Citric Acid Inhibits Against Trichothecium Roseum In Vitro And Its Partial Mechanism

Trichothecium roseum is one of important pathogens that causes post-harvest losses at high frequency which holds a wide range of host. It was reported that citric acid possessed antimicrobial, antifungicidal and harmless characteristics which were considered to have great potential as antimicrobial agents. There were little reports available on the underlying mechanism of antifungal function of citric acid against T. roseum. This study investigated the antifungal action and potential anti- fungal mechanisms of citric acid. The study use traditional biological methods evaluated the antifungal activity of citric acid on spore germination, expansion of the colony diameterand and hyphal biomass. The cytological method detect the release of cell constituents, ergosterol content and PI staining indicators of citric acid on plasma membrane integrity and observe the morphological modification ultrastructural changes. The biochemistry methods used to evaluate the changes of ROS metabolism in spores. The results showed as below:1. 10 m M and 20 m M citric acid treatment inhibited the spore germination, colony diameter and hyphal biomass effectivly. The inhibition effect was related to the concentration of citric acid. 20 m M citric acid treatment showed remakeble inhibition, compard with control spore germination, colony diameter and hyphal biomass declined 84.81%, 89.20% and 86.46%, respectively.2. Citric acid reduce the release of cell constituents. Total ergosterol content of hyphae dclined, 10 m M and 20 m M citric acid treatment only occupy for 38.48% and 23.66% of control. PI staining also examined the integrity of plasma membrane of spores detected by flow cytometry revealed significant increase in PI fluorescence incubation with the higher concentration citric acid.3. SEM and TEM showed that citric acid treatment lead to large alterations in morphology and structure which contrbuite to induce the destruction of the plasma membrane. The morphological modifications, including mycelium asymmetry, hyphal swelling, and a irregular branch under the citric acid treatment. TEM observation results show that the treatment spore ultrastructural changes dramatically, 20 m M treatment lead to plasma membrane damage degree deepening, and even appear plasmolysis. Normal physiological metabolism of T. roseum was destroyed by citric acid.4.Citric acid- induced endogenous production of ROS in T. roseum spore. The accumulation of ROS result in content of H2O2 elevated and protein, lipid oxidative damage. Under the treatment of citric acid SOD、CAT and POD enzyme activity elevated. When adding exogenous antioxidant vitamin C, citric acid spores H2O2 accumulation significantly reduced in the spores, lipid peroxidation and carbonylation of protein level was significantly reduced.These results suggest that the mechanisms of the antifungal action of citric acid against T. roseum can be attributed to the disruption of the membrane integrity and accumulation of ROS.