From genomics to treatment: overcoming pan-drug-resistant Klebsiella pneumoniae in clinical settings

Abstract

Introduction: The spread pan-drug resistant pathogens pose a critical challenge to current therapies, resulting in high mortality and necessitating alternative approaches.

Methods: We report pan-drug resistant Klebsiella pneumoniae isolates from five patients in a single hospital, including resistance to cefiderocol and cefepime-zidebactam in one isolate.

Results: Whole-genome sequencing identified blaNDM-5 and blaCTX-M-15 genes in all isolates, explaining carbapenemase and extended-spectrum β- lactamase phenotypes, with blaKPC-2 in one isolate. A novel sulfhydryl variable β-lactamase (SHV) variant, blaSHV-231, was present in all isolates under a strong promoter. Two isolates exhibited a non-synonymous mutation in fstI encoding PBP3, the primary target of aztreonam in Gram-negative bacteria. Genomic and phenotypic characterization guided successful compassionate treatment using aztreonam, ceftazidime-avibactam, and amoxicillin-clavulanate at maximum doses.

Discussion: Dissection of the roles of the substitutions present in blaSHV-231 revealed that this variant was responsible for the reduced susceptibility to aztreonam-avibactam, at the expense of a higher susceptibility to clavulanate. Targeted therapy can be successful upon dissection of unexpected mechanisms of resistance that enhance the contribution of endemic β-lactamase.

Keywords: Klebsiella pneumoniae; NDM carbapenemase; aztreonam-avibactam; cefepime-zidebactam; cefiderocol; emerging pathogens; multi-drug resistance; pan-drug resistance.

FIGURE 1

Resistance genes and/or chromosomal mutations in pan-drug resistant isolates obtained by WGS. Red dots indicate the presence of the indicated gene or mutation.

FIGURE 2

Alignment of the amino acid sequences of selected SHV variants. Conserved residues of the ß-lactamase are denoted in blue while mutations are highlighted in white.

FIGURE 3

Susceptibility to selected antimicrobial agents alone or in combinations against PDR K. pneumoniae isolates. Avibactam, relebactam, clavulanic acid and rifampin were tested at fixed concentration of 4 mg/L, ceftazidime at fixed concentration of 8 mg/L, ertapenem and fosfomycin at fixed concentration of 10 mg/L and dipicolinic acid at fixed concentration of 1,000 µM. ND: not determined. Abbreviations. XDR: pan-drug resistant. Kpn: Klebsiella pneumoniae. The color-coded scale in the table indicates the log₂ (dils.) difference between MICs for β-lactam with the inhibitor in relation to MICs for β-lactam alone (aztreonam or cefiderocol) for each assessed clinical strain, following the provided reference in Table.

FIGURE 4

Susceptibility testing of recombinant E. coli strains Minimum Inhibitory Concentrations (MICs) were determined for E. coli DH5α recombinants carrying either the empty pMBL plasmid or bla _SHV-1 with the specified mutations. The mutation combinations resulting in bla _SHV-1, bla _SHV-5, bla _SHV-84 and bla _SHV-231 are denoted in parentheses. Clavulanate and avibactam were held at a fixed final concentration of 4 mg/L. The color-coded scale in the table indicates the log₂ difference (dils.) between MICs for β-lactam with the inhibitor in relation to MICs for β-lactam alone for each assessed recombinant strain, following the provided reference in Table.