Global phylogeography and genetic characterization of carbapenem and ceftazidime-avibactam resistant KPC-33-producing Pseudomonas aeruginosa.

Abstract

Ceftazidime-avibactam (CZA) is currently one of the last resorts used to treat infections caused by carbapenem-resistant Enterobacteriaceae and Pseudomonas aeruginosa. However, KPC variants have become the main mechanism mediating CZA resistance in KPC-producing gram-negative bacteria after increasing the application of CZA. Our previous study revealed that CZA-resistant KPC-33 had emerged in carbapenem-resistant P. aeruginosa (CRPA) and had resulted in death due to hypervirulence and extensive drug resistance; however, the evolutionary path of KPC-33-producing CRPA has not been investigated. Here, we observed the emergence of bla_KPC-33 in CRPA under drug pressure, leading to resistance to CZA. We further elucidated the pathway of resistance development due to bla_KPC mutations in P. aeruginosa. Three KPC-producing P. aeruginosa (KPC-PA) strains (including one bla_KPC-33-positive strain and two bla_KPC-2-positive strains) were successively isolated from a hospitalized patient. The bla_KPC-33-positive CZA-resistant strain SRPA0656 (CZA MIC >128 μg/mL, imipenem MIC = 32 μg/mL) was isolated after the bla_KPC-2-positive P. aeruginosa SRP2863 (CZA MIC = 1 μg/mL, imipenem MIC >128 μg/mL) was treated with CZA. The subsequent use of carbapenems to treat the infection led to the re-emergence of the KPC-2-producing strain SRPA3703. Additionally, we collected four other KPC-33-producing P. aeruginosa strains. Antimicrobial susceptibility testing revealed that all the KPC-33-bearing P. aeruginosa strains in this study were multidrug-resistant but susceptible to colistin and amikacin. Whole-genome sequencing indicated that bla_KPC-33 was located on two Tn4401-like transposons contained in the plasmids and that most of these plasmids could be transferred into P. aeruginosa PAO1^Rif isolates. Growth rate determination demonstrated that the relative growth rate of P. aeruginosa harboring bla_KPC-33 was faster than that of P. aeruginosa harboring bla_KPC-2 in the logarithmic phase. Global phylogenetic analysis revealed that most KPC-PA strains were isolated from China and the USA. MLST revealed that the most common ST in KPC-PA was ST463, which was detected only in China, and that all the strains carried bla_KPC-2 or its derivatives. These results indicated that the use of CZA for the treatment of KPC-2-producing P. aeruginosa may have contributed to the evolution of KPC-33. The widespread dissemination of KPC-PA (especially the ST463) and Tn4401 transposons may increase the spread of CRPA isolates carrying bla_KPC-33. Close attention to the development of resistance to CZA during clinical treatment of CRPA infection and monitoring CZA-resistant strains is necessary to prevent further spread.