Emergence of Ceftazidime-Avibactam Resistance Due to Plasmid-Borne blaKPC-3 Mutations during Treatment of Carbapenem-Resistant Klebsiella pneumoniae Infections

Abstract

Ceftazidime-avibactam is a novel β-lactam/β-lactamase inhibitor with activity against carbapenem-resistant Enterobacteriaceae (CRE) that produce Klebsiella pneumoniae carbapenemase (KPC). We report the first cases of ceftazidime-avibactam resistance to develop during treatment of CRE infections and identify resistance mechanisms. Ceftazidime-avibactam-resistant K. pneumoniae emerged in three patients after ceftazidime-avibactam treatment for 10 to 19 days. Whole-genome sequencing (WGS) of longitudinal ceftazidime-avibactam-susceptible and -resistant K. pneumoniae isolates was used to identify potential resistance mechanisms. WGS identified mutations in plasmid-borne bla_KPC-3, which were not present in baseline isolates. bla_KPC-3 mutations emerged independently in isolates of a novel sequence type 258 sublineage and resulted in variant KPC-3 enzymes. The mutations were validated as resistance determinants by measuring MICs of ceftazidime-avibactam and other agents following targeted gene disruption in K. pneumoniae, plasmid transfer, and bla_KPC cloning into competent Escherichia coli In rank order, the impact of KPC-3 variants on ceftazidime-avibactam MICs was as follows: D179Y/T243M double substitution > D179Y > V240G. Remarkably, mutations reduced meropenem MICs ≥4-fold from baseline, restoring susceptibility in K. pneumoniae from two patients. Cefepime and ceftriaxone MICs were also reduced ≥4-fold against D179Y/T243M and D179Y variant isolates, but susceptibility was not restored. Reverse transcription-PCR revealed that expression of bla_KPC-3 encoding D179Y/T243M and D179Y variants was diminished compared to bla_KPC-3 expression in baseline isolates. In conclusion, the development of resistance-conferring bla_KPC-3 mutations in K. pneumoniae within 10 to 19 days of ceftazidime-avibactam exposure is troubling, but clinical impact may be ameliorated if carbapenem susceptibility is restored in certain isolates.

Keywords: Klebsiella pneumoniae; Klebsiella pneumoniae carbapenemase; carbapenem-resistant Enterobacteriaceae; ceftazidime-avibactam; resistance; sequence type 258.

FIG 1

Time courses of infection and treatment among patients in whom ceftazidime-avibactam-resistant K. pneumoniae emerged. Black and shaded bars represent duration of treatment with respective antibiotics. *, interpretation of K. pneumoniae phenotype, as reported by the clinical microbiology laboratory. Abbreviations: CRE, carbapenem-resistant Enterobacteriaceae; CTB, ceased to breathe; ESBL, extended-spectrum β-lactamase; IAI, intra-abdominal infection; PNA, pneumonia; UTI, urinary tract infection.

FIG 2

Phylogenetic comparison of ST258, clade II K. pneumoniae isolates from our center and others in the United States. The phylogenetic tree, based on whole-genome SNP analysis, was generated with the use of the maximum-likelihood optimality criterion. Branch lengths are proportional to the number of evolutionary changes, and all nodes were supported by 100% bootstrap. Ten K. pneumoniae isolates from the present study and 22 isolates collected from hospitals in New York, New Jersey, Pennsylvania, Maryland, and Michigan in previous studies were included (15, 37, 39). Isolates from our medical center are presented in yellow ovals. Isolates in cluster a are from the present study, and isolates in cluster b were collected from our center in 2010 to 2011.

FIG 3

SNP matrix of the 10 K. pneumoniae isolates from the present study. Numbers of SNPs for each pairwise comparison of isolates are shown.