毛霉诱导脐橙产抗病物质对指状青霉和酸腐菌的抑制

绿霉和酸腐是柑橘果实采后主要的病害,生产上迫切需要化学杀菌剂的替代方法。该研究对毛霉诱导脐橙果皮产生的具有抗病活性的红色物质(Citru Peel Red-substance Extract,CPRE)进行部分分离纯化,评价了CPRE对指状青霉和酸腐菌的抑菌活性,分析活性成分对两病原真菌菌丝细胞壁与膜功能的影响。结果表明,在50~200μg/mL的质量浓度下,CPRE对指状青霉和酸腐菌的孢子萌发,芽管伸长,菌丝生长有强烈的抑制作用,并且呈现剂量效应,最低抑菌浓度均为200μg/mL。进一步研究表明,与空白对照比较,50、100μg/mL抗菌活性成分处理明显改变指状青霉和酸腐菌菌丝细胞壁中几丁质分布,降低了细胞壁中几丁质含量、促进了质外体碱性磷酸酶的释放,表明CPRE严重损伤两致病真菌菌丝的细胞壁功能;CPRE处理还明显损伤两病菌菌丝的细胞膜通透性(P<0.05),刺激菌丝膜外电导率显著上升(P<0.05),加剧细胞内蛋白和核酸的丢失,降低了菌丝细胞膜总脂质含量,影响了细胞膜的正常功能。上述结果表明,红色抗菌成分在防控柑橘采后绿霉病和酸腐病中有潜在应用前景。

Inhibition of induced citrus peel disease-resistant components against Penicillium digitatum and Geotrichum citri-aurantii

Citrus are popular worldwide fruits due to the high nutritional value and special flavor,but they are easy to decay during storage and transportation,particularly on green mold,blue mold and sour rot.Alternatively,disease controlling methods are also necessary to protect from the bandage of synthetic chemical fungicides due to the health and environmental concerns.In a previous study,the fruits cannot decay when red pigment substances were accumulated in the fruit peels around wounding sites after inoculation with Actinomucor elegan,indicating the Citrus Peel Red-substances Extract(CPRE)has strong antifungal activity against Penicillium italicum.Here,this study aims to assess antifungal effects of the partially purified CPRE on Penicillium digitatum and Geotrichum citri-aurantii,the casual agents for green mold and sour rot,respectively.To further explore the possible inhibition mechanism of the substance against the pathogens,the damage on cell wall was evaluated by chitin content and distribution,and leakage of alkaline phosphatase,while the interferences on cell membrane were investigated to assess membrane integrity,release of proteins and nucleic acids,extracellular electronic conductivities and contents of lipids in the hyphae cells of both paghogens.The results showed that at the concentration of 50 to 200μg/mL,CPRE exhibited strong inhibitory effects on spore germinations,elongations of germ tubes,and mycelia growth of P.digitatum and G.citri-aurantii,in a concentration dependent manner,with the Minimal Inhibitory Concentration(MIC)at 200μg/mL for both pathogens.Calcofluor white staining,an agent which specifically binds to chitin exposed,revealed that the chitin distribution and cell wall integrities of the hyphae of both pathogens were markedly interfered when incubated with CDPC at 1/2 MIC for 24 hours.CPRE treatments also made chitin contents decrease by 30.29%,52.64%,and accelerate the release of alkaline phosphatase in the mycelia of both pathogens,indicating that cell wall structures and functions were disturbed by the active components in CPRE.Furthermore,membranes of the hyphae cells of both pathogens can be deteriorated after CPRE treatments at the concentration of 100μg/mL,as indicated by propidium iodide staining,which emits red fluorescence when entered into dead cells and bind with nucleic acids.The extracellular electronic conductive of the hyphae cells of both pathogens increased significantly after 6 hours of incubation with CPRE within the concentrations tested,indicating that the metal ion balance was disrupted in the treated cells.There were significant differences in leakages of soluble proteins and nucleic acids from hyphae cells with CPRE treatments,showing a consistence with that of extracellular conductivity assays.Moreover,CPRE treatments reduced the total contents of cell lipid in hyphae of both pathogens.These results demonstrated that CPRE can effectively inhibit both pathogens by damaging or interfering with the cell membranes and cell walls of the hyphae.The powerful antifuangal effects of CPRE against P.digitatum and G.citri-aurantii can offer the great potential application prospects in control of postharvest decay of citrus fruits,and futher provide a theoretical basis for the development of natural products with antifungal effects.