Molecular Epidemiological Insights into Colistin-Resistant and Carbapenemases-Producing Clinical Klebsiella pneumoniae Isolates

Purpose: Carbapenemases-producing Klebsiella pneumoniae are challenging antimicrobial therapy of hospitalised patients, which is further complicated by colistin resistance. This study describes molecular epidemiological insights into colistin-resistant and carbapenemases-producing clinical K. pneumoniae. Patients and methods: Cultures collected from 26 hospitalised patients during 2014-2017 in the main hospital in Molise Region, central Italy, were characterized. The minimum inhibitory concentration for 19 antibiotics was determined, including carbapenems and colistin. Prevalence of resistance-associated genes was investigated through PCR, detecting bla(KPC), bla(GES), bla(VIM), bla(IMP), bla(NDM), bla(OXA-48), bla(CTX-M), bla(TEM), bla(SHV), and mcr-1,2,3,4,5,6,7,8. The mgrB gene was also analysed in colistin-resistant strains by PCR and sequencing assays. K. pneumoniae were typed by pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST). Results: Twenty out of 26 K. pneumoniae were phenotypically resistant to carbapenems and 19 were resistant to colistin. All isolates harbored bla(KPC), and bla(SHV), bla(TEM) and bla(VIM) were further the most common resistance-associated genes. In colistin-resistant strains, mcr-1,2,3,4,5,6,7,8 variants were not detected, while mutations and insertion elements in mgrB were observed in 68.4% (n=13) in 31.6% (n=6) isolates, respectively. PFGE revealed 12 clusters and 18 pulsotypes at 85% and 95% cut-off, while the Sequence Types ST512 (n=13, 50%), ST101 (n=10, 38.5%), ST307 (n=2, 7.7%) plus a novel ST were detected using MLST. Conclusion: All K. pneumoniae showed a multidrug-resistant phenotype, particularly to carbapenems and colistin. According to national data, b/a(KPC) was the prevailing carbapenemase, followed by bla(VIM), while bla(TEM) and bla(SHV) were among the most frequent beta-lactamases. Consistent with previous reports in Italy, ST512 was the most common clone, particularly during 2014-15, whilst ST101 became dominant in 2016-17. Colistin resistance was mainly associated with deleterious mutations and transposon in the mgrB gene. Improvements of surveillance, compliance with infection prevention procedures and antimicrobial stewardship are essential to limit the spread of resistant K. pneumoniae.