Mechanisms of colistin resistance and the antimicrobial effects of antibiotic and adjuvant combination on colistin-resistant Klebsiella pneumoniae

Abstract

The prevalence of colistin-resistant Klebsiella pneumoniae (ColRkp) has progressively increased globally. The purposes of this study were to characterize the molecular mechanisms responsible for colistin resistance among ColRkp clinical isolates in Thailand, to determine the expression levels of virulence factors associated with ColRkp clinical isolates, and to evaluate in vitro and in vivo synergistic activities of novel combination of antibiotic and adjuvant against ColRkp clinical isolates. A total of 165 carbapenem-resistant K. pneumoniae clinical isolates were obtained from King Chulalongkorn Memorial Hospital between 2016 and 2021. We discovered a rising trend of ColRkp displaying extensively drug-resistant (XDR) and pandrug-resistant (PDR) characteristics, with a prevalence of 28.5% (n=47). Both chromosomal mgrB, pmrB, or phoQ genes mutations (91.5%) and plasmid-mediated mcr-1.1, mcr-8.1, or mcr-8.2, alone or in combination with R256G PmrB (8.5%), were responsible for colistin resistance in these ColRkp isolates. Several independent insertions, deletions, or substitutions in mgrB (85.1%) associated with increased expressions of Ara4N-related phoPQ and pmrK transcripts were observed to be crucial in establishing colistin resistance in our isolates. Since mgrB, pmrAB, and phoPQ are involved in supporting bacterial virulence, we observed a significant association between XDR ColRkp and increased biofilm production (p<0.0001). Moreover, significantly altered bacterial virulence factors expressions were found to be associated with XDR ColRkp clinical isolates. A novel colistin-EDTA combination exhibited potent synergistic activity in both planktonic and mature biofilms of all tested XDR and PDR ColRkp isolates in vitro. A combination of colistin and EDTA also exhibited significant therapeutic effectiveness in eradicating ColRkp catheter-related biofilm infections and eliminating the risk of recurrence both in vitro and in vivo. Furthermore, colistin-EDTA combination demonstrated its significant therapeutic efficacy and safety in decreasing bacterial loads in internal organs, reducing serum creatinine, and enhancing animals survival in vivo. This is the first in vitro and in vivo study to highlight that a novel colistin-EDTA combination is a promising therapeutic strategy for successfully overcoming colistin resistance in ColRkp catheter-related biofilm infections.