The Study On Biocontrol And Mechanism Of Blue And Green Mold Of Citrus Fruits By Pichia Membranefaciens

Citrus fruit are the most important worldwide commercial fruit. Penicillium digitatum (green mold) and P. italicum (blue mold) are the major pathogens of citrus fruit after harvesting. Though synthetic chemical fungicides can control several diseases effectively, the use of fungicides to control postharvest deterioration has been restricted due to their high and acute residual toxicity, long degradation period, environmental pollution and their effects on food and possible side-effects on human health. Thus, it has become a worldwide trend to explore new alternatives to reduce the use of synthetic fungicides. The biological control of postharvest diseases by using antagonistic yeasts is one of the most promising non-fungicidal means.In the present study, we evaluated the biocontrol activity of P. membranaefaciens against the major postharvest pathogen (Penicillium digitatum and Penicillium italicum) on citrus fruits and studied the biocontrol mechanisms. In the study, we discussed the effect of some factors on the inhibiting activity of P. membranaefaciens. The factors are as below:the different treatments of P. membranaefaciens; the concentrations of cells the yeast and application time of the yeast. Population dynamics of P. membranaefaciens in wounds of citrus fruits and the effect of the yeast antistaling agent which composite exogenous chemicals such as chitosan, salicylic acid, calcium salt on citrus storage quality was also discussed. In addition, possible mechanisms such as compitition of nutrition and induced resistance by the antagonistic yeasts in Citrus fruit were demonstrated.The main results were as following:(1) In vitro experiment, the yeast has obvious inhibited the developing of P. digitatum and P. italicum whether it vegetated with pathogen in same hole or different holes. That means there was a strong competition relationship between yeast and pathogen, the antibacterial mechanism may be the competitive nutrient space. In addition, the yeast colonies without bacteriostatic laps around on the a flat plate can be concluded that the yeast cannot produce bacteriostasis of secondary metabolites, that means the yeast has not antibiosis mechanism and hyperparasitism mechanism.(2) citrus fruit treated with 1×108 CFU·mL-1 antagonistic yeast showed lower disease incidence and smaller lesion diameters caused by P. italicum and P. digitatum, immersion treatments by yeast reduced the natural incidence rate of citrus, these results indicated that both immersion and inoculation treatments significantly reduced postharvest disease of citrus. Moreover, the natural incidence rate of immersion yeast fruit was significantly lower than the control in the later storage time. This suggest that the P. membranaefaciens has a great potential to control bule and green mold of citrus fruit, and the biocontrol mechanism may be the compitition of nutrition and induced resistance.(3) The biocontrol efficacy of P. membranaefaciens was related with different formulations of yeast, the concentrations of cells the yeast and application time of the yeast.The biocontrol mechanism of P. membranaefaciens was detected by comparing the efficacy of different treatments.The results demonstrated that the disease incidence for the treatments of autoclaved cell culture and culture filtrate of P. membranaefaciens was not effctive in preventing the decay and even aggravated the disease. Living cell suspension and washed cell suspension of the yeast was significantly lower than that for control experiment, and the washed cell suspension of P. membranaefaciens at 1×108 CFU(colony-forming unit)·mL-1 provided the best biocontrol capacity of postharvest rot on citrus. This also suggest that the biocontrol mechanism of P. membranaefaciens may be the compitition of nutrition.(4) P. membranaefacien could acclimatize itself to the environment of Citrus fruit wounds and occupy the living space quickly whether at room temperature or cold storage temperature. The efficacy of P. membranaefaciens was consistent when applied 24 h or 48 h before the pathogen. Rapid colonization of the wounds by P. membranaefaciens was observed at both at 4℃or 20℃. After 48 h inoculation,the population of the yeast on the inoculated wounds was increased by more than 4 and 11 folds. That is mean the yeast has the prerequisite for commercial application.(5) The fruit immersed or inoculated with P. membranaefacien demonstrated changes in phenolic and flavonoids compounds content, peroxidase (POD), polyphenoloxidase (PPO), phenylalanine ammonia-lyase (PAL), chitinase (CHI) andβ-1,3-glucanase activities(GLU) in Citrus fruit peel, all of which were correlated with the onset of induced resistance. This result suggests that Citrus fruit is capable of responding to the yeast P. membranaefacien which could activate defensive enzymes and thereby induce host disease resistance.(6) SOD, APX and GR activity as well as the content of H2O2, O2-, GHS and ASA was enhanced in yeast-treated fruit, but activities of catalase (CAT) showed a decrease in the same fruit. These results indicated that yeast-treatments could induce the fruit disease resistance by regulating the active oxygen metabolism and antioxidant system.(7) Exogenous chemicals mixed with yeast can significantly improve the antimicrobial activity of the antagonistic yeast. Meanwhile, the yeast and yeast compound preservatives which mixed with other substances have no adverse effects on storage quality of citrus fruit, and to some extent, delayed maturation and senescence of citrus, improve the storage quality.