Proteção de citros com peptídeos antibacterianos e galatos
Arquivos
Data
2021-12-20
Autores
Dilarri, Guilherme [UNESP]
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Editor
Universidade Estadual Paulista (Unesp)
Resumo
O Brasil é o maior exportador de suco de laranja concentrado do mundo. Apesar do sucesso da citricultura no país, uma das principais doenças bacterianas que causam grandes perdas na produtividade é o cancro cítrico. Essa doença é causada pela bactéria Gram-negativa Xanthomonas citri subsp. citri (X. citri). Para fazer o controle da doença realiza-se o manejo integrado com aplicação constante de soluções cúpricas nos pomares. Porém, o cobre é um metal pesado tóxico para o meio ambiente e para a saúde humana, sendo importante encontrar alternativas, diminuindo assim a aplicação desse metal na citricultura. Logo, o objetivo deste trabalho foi testar dois novos protetivos para folhas de citros contra infecção por X. citri. A primeira plataforma desenvolvida foi baseada em microgéis para liberação controlada de agentes antibacterianos e que contém um peptídeo de ancoragem em sua superfície capaz de promover sua adesão às folhas de citros. Tais géis possuem o galato G6 como agente bactericida imobilizado em sua matriz polimérica. A segunda plataforma proposta foi a de peptídeos antimicrobianos em fusão com peptídeo de ancoragem também para fixação foliar. Os resultados mostraram que de oitos diferentes microgéis sintetizados apenas GMA_hDerm_G6 e GMA_hDerm_AT_G6 apresentaram inibição de X. citri in vitro, com atividade bactericida nas concentrações de 1,375 e 0,343 mg mL-1, respectivamente. Dos peptídeos produzidos, apenas o Melittin_Cg-Def na concentração de 125 µg mL-1 apresentou efeito bactericida em ensaios in vitro. Ensaios de microscopia mostraram que o G6 presente no microgel era o principal agente antimicrobiano, mantendo seu mecanismo de ação mesmo após ter sido imobilizado na matriz polimérica do microgel, afetando a estrutura da membrana citoplasmática e desfazendo o anel divisional da X. citri. O Melittin também continuou apresentando seus mecanismos de ação mesmo após ser fusionado com o peptídeo de ancoragem Cg-Def, apresentando ainda ação na perturbação e rompimento da membrana citoplasmática bacteriana. Os testes de fitotoxicidade e citotoxicidade mostraram que os microgéis e o peptídeo bifuncional eram tóxicos para as sementes de Eruca sativa e para as células J774A.1. Porém, suas toxicidades eram menores que o composto cúprico comercial (Difere). Os resultados em casa de vegetação confirmaram que os microgéis na concentração de 2,75 mg mL-1 e o Melittin_Cg-Def na concentração de 250 µg mL-1 apresentavam uma proteção efetiva contra a infecção da X. citri, reduzindo significantemente o número de lesões por área foliar. Destacou-se nos testes o microgel GMA_hDerm_G6, que mesmo após uma simulação de chuva, continuava protegendo eficientemente as plantas contra a X. citri. Pode-se concluir que ambas as plataformas propostas, utilizando as respectivas construções mostradas no presente trabalho, são uma possível alternativa aos compostos cúpricos aplicados em campo para o controle do cancro cítrico, podendo vir a ser uma alternativa menos tóxica e mais sustentável no manejo da doença.
