Soil pH could be greatly affected by agriculture practices in paddy soils, in which microbial Fe(III) reduction is a prevalent and important biogeochemical process. There is increasing evidence that microbial fermentative processes can play an important role in Fe(III) reduction. In this study, the response of metabolically active Clostridium community to initial pH adjustment was investigated to elucidate the mechanism of the effects of soil pH on microbial Fe(III) reduction through fermentative microbes.

针对传统实验教学方法的弊端,对基因工程实验课程进行了系统改革,通过设计综合型大实验体系,引入多媒体、讨论式和总结式教学方法,充分调动了学生主动学习的积极性,提高了学生的综合实验技能,同时将科研实践融入实验教学中,通过几年的教学实践,受到各届学生和国内同行的一致好评.

Microbial Fe(III) reduction can make an excellent contribution to the bioremediation of contaminated environments and potentially reduce methanogenesis. Excessive input of phosphorus (P) by P fertilizer application and eutrophied irrigation water might have a substantial influence on the process of microbial Fe(III) reduction in flooded paddy soils. To evaluate the effect of P application on microbial Fe(III) reduction, the responses of Clostridium and Geobacteraceae communities to different concentrations of P addition (CK: 0mmolPkg-1 soil; P1: 3.3mmolPkg-1 soil; P2: 20mmolPkg-1 soil) were investigated in anaerobically incubated paddy slurries. P addition significantly inhibited Fe(III) reduction during the early stage of incubation (from days 0 to 20). Compared with the CK treatment, the maximum Fe(III) reduction rate (Vmax) in treatments P1 and P2 remarkably decreased by 0.281 and 0.439mg·g-1·d-1, respectively. However, the addition of P had no significant effect on Fe(III) reduction during the later stage of incubation (after 20days). The abundances of Clostridium and Geobacteraceae were suppressed by P addition, and the suppression effect was more obvious with higher P concentration. P addition significantly changed the community structures of Clostridium and Geobacteraceae during the entire incubation. The communities of Clostridium and Geobacteraceae were closely correlated with the process of Fe(III) reduction. In conclusion, P addition could inhibit the microbial reduction of Fe(III) during the early stage of incubation by reducing the abundances and altering the community structures of Clostridium and Geobacteraceae, however, the inhibition could be eliminated with increased incubation time. This study demonstrates that soil microbial communities are sensitive to excessive P application, which can jointly impact relevant biogeochemical processes in flooded paddy soils.

Microbial Fe (III) reduction is an important biogeochemical process in anaerobic environments, and it can be regulated by adding organic carbon. Many researchers have studied the unique mechanisms and communities of respiratory Fe (III) reducers. However, the contributions and communities of fermentative microbes have rarely been studied. This study aimed to investigate the response of the metabolically active Clostridium community to glucose additions and to assess its contribution to microbial Fe (III) reduction in flooded paddy soils.

为了营造一个良好的高校生物化学优质课堂教学,创新教育教学理念,激发和调动学生学习化学的积极性和主动性. 要从四个方面进行改革:教学内容,教学方法,教学手段,教学评价进行探讨,以不断推动和提高高校生物化学教学和化学学科素质水平.

基因工程是生物技术专业的必修课程,为了满足创新型人才培养的需要,我们应对基因工程教学内容体系、教学方法、教学手段和考核方式等方面进行改革,从而提高了基因工程课程的教学质量,全面提升学生的综合素质和创新能力.

