Tracking emergence and outbreak of Klebsiella pneumoniae co-producing NDM-1 and KPC-2 after sulfamethoxazole-trimethoprim treatment: Insights from genetic analysis

The co-production of KPC and NDM carbapenemases in carbapenem-resistant Klebsiella pneumoniae (CRKP) complicates clinical treatment and increases mortality rates. The emergence of KPC-NDM CRKP is believed to result from the acquisition of an NDM plasmid by KPC CRKP, especially under the selective pressure of ceftazidime-avibactam (CZA). In this study, a CRKP-producing KPC-2 (JNP990) was isolated from a patient at a tertiary hospital in Shandong Province, China. Following sulfamethoxazole-trimethoprim (SXT) treatment, the isolate evolved into a strain that co-produces KPC and NDM (JNP989), accompanied by resistance to SXT (minimum inhibitory concentration >2/38 µg/mL) and CZA (dd ≤14 mm). Whole-genome sequencing and S1 nuclease pulsed-field gel electrophoresis revealed that JNP989 acquired an IncC plasmid (NDM plasmid) spanning 197 kb carrying sul1 and bla_NDM-1 genes. The NDM plasmid could be transferred successfully into Escherichia coli J53 at a conjugation frequency of (8.70±2.47) × 10^-4. The IncFⅡ/IncR plasmid carrying the bla_KPC-2 gene in JNP990 could only be transferred in the presence of the NDM plasmid at a conjugation frequency of (1.93±0.41) × 10^-5. Five CRKP strains with the same resistance pattern as JNP989, belonging to the same clone as JNP989, with sequence type 11 were isolated from other patients in the same hospital. Two strains lost resistance to CZA due to the loss of the bla_NDM-1-carrying fragment mediated by insertion sequence 26. Plasmid stability testing indicated that the IncC plasmid was more stable than the bla_NDM-1 genes in the hosts. This study describes the evolution of KPC-NDM CRKP and its spread in hospitalized patients following antibiotic treatment, highlighting the severity of the spread of resistance.