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Liposome delivery system of antimicrobial peptides against Huanglongbing

Jeanette Velásquez Guzmán: New Mexico Consortium

<div>Huanglongbing (HLB), caused by <em>Candidatus</em> Liberibacter spp, is the most devastating citrus disease and threatens citrus production worldwide. Current HLB control strategies are sorely inadequate to prevent spread of the disease. Although both streptomycin and oxytertracycline have been used in foliar sprays, each compound only moderately reduces Liberibacter load, and antibiotic resistant pathogens remain a concern. Here, membrane-targeting, positively-charged amphipathic helical antimicrobial peptides were designed and tested against several bacterial strains <em>in vitro</em> and against Liberibacter<em> in vivo</em>. Peptide P11 exhibit higher antimicrobial activity against culturable bacterial strains, and also exhibit higher activity than streptomycin against Liberibacter. Based on our study of the resistance mechanism of <em>Escherichia</em><em> coli </em>against P11, we designed a 2<sup>nd</sup> generation helical amphipathic peptide, P26, in which two P11 peptides are connected by a 4-amino acid beta turn. P26 is not only more active against <em>E. coli</em>, but it is also not susceptible to bacterial resistance. Importantly, P26 has higher anti-Liberibacter activity than either P11 or streptomycin. To effectively deliver these peptides into citrus phloem, we developed virus-like liposome vesicles that encapsulate P11 and P26, using an outer shell of <em>Citrus tristeza </em>virus (CTV) capsid protein. Attachment of virus coat protein to liposome vesicles should ensure transport into and within the plant cells, as well as a controlled release of peptides. We are now testing the efficacy of P11 and P26 by <em>in planta</em> assays using this delivery system. Successful completion of these studies should enable using P11 and P26 for HLB treatment.</div>