Analysis of drug resistance genes and molecular typing of carbapenem-resistant Klebsiella variicola strains
Objective 
To investigate the prevalence and drug resistance of clinical Klebsiella variicola (K.variicola) isolates and to illuminate the mechanism of drug resistance in carbapenem-resistant strains. 
 
 
Methods 
Clinical K. variicola isolates were identified with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). The antimicrobial susceptibility profile of these strains was determined using broth microdilution. Resistance genes carried by carbapenem-resistant K. variicola strains were detected by PCR with specific primers. Multilocus sequence typing (MLST) was used for molecular typing. A pan-drug resistant strain which was isolated from cerebrospinal fluid sample was analyzed with whole genome sequencing (WGS). 
 
 
Results 
Twenty-six isolates were identified as K. variicola by MALDI-TOF MS. Results of the antimicrobial susceptibility test showed that there were 15.4% (4/26) resistant to carbapenem and 11.5% (3/26) unsusceptible to tigecycline. These strains were highly susceptible to amikacin and gentamicin, which accounted for 96.2% (25/26). As for the third- and fourth-generation cephalosporins, the resistance rate was 23.1% (6/26). All of the four carbapenem-resistant isolates carried the resistance genes of blaIMP-4, qnrA/B and blaTEM, and one of them was also positive for blaNDM-1 gene. The fosfomycin resistance gene, fosA, was detected in three of them. Molecular typing analysis indicated these isolates belonged to two sequence types (ST) of ST357 (three strains) and ST1737 (one strain). Two plasmids were obtained from the pan-drug resistant strain by WGS, including IncFⅡ/FIB(k) type plasmid (160 kb) that was highly homologous to LMG 23571 plasmid (GenBank: CP013986.1) and IncHⅠ1B/FIB type plasmid (260 kb) sharing high homology with pIMP4_LL34 (GenBank: CP025964.1). Besides the resistance genes mentioned above, the two plasmids also carried a variety of other genes that mediated the resistance to aminoglycosides (strB, strA, armA, aac3-Ⅱd, aadA2), macrolides (msrE, mphE), chloramphenicol (catA2), sulfonamides (sulⅠ) tigecycline (tetA variant) and trimethoprim (dfrA16). However, no virulence genes were detected. 
 
 
Conclusions 
In general, the resistance profile of K. variicola was similar to that of Klebsiella pneumoniae, but the differences were that carbapenem-resistant K. variicola strains mainly belonged to ST357 and the leading causes of resistance were carrying the genes encoding IMP-4 and NDM-1 metal β-lactamases. WGC analysis revealed that the pan-drug resistant K. variicola strain carried multiple drug resistance genes without virulence determinants, which might be resulted from the evolution of drug resistance. 
 
 
Key words: 
Klebsiella variicola; Whole genome sequencing; Carbapenem resistance