RamA: The hidden regulator of virulence transformation in Klebsiella pneumoniae

Klebsiella pneumoniae is a major opportunistic pathogen responsible for hospital-acquired infections, often linked to high mortality rates and extended hospital stays. RamA, a member of the AraC transcription factor family, can induce multidrug resistance by upregulating the expression of efflux pumps, such as AcrAB and OqxAB. However, the effect of RamA on K. pneumoniae virulence remains unreported. In this study, we focused on the regulation of virulence of K. pneumoniae through ramA, in vitro and in vivo. We constructed ramA-knockout (KP-ΔramA) and ramA-complemented strains for scanning electron microscopy, serum resistance tests, and pathogenicity studies in mice. In vivo virulence tests using an intraperitoneal injection model and serum resistance assays demonstrated that the KP-ΔramA strains exhibited significantly higher virulence than the wild-type (KP-WT) strains (P<0.0001). Scanning electron microscopy revealed that the biofilm formed by the KP-ΔramA strains displayed significantly greater density and completeness than that of KP-WT strains. Enzyme-linked immunosorbent assays and immunohistochemistry revealed that the KP-ΔramA infected murine model exhibited attenuated levels of proinflammatory cytokines, including TNF-α and IL-6, in both serum and lung tissues compared to those infected with KP-WT. These findings suggest that KP-ΔramA infection suppresses the immune response in mice. Histopathological examination revealed that the lung tissues of KP-ΔramA infected mice exhibited more severe consolidation, inflammatory infiltration, and hyperemic exudation than those infected with KP-WT. In conclusion, our findings indicate that the loss of ramA leads to upregulation of K. pneumoniae virulence by increasing serum resistance and biofilm formation, while suppressing the host immune response.