Whole genome sequencing snapshot of multi-drug resistant Klebsiella pneumoniae strains from hospitals and receiving wastewater treatment plants in Southern Romania

• Published in: PloS one 2020-01, Vol.15 (1), p.e0228079-e0228079
• Main Authors: Surleac, Marius, Czobor Barbu, Ilda, Paraschiv, Simona, Popa, Laura Ioana, Gheorghe, Irina, Marutescu, Luminita, Popa, Marcela, Sarbu, Ionela, Talapan, Daniela, Nita, Mihai, Iancu, Alina Viorica, Arbune, Manuela, Manole, Alina, Nicolescu, Serban, Sandulescu, Oana, Streinu-Cercel, Adrian, Otelea, Dan, Chifiriuc, Mariana Carmen
• Format: Article
• Language: English
• Subjects: Aminoglycoside antibiotics; Aminoglycosides; Anti-Bacterial Agents - pharmacology; Antibacterial agents; Antibiotic resistance; Antibiotics; Antimicrobial agents; Antiseptics; Bacteria; Beta lactamases; beta-Lactams - metabolism; Biology; Biology and Life Sciences; Carbapenemase; Characterization; Deoxyribonucleic acid; DNA; DNA sequencing; Drug resistance; Drug Resistance, Multiple, Bacterial - drug effects; Drug Resistance, Multiple, Bacterial - genetics; Earth Sciences; Ecology and Environmental Sciences; Effluents; Enzymes; Epidemiology; Fosfomycin; Functional groups; Gene sequencing; Genes; Genes, Bacterial; Genomes; Genomics; Hospitals; Humans; Imipenem; Infectious diseases; Integrons - genetics; Iron; Klebsiella; Klebsiella pneumoniae - classification; Klebsiella pneumoniae - drug effects; Klebsiella pneumoniae - genetics; Klebsiella pneumoniae - pathogenicity; Laboratories; Markers; Medical wastes; Medicine; Medicine and Health Sciences; Meropenem; Microbial drug resistance; Microbial Sensitivity Tests; Multidrug resistance; Multilocus Sequence Typing; Outer membrane proteins; Pathogens; People and places; Pharmacy; Plant sciences; Pneumonia; Quinolones - pharmacology; R&D; Research & development; Rifampin; Romania; Sewage treatment; Strains (organisms); Sulfamethoxazole; Tetracyclines; Virulence; Virulence (Microbiology); Virulence - genetics; Waste Water - microbiology; Wastewater facilities; Wastewater treatment; Wastewater treatment plants; Water; Water analysis; Water Purification; Water quality; Water sampling; Water treatment; Water treatment plants; Whole Genome Sequencing
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0228079

We report on the genomic characterization of 47 multi-drug resistant, carbapenem resistant and ESBL-producing K. pneumoniae isolates from the influent (I) and effluent (E) of three wastewater treatment plants (WWTPs) and from Romanian hospital units which are discharging the wastewater in the sampled WWTPs. The K. pneumoniae whole genome sequences were analyzed for antibiotic resistance genes (ARGs), virulence genes and sequence types (STs) in order to compare their distribution in C, I and E samples. Both clinical and environmental samples harbored prevalent and widely distributed ESBL genes, i.e. blaSHV, blaOXA, blaTEM and blaCTX M. The most prevalent carbapenemase genes were blaNDM-1, blaOXA-48 and blaKPC-2. They were found in all types of isolates, while blaOXA-162, a rare blaOXA-48 variant, was found exclusively in water samples. A higher diversity of carbapenemases genes was seen in wastewater isolates. The aminoglycoside modifying enzymes (AME) genes found in all types of samples were aac(6'), ant(2'')Ia, aph(3'), aaD, aac(3) and aph(6). Quinolone resistance gene qnrS1 and the multi-drug resistance oqxA/B pump gene were found in all samples, while qnrD and qnrB were associated to aquatic isolates. The antiseptics resistance gene qacEdelta1 was found in all samples, while qacE was detected exclusively in the clinical ones. Trimethroprim-sulfamethoxazole (dfrA, sul1 and sul2), tetracyclines (tetA and tetD) and fosfomycin (fosA6, known to be located on a transpozon) resistance genes were found in all samples, while for choramphenicol and macrolides some ARGs were detected in all samples (catA1 and catB3 / mphA), while other (catA2, cmIA5 and aac(6')Ib / mphE and msrE) only in wastewater samples. The rifampin resistance genes arr2 and 3 (both carried by class I integrons) were detected only in water samples. The highly prevalent ARGs preferentially associating with aquatic versus clinical samples could ascribe potential markers for the aquatic (blaSHV-145, qacEdelta1, sul1, aadA1, aadA2) and clinical (blaOXA-1, blaSHV-106,blaTEM-150, aac(3)Iia, dfrA14, oqxA10; oqxB17,catB3, tetD) reservoirs of AR. Moreover, some ARGs (oqxA10; blaSHV-145; blaSHV-100, aac(6')Il, aph(3')VI, armA, arr2, cmlA5, blaCMY-4, mphE, msrE, oqxB13, blaOXA-10) showing decreased prevalence in influent versus effluent wastewater samples could be used as markers for the efficiency of the WWTPs in eliminating AR bacteria and ARGs. The highest number of virulence genes (75) was recorded f