Within-host mechanisms of resistance evolution in polymyxin B-resistant Klebsiella pneumoniae without prior exposure.

Abstract

BACKGROUND: Polymyxin B-resistant Klebsiella pneumoniae (PBR-KP) has been emerging as a significant concern. This study investigated the within-host mechanisms of resistance evolution in PBR-KP without prior PB exposure. METHODS: Six groups were enrolled [PB-sensitive K. pneumoniae (PBS-KP) evolved into PBR-KP from the same patient], and antimicrobial susceptibility testing was conducted using the microdilution broth method. Whole-genome sequencing was performed to analyze molecular characteristics, and to investigate the resistance mechanisms, gene expression analysis of pmrAB, phoPQ, crrAB, and eptA was performed using RT-qPCR. Fitness costs were assessed through biofilm formation, in vitro growth rate, and virulence analysis. RESULTS: All 12 strains belonged to ST11-KL64, except for PB9 and PB10, which were classified as ST11-KL47. Each resistant strain was highly related to its corresponding sensitive strain. The relative expression levels of phoPQ, eptA, and pmrAB were significantly upregulated in most PBR-KP strains. PBR-KP strains exhibited no fitness cost regarding in vitro growth rate and biofilm formation. Notably, the eptA gene was highly expressed in the four PBR-KP strains with enhanced virulence, indicating its association with virulence. Furthermore, PBR-KP strains with enhanced virulence were highly associated with KL64, whereas those without enhanced virulence were linked to KL47, except for PB12, indicating a potential relationship between virulence and serotype. CONCLUSIONS: PB resistance can develop in KP strains without prior exposure to the drug, possibly driven by upregulation of phoPQ, pmrAB, and eptA, as supported by validated transcriptomics data. These resistant strains did not exhibit fitness costs related to biofilm formation or growth rate. Enhanced virulence, found in most PBR-KP strains, may be closely associated with eptA overexpression and specific serotypes.