Investigation of the characterization of polymyxin-and tigecycline-resistant Klebsiella pneumoniae based on whole genome sequencing
Objective This study aimed to investigate the genomic feature of one Klebsiella pneumoniae strain that was resistant to polymyxin and tigecycline.Methods K.pneumoniae KP2016 was isolated from clinical sputum samples from our hospital.The minimum inhibitory concentrations of KP2016 against imipenem,meropenem,polymyxin and tigecycline were determined by the broth microdilution method.The second generation of Illumina and the third generation of Oxford Nanopore whole-genome sequencing were performed.Hybrid assembly of second-and third-generation sequences was performed by Unicycler.ABRicate v0.8.13 was used to analyze drug resistance genes with the Resfinder database.Mobile elements were analyzed via ISFinder.The ST type and plasmid replicon type of the strain were analyzed using the CGE(Center for Genomic Epidemiology)website.Plasmid structures were visualized using Proksee.Results The KP2016 strain was resistant to polymyxin and tigecycline,with MICs of 512 and 16 mg/L,respectively.MLST analysis showed KP2016 belonged to ST656,carrying the polymyxin resistance-related mcr-1 and mcr-8 genes and the tigecycline resistance-related tmexCD1-toprJ1 gene cluster.KP2016 carried one chromosome and five circular plasmids;the size of the plasmid was 3,991～275,345 bp,and the GC content was 44.35％～52.20％.The mcr-1 gene was located on the plasmid p1 of about 44 kb size,and the upstream and downstream environment was ISKpn26-mcr-1-pap2-ISApi1;the mcr-8 gene was located on the IncR/IncN type plasmid p2,and the genetic structure was ISEcl1-mcr-8-orf-ISKpn26;the structure of the tmexCD1-toprJ1 gene cluster was IS26-tnfxB1-tmexC1-tmexcD1-toprJ1.In addition,the strain also carried chromosomal-mediated CrrB mutations(L204V,V237I)related to polymyxin resistance.Conclusion The polymyxin resistance of KP2016 was mediated by mcr-1,mcr-8 and crrB mutations,and the tigecycline resistance was mediated by the tmexCD1-toprJ1 gene cluster.Reasonable surveillance should be strengthened to prevent its further spread in healthcare settings.
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