| Rice is one of the main food crops in the world.The rice planting area in the world accounts for 23%of the grain planting area,and the rice yield accounts for 29%of the total grain yield.The rice planting area in China accounts for 31%of Asia.The paddy field ecosystem is an extremely special ecosystem.The formation,evolution and development of paddy field ecosystem are inseparable from by flooding irrigation,artificial tillage,rice cultivation and the required environment of rice growth.The paddy soil is developed under special soil management practices,including regular flooding,drainage,tillage,ploughing and fertilization."Tillage under flooding conditions"has always been the greatest difficult problem in the utilization of paddy soil,which leads to the destruction of soil aggregates,loss of soluble nutrients,changes in soil microbial abundance and community structure,which makes the nitrogen utilization rate of paddy soil less than half of that of dry land.In recent years,the application of a large number of chemical fertilizers in paddy fields has intensified the major ecological and environmental problems such as ammonia volatilization,N2O emission and nitrogen leaching.Combined ridge with no tillage(RNT)in paddy field is a kind of conservation tillage practices,which has changed the soil ventilation conditions and water regime,which can effectively improve the fertility status of low and medium yield paddy fields and increase crop yield in Southwest China.However,its mechanism is completely unclear,especially the effect of RNT on soil nitrogen transformation and nitrogen transformation microorganisms is still lacking in-depth research.The purpose of this study is to explore the mechanism of tillage managements affecting paddy soil fertility,to establish the quantitative relationship between soil comprehensive fertility index and crop productivity under long-term RNT,and to take nitrogen as an example to study the effect of tillage regimes on nitrogen use efficiency in paddy fields,and nitrogen loss caused by denitrification and anammox,and the abundance,activity and community structure of related microorganisms,using metagenomics method to study the effects of tillage regimes on the characteristics of nitrogen metabolism pathways in paddy soils,preliminarily explain part of the mechanism of RNT to improve soil fertility.In this study,Fuzzy comprehensive evaluation method,15N isotope labeling method(in situ and incubation),quantitative PCR and high-throughput sequencing were used to study the temporal and spatial evolution of paddy soil fertility under RNT,the relationship between soil fertility and crop productivity,the influence mechanism of denitrification and anammox,and the improvement of nitrogen use efficiency have been preliminarily studied.The major results are summarized as follows:1.After nearly 30 years of long-term tillage,there are great differences in the changes in paddy soil fertility and rice yield.Under different tillage regimes,the integrated fertility index(IFI)and rice yield of paddy fields are all expressed as:RNT>CT>FPF.The rice yield increased from 7000 kg·hm-2 in 1990 to 8993.3 kg·hm-2,8572.7kg·hm-2 and 8312.4 kg·hm-2 under RNT,CT and FPF,respectively.The average value of comprehensive soil fertility index of paddy field under RNT was 0.66,which was 0.09and 0.18 higher than that under CT and FPF respectively.It proves that long-term RNT could significantly improve the soil fertility and rice yield.2.According to the characteristics of soil nutrient interannual variation in paddy field,after nearly 30 years of long-term tillage,we found that RNT could effectively increase the contents of soil organic matter(SOM),total nitrogen(TN),alkali-hydrolyzable nitrogen(AN)and available phosphorus(AP)content in paddy field.The average contents of SOM,TN,ANand AP in paddy soil under RNT were 32.86 g·kg-1,2.20 g·kg-1,143.35 mg·kg-1 and 26.61 mg·kg-1,respectively,which were significantly higher than those under flooding paddy field(FPF)and conventional tillage(CT).The changes of SOM,TN and AK under the three tillage regimes were basically consistent.From the seasonal variation characteristics of soil nutrients in paddy field,compared with CT,FPF and RNT can effectively utilize soil nutrients such as nitrogen,phosphorus and potassium,and the seasonal variation trend of soil fertility is consistent.Tillage practices does not significantly affect the seasonal variation in soil p H and CEC,while the seasonal variation in the free iron oxide content of the paddy soil are relatively large.According to the spatial variation characteristics of paddy soil nutrients,the contents of SOM,TN,AP and free iron oxide in paddy soil all gradually decreased with the increase of soil depth,showing 0-10 cm>10-20 cm>20-40 cm.It showed that the nutrients have a tendency to enrich and accumulate towards the surface layer of the soil(0-10cm).3.It was found that ridge tillage could increase the nitrogen use efficiency(NUE)by 15N isotope labeling method(in situ and incubation).For ridge tillage,the NUE ranged from 29%to 33%of the