Convergence of MCR-8.2 and Chromosome-Mediated Resistance to Colistin and Tigecycline in an NDM-5-Producing ST656 Klebsiella pneumoniae Isolate From a Lung Transplant Patient in China.

We characterized the first NDM-5 and MCR-8.2 co-harboring ST656 Klebsiella pneumoniae clinical isolate, combining with chromosomal gene-mediated resistance to colistin and tigecycline. The K. pneumoniae KP32558 was isolated from the bronchoalveolar lavage fluid from a lung transplant patient. Complete genome sequences were obtained through Illumina HiSeq sequencing and nanopore sequencing. The acquired resistance genes and mutations in chromosome-encoded genes associated with colistin and tigecycline resistance were analyzed. Comparative genomic analysis was conducted between mcr-8.2-carrying plasmids. The K. pneumoniae KP32558 was identified as a pan-drug resistant bacteria, belonging to ST656, and harbored plasmid-encoded blaNDM-5 and mcr-8.2 genes. The blaNDM-5 gene was located on an IncX3 type plasmid. The mcr-8.2 gene was located on a conjugative plasmid pKP32558-2-mcr8, which had a common ancestor with another two mcr-8.2-carrying plasmids pMCR8_020135 and pMCR8_095845. The MIC of KP32558 for colistin was 256 mg/L. The mcr-8.2 gene and mutations in the two-component system, pmrA and crrB, and the regulator mgrB, had a synergistic effect on the high-level colistin resistance. The truncation in the acrR gene, related to tigecycline resistance, was also identified. K. pneumoniae has evolved a variety of complex resistance mechanisms to the last-resort antimicrobials, close surveillance is urgently needed to monitor the prevalence of this clone.

Emergence of an ST1326 (CG258) Multi-Drug Resistant Klebsiella pneumoniae Co-harboring mcr-8.2, ESBL Genes, and the Resistance-Nodulation-Division Efflux Pump Gene Cluster tmexCD1-toprJ1 in China.

CG258 is the dominant carbapenemase-producing Klebsiella pneumoniae clone worldwide and treatment of infections caused by this clone relies largely on the last-line antibiotics, colistin, and tigecycline. However, the emergence and global dissemination of mcr and tmexCD1-toprJ1 genes have significantly compromised their clinical applications. CG258 K. pneumoniae carrying both mcr and tmexCD1-toprJ1 have not been reported. A colistin-resistant strain T698-1 belonging to ST1326, a member of CG258, was isolated from the intestinal sample of a patient and characterized by the antimicrobial susceptibility testing, conjugation assay, WGS and bioinformatics analysis. It was resistant to colistin, tetracycline, aminoglycoside, fluoroqinolone, phenicols, sulfonamide, and some ß-lactams, and positive for mcr-8.2, tmexCD1-toprJ1, and ESBL genes (bla DHA-1 and bla CTX-M-15). The tmexCD1-toprJ1 gene cluster was located in an multi-drug resistant (MDR) region flanked by TnAs1 elements on an IncHI1B/FIB plasmid. The genetic context of tmexCD1-toprJ1 was slightly distinct from previously reported Tn5393-like structures, with an IS26 element disrupting the upstream Tn5393 and its adjacent genetic elements. The mcr-8.2 gene was inserted into the backbone of an IncFII/FIA plasmid with the genetic context of ISEcl1-mcr-8.2-orf-ISKpn26. To our knowledge, this is the first report of co-occurrence of mcr-8.2 and tmexCD1-toprJ1 in a CG258 K. pneumoniae strain. Though this strain is tigecycline sensitive, the acquisition of colistin and tigecycline resistance determinants by the endemic CG258 K. pneumoniae clone still poses a serious public health concern. CG258, which became resistant to multiple last resort antibiotics, would be the next emerging superbug.

Characterization of a Novel mcr-8.2-Bearing Plasmid in ST395 Klebsiella pneumoniae of Chicken Origin.

The emergence of mobile colistin resistance mcr genes undermines the efficacy of colistin as the last-resort drug for multi-drug resistance infections and constitutes a great public health concern. Plasmids play a critical role in the transmission of mcr genes among bacteria. One colistin-resistant Klebsiella pneumoniae strain of chicken origin was collected and analyzed by antimicrobial susceptibility testing, PCR, conjugation assay and S1-PFGE. Whole-genome sequencing (WGS) approach combining Illumina and MinION platforms was utilized to decipher the underlying colistin resistance mechanism and genetic context. A novel mcr-8.2-bearing plasmid p2019036D-mcr8-345kb with 345 655 bp in size encoding various resistance genes including floR, sul1, aadA16, aadA2, bla CTX-M-27, bla DHA-1, tet(D), dfrA12 and qnrB4 was identified responsible for the colistin resistance phenotype. Plasmid comparison has shown that the mcr-8.2-bearing plasmid differed from other reported plasmids positive for mcr-8.2 but shared the same core mcr-8.2-bearing conserved region. This study demonstrates the emergence of mcr-8.2-bearing K. pneumoniae of animal origin is a potential risk to humans.