水稻土是非常复杂又典型的生态系统,分析淹水培养过程中水稻土细菌的丰度和群落结构变化规律,可以客观反映水稻土中细菌群落结构信息,为深入探讨水稻土细菌微生物对稻田的影响和在生态系统中的作用(营养元素转换、重金属还原与抑制甲烷生成过程等)提供实验基础与理论依据.作者采用淹水非种植水稻土微环境模式系统,提取水稻土淹水培养1h和1、5、10、20、30、40、60 d后的微生物总DNA,利用Real-time PCR和PCR-DGGE(denaturing gradient gel electrophoresis)技术检测了淹水培养过程中细菌丰度与群落结构的变化.结果表明:淹水水稻土中细菌的丰度在1d时最大,并在40 d到达第二个峰值,说明淹水过程改变了细菌的丰度.基于16S rRNA基因V3区的DGGE图谱分析显示,淹水过程中细菌的群落结构发生了演替性变化:r-策略生存的细菌仅存在于淹水初期;k-策略生存的细菌存在于淹水后期;r-和k-策略共生存的细菌存在于整个淹水过程中,淹水后期k-策略的细菌占据优势.淹水培养过程中优势种群多样性指数大体呈现先上升后减小的趋势.主成分分析(PCA)将淹水处理过程分成几类不同的生境,反映出中、后期细菌群落结构较为稳定;测序结果表明,32个优势条带所代表的细菌分别属于厚壁菌门、绿弯菌门、拟杆菌门、变形菌门和酸杆菌门,且与来自不同地域的水稻土、其他类型土壤、活性污泥以及湖泊沉积物等生态系统的细菌关系密切.

Ammonia-oxidizing archaea(AOA)play an important role in ammonium oxidation in soil ecosystem, and predominate among ammonia-oxidizing prokaryotes in paddy soils. In this study, dynamic changes of abundance and community structures of ammonia-oxidizing archaea were investigated in paddy soils that were flooded for 1 h, 1 d, 5 d, 10 d, 20 d, 30 d, 40 d and 60 d, using sequential analysis and real-time PCR. The abundance of bacteria was 29 times that of crenarchaeota, while AOA was 4 times ammonia-oxidizing bacteria(AOB). Based-on arch-amoA gene, OTU analysis showed that the AOA community structures shifted at different flooding times:T12, a type of AOA and r-strategist organism, was present only at the early flooding time. T4, T5 and T9, k-strategist organisms, existed at the late flooding time. T1, T8 and T16, r-k-strategist symbiotic organisms, appeared during whole flooding period. AOA predominated at the late flooding time. The diversity index of dominant groups was larger at the early flooding than at the middle and late flooding times. Sequencing results showed that all 16 dominant OTU types belonged to crenarchaeota, and had a close relationship with AOA from paddy soil, dry highland soil, red soil and sediments in different regions.