Brazil is the largest exporter of concentrated orange juice in the world. Despite the success of citriculture in the country, one of the main bacterial diseases that cause great losses in productivity is citrus canker. This disease is caused by the Gram-negative bacterium Xanthomonas citri subsp. citri (X. citri). To control the disease, integrated management is carried out with the constant application of cupric solutions in the orchards. However, copper is a heavy metal that is toxic to the environment and to human health, and it is important to find alternatives, thus reducing the application of this metal in citriculture. Therefore, the aims of this work were to test two new protective compounds for citrus leaves against X. citri infection. The first platform developed was based on microgels for controlled release of antibacterial agents and which contain an anchoring peptide on its surface capable of promoting its adhesion to citrus leaves. These gels have the G6 gallate as a bactericidal agent immobilized in their polymer matrix. The second platform proposed was that of antimicrobial peptides fused with an anchor peptide for foliar fixation. The results showed that from eight different microgels synthesized, only GMA_hDerm_G6 and GMA_hDerm_AT_G6 showed inhibition of X. citri in vitro, with bactericidal activity at concentrations of 1.375 and 0.343 mg mL-1, respectively. From the synthesized peptides, only Melittin_Cg-Def at the concentration of 125 µg mL-1 showed bactericidal effect at in vitro assays. Microscopy tests showed that the G6 present in the microgel was the main antimicrobial agent, maintaining its mechanism of action even after being immobilized in the polymeric matrix of the microgel, affecting the structure of the cytoplasmic membrane and undoing the divisional ring of X. citri. Melittin also continued to show its mechanisms of action even after being fused with the Cg-Def peptide, still acting on the disruption of the bacterial cytoplasmatic membrane. Phytotoxicity and cytotoxicity tests showed that microgels and bifunctional peptide were toxic to Eruca sativa seeds and J774A.1 cells. However, its toxicities were lower than the commercial cupric compound (Difere). The results in a greenhouse confirmed that microgels at a concentration of 2.75 mg mL-1 and Melittin_Cg-Def at a concentration of 250 µg mL-1 provided effective protection against X. citri infection, significantly reducing the number of lesions caused by leaf area. The microgel GMA_hDerm_G6 was highlighted out in the tests, which even after a rain simulation, continues efficiently protective for plants against X. citri. It can be concluded that both proposed platforms, using the respective constructions showed in this work, are a possible alternative to cupric compounds applied in the field for the control of citrus canker, and may become a less toxic and more sustainable alternative in disease management.
Brazil is the largest exporter of concentrated orange juice in the world. Despite the success of citriculture in the country, one of the main bacterial diseases that cause great losses in productivity is citrus canker. This disease is caused by the Gram-negative bacterium Xanthomonas citri subsp. citri (X. citri). To control the disease, integrated management is carried out with the constant application of cupric solutions in the orchards. However, copper is a heavy metal that is toxic to the environment and to human health, and it is important to find alternatives, thus reducing the application of this metal in citriculture. Therefore, the aims of this work were to test two new protective compounds for citrus leaves against X. citri infection. The first platform developed was based on microgels for controlled release of antibacterial agents and which contain an anchoring peptide on its surface capable of promoting its adhesion to citrus leaves. These gels have the G6 gallate as a bactericidal agent immobilized in their polymer matrix. The second platform proposed was that of antimicrobial peptides fused with an anchor peptide for foliar fixation. The results showed that from eight different microgels synthesized, only GMA_hDerm_G6 and GMA_hDerm_AT_G6 showed inhibition of X. citri in vitro, with bactericidal activity at concentrations of 1.375 and 0.343 mg mL-1, respectively. From the synthesized peptides, only Melittin_Cg-Def at the concentration of 125 µg mL-1 showed bactericidal effect at in vitro assays. Microscopy tests showed that the G6 present in the microgel was the main antimicrobial agent, maintaining its mechanism of action even after being immobilized in the polymeric matrix of the microgel, affecting the structure of the cytoplasmic membrane and undoing the divisional ring of X. citri. Melittin also continued to show its mechanisms of action even after being fused with the Cg-Def peptide, still acting on the disruption of the bacterial cytoplasmatic membrane. Phytotoxicity and cytotoxicity tests showed that microgels and bifunctional peptide were toxic to Eruca sativa seeds and J774A.1 cells. However, its toxicities were lower than the commercial cupric compound (Difere). The results in a greenhouse confirmed that microgels at a concentration of 2.75 mg mL-1 and Melittin_Cg-Def at a concentration of 250 µg mL-1 provided effective protection against X. citri infection, significantly reducing the number of lesions caused by leaf area. The microgel GMA_hDerm_G6 was highlighted out in the tests, which even after a rain simulation, continues efficiently protective for plants against X. citri. It can be concluded that both proposed platforms, using the respective constructions showed in this work, are a possible alternative to cupric compounds applied in the field for the control of citrus canker, and may become a less toxic and more sustainable alternative in disease management.
Descrição
Palavras-chave
Cancro cítrico, Defensivos agrícolas, Melittin, Microgéis, Xanthomonas citri, Citrus cancer, Pesticides, Microgels