| Coastal mudflat resources are abundant in Jiangsu Province,and its rational development and utilization is an important strategic measure to achieve long-term stable development of agriculture in Jiangsu.However,low fertilization and high salinity of mudflats severely limit their extensive use in agricultural processes.Rice cultivation is a trinity of "utilization,improvement and fertilization",which has the special function of continuously optimizing and accelerating the improvement of mudflats.Previous studies have focused on the effect of rice cultivation on the soil physical-chemical properties,while few studies have been reported on the succession of microbial community structure.In this paper,we analyzed the bacterial and major N-cycling related communities in mudflats with different rice cultivation years using Illumina Miseq sequencing,quantitative real-time PCR and terminal restriction fragment length polymorphism,aimed to clarify the succession rule of bacterial and major N-cycling related communities in mudflats under long-term rice cultivation and their relationship with the accumulation of carbon and nitrogen.On this basis,the regulation of nitrogen application on bacterial community structure in newly reclaimed mudflat was studied through short-term experiment,so as to provide a scientific basis for accelerating the transformation and improvement from mudflat to cultivated land.The main conclusions are as follows:1)Long-term rice planting promoted desalination of mudflats,and increased the content of organic matter,total nitrogen,and ammonium,but the changes of pH was not obvious.The activities of alkaline phosphatase,urease,invertase,protease,and catalase were significantly increased with the increase of rice cultivation years,while the activity of polyphenol oxidase significantly decreased.The bacterial community structure in mudflats showed a significant tendency succession with the increase of rice cultivation years,and the relative abundances of Proteobacteria,Chloroflexi,and Acidobacteria gradually increased,the relative abundances of Bacteroidetes,Cyanobacteria,and Planctomycetes decreased significantly.In addition,rice cultivation significantly increased the number of anaerobic and iron reducing bacterial,such as Anaerolineae,Geobacter,Anaeromyxobacter,which may play an important role in the accumulation of organic matter in mudflats.And further promote the enhancement of microbial metabolism and the trend succession of bacterial communities in mudflat in coordination with salt desalinization2)Rice cultivation in mudflats increased the abundance of N-cycling genes,while the relative abundance showed a downward trend.The relative richness of nifH,nirK,narG in the N-cycling network also increased with rice cultivation,while the relative richness of AOA,nirS,and nosZ reduced.Moreover,the ratio of nifH to AOA and norB to nosZ increased with prolonged rice cultivation years,indicating rice planting significantly increased soil nitrogen fixation potential,nitrate consumption potential and N2O production potential.Ammonia-oxidizing archaea dominated the ammonia oxidation in mudflats;while the dominant denitrite-reducing bacteria changed from nirS-microbe to nirK-microbe.The community structure of nifH-,AOA-,nirS-,nirK-,and nosZ-microbe showed a trend changes with prolonged rice cultivation years based on T-RFLP analysis.Moreover,most of the nifH-,AOA-,nirS-,and nirK-microbes were belonged to uncultured bacteria,and the dominant nosZ-denitrifying microbes evolved from Azoarcus to uncultured bacteria.Redundancy analysis and Mantel test showed that community structure and distribution of genes in the N-cycling network were mainly affected by electrical conductivity,organic matter,total nitrogen,available phosphorus,and available potassium.3)Nitrogen increased the content of organic matter,total nitrogen,nitrate,and ammonium in newly-reclaimed mudflat paddies,and had regional(rhizosphere,bulk)differences on pH and electrical conductivity,wheres it decreased a-diversity.Nitrogen increased the relative abundance of Bacteroidetes and Actinobacteria,decreased the relative abundance of Chloroflexi,Acidobacteria,Gemmatimonadetes,Planctomycetes,a-and 8-Proteobacteria,Firmicutes showed an increasing-decreasing trend,and y-Proteobacteria showed regional differences.Principal component analysis showed that the bacterial community and its functional characteristics under high-N treatment(345-390 kg·N·ha-1)were stronger than that of low-N treatment(0-300 kg·N·ha-1),while low-N treatment in newly-reclaimed mudflat paddies was more similar to that of rice planting for nearly 20 years.Correlation analysis showed that the increase of ammonium,nitrate and organic matter caused by nitrogen application were the main driving factor of bacterial community diversity and structure.4)Net N mineralization rate and nitrification rate in the newly-reclaimed mudflat paddies decreased with the increase of nitrogen application rate,while denitrification rate showed decreased-increased in rhizosphere and showed a downward trend in bulk.Nitrogen application increased the abundance of nitrogen-fixing bacteria and denitrifying bacteria,while decreased the abundance of ammonia-oxidizing archaea;however,the response of gene proportion in the N-cycle to nitrogen application was more complex,and the response mechanism to nitrogen fertilizer varies in different regions.In addition,the ratio of nifH to amoA and {narG+napA)to(narG+napA+nirK&S+norB+nosZ)showed increasing-stable trend with the increase of nitrogen application rate;and the ratio of norB to nosZ generally remains stable,and increased sharply in rhizosphere area under high-N treatment;implying nitrogen fertilizer increased the potential of ammonium accumulation and nitrate consumption in the newly-reclaimed mudflat paddies,and increased the potential of N2O production in rhizosphere area under high-N treatment.Ammonia-oxidizing archaea dominated the ammonia oxidation in the newly-reclaimed paddies,nirS-microbe dominated the nitrite reduction.From the perspective of community structure,the community structure of nifH-,AOA-,nirS-,nirK-,nosZ-,and norB-microbe has changed correspondingly by nitrogen fertilizers,and the responses to nitrogen fertilizers was different in different regions.Phylogenetic tree showed that these N-cycling microbes were belonged to uncultured bacteria.Correlation analysis showed that the changes of nutrient content caused by nitrogen application were the main factors affecting the gene distribution in the N-cycling network,and ammonium and nitrate were the main factors driving the variation in the community structure of nifH-,AOA-,nirS-,nirK-,nosZ-,and norB-microbe.5)Correlation analysis between the characteristics of bacterial community with rice yield and nitrogen use efficiency was analyzed.?-diversity of bacterial community in rhizosphere negative effect on rice yield,possibly through nitrogen fertilizer selectively enriched the organic matter degradation bacterial,indirectly provided nutrients for rice growth,meanwhile,some bacterial that can improve the nutritional status and enhance plant resistance are enriched in rice rhizosphere,such as Nocardioides,Pontibacter,Salegentibacter,Psychroflexus,Jeotgalibacill us,Salinarimonas,Bacillus,Hydrogenophaga,Lysobacter.N-cycling genes in the newly-reclaimed paddies are closely related to nitrogen use and loss,but the influence mechanisms are different in different regions.Correlation analysis shows that the increase of N2O emission potential may be the main reason for N loss in rhizosphere of newly-reclaimed paddies,and ammonia volatilization caused by the capacity of ammonium accumulation enhanced may be the main path of N loss in bulk.To sum up,long-term rice cultivation increased soil microbial activity and material metabolism capacity of microorganisms,and drove the trend succession of bacterial and major N-cycling related communities in mudflat paddies,this was mainly related to the change of soil physical-chemical properties with the prolonged rice cultivation.The increase of ammonium and nitrate content caused by short-term nitrogen application were the main driving factor for the corresponding changes in bacterial and major N-cycling related communities in newly-reclaimed mudflat paddies.300 kg·N·ha-1 nitrogen application help to realize double safeguard called economic benefits and ecological benefits,and accelerate the ripening of newly-reclaimed paddies. |
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