Plant defensins are antimicrobial host defense peptides expressed in all higher plants. Performing a significant role in plant innate immunity, plant defensins display potent activity against a wide range of pathogens. Vertebrate and invertebrate defensins have well-characterized antibacterial activity, but plant defensins are commonly considered to display antimicrobial activity against only fungi. In this review, we highlight the often-overlooked antibacterial activity of plant defensins. Also, we illustrate methods to evaluate defensins for antibacterial activity and describe the current advances in uncovering their antibacterial modes of action.