Emergence of a ST307 clone carrying a novel insertion element MITEKpn1 in the mgrB gene among carbapenem-resistant Klebsiella pneumoniae from Moscow, Russia.

Carbapenem-resistant Klebsiella pneumoniae (CRKP) represents a major nosocomial pathogen with only a few antimicrobial agents, including colistin, remaining active. However, the emergence of colistin-resistant (Col-R) isolates is compromising the activity of colistin. In this study, a collection of 159 CRKP recovered from three hospitals in Moscow (Russian Federation) was examined. The isolates demonstrated resistance to cephalosporins (100%), ciprofloxacin (92.5%), fosfomycin (90.1%), netilmicin (81.1%), gentamicin (84.3%) and amikacin (49.7%). The rate of colistin resistance (MIC > 2 mg/L by broth microdilution) was 44.7%; moreover, 6.7% of isolates were tigecycline-resistant. Among 18 sequence types (STs) discovered, isolates of five lineages including ST307 (n = 46; 28.9%), ST395 (n = 40; 25.2%), ST377 (n = 17; 10.7%), ST48 (n = 17; 10.7%) and ST23 (n = 16; 10.1%) dominated. Carriage of a blaOXA-48-like carbapenemase gene was detected in 146 CRKP (91.8%); 11 (6.9%) and 2 (1.3%) isolates harboured blaNDM-1 and blaKPC-3, respectively. Among 71 Col-R isolates, colistin MICs ranged from 4 mg/L to >1024 mg/L (MIC50/90, 2/512 mg/L). All Col-R isolates were mcr-1-negative. In 19 (26.8%) Col-R isolates, inactivation of mgrB by insertion sequences IS1A, IS1R, ISKpn14 and ISKpn26 and a novel miniature inverted-repeat transposable element (MITE) Kpn1 was observed. Carriage of MITEKpn1 was restricted to six ST307 isolates and affected mgrB at nucleotide position 75. mgrB deletion was observed in four (5.6%) Col-R isolates. Moreover, PmrA and/or PmrB were altered in three (4.5%) Col-R isolates with wild-type mgrB. Thus, blaOXA-48-like-carrying Col-R ST307 K. pneumoniae is emerging as a dominant clone in Moscow.

[The comparison of methods for determination of colistin susceptibility in carbapenem-resistant Klebsiella pneumoniae.]

In recent years, because of carbapenemase spreading in Klebsiella pneumoniae strains, the antibiotic of reserve group, colistin, is increasingly prescribed. In vitro testing of colistin susceptibility in everyday practice has a number of difficulties due to the cationic properties of molecule and weak diffusion into agar. Therefore it is recommended to use the reference Broth Microdilution Method (BMD) for determination of the Minimum Inhibitory Concentration (MIC) for colistin. The purpose of the study was to determine susceptibility to colistin in 119 carbapenem-resistant K. pneumoniae (CRKp) which were isolated from the patients at three hospitals in Moscow in 2012-2016 by the broth microdilution method (BMD) and to compare these data with the ones obtained by epsilometer test (E-test) and VITEK 2 Compact. The proportion of resistant isolates (MIC>2 mg/L) was 52%, 39%, 35% respectively. Both commercial methods demonstrated a high level of the very major error (VME) that was 26% for the E-test method and 34% for the VITEK 2 Compact. The values of categorical agreement and essential agreement (CA, EA) were less than 90%. A single major error (ME) was detected for the VITEK 2 Compact. In conclusion, results of both commercial tests for determination of MIC for colistin showed differences with the results of the reference BMD. It is necessary for clinical laboratories to be aware about this discrepancy and to use E-tests and VITEK 2 Compact with caution to determine colistin susceptibility.