Islam, M. M.; Beckman, R. L.; Nutter, N. A.; Valencia Bacca, J.; Hernandez, G. E.; Fleeman, R. N.; Haas, K.; Zafar, M. A.

Successful gastrointestinal colonization (GI) by bacterial pathogens requires adaptation to nutrient competition and host-derived stresses in the gut, with adaptation via the bacterial stringent stress response playing a critical role. Epidemiological data suggest that the GI tract serves as a reservoir from where K. pneumoniae can spread and cause invasive disease or transmit to another host. DksA is a conserved stringent response transcriptional regulator that was identified in an in vivo transposon mutagenesis screen as an important K. pneumoniae gut determinant. However, its role in K. pneumoniae pathogenesis and gut colonization remains uncharacterized. Here, we demonstrate that DksA is required for survival against membrane-targeting antibiotics, consistent with a role in cell envelope stress tolerance. In addition, DksA positively regulates capsule biosynthesis gene expression and hypermucoviscosity and is essential for robust biofilm formation. Using a murine model, we show that DksA functions as a determinant of GI colonization independently of the resident gut microbiota. Furthermore, we demonstrate that DksA is important for environmental survival and transmission by regulating RpoS, thereby providing a mechanistic link between the stringent stress response, environmental survival, and subsequent transmission. Together, these findings establish DksA as a central integrator of the stringent response, coordinating membrane stress resistance, virulence traits, and gastrointestinal colonization in K. pneumoniae.