Characterization of an IncFIB/IncHI1B Plasmid Encoding Efflux Pump TMexCD1-TOprJ1 in a Clinical Tigecycline- and Carbapenem-Resistant Klebsiella pneumoniae Strain

Abstract

A novel plasmid-encoded resistance-nodulation-division (RND) efflux pump, TMexCD1- TOprJ1, conferring resistance to multiple drugs, including tigecycline, has recently been identified in Klebsiella pneumoniae strains (1, 2).However, the gene cluster tmexCD1-toprJ1 was rarely identified in clinical Enterobacteriaceae isolates (1).Here, we report a novel plasmid from a clinical K. pneumoniae strain isolated in Hong Kong that harbored this gene cluster and exhibited resistance to tigecycline.A K. pneumoniae strain HKU57 isolate was recovered from the blood culture of a clinical patient in a hospital in Hong Kong in 2017.Antimicrobial susceptibility testing showed that it was resistant to tetracycline and tigecycline; the b-lactam antibiotics aztreonam, cefotaxime, ceftazidime, meropenem, imipenem, and ertapenem; the aminoglycoside antibiotics gentamicin and amikacin; ciprofloxacin; chloramphenicol; and azithromycin but remained susceptible to colistin (see Table 1).Strain HKU57 was subsequently characterized by whole-genome sequencing and in silico data analysis.The complete genome of strain HKU57 was obtained by hybrid assembly of both Illumina and Nanopore reads using Unicycler (3).The isolate harbored a 5.11-Mb chromosome and three plasmids, i.e., pHKU57_1 (273, pHKU57_2 (111, and pHKU57_3 (59,.This strain was found to belong to ST15 by Kleborate based on multilocus sequence typing (MLST) (https://github.com/katholt/Kleborate) (4) and KL48 and O1v1 serotype by Kaptive based on capsule and lipopolysaccharide synthesis loci (5).BLASTn against the virulence genes database (http:// bigsdb.pasteur.fr/klebsiella/klebsiella.html) showed that strain HKU57 harbored virulence genes mrkABCDFHIJ (type 3 fimbriae) and kfuABC (ferric uptake system).This strain did not harbor virulence plasmid-bearing virulence genes, such as iro, iuc, rmpA/ rmpA2.The newly reported RND efflux pump gene cluster tnfxB1-tmexCD1-toprJ1 was located on plasmid pHKU57_1 (Fig. 1a).The MIC of tigecycline testing strain HKU57 was 8 mg/ml (Table 1), which was consistent with the previously reported tnfxB1-tmexCD1-toprJ1 harboring strains (1, 2).Plasmid pHKU57_1 was an IncFIB/IncHI1B plasmid containing multiple resistance genes, including strAB, armA, aph(39)-Ia, qnrB4, sul1, mphA, mphE, msrE, and bla DHA-1 .In BLAST against the NCBI database, this plasmid was found to show the highest similarity (82% coverage, 99% identity) to plasmid pHNWH61-1, a 256,606-bp IncFIB/IncHI1B plasmid recovered from a K. pneumoniae strain, which also harbored the tnfxB1-tmexCD1-toprJ1 gene cluster (Fig. 1a).Another similar plasmid was pF10AN_1, with 73% coverage and 99% identity, from a K. pneumoniae strain, which did not harbor this gene cluster.Plasmid pHKU57_1 shared a

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