Abstract

Host-directed strategies that enhance early innate immune defense are increasingly recognized as important adjunct approaches for controlling mycobacterial infections. Flagellin, a classical ligand of Toll-like receptor 5 (TLR5), has been widely studied as a vaccine adjuvant. However, whether it can transiently modulate innate immunity to enhance host resistance against mycobacterial infection without inducing sustained inflammation remains unclear in vivo. In this study, we systematically evaluated this question using a zebrafish– Mycobacterium marinum infection model. Our results demonstrate that a single exposure to Escherichia coli -derived flagellin significantly reduces bacterial burden and improves host survival upon subsequent infection, without disrupting immune homeostasis. Mechanistically, this protective effect depends on the TLR5–MyD88 signaling pathway and is associated with enhanced early inflammatory responses, as well as accelerated recruitment of neutrophils and macrophages. Notably, even after the resolution of initial inflammation, the host exhibits an enhanced responsiveness to secondary stimulation, resembling features of trained immunity, suggesting that flagellin may act as an inducer of innate immune memory. Collectively, our findings provide in vivo evidence that flagellin enhances early host defense against mycobacterial infection by modulating innate immunity and support the utility of the zebrafish model for evaluating immunomodulator-induced protection.