectively, on the first day of subcultured. On the other hand, we isolated and characterized the OMVs from native and evolved E. coli M19736 by transmission electron microscopy, and dynamic light scattering that showed particles with a size between 100–300 nm in diameter. By PCR the mcr-1 gene was detected within the OMV in both native and evolved strains. OMV from evolved E. coli M19736 also harbored blaCTX-M-15 and aadB genes. Besides, by Liquid chromatography–MS/MS we studied the proteins that are inside of the OMVs. Proteomics analysis showed that the triplicate of the samples was enriched by biological processes and molecular functions sharing 31,4% of the proteins found including the presence of the MCR-1 protein in one of them. These results showed the genetic plasticity of a sporadic clone of E. coli showing that this kind of isolates could play a very important transitional link in the clinical dynamics and evolution to XDR phenotypes within the nosocomial niche by acting as reservoir of ARG that in turn could disseminate by several mechanisms of the Horizontal Genetic Transfer.