Research On Characteristics Of Nitrification In Rhizosphere Soil Growing With Different Rice Cultivars

More and more evidence shows that rice plants may take up more nitrate (NO3) than we expected. Hydroponic experiments show that rice growth, yield and nitrogen (N) use efficiency (NUE) can be much improved when the plant is provided with both NO3" and ammonium (NH₄⁺) compared with that fed solely on NH₄⁺. Rice plants have aerenchyma in their shoots and roots that allow oxygen to diffuse down into the rice roots, and partial oxygen is released into the soil, and thus nitrification occurs immediately in niche of the rhizosphere or on the surface of the roots. Therefore, even in a flooded paddy soil, rice roots are actually exposed to a mixed N supply (NH₄⁺+ NO_<sup>-), although the predominant species of mineral N in bulk soil in paddy rice field is likely to be NH₄⁺. This raised several questions such as whether there is a difference of nitrification activity happened in rhizosphere soil growing with different rice cultivars, or whether there is a relationship between the NUE and the nitrification activity in the rhizosphere of different rice cultivars; what is the effect on nitrification in the root rhizosphere to N nutrition of rice plants in soil culture condition; whether there has an effect of N fertilizer application on nitrification; whether there is difference of nitrification activities happened in different place, such as root surface, rhizosphere and bulk soil, or which place is more important to rice N nutrition; what is the reason to cause the difference of nitrification activities among different rice cultivar. To answer these questions, a rhizobox with three compartments and subsequent soil-slicing after quick freezing was used to measure simultaneously the spatiotemporal variations of mineral nitrogen, nitrification and AOB in the rhizosphere soil of two group of rice cultivars Yangdao 6 (Indica) and Nongken 57 (Japonica). Rice growth, N accumulation, NUE and root porosity (POR) were also examined to clarify the importance of nitrification in rhizosphere soil to rice N nutrition Three Japonica rice cultivars (4007, Wuyunjing7 and Elio) with different NUE were used to study rice grain yields, total N accumulation and the nitrification characteristics under three N treatments, such as zero N level (0 kg N hm^-2), moderate N level (180 kg N hm^-2) and high N level (300 kg N hm^-2) in field conditions. This is to further examine the effect of N application on fertilizer-NUE, nitrification and nitrifying microorganisms in rhizosphere soil growing with different rice cultivars. Finally, the high NUE cultivar (4007) and low NUE cultivar (Elio) were used to illustrate the relationship between characteristics of nitrification and NUE in field condition. The main results obtained were as followed.1. The fresh and dry weights of Indica were significantly higher than those of Japonica at the whole incubation periods. The variation trends of the root/shoot ratio of dry weight were different between the two rice cultivars and the root/shoot ratio of Indica, for instance, decreased with the development of the incubation time, while the reverse was true for the Japonica. The root/shoot ratio of Indica was significantly higher than Japonica. N accumulations of the Indica and Japonica were significantly increased with the incubation time, while the NUE were significantly decreased with the time passed. The N accumulations and NUE of Indica were significantly higher than those of Japonica during the three sampling dates. GSA and NRA of leaves were always higher than those of roots for the two rice cultivars. The leaf GSA and NRA of Indica were significantly higher than those of Japonica, while there were no significant differences of the root GSA and NRA between Indica and Japonica.2. The main nitrogen form was NH₄⁺-N in flooded paddy soil and NH₄⁺-N concentrations in bulk soil showed almost no changes with incubation time, but NH₄⁺-N concentrations increased with the distance from root surface of both rice cultivars. The NH₄⁺-N concentration of both Yangdao 6 and Nongken 57 in the zone 40 mm from the root at 51 days after sowing, for example, achieved 13.8 and 14.6 mg kg^-1 soil, respectively. However, the NO₃^--N concentration decreased significantly with the development of the incubation time, although the distribution of NO₃^--N was even in the bulk soil. The average NO₃^--N concentration for both cultivars was 0.05 mg kg^-1 soil. When the two varieties were compared, the NH₄^--N concentration was almost the same, while NNO₃^--N concentration was significantly different at every sampling time.The nitrification activities of both rice cultivars increased with incubation time. Maximal nitrification activities were found in rhizosphere soil, followed by those in the bulk soil and in the root surface in every sampling. In the rhizosphere the nitrification activities decreased with increased distance from the root. The maximal nitrification activity measured at 44, 51 and 58 days after sowing of Yangdao 6 and Nongken 57 rice cultivars was at a distance of 6 mm and 2 mm from root surface, respectively. The maximal nitrification activity values measured were 0.88 and 0.73 mg kg^-1 h^-1, respectively. The AOB in the root surface, rhizosphere (0-4 mm away from root surface) and bulk soil (>4 mm away from root surface) for both rice cultivars increased during the growth periods. The AOB in the root surface was always the lowest, while that in the rhizosphere soil was the highest in both cultivars. In these experiments, the nitrification activities measured were significantly proportional to AOB (r=0.86, p<0.01). The nitrate concentration, nitrification activities and AOB of Indica were always higher than those of Japonica rice. Therefore, nitrification in rhizosphere had more important significance for rice N nutrition, especially for the Indica rice cultivars.3. The root biomass (fresh and dry weights), POR, and rhizosphere soil nitrification activity of Yangdao 6 were higher than Nongken 57 at 44, 51 and 58 days after sowing. The POR values were significantly and positively correlated with the nitrification activity (r=0.763, p<0.01). Scanning electron microscope (SEM) results indicated that aerenchyma was not fully developed in the tissues close to the roots apex (0-5mm behind the root tip) of both Yangdao 6 and Nongken 57. However, at the point of 10mm behind the root tip of Yangdao 6, developing aerenchyma was observed but not yet for