Induced Effect Of γ-Aminobutyric Acid On Host Resistance Against Blue Mold And Defense-Related Mechanism In Pear Fruit

Pathogen infection is an important factor affecting fruit postharvest physiology, metabolism. Postharvest fruit losses caused by diseases annually are very considerable. At present, the theory of plant induced disease resistance and technology has become a common method instead of chemical fungicides for the control of postharvest diseases in fruits and vegetables. Gamma-aminobutyric acid (GABA), which has become.a new type of fodd functional components, is widely used in food industry. GABA, considered as the metabolite in plant, plays a key role of metabolic regulation, stress resistance, response to environmental stress in postharvest fruit. In recent years, the researches have not yet reported the effect of GABA on the resistance to disease in fruit. The objective of this research was to analyze the function of GABA induced resistance and the key influence factors; preliminarily determine the mechanism of GABA action on pathogenic bacteria; study the related physiological and biochemical mechanism about regulating fruit resistance by GABA and the impact on the ultrastructure and fruit quality; further identify the expression of fruit resistane-related gene by GABA; investigate the effect of GABA on the metabolism of resistance substances and the GABA metabolic pathway in fruit. The main research results are as follows:1. Effect of GABA on induction of resistance to P. expansum in pear fruit1～1000μg/ml GABA had no direct fungitoxic activity against P. expansum in pear fruit wounds but induced fruit resistance to blue mold. The efficiency was might be closely associated with the concentration of GABA, the time interval between GABA treatment and P. expansum inoculation, and the spore concentration of P. expansum.100μg/ml GABA significantly inhibited the incidence of the blue mold rot caused by104spores/ml P. expansum by54.5%when the time interval was increased to24h with respect to control at room temperature.100μg/ml GABA completely inhibited the blue mold rot16days after inoculation of P. expansum at low temperature. The good control efficiency was observed in intact pear fruit and disease incidence reduced by22.2%compared to control. The effectiveness of induced resistance increased with the decrease of P. expansum concentration.In addition, there were no significant differences of postharvest quality parameters, among all treatments during1month of storage. GABA treatment had no adverse impact on the pear fruit quality.2. Effect mechanism of GABA on P. expansum and pear fruit2.1Effect of GABA on germination and survival of P. expansum spores in vitro and in vivo1～1000μg/ml GABA was not significantly inhibited P. expansum growth in vitro tests. Howerer,100μg/ml GABA had a significant and strong inhibition effect on spore germination of P. expansum when the time interval was increased to24h in pear fruit wounds. These results suggested that GABA reduced postharvest diseases of pear fruit by induction of host resistance.2.2Effect of GABA on ultrastructure of pear fruitThe effect of GABA on the ultrastructure of pear fruit was observed by the scanning electron microscopy (SEM) and transmission electron microscopy (TEM). The result showed that the cell wall of fruit tissue was thicker, tissue morphology and organelles was preservated better by GABA treatment after inoculated with P. expansum. Whereas, a large amount of inclusion seepaged and became fragmented. Meanwhile, the collapsed and died fruit tissue and cell occurred in the control group. The study demonstrated that GABA could increase pear fruit resistance to P. expansum at the cellular level.2.3Effect of GABA on physiological indexes related to resistance in pear fruitBy measuring8physiological and biochemical indexes related to fruit defense system and using laser confocal microscope to observe ROS in tissue cells, the result showed that a pronounced and rapid increase in the activities of enzymes (PPO, PAL, SOD, POD, CAT activity) and after treatment with GABA followed by P. expansum in contrast to the treatment with the pathogen or GABA alone, excessive ROS in cell was effectively removed, and MDA content reduced. At the same time GABA also induced the CHI and GLU activity with the direct effect on pathogen. These changed indicators were consistented with the performance of fruit resistance to P. expansum. The result explained the mechanism of induced resistance with GABA treatment was linked to the stimulation of defense-related enzyme and pathogenic-related proteins and the mechanism of priming that achieved suppression on fruit disease.2.4Effect of GABA on resistance-related genes expression in pear fruit woundsUsing RT-PCR to amplify PpPPO, PpPOD family genes, PpPAL, PpCHI family genes, PpGLU, the result indicated that single objective strap size. The expression of PpPOD3down-regulted after GABA+P. expansum treatment. Moreover, PpPOD4, PpCHI2and PpCHB expression did not markedly change. Family genes wih different structural might result in the different function. The experiment suggested PpPOD3, PpPOD4, PpCHI2, PpCHI3might not regulate the enzyme activity of POD, CHI in fruit defence system. Therefore, PpPOD1and PpCHI4genes as the target gene were quantitively determinated by qPCR.qPCR results showed GABA+P. expansum treatment remarkedly elevated the gene expression level of PpPPO, PpPOD1, PpPAL, PpCHI4and PpGLU in pear fruit, which exhibited a stronger and faster excitation than P. expansum alone. In contrast, GABA alone did not induce the expression of defense gene. Theses results further implied that the GABA induced pear fruit resistance by regulating resistance-related gene expression at the molecular level and possible priming mechanism.2.5Effect of GABA on resistance materials metabolism and GABA shunt in pear fruit Cinnamic acid and coumarin contents were significantly higher than that of the other three treatments (control; GABA or P. expansum alone) by GC-MS analysis in pear fruit tissue with GABA+P. expansum treatment. It was consistent with the resultes obtained by physiological and biochemical analysis that GABA+P. expansum could significantly induce gene expression PpPAL in pear fruit tissue and regulate the PAL activity. Therefore, PAL metabolism might be closely related to the mechanism of GABA induced resistance of pear fruit.Exgenous GABA treatment strongly activated GABA metabolism pathways under biological stress casued by P. expansum. GABA+P. expansum treatment enhanced succinic acid, glutamate and putrescine contents compared to, control, GABA or P. expansum alone treatment, followed by the generation of fumaric acid that was not detected in that of the other treatments. Moreover, the fructose content significantly increased after the infection of P. expansum, which might be correlated with the pathogenic factor (cell wall degradation enzymes); the similar result of xylitol was found, but no other polyols had the same trend. Above results might be involed in rhe different reponse mechanism of polyols under stress signals...