Antimicrobial peptides (AMPs) form part of the innate immunity establishing first line of defense against pathogen(s). Plant AMPs are a component of plant barrier defense system. They have been isolated from different parts of a variety of plant species and are known to have activities toward phytopathogens and also against human pathogens. All plant organs express AMPs constitutively or in response to microbial challenges. Plant AMPs are structurally and functionally diverse. Plant AMPs are grouped into several families and share general features such as positive charge and the presence of disulfide bonds which stabilize the structure. Thionins, defensins, lipid transfer proteins (LTPs), snakins, knottins, hevein-like AMPs and cyclotides represent the different plant AMPs. Besides targeting known bacterial, fungal, and other pathogens, some of them can be directed against other organisms, like insects. They have anticancer and antiviral activities. The biological activity of plant AMPs primarily depends on the interactions with membrane lipids, using different mechanisms to kill the targeted pathogens, but other modes of action exist as in the case of defensins with a-amylase activity or a defensin-like peptide that interacts with a receptor kinase. Plant AMPs are considered as promising antibiotic compounds with important applications in biotechnology, pharmacy and agriculture. AMPs, therefore, possess a high potential for therapeutic use in healthcare and can be used as natural antibiotics as an alternative for their chemical counterparts, protection of plants or animals against diseases. Six classes of plant AMPs will be discussed in this chapter and special attention will be given to their mechanism of action and applications. Therefore, this review can be considered as a snapshot of the progress in this field of research.