Nongken 57. At the point of 15 mm behind the root tip, well-developed aerenchyma was found in both rice cultivars. There were two distinct patterns of lysigenous origin of lacunae between two rice cultivars: radial lysigeny (Yangdao 6) and tangential lysigeny (Nongken 57). The POR and rhizosphere nitrification activity increased with the rice plant development. Therefore, rice cultivar with well-developed root system (having high root biomass) and aerenchyma (having high POR) lead to more root oxygen release to the soil, which resulted in more AOB survived in the rhizosphere soil and consequently led to stronger nitrification activity in the rhizosphere.4. There were significant differences of rice grain yields among the three rice cultivars under different N application rates. The maximal and minimal grain yields were obtained in 4007 in the moderate N level and in Elio in the zero N level, achieving 11117 kg hm^-2 and 5322 kg hm^-2, respectively. There were significant differences of the total N accumulation among the three N treatments, and the total N accumulation increased with the increase of the N fertilizer application rates. Significant differences were found in the fertilizer-NUE (FNUE) and rice grain yields among the three rice cultivars under different N application rates. For example, the FNUE of 4007 was always significantly higher than those of the Wuyunjing 7 and Elio in both moderate and high N level treatments, and the average FNUE in the high N level treatment was 42.2% lower than that of the moderate N level treatment. Based on the FNUE and grain yield at zero N fertilization level, the three rice cultivars could be classified into efficient and responsive (4007), efficient and nonresponsive (Wuyunjing7) and nonefficient and nonresponsive (Elio) to N fertilizers.Under the water management of alternation of wetting and drying during the middle-late rice growing stages, the main N form in the rice growing rhizosphere soil was NO₃^--N in the zero and moderate N level treatments, while NH₄⁺-N was the main N form in the high N level treatment. The contents of NH₄⁺-N and NO₃^--N in the rhizosphere soil increased with the increase of the N fertilizer applications. For example, the average contents of NH₄⁺-N at the zero, moderate and high N level conditions were 0.88, 0.94 mg kg^-1 and 13.5 mg kg^-1, respectively, while those of NO₃--N were 1.61,1.73 mg kg^-1 and 2.33 mg kg^-1, respectively. The nitrification potential in the rice growing rhizosphere soil represented significant differences among the three rice cultivars, and the average values were 6.94, 5.46μg kg^-1 h^-1 and 2.42μg kg^-1 h^-1 for 4007, Wuyunjing 7 and Elio, respectively, under all the N application levels. The abundance of AOB in the rhizosphere soil of Elio was significantly lower than those of 4007 and Wuyunjing 7 under the different N application rates. The maximal abundance of AOB was 2.02×10⁶ g^-1 soil in the rhizosphere soil of 4007 at high N level, while the minimal one was 1.89×10⁶ g^-1 soil in the rhizosphere soil of Elio at moderate N level. The nitrification potentials in rhizosphere soil were significantly correlated with the rice grain yield at zero, moderate and high N levels (r=0.799^** 0.877^** and 0.934^**, respectively), and also they were significantly correlated with the N physiological efficiency at moderate N level (r=0.735^*). Furthermore, the abundances of AOB in the rhizosphere soil were correlated with the nitrification potentials and the grain yields. These results inferred that there should be a relationship among rice yields, FNUE and nitrification potential in rhizosphere of rice plants. 5. The pH values in root surface soil were significantly lower than those in rhizosphere and bulk soil in Heading, filling and harvesting stages, ranging from 5.95 to 6.84. NH₄⁺-N concentration decreased but NO₃^--N increased with the time. N application increased NH₄⁺-N and NO₃^--N concentrations. Depletion sections of both NH₄⁺ and NO₃^- were found in root surface soil. The NH₄⁺-N concentration increased with increasing distance from the root surface. The maximal NO₃^- concentration was in rhizosphere soil, then the bulk soil and the lowest was in root surface soil. Nitrification activities in both root surface and rhizosphere soils significantly decreased with the incubation time, but the reverse was true with the bulk soil. N application improved nitrification activities in root surface soil growing with 4007 both at heading and harvesting stages, and also improved nitrification activity in rhizosphere soil growing with Elio at heading stage. But there was no significant difference between N180 and N300 treatments. The nitrification activity showed such order as rhizosphere>root surface>bulk soil in the whole sampling stages. AOB abundance in both root surface and rhizosphere soils significantly decreased with the incubation time, while those in the bulk soil indicated no difference as the time passed. For example, the AOB abundances in root surface soil at heading, filling and harvesting stages were 16.7, 8.77 and 8.01×10⁵ g^-1 dry soil, respectively. There was no significant difference of AOB abundance between root surface soil and rhizosphere soil, but they were all significantly higher than those in the bulk soil. As far as the two rice cultivars were concerned, there was no difference with the soil pH values. The 4007 growing soil NH₄⁺-N concentration was higher than Elio. NO₃^--N concentrations in root surface, rhizosphere and bulk soils for Elio growing treatment under N180 level at heading stage were higher than those for 4007. But NO₃^--N concentrations in root surface, rhizosphere and bulk soils for Elio growing treatment at filling and harvesting stages were significantly lower than those for 4007. Nitrification activities and AOB abundance in bulk had no difference among the three N treatments. Nitrification activity and AOB abundance in root surface and rhizosphere soil for Elio growing treatment were significantly higher than those for 4007 before filling stage, while the reverse was true after filling stage, and the rice yield and NUE for Elio were much lower than 4007. That might be due to the higher nitrification and higher NO₃^--N concentration in thizosphere soil for 4007 than Elio after filling stage, which caused more NO₃^--N absorption by 4007 than Elio. Thereby nitrification and AOB abundance in root surface and rhizosphere soil at filling and harvesting stages were important to a high rice yield and high NUE.