A novel self-transmissible mega plasmid from extensively drug-resistant Klebsiella oxytoca carries multiple antimicrobial resistance genes and acts as a resistance reservoir.

Klebsiella oxytoca is an opportunistic pathogen linked to hospital-acquired infections, with extensively drug-resistant (XDR) strains posing serious treatment challenges. Here, an XDR K. oxytoca 611 isolated from hospital wastewater was found to carry 59 antimicrobial resistance genes (ARGs) compared to the average of six ARGs for K. oxytoca. Twenty-eight of these ARGs were on a mega hybrid plasmid (pKO611.1) carrying C, HI2 and HI2A replicons. Among the resistance genes carried on pKO611.1 was a new tetracycline efflux pump tet(E) variant and an uncharacterised AmpC β-lactamase gene. Cloning and characterising of these genes in Escherichia coli confirmed their ability to confer resistance to tetracyclines and β-lactams, respectively. Conjugation between K. oxytoca and E. coli transferred pKO611.1 with high efficiency. Propagation of plasmid pKO611.1 conferred multi-drug resistance to E. coli. Attempts to cure pKO611.1 from K. oxytoca produced a shortened version of the plasmid where a 63,815 bp segment, containing several resistance genes, was excised. Accordingly, K. oxytoca harbouring the shortened version of the plasmid, did not display resistance to the associated antibiotics. The ease of inter-genera transmission of pKO611.1 and its ability to recombine indicates its potential role as a reservoir for dissemination of multiple ARGs.