Abstract
In this paper we describe the transmission of a multi-drug resistant Klebsiella pneumoniae ST101 clone from hospital to wastewater and its persistence after chlorine treatment. Water samples from influents and effluents of the sewage tank of an infectious diseases hospital and clinical strains collected from the intra-hospital infections, during a period of 10 days prior to wastewater sampling were analyzed. Antibiotic resistant K. pneumoniae strains from wastewaters were recovered on selective media. Based on antibiotic susceptibility profiles and PCR analyses of antibiotic resistance (AR) genetic background, as well as whole-genome sequencing (Illumina MiSeq) and subsequent bioinformatic analyses, 11 ST101 K. pneumoniae strains isolated from hospital wastewater influent, wastewater effluent and clinical sector were identified as clonally related. The SNP and core genome analyses pointed out that five strains were found to be closely related (with ≤18 SNPs and identical cgMLST profile). The strains belonging to this clone harbored multiple acquired AR genes [blaCTX–M–15, blaOXA–48, blaOXA–1, blaSHV–106, blaTEM–150, aac(3)-IIa, aac(6′)-Ib-cr, oqxA10, oqxB17, fosA, catB3, dfrA14, tet(D)] and chromosomal mutations involved in AR (ΔmgrB, ΔompK35, amino acid substitutions in GyrA Ser83Tyr, Asp87Asn, ParC Ser80Tyr). Twenty-nine virulence genes involved in iron acquisition, biofilm and pili formation, adherence, and the type six secretion system – T6SS-III were identified. Our study proves the transmission of MDR K. pneumoniae from hospital to the hospital effluent and its persistence after the chlorine treatment, raising the risk of surface water contamination and further dissemination to different components of the trophic chain, including humans.
Highlights
Due to the worldwide use of antibiotics in the treatment of human and animal infectious diseases, and in livestock and agriculture, a large amount of antibiotics of pharmaceutical origin are found in anthropic environments, such as sewage and wastewater treatment plants (WWTPs) and ends up being discharged in the natural environment (Kraemer et al, 2019; Zhang T. et al, 2019; Zhang Z. et al, 2019)
For the 31 strains selected, the antibiotic susceptibility profiles, the genetic background of β-lactam resistance and ST-types were compared, (Supplementary Table 3) revealing the presence of matching patterns of antibiotic resistance (AR) profiles, the presence/absence of ARGs and the abundance of K. pneumoniae ST101 The PCR for detection of β-lactam resistance genes revealed the presence of carbapenemases genes, which were, in decreasing frequency order, blaOXA−48, blaNDM, and blaKPC as well as of the blaCTX−M encoding for ESBLs
The main characteristics of the selected K. pneumoniae ST101 strains were: the presence of the multi-drug resistance (MDR) phenotype, three wastewater-sourced strains being resistant to all tested antibiotics (22 bac, 23 bac, 34bac), while the other two were susceptible only to aztreonam (29 bac, 82 bac)
Summary
Due to the worldwide use of antibiotics in the treatment of human and animal infectious diseases, and in livestock and agriculture, a large amount of antibiotics of pharmaceutical origin are found in anthropic environments, such as sewage and wastewater treatment plants (WWTPs) and ends up being discharged in the natural environment (Kraemer et al, 2019; Zhang T. et al, 2019; Zhang Z. et al, 2019). Water plays a crucial role in the spreading of AR, through inappropriate release of human and animal effluents in the surface waters through hospital wastewaters, WWTPs, aquaculture farms, surface, and groundwater. Hospital wastewaters are highly complex effluents, carrying a wide range of micro- and macropollutants, including antibiotic compounds, metabolized drugs, disinfectants, patient excrements, and microorganisms. The presence of ARB as well as of antibiotic residues, which could inhibit the growth of susceptible bacteria, are thereby increasing the population of resistant bacteria in the receiving water (Kaur et al, 2020; Rozman et al, 2020)
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