applied N fertilizer,with an average of 31%,which was 11%and 14%higher than for FPF and CT,respectively.Water loss of applied N fertilizer was significantly different under three tillage regimes(P<0.05)and ranged from 19.5%to53.9%.The NH3 volatilization loss under ridge tillage was highest(25.4%),and was 7.1%and 12.1%higher than for flooded paddy field and conventional tillage,respectively.It shows that nitrogen loss with water and ammonia volatilization loss is the most important ways of nitrogen loss.Ridge tillage was demonstrated to be a better tillage regime for increasing NUE in the rice-based cropping systems compared to conventional tillage and the year-long flooded paddy field.4.The denitrification rate and anammox rate of paddy soil under different tillage regimes were determined by 15N isotope labeling technique.It was found that denitrification and anammox reactions occurred in paddy soils,and the order of denitrification rate and anammox rate was as follows:FPF>CT>RNT.The average value of denitrification rate ranged from 2.85 to 4.20 nmol N g-1 dry soil h-1,and the anammox rate ranged from 0.42 to 1.09 nmol N g-1 dry soil h-1.It shows that there is no significant effect on the contribution of anammox to nitrogen production in paddy soil between different tillage regimes,and it is proved that anammox is one of the important ways of nitrogen loss in paddy field.We found that denitrifying and anammox microorganisms were detected in the paddy soils under three tillage regimes by Quantitative-PCR and Illumina Hi Seq high-throughput sequencing technology.We found that the abundance of nos Z gene(DNA level)was 4.86×107-7.56×107 gene copies·g-1(dry soil),and the abundance of anammox 16S r RNA gene(DNA level)was 6.91×105-8.52×105 gene copies·g-1(dry soil).The abundance of nos Z gene was two orders of magnitude higher than that of anammox 16S r RNA gene.The transcriptional abundances of nos Z genes(c DNA level)ranged from 4.21×105-7.43×105 gene copies·g-1(dry soil)and the transcriptional abundances of anammox 16S r RNA genes(c DNA level)ranged from 3.07×105-7.44×105gene copies·g-1(dry soil).The highest abundance of the denitrifying microorganism nos Z gene and the anammox microorganism 16S r RNA gene in paddy soils under three tillage regimes were FPF,and the lowest were RNT.The nos Z gene community of denitrifying microorganisms in paddy field is closely related to Azospirillum and Rhizobium.The contents of nitrate and ammonium nitrogen are the main environmental factors affecting the community structure of denitrifying microorganisms.The dominant population of the anammox microbial community in paddy field is Candidatus Brocadia.The content of TN and p H are the main environmental factors that affect the structure of the anammox microbial community.The community structure of denitrifying microorganism nos Z gene and anammox microorganism 16S r RNA gene was not significantly affected by different tillage regimes.It is suggested that RNT may affect the denitrification and anammox of nitrogen by affecting the number of denitrifying bacteria and anammox bacteria,rather than the community composition.5.Combined with hiseq high-throughput sequencing technology,a metagenomic sequencing of paddy soils was conducted under three different tillage practices.It was found that the paddy soils under three different tillage practices have the same characteristics of nitrogen metabolism pathways.The ammonia assimilation process was the most frequently detected nitrogen metabolism pathway,while anammox process was the least frequently detected nitrogen metabolism pathway.The microorganisms of Proteobacteria can simultaneously participate in seven nitrogen metabolism pathways including ammonia assimilation,nitrate dissimilation reduction and nitrate assimilation reduction.Microorganisms from Proteobacteria,Nitrospirae,Acidobacteria and Actinobacteria can simultaneously participate in four nitrogen metabolism pathways including ammonia assimilation,denitrification,nitrate dissimilatory reduction,and nitrate assimilation reduction,reflecting the diversity of nitrogen metabolism in paddy soils,reflecting the functional diversity of nitrogen metabolism in paddy soil.Under different tillage practices,the same nitrogen metabolism process in paddy soil can be participated by different microorganisms,and the community structure of microbial responsible for the entire nitrogen metabolism pathway is not the same completely.In summary,compared with FPF and CT,long-term RNT significantly improved soil fertility and crop productivity in purple paddy soil ecosystem,reduced nitrogen loss and improved nitrogen use efficiency,and achieved the purpose of soil fertility by using tillage regimes.Although the present study proposes a promising tillage for improving N fertilizer use efficiency in rice-based cropping systems,more information is necessary before widespread recommendation.Further research at a larger field scale across multiple growing seasons,and for more diverse soil types,would be necessary. |
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