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Expression of antimicrobial peptides in plants to control pathogenic bacteria and fungi [Image: photo of fungi] [Image: photo of fungi on a control plant] [Image: photo of fungi on a transgenic plant] Fungal and bacterial diseases cause millions of dollars of crop damage and present an ongoing challenge for farmers. Breeding crops resistant to multiple microbial diseases by conventional methods has had limited success. Introduction of antimicrobial peptides through plant transformation could offer a solution for creating crops resistant to a wide range of bacterial and fungal pathogens. Suggested sources to read more about antifungal and antimicrobial peptides are: Hancock R.E., Patrzykat A. (2002) Clinical development of cationic antimicrobial peptides: from natural to novel antibiotics. Curr Drug Targets Infect Disord. 2(1):79-83. Review. Devine D.A., Hancock R.E. (2002) Cationic peptides: distribution and mechanisms of resistance. Curr Pharm Des. 8(9):703-14. Review. Using an in vitro assay, we identified antimicrobial peptides effective against Rhizoctonia solani. Among 11 antimicrobial peptides tested, several natural and synthetic antimicrobial peptides such as cecropin B, melittin, D4E1, and phor21 showed very similar antifungal activity. A plant defensin from wheat seed called purothionin showed inhibitory activity comparable to the antifungal antibiotics nystatin and nikkomycin. We found that all active peptides rapidly permeabilize fungal membranes. Membrane permeabilization can be detected using the nonpermeable fluorescent dye Sytox Green in 1 min after the peptide addition of cecropin B, melittin, phor21, or D2A21 and reaches maximum in 10 min. Noticeable increase of membrane permeabilization and antifungal activity are observed at concentrations 2-4 µM (Fig. 1). Levels of permeabilization strongly correlate with antifungal activity (Oard, S. et al., 20041). Three antimicrobial peptides exhibiting in vitro antifungal activity were expressed in Arabidopsis to compare their in planta activity. β-Purothionin, cecropin B, and phor21 were expressed under an endogenous promoter with moderate-level activity and excreted extracellularly. Expression of β-purothionin rendered the greatest antibacterial and antifungal resistance while cecropin B enhanced only antibacterial activity, and phor21 did not improve antimicrobial resistance. The transgenic β-purothionin arrested fungal growth on leaf surfaces and infection of stomata. Leaf extracts from plants producing β-purothionin and cecropin B displayed membrane permeabilizing activity (Fig. 2, 3). The in planta antimicrobial activity of the tested peptides was consistent with previously reported in vitro experiments. Our expression strategy allowed enhanced antifungal resistance without high-level transgene expression (Oard, S. and Enright, F., 20062). In collaboration with Dr. Chuck Rush and Dr. Jim Oard, our group applies molecular breeding as a potential method to control sheath blight disease. This disease is caused by the most notable of rice pathogenic fungi, R. solani. This pathogen causes sheath blight disease in rice resulting in millions of dollars in damage in the USA and around the world. It produces a toxin designated as RS-toxin, a carbohydrate compound containing mainly alpha-glucose and mannose. RS-toxin is translocated to the rice cells during infection that causes electrolyte leakage followed by cell death. Fungi thrive on host cells killed with RS-toxin.