Ceftazidime-avibactam tolerance and persistence among difficult-to-treat KPC-producing Klebsiella pneumoniae clinical isolates from bloodstream infections

Eur J Clin Microbiol Infect Dis. 2025 Feb;44(2):343-353. doi: 10.1007/s10096-024-05005-4. Epub 2024 Nov 30.

Abstract

Purpose: Tolerance and persistence occur "silently" in bacteria categorized as susceptible by antimicrobial susceptibility testing in clinical microbiology laboratories. They are different from resistance phenomena, not well-studied, and often remain unnoticeable. We aimed to investigate and characterize ceftazidime-avibactam (CZA) tolerance/persistence in 80 Klebsiella pneumoniae isolates from bloodstream infections.

Methods: We used the Tolerance Disk Test (TDtest) to detect CZA tolerance/persistence and investigate the avibactam (AVI) influence on them, and time-kill assays with minimal duration for killing (MDK) determination to characterize/differentiate CZA tolerance from persistence, for selected isolates. Whole genome sequencing was performed for 49/80 selected isolates to investigate genes related to beta-lactam tolerance/persistence and resistance as well as phylogeny studies.

Results: Tolerance/persistence to CZA was detected in 48/80 (60%) isolates, all extensively drug-resistant (XDR) or multidrug-resistant, carbapenem-resistant K. pneumoniae (CRKp), KPC producers, and previously categorized as susceptible (not resistant) to CZA. No heteroresistance was detected. CZA tolerance/persistence occurred due to ceftazidime tolerance/persistence and was not related to AVI in the CZA combination. 5/11 isolates were characterized as CZA-tolerant and 5/11 as CZA-persistent. The single (1/11) XDR and CRKp non-KPC producer was truly susceptible. All the CZA-tolerant/persistent isolates (ST11, ST258, ST340, ST437, ST16, ST17, and ST307) harbored the carbapenemase-encoding gene blaKPC-2. Mutation in only two genes (rpoS and degQ) related to beta-lactam tolerance/persistence was found in only 7/49 CZA-tolerant/persistent isolates, suggesting the presence of yet unknown beta-lactam tolerance/persistence genes.

Conclusion: Among the K. pneumoniae bloodstream isolates studied, 60%, previously categorized as susceptible to CZA, were, actually, tolerant/persistent to this antibiotic, all these KPC producers.

Keywords: Antibiotic resistant bacteria; Minimal duration for killing (MDK); Time-kill assays; Tolerance disk test (TDtest).

MeSH terms

  • Anti-Bacterial Agents* / pharmacology
  • Azabicyclo Compounds* / pharmacology
  • Bacteremia* / drug therapy
  • Bacteremia* / microbiology
  • Bacterial Proteins / genetics
  • Ceftazidime* / pharmacology
  • Drug Combinations
  • Drug Resistance, Multiple, Bacterial / genetics
  • Humans
  • Klebsiella Infections* / drug therapy
  • Klebsiella Infections* / microbiology
  • Klebsiella pneumoniae* / drug effects
  • Klebsiella pneumoniae* / enzymology
  • Klebsiella pneumoniae* / genetics
  • Klebsiella pneumoniae* / isolation & purification
  • Microbial Sensitivity Tests
  • Whole Genome Sequencing
  • beta-Lactamases* / genetics
  • beta-Lactamases* / metabolism

Substances

  • Ceftazidime
  • Azabicyclo Compounds
  • avibactam, ceftazidime drug combination
  • Drug Combinations
  • Anti-Bacterial Agents
  • beta-Lactamases
  • Bacterial Proteins