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

CARRERA PAEZ LAURA; KNECHT CAMILA; GONZALEZ MACHUCA ADRIAN; CARPIO DIAZ, EDUARDO; VARGAS, CLAUDIA; ALVAREZ, VERONICA ELIZABETH; PIEKAR MARIA; DONIS NICOLAS; GAMBINO ANAHI SAMANTA; QUIROGA MARIA PAULA; CENTRON DANIELA

Congreso; 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 ofmultidrug, extreme and pandrug resistance (MDR, XDR, PDR). These geneticelements can be transferred by conjugation, transformation, transduction, and bythe most recent mechanism discovered named outer membrane vesicles (OMV)within bacterial communities. In the present study, we used as biological modelone of the first clinical MDR colistin resistant isolate from Argentina, E. coli M19736which harbors a plasmid with a mcr-1 gene. We studied its ability to receive crucialARG by transformation and conjugation. We used as donors several plasmids fromdifferent genera from clinical isolates sequenced by Illumina technology: i) pDCAG-1, (>112.000 pb, IncFII) that harbors bla CTX-M-15, ii) pDCCK 1 -KPC (>77.218 pb, IncM1)that harbors bla KPC-2 , iii) pDCVA3 (IncFII) that harbors bla NDM-5, iv) pDCASG6-NDM(137.269 pb, IncC) that harbors bla NDM-1 , and v) paadB (5877 bp) . Gene acquisitionwas verified by PCR, minimum inhibitory concentration, and phenotypic detectionof beta-lactamases. Then, each transconjugant and transformant was evaluated forits ability to maintain the ARG on the first and 10 th days after to being subculturedwithout antibiotic pressure. E. coli M19736 acquired bla CTX-M-15, bla KPC-2 , bla NDM-5, bla NDM-1,aadB and maintained these ARG 100%, 73,3%, 82,20, 100% and 100%, at the 10 thday of subculture, respectively. When evolved PDR E. coli M19736 acquiredprogressively bla CTX-M-15, bla NDM-1 and aadB genes, a different pattern of maintenancewas found with 6,6%, 98,8% and 100%, respectively, on the first day ofsubcultured. On the other hand, we isolated and characterized the OMVs fromnative and evolved E. coli M19736 by transmission electron microscopy, anddynamic light scattering that showed particles with a size between 100–300 nm indiameter. By PCR the mcr-1 gene was detected within the OMV in both native andevolved strains. OMV from evolved E. coli M19736 also harbored bla CTX-M-15 andaadB genes. Besides, by Liquid chromatography–MS/MS we studied the proteinsthat are inside of the OMVs. Proteomics analysis showed that the triplicate of thesamples was enriched by biological processes and molecular functions sharing31,4% of the proteins found including the presence of the MCR-1 protein in one ofthem. These results showed the genetic plasticity of a sporadic clone of E. colishowing that this kind of isolates could play a very important transitional link in theclinical dynamics and evolution to XDR phenotypes within the nosocomial niche byacting as reservoir of ARG that in turn could disseminate by several mechanismsof the Horizontal Genetic Transfer.