1. Evaluation of cytotoxicity features of antimicrobial peptides with potential to control bacterial diseases of citrus
Rosangela Naomi Inui Kishi, et al. PLoS One. 2018 Sep 7;13(9):e0203451. doi: 10.1371/journal.pone.0203451. eCollection 2018.
Antimicrobial peptides (AMPs) can be found in various organisms, and could be considered an alternative for pesticides used to control plant pathogens, including those affecting citrus. Brazil is the largest producer and exporter of frozen concentrated orange juice in the world. However, the citrus industry has been affected by several diseases such as citrus canker and huanglongbing (HLB), caused by the bacteria Xanthomonas citri subsp. citri (X.citri) and Candidatus Liberibacter asiaticus (CaLas), respectively. In order to control these pathogens, putative AMPs were prospected in databases containing citrus sequences. Furthermore, AMPs already reported in the literature were also used for in vitro and in vivo assays against X.citri. Since CaLas cannot be cultivated in vitro, surrogates as Sinorhizobium meliloti and Agrobacterium tumefaciens were used. This study reports the evaluation of six AMPs obtained from different sources, two of them from Citrus spp. (citrus-amp1 and citrus-amp2), three from amphibians (Hylin-a1, K0-W6-Hy-a1 and Ocellatin 4-analogue) and one from porcine (Tritrpticin). Peptides K0-W6-Hy-a1, Ocellatin 4-analogue, and citrus-amp1 showed bactericidal activity against X.citri and S. meliloti and bacteriostatic effect on A. tumefaciens. These results were confirmed for X.citri in planta. In addition cytotoxicity evaluations of these molecules were performed. The AMPs that showed the lowest hemolytic activities were Triptrpticin, citrus-amp1 and citrus-amp2. Citrus-amp1 and citrus-amp2 not presented toxicity in experiments using in vivo model, G. mellonella and U87 MG cells. To verify the interaction of these AMPs with bacteria and erythrocyte cell membranes, vesicles mimicking these cells were built. Citrus-amp1 and Tritrpticin exhibited higher affinity to bacterial membranes, while Ocellatin 4-analogue and Hylin-a1 showed higher affinity to erythrocyte membranes; exclude their use in citrus. This work demonstrates an essential alternative, trough AMPs obtained from Citrus spp., which can be feasibly used to control bacterial pathogens.
2. Purification, characterization and homology analysis of ocellatin 4, a cytolytic peptide from the skin secretion of the frog Leptodactylus ocellatus
Anna Nascimento, Alex Chapeaurouge, Jonas Perales, Antonio Sebben, Marcelo V Sousa, Wagner Fontes, Mariana S Castro Toxicon. 2007 Dec 15;50(8):1095-104. doi: 10.1016/j.toxicon.2007.07.014. Epub 2007 Aug 3.
Neobatrachia is the amphibian suborder with the largest number of species and a most important source of bioactive peptides from frog skin secretions. However, 90% of the studies on this subject have been focused on the frog families Hylidae and Ranidae, while very little is known about peptides of other families, like Leptodactylidae. Our work reports for the first time the isolation and characterization of ocellatin 4 (GLLDFVTGVGKDIFAQLIKQI-NH(2)), a cytolytic peptide from the skin secretion of the South American frog Leptodactylus ocellatus. While most cytolytic amphibian skin peptides are selective for microorganisms and harmless for mammalian cells, the HC(50) of ocellatin 4 against human erythrocytes is 14.3muM. The interaction between ocellatin 4 and zwitterionic phospholipids in mammalian plasma membranes may be favored by its neutral charge at pH 7.0. Ocellatin 4 also shows some antibacterial activity (MICs(E. coli)(and)(S. aureus)=64muM) and its sequence shares similarities with the only six leptodactylid peptides previously known and with four peptides from Australian hylid frogs of the genus Litoria.
