Changes of key enzymes and secondary metabolites in the phenylpropane metabolic pathway of fruit black spot disease in walnut

【Objective】Walnut bacterial blight (Xanthomonas arboricola pv. juglandis) poses a serious threat to the yield and quality of Juglans regia L. In the process of disease resistance, the plant's own metabolic system plays a crucial role. The study aimed to investigate the effects of black spot disease on osmoregulatory substances, oxidative substances, key enzymes and secondary metabolites in phenylpropane metabolism pathways during different stages of fruit development in walnut.【Methods】The fruits of Juglans regia L.‘Qingxiang’was used as the experimental materials. The disease- free (CK) and black spot infected (Xaj) fruits were collected at 30, 60 and 90 days after fruit setting, respectively. The contents of osmoregulation substances (SS, SP, pro) and oxidation substances (MDA) in fruits, the activities of key enzymes (PAL, C4H, 4CL) in phenylpropane metabolic pathway and the contents of secondary metabolites (lignin, flavonoids, TP) were determined.【Results】The results showed that as the disease progressed, the accumulation of soluble sugar (SS), soluble protein (SP), proline (Pro), and malo-ndialdehyde (MDA) in walnut fruits increased as the diaease progresses of the black spot after infection; The activities of PAL and 4CL, as well as the contents of lignin and TP, increased significantly (p＜0.01), while the activities of C4H and the content of flavonoids increased extremely significantly (p＜0.05) during the course of the disease infection, reflecting the role of the phenylpropane metabolic pathway in the resistance of walnuts to black spot disease. The SS content of the fruits infected by the disease decreased by 24.36%, increased by 30.48% and decreased by 21.00%, compared with that of the CK 30 days, 60 days and 90 days after infection. The SP content of the fruits infected by the disease increased by 30.46%, increased by only 2.23% and decreased by 77.74% compared with that of the CK 30 days, 60 days and 90 days after infection. The MDA content of the fruits infected by the disease was 200.98% higher than that of the CK 30 days after infection, 2.34% higher than that of CK 60 days after infection, and 16.33% lower than that of the CK 90 days after infection. The results indicated that black spot disease had a significant periodic effect on the activities of PAL, C4H, and 4CL key enzymes in the phenylpropanoid metabolism pathway of walnut fruit, as well as on the content of lignin, flavonoids, and TP in secondary metabolites. The activity of PAL and 4CL, as well as the content of lignin and TP, showed a significant increase (p＜0.01) in the infected fruit, reflecting the key role of these enzymes in resistance to black spot disease. The activity of C4H enzyme and the content of flavonoids also significantly increased in the subsequent course of the disease progress (p＜0.05), indicating that they would play an important role in the walnut fruit’s response to pathogen infection.【Conclusion】Under the stress of black spot disease, Qingxiang walnut fruit responded to the disease by adjusting the content of osmotic adjustment substances. The changes of key enzyme activities in phenylpropane metabolic pathway promoted the production of a variety of secondary metabolites, which jointly participated in the response and adaptation process of the fruit to black spot disease. This pathway played an important role in walnut resistance to black spot disease.