| Nitrogen is one of the major elements essential for the growth and development of Potato (Solanum tuberosum), therefore adequate nitrogen nutrition must be supplied to achieve high yield, whereas high dosages of nitrogen application is easy to cause the serious occurrence of potato late blight, resulting in a substantial reduction of the yield. From the health and green plant protection point of view, it is necessary to explore the appropriate nitrogen level for both obtaining a higher yield and giving full play to the nitrogen fertilizer to plant resistance. In this study, indoor potted and field plots controlled experiments were combined, integrated research methods such as:PCR-DGGE technique, physiological and biochemical detection means,15N isotope tracer technique, microbial isolation and culture method were used, the effects of different nitrogen levels on potato rhizosphere microbial quantity and richness, soil enzyme activities, in vivo resistance related enzyme activities and the content of resistance related substances, the degree of Phytophthora infestans and its effect on yield were discussed, and the appropriate level of nitrogen application range of 90～135 kg · hm-2 and the optimal nitrogen application level of 135 kg · hm-2 were determined. The results of this study are as follows:1. There are 46.4%～56.3% bacteria,69.0%～82.1% fungi, and 53.6%～61.2% actinomycetes were distributed in 0～10cm soil layer, only 17.2%～21.8% bacteria,4.6%～8.1% fungi, and 9.2%～18.1% actinomycetes distributed in 20～30 cm soil layer. The vertical distribution of soil enzyme activity is consistent with the distribution of microorganisms in the soil. During the potato growth period, the amount of soil bacteria, Bacillus spp. and Actinomycetes increased first and then decreased, soil bacteria peaked at 61 d after sowing and Actinomycetes peaked at 75d after sowing, respectively; total number of fungi and Trichoderma spp. maximized on 43d after sowing, and then gradually reduced; the number of Fusarium spp. and Rhizoctonia solani was gradually increased, but the changes were not significant. The results of PCR-DGGE of potato rhizosphere soil bacterial communities showed that each different growth stages of potato had their own particular DGGE bands profiles. This vertical distribution of soil microorganisms and its dynamic changes over time were unrelated to nitrogen levels.2. The appropriate amount of nitrogen was helpful to increase beneficial soil microorganisms and inhibit the number of plant pathogens, to increase the richness of the soil bacteria, and to improve the activities of soil urease, invertase, phosphatase and catalase. When the supplied nitrogen level was 90～135 kg·hm-2, the total amount of bacteria, fungi, Actinomycetes, Bacillus spp and Trichoderma spp. was significantly increased in each potato rhizosphere soil tilth layers at different growth stages, while the number of Fusarium spp. and Rhizoctonia solani significantly reduced, especially for deep soil layer where total number of bacteria and Bacillus spp spores was smaller under natural conditions, and the improving effects on early and late growth stages of potato were more obvious, but fluctuated at different growth stages. When the nitrogen level was above 157.5 kg · hm-2, the total number of bacteria and Bacillus spp showed fluctuations, the total number of fungi, Fusarium spp. and R. solani increased greatly, and a relatively small increase in the amount of Trichoderma spp. was presented. There were big differences among band numbers and brightness of DGGE profiles of rhizosphere soil bacteria communities under different nitrogen treatments, the number of DGGE bands was at its maximum when the nitrogen level was at 135 kg · hm-2, the same as the richness, diversity and evenness index, and these parameters were significantly higher on 61 d and 75d after sowing than other nitrogen levels. Nitrogen level at 67.5～135.0 kg · hm-2 was helpful to improve the soil urease activity, nitrogen of 112.5～157.5 kg· hm-2 was helpful to improve the soil phosphatase and catalase activities at all growth stages and sucrase activity at early growth stage. During the potato growth and development, a variety of soil enzyme activities showed a trend of increased first and then decreased.3. The vertical distribution of 15N fertilizer in the soil showed dynamic changes over time. The content of nitrogen fertilizer in the soil (Nspf) and the percentage of nitrogen fertilizer in total soil nitrogen (Ndffs) was high in 0～10cm shallow soil layer on 45d after sowing, and smaller on 75d after sowing; Nspf and Ndffs decreased slowly in 10-20cm soil layer; while in 20-30cm soil layer, Nspf and Ndffs increased first and then decreased. The value of Nspf and Ndffs had influences on the total number of the soil bacterium, Bacillus spp quantity, total fungi, Trichoderma spp. quantity and phosphatase activity, while had little effects on Actinomycetes and other soil enzyme activities. Total nitrogen content of the soil had a certain impact on soil phosphatase and sucrase activities.4. Under the natural occurrence of diseases, nitrogen was applied to increase the plants PPO, PAL, POD,β-1,3- glucanase and chitinase activities, and to improve the total phenols, chlorogenic acid, flavonoids, soluble protein and soluble sugar content. The effect of nitrogen application of 135 kg·hm-2 was stronger and more stable, while the effect of nitrogen application of 180 kg·hm-2 was also very strong but unstable. The different levels of nitrogen fertilizer application had certain effects on CAT and SOD activities and MDA content, but not stable. After artificial inoculation of P. infestans, there were differences among the testing potato varieties. The potato variety Daxiyang had a rapid response in POD activity, and the activity of CAT was significantly higher than that without nitrogen application, the response of total phenol content was slower but stable, and the increase of soluble protein content was relatively high; the variety Kexin 18 also had a fast response in POD activity, the activity of CAT was significantly lower than that of no nitrogen, the response of total phenol content was fast and stable, and the increase of soluble protein content was significant; whereas for variety Kexin 1, the POD response was slow, the activity of CAT was significantly higher than that of no nitrogen, the response of total phenol content was fast and but less stable, and the increase of soluble protein content was relatively low.5. Potted plants and field experiments showed that the pure nitrogen application of 135 kg· hm-2 resulted in lower severity of disease incidence, the highest yield and larger tubers; nitrogen of 180 kg · hm-2 resulted in higher severity of disease incidence, lower yield and smaller tubers. More appropriate nitrogen application range was 90～135 kg·hm-2, and the optimum nitrogen application level was 135 kg· hm-2. |
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