Stress-induced iro gene expression in hypervirulent Klebsiella pneumoniae

Introduction: Klebsiella pneumoniae (K. pneumoniae) is a Gram-negative bacterium that poses a significant threat to public health. The virulence factors that enable its colonization and invasion of different anatomical sites, particularly iron acquisition, are critical, as iron limitation within the host environment necessitates specialized uptake mechanisms. In this study, the transcriptional expression of iron acquisition-related virulence genes in K. pneumoniae from environmental interfaces was investigated. Materials and methods: A total of 302 drainage water samples surrounding primary, secondary, and tertiary care hospitals were collected. Phenotypic characterization was done using the string test to assess the hypermucoviscosity. Genotypic characterization was done to detect the presence of iron acquisition-related virulence genes using polymerase chain reaction (PCR). Expression analysis under varying iron conditions and antibiotic stress was conducted through quantitative real-time PCR, and antibiotic susceptibility testing (AST) was done using the Kirby–Bauer disc diffusion method. Results: In total, 76 K. pneumoniae isolates were identified, out of which 18 isolates showed positive results on the string test. Out of them, 15 isolates were found to be hypervirulent and harbored iron acquisition genes such as iroN (PB66, PB77, PB109, PC7, and PC15); iroB (PB60, PB77, PB109, HN72, and PC10); iroC (PB77, PB107, PC29, PC34, and PC40); and iroD (HN21, PC7, PC8, PC39, and PC40). In all the isolates harboring iroB and iroC, expression was upregulated under combined FeCl3 + ethylenediaminetetraacetic acid (EDTA) and imipenem stress, whereas in all the isolates harboring iroN, expression was downregulated. In iroD-harboring isolates, expression was upregulated under FeCl3 and FeCl3 + EDTA but suppressed under EDTA and imipenem. The AST results revealed the highest resistance was to ampicillin (87%), followed by norfloxacin (83%) and ciprofloxacin (76%). Conclusions: This study revealed that iron availability and antibiotic stress significantly influence the transcriptional regulation of key virulence genes in K. pneumoniae. By integrating molecular insights with environmental context, this work contributes to a broader understanding of how hypervirulent K. pneumoniae (hvKp) adapts and persists in environmental interfaces.