In vivo Selection of Imipenem Resistance Among Ceftazidime-Avibactam-Resistant, Imipenem-Susceptible Klebsiella pneumoniae Isolate With KPC-33 Carbapenemase

We describe in vivo evolution of carbapenem and ceftazidime-avibactam resistance by analyzing four longitudinal Klebsiella pneumoniae clinical isolates from a patient with pneumonia following antimicrobial treatment. The patient had fever, cough associated with expectoration, and new infiltration was found on the chest CT. Antimicrobial susceptibility was determined, and whole genome sequencing (WGS) was performed to investigate its dynamic change of resistance phenotype. Population analysis profile was performed to investigate the population of Klebsiella pneumoniae. The infection started with a KPC-2-producing K. pneumoniae (ZRKP01, ceftazidime-avibactam-S/carbapenem-R). Then, after ceftazidime-avibactam treatment, the strain switched to D179Y mutant that is KPC-33 (ZRKP02, ceftazidime-avibactam-R/carbapenem-S), which restored carbapenem susceptibility. However, the restored carbapenem susceptibility in vivo was not stable and the subsequent use of imipenem against KPC-33-producing K. pneumoniae infection resulted in a reversion of KPC-2 producers (ZRKP03 and ZRKP04, ceftazidime-avibactam-S/carbapenem-R). Genetic analysis demonstrated that all four K. pneumoniae isolates belonged to sequence type 11and had identical capsular polysaccharide (KL47), identical porin genes, and same plasmid replicon types. Phylogenetic analysis indicated that four K. pneumoniae isolates showed a high degree of relatedness. Single nucleotide polymorphisms analysis indicated that the number of mutations observed in the KPC-33 isolate was more than in the wild-type KPC-2 isolates and the four KPC-Kp isolates evolved from a longitudinal evolution of K. pneumoniae harboring bla(KPC-2) gene. This is the first report to observe the in vivo evolution of wild-type KPC-2 to KPC-33 and then the reversion to its original wild-type KPC-2. Through WGS, we demonstrated the role of selective pressure of antibiotic in the mutation and reversion of bla(KPC) genes, which leading to the dynamic change of KPC enzymes and the dynamic emergence of resistance to ceftazidime-avibactam and carbapenems. Statement: Recently, studies reported the emergence of ceftazidime-avibactam-resistant strains. The KPC mutations mediating ceftazidime-avibactam resistance are generally associated with the restoration of carbapenem susceptibility. However, clinical significance of this observation is unclear. In this manuscript, we demonstrate the role of selective pressure of antibiotic in the mutation and reversion of bla(KPC) genes, which leading to the dynamic change of KPC enzymes and the dynamic emergence of resistance to ceftazidime-avibactam and carbapenems. To the best of our knowledge, this is the first report to observe the in vivo evolution of wild-type KPC-2 to KPC-33 and then the reversion to its original wild-type KPC-2. It should be noted that understanding the clinical significance of this observation is of critical importance, and reversion to carbapenem susceptibility would not imply a potential role for carbapenems monotherapy. We hope our study will draw attention to clinicians, so that this agent can be used most effectively for the longest period of time.