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Citrus is one of the most important fruits in China, and its post-harvest occurrence of citrus green mold is a serious threat to the quality and safety of citrus, and also causes mycotoxin contamination in the fruit. A strain of Streptomyces sp. R2A-15 with good antagonistic effect on the pathogen of citrus green mold was isolated from the root soil of red orange plants in the red orange orchard of Jiuzhoujiang, Lianjiang, Zhanjiang city, Guangdong province. The optimization of the fermentation conditions and stability analysis of the bacterial inhibition activity were studied to provide reference for the preparation and application of the biocontrol agent of citrus green mold. In this study, Penicillium digitatum was used as the target fungus, and the fermentation broth was used as the inhibitory activity according to the size of the diameter of the inhibition zone by paper diffusion method. The fermentation medium of strain R2A-15 was optimized using one-way and cross-combination tests, and the fermentation conditions were optimized by response surface analysis to improve the inhibitory activity of the fermentation broth, and the stability of the fermentation broth was investigated under different acid-base, temperature and ultraviolet light conditions. The optimum carbon source for the fermentation medium of the strain was cornmeal and the optimum nitrogen source was acid-hydrolyzed casein in the one-way and cross-combination tests. Based on response surface analysis, it was concluded that cornmeal and fermentation time had a significant effect on the bacteriostatic activity of the fermentation broth. The optimal fermentation medium for maximum inhibitory activity of the fermentation broth, as determined by validation tests, was 12.09 g of cornmeal, 4.00 g of acid-hydrolyzed casein, 2.00 g of calcium carbonate, 2.00 g of magnesium sulfate, 2.00 g of dipotassium hydrogen phosphate, 1.00 g of sodium chloride, and 1 L of distilled water. The optimal fermentation conditions were: initial fermentation pH 7.0, fermentation time 37.50 h, fermentation temperature 34 ℃, inoculum volume 3%, and shake flask filling volume 100 mL/250 mL. The bacteriostatic activity of the fermentation broth of the strains increased by 39% after optimization. The overall bacteriostatic activity of the fermentation broth of the strain could be maintained at a relatively stable and high activity level after treatment with different temperatures and ultraviolet light conditions, but the bacteriostatic activity of the fermentation broth decreased under strong acid and alkali. The results indicate that Streptomyces sp. R2A-15 has a good and stable inhibitory effect on the citrus green mold pathogen, and has a good prospect for practical application, which lays a foundation for the later scale fermentation as well as the development of bacterial agents.
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| 科 Family | 属数 Number of genus | 种数 Number of species | 占总种数比例 Percentage of total species (%) | 属 Genus | 种数 Number of species | 占总种数比例 Percentage of total species (%) |
|---|---|---|---|---|---|---|
| 鹅膏菌科Amanitaceae | 2 | 11 | 5.26 | 鹅膏菌属 Amanita | 10 | 4.78 |
| 小菇科 Mycenaceae | 2 | 12 | 5.74 | 丝盖伞属 Inocybe | 5 | 2.39 |
| 多孔菌科 Polyporaceae | 8 | 14 | 6.70 | 蜡蘑属 Laccaria | 5 | 2.39 |
| 红菇科 Russulaceae | 3 | 23 | 11.00 | 小皮伞属 Marasmius | 6 | 2.87 |
| 小菇属 Mycena | 11 | 5.26 | ||||
| 光柄菇属 Pluteus | 5 | 2.39 | ||||
| 红菇属 Russula | 17 | 8.13 | ||||
| 栓菌属 Trametes | 5 | 2.39 |
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