Evaluation on bacterial canker disease and analysis of physiological indexes in main kiwifruit cultivars

Abstract: 【Objective】Kiwifruit bacterial canker disease is a devastating disease with a wide range, strong pathogenicity, rapid spread and difficult eradication, which is caused by Pseudomonas syringae pv. actinidiae. Although extensive research has conducted on the classification, biological characterization, pathogenicity analysis and rapid detection methods, there is still a lack of effective control technology in the production. Screening and analyzing the resistance mechanism of high- resistant varieties is crucial for the prevention and control of kiwifruit bacterial canker disease.【Methods】Seventy-five varieties of Actinidia chinensis var. chinensis and Actinidia chinensis var. deliciosa were used as materials. Healthy canes were collected after leaf fall. Canes with a diameter of about 0.8 cm and consistent growth were selected and cut into 10 cm stem. Both ends were sealed with paraffin to minimize water loss. A perforator was used to make a 3-mm-width xylem-deep wound on the cane. The wounded cane was inoculated with bacterial suspension (OD600=1.0), and sterile water was used as a negative control. Forty-two days after inoculation, the surface was removed with a knife and the length of the lesions was measured with a vernier caliper for preliminary evaluation of resistance. Then, nine varieties with significant resistance differences were further identified by the isolated leaf disc method. New healthy kiwi-fruit leaves of uniform size and growth were collected and leaf discs were prepared by the perforation method. After suspension vacuum infiltration and washing with sterile water, the leaves were placed with dorsal side up in petri dishes in an artificial climate chamber with L/D=16 h /8 h at 16 ℃, and the lesions were counted after 5 days of inoculation. Finally, resistance-related physiological and biochemical indices of the nine varieties were determined. The total phenol content was determined spectrophotometrically by adding reagents and comparing the absorbance values of different varieties’solutions at 760 nm; soluble sugar contents were measured by anthrone colorimetry at 630 nm for each variety; lignin content was determined by double-antibody one-step sandwich enzyme-linked immunosorbent assay (ELISA) at 450 nm.【Results】Brown lesions could be observed at the inoculation site forty-two days after inoculation, extending from the incision site to both ends. The length of the lesions of different varieties showed significant differences, ranging from 5 mm to 16 mm. 75 kiwifruit germplasm resources could be categorized into four classes, i.e., resistant, tolerant, susceptible and highly-susceptible according to the lesion length, which contained 16 resistant resources, 30 tolerant resources, 19 susceptible resources and 10 highly-susceptible resources. According to the results of the resistance evaluation of isolated canes, nine varieties with different resistance were selected for the isolated leaf disc resistance evaluation experiment to further verify their resistance. Leaf discs inoculated with the suspension would gradually produce black-brown spots after incubation, whereas the negative control of leaf discs inoculated with water basically showed no symptoms. After 5 days of light incubation, there was a difference in the area of leaf discs affected by different varieties. Among the nine varieties, Jin Kui and Cui Yu had the smallest percentage of spots after inoculation, being 3.51% and 5.48% , respectively, which were below 6% . Donghong and Hongyang had the most serious disease, with 45.79% and 46.75% of spots. The correlation analysis between the percentage of leaf disc spots and the lesion length on cane spots of different varieties showed a highly significant correlation of 0.948, confirming the reliability of both methods for resistance identification. Physiological indexes were measured on leaves of the above nine varieties with differences in resistance, and differences were found in the physiological data among different varieties. The values of total phenol content ranged from 3 to 6 mg · g- 1 , with the resistant variety Cuiyu having the highest total phenol content of 5.7 mg · g- 1 , and the highlysusceptible variety Hongyang having the lowest total phenol content value of 3.07 mg·g-1 . With the variety resistance decreased, the total phenol content showed a decreasing trend. Similarly, the resistant variety Cuiyu had the highest soluble sugar content, with roughly 29.88 mg · g-1 , but the highly-susceptible variety Hongyang had the lowest soluble sugar content of 18.34 mg · g- 1 ; the soluble sugar content also showed a decreasing trend with the weakening of resistance. Regarding lignin content, the resistant variety Jinmei had the highest content of 9.59 mg of lignin per 100 g of fresh leaves, followed by Cuiyu and Jinkui, which were all resistant varieties. As the varieties’resistance decreased, the lignin content also reduced, and Hongyang had the lowest lignin content, with only 8.87 mg per 100 g of fresh leaves. In summary, it can be seen that the contents of total phenolics, soluble sugars and lignin differed among the different varieties, but the contents of resistant varieties were significantly higher than the tolerant and sensitive varieties.【Conclusion】The screened resistant germplasm lays a material foundation for the breeding and mechanism research on resistance to kiwifruit bacterial canker disease. There was a significant negative correlation between disease resistance and in vitro leaf disc resistance, and a significant positive correlation with physiological indicators, which can be used as a rapid identification method for resistant germplasm.