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Background: Romania is one of the European countries reporting very high antimicrobial resistance (AMR) rates and consumption of antimicrobials. We aimed to characterize the AMR profiles and clonality of 304 multi-drug resistant (MDR) Acinetobacter baumannii (Ab) and Pseudomonas aeruginosa (Pa) strains isolated during two consecutive years (2018 and 2019) from hospital settings, hospital collecting sewage tanks and the receiving wastewater treatment plants (WWTPs) located in the main geographical regions of Romania. Methods: The strains were isolated on chromogenic media and identified by MALDI-TOF-MS. Antibiotic susceptibility testing and confirmation of ESBL- and CP- producing phenotypes and genotypes were performed. The genetic characterization also included horizontal gene transfer experiments, whole-genome sequencing (WGS), assembling, annotation and characterization. Results: Both clinical and aquatic isolates exhibited high MDR rates, especially the Ab strains isolated from nosocomial infections and hospital effluents. The phenotypic resistance profiles and MDR rates have largely varied by sampling point and geographic location. The highest MDR rates in the aquatic isolates were recorded in Galați WWTP, followed by Bucharest. The Ab strains harbored mostly blaOXA-23, blaOXA-24, blaSHV, blaTEM and blaGES, while Pa strains blaIMP, blaVIM, blaNDM, blaVEB, blaGES and blaTEM, with high variations depending on the geographical zone and the sampling point. The WGS analysis revealed the presence of antibiotic resistance genes (ARGs) to other antibiotic classes, such as aminoglycosides, tetracyclines, sulphonamides, fosfomycin, phenicols, trimethoprim-sulfamethoxazole as well as class 1 integrons. The molecular analyses highlighted: (i) The presence of epidemic clones such as ST2 for Ab and ST233 and ST357 for Pa; (ii) The relatedness between clinical and hospital wastewater strains and (iii) The possible dissemination of clinical Ab belonging to ST2 (also proved in the conjugation assays for blaOXA-23 or blaOXA-72 genes), ST79 and ST492 and of Pa strains belonging to ST357, ST640 and ST621 in the wastewaters. Conclusion: Our study reveals the presence of CP-producing Ab and Pa in all sampling points and the clonal dissemination of clinical Ab ST2 strains in the wastewaters. The prevalent clones were correlated with the presence of class 1 integrons, suggesting that these isolates could be a significant reservoir of ARGs, being able to persist in the environment. [ABSTRACT FROM AUTHOR]

Background: The present study aims the characterization of antibiotic resistance phenotypes and encoding genes in bacterial strains isolated from some Romanian aquatic fishery lowland salted lakes. Material/Methods: This study was conducted on 44 bacterial strains, mainly belonging to species used as microbiological indicators of fecal pollution isolated from four natural fishery lakes. All strains were tested for their antibiotic susceptibility by disk diffusion method. Simplex and multiplex PCR were performed to identify the β-lactams antibiotic resistance genes (blaNMD, blaOXA−48, blaVIM, blaIMP, blaCTX−M, blaTEM), sulfonamides (Sul1, Sul2), tetracyclines (TetA, TetB, TetC, TetD, TetM), aminoglycosides (aac3Ia), vancomycin (VanA, VanB, VanC), macrolides (ermA, ermB, ermC) as well as the plasmid-mediated quinolone resistance (PMQR) markers (QnrA, QnrB, QnrS), and class 1 integrons (Int1, drfA1-aadA1). Results: The Enterococcus spp. isolates exhibited phenotypic resistance to vancomycin (35 %) and macrolides (erythromycin) (75 %); from the vancomycin – resistant strains, 5 % harboured VanA (E. faecalis), while the erythromycin resistant isolates were positive for the ermA gene (E. faecalis − 10 %, E. faecium − 5 %). The Gram- negative rods (GNR) exhibited a high level of resistance to β-lactams: cefuroxime (63 %), cefazolin (42 %), ceftriaxone (8 %), ceftazidime and aztreonam (4 % each). The genetic determinants for beta-lactam resistance were represented by blaCTX−M−like (33 %), blaNDM−like and blaIMP−like (8.33 %) genes. The resistance to non-β-lactam antibiotics was ascertained to the following genes: quinolones (QnrS − 4.16 %); sulfonamides (Sul1–75 %, Sul2–4.16 %); aminoglycosides (aac3Ia − 4.16 %); tetracyclines (tetA – 25 %, tetC − 15 %). The integrase gene was found in more than 50 % of the studied strains (58.33 %). Conclusions: The cultivable aquatic microbiota from fishery lakes is dominated by enterococci and Enterobacterales strains. The GNR strains exhibited high levels of β-lactam resistance mediated by extended spectrum beta-lactamases and metallo-β-lactamases. The Enterococcus sp. isolates were highly resistant to macrolides and vancomycin. The high level and diversity of resistance markers, correlated with a high frequency of integrons is suggesting that this environment could act as an important reservoir of antibiotic resistance genes with a great probability to be horizontally transmitted to other associated species from the aquatic sediments microbiota, raising the potential zoonotic risk for fish consumers. [ABSTRACT FROM AUTHOR]