Unidirectional liquid transport is critical in achieving high-performance moisture-management fabrics for medical care. However, realizing unidirectional liquid transport while simultaneously satisfying other requirements, such as antibacterial function, adhesiveness, low cytotoxicity, and adequate mechanical strength remains a challenge. In this study, Janus nanofibrous membranes exhibiting both unidirectional liquid transport and antibacterial activity were fabricated via electrospinning and a mild crosslinking procedure. This membrane provides continuous and spontaneous unidirectional water transport with a high one-way transport value (R) of 1483%. The membrane achieved antibacterial rates of 99.2% and 98.7% against E. coli and S. aureus, respectively, without leaching antibacterial agents. In addition, it has high elasticity and self-adhesive properties, which facilitates its use in a range of applications. The design of this versatile Janus nanofibrous membrane provides a new strategy for developing novel moisture-wicking systems, particularly in the field of medical dressings.