SummarySoil interrill erodibility is a key component of soil erosion models. However, when using aggregate stability to assess soil erodibility, samples are usually collected from the plough layer, while soil erosion occurs at the soil surface. Hence, the potential changes in erodibility caused by crusting are ignored. Moreover, soil interrill erodibility is difficult to predict accurately. This lack of predictability means that current erosion models use a constant erodibility value for a given soil, and thus do not consider potential heterogeneity of erodibility. This study was conducted to (i) assess the heterogeneity of aggregate stability for a crusted soil and (ii) relate this heterogeneity to the aggregate stability of the underlying material (sub‐crust) and to standard soil properties. A field study was conducted in a small area of the Loess Plateau in China in which the crust and the sub‐crust soils were sampled. Standard soil properties (organic matter content, sand content, silt content, clay content, cation exchange capacity (CEC), pH in water, and water content at the time of sampling) were measured as potential explanatory factors of aggregate stability. The results showed a large heterogeneity in aggregate stability among the sites, even though the sites had the same soil type. The mean weight diameter (MWD) of the crust varied between 0.33 and 2.04 mm while the MWD of the sub‐crust varied between 0.23 and 1.42 mm. Soil texture and pH were uniform among the sampling sites, whereas water content, organic matter content and CEC varied more. Even though some correlations existed (for example r = 0.57 between MWD for the slow wetting test and organic matter content), none of the standard soil properties was able to predict aggregate stability accurately. The aggregate stability of the crust was significantly greater than that of the sub‐crust. The large differences in aggregate stability imply large differences in soil interrill erodibility. Because a single soil type was investigated, this finding proves that erodibility can vary greatly in space even for a given soil type. Soil interrill erodibility should be estimated from the exact material exposed to erosive forces, the soil surface material. Using the sub‐crust would have led to greatly over‐estimated erodibility and thus to a marked bias in erosion model predictions.RésuméLa stabilité structurale d'un sol encroûté : différences entre la croûte et le matériau sous‐jacent, et conséquences pour l'estimation de l'érodabilité inter‐rigole. Un exemple dans le Plateau de Loess (Chine)L'érodabilité inter‐rigole est un paramètre clef des modèles d'érosion du sol. Cependant, lorsque des tests de stabilité structurale sont utilisés pour évaluer l'érodabilité, les mesures sont habituellement réalisées sur des échantillons prélevés dans l'horizon labouré alors que l'érosion a lieu à la surface du sol. Ainsi, les changements potentiels d'érodabilité causés par la formation de croûte sont ignorés. De plus, l'érodabilité inter‐rigole reste encore difficile à prédire avec précision. Ces difficultés conduisent les modèles d'érosion à utiliser une érodabilité constante pour un type de sol donné, et donc à ne pas considérer l'hétérogénéité potentielle de l'érodabilité. Cette étude a été conduite pour (i) évaluer l'hétérogénéité de la stabilité structurale pour un sol encroûté et (ii) relier cette hétérogénéité à la stabilité structurale du matériau sous‐jacent (sous‐croûte) et aux propriétés standards du sol. Une étude de terrain a été réalisée sur un secteur de surface limitée du Plateau de Lœss (Chine). Des échantillons provenant de la croûte et de la sous‐croûte ont été collectés. Les propriétés standards (teneur en carbone organique, teneurs en sable, limon et argile, CEC, pH, et teneur en eau au prélèvement), ont été mesurées en tant que facteurs explicatifs potentiels de la stabilité structurale. Les résultats ont montré une grande hétérogénéité de la stabilité structurale entre les différents sites alors que ces derniers présentaient le même type de sol. Le MWD de la croûte variait entre 0.33 et 2.04 mm tandis que le MWD de la sous‐croûte variait entre 0.23 et 1.42 mm. La texture du sol et le pH étaient très homogènes entre les sites étudiés, tandis que la teneur en eau, la teneur en matière organique et la CEC variaient plus fortement. Bien que certaines corrélations aient été identifiées (par exemple r = 0.57 entre le MWD du test à l'humectation lente et la teneur en carbone organique), aucune de ces propriétés n'a permis de prédire précisément la stabilité structurale. La stabilité structurale de la croûte était significativement supérieure à celle de la sous‐croûte. Les grandes différences de stabilité structurale mesurées impliquent des érodabilités très contrastées. Comme un seul type de sol a été étudié, ce résultat prouve que l'érodabilité peut être très variable spatialement pour un type de sol donné. L'érodabilité inter‐rigole du sol devrait être mesurée sur le matériau exact qui subit l'érosion, c'est‐à‐dire le matériau de surface. L'utilisation du matériau sous‐jacent aurait engendré une forte surestimation de l'érodabilité et donc un biais important dans les prédictions d'un modèle d'érosion.

Bacillus is one of the best characterized bacterial genera.It is defined as a gram-positive,rod-shaped bacterium which can be aerobic or facultative anaerobic.Bacillus can produce highly resistant dormant endospores in response to nutritional or environmental stresses.Since the late 19t h century,the long history of Bacilli research has included classical microbiology,biochemistry,and more modern genomic and proteomic approaches.Flooded rice field has become a model system for the study of soil microbial ecology.In this study,the total DNA was extracted from six paddy soil slurries including 1 h,1 d,5 d,10 d,20 d and 30 d of flooding incubation.The changing characteristics of community diversity and succession of Bacillus were analyzed by using Denatured Gradient Gel Electrophoresis(DGGE) based on 16S rRNA gene(rDNA) of Bacillus.The influence factors on characteristics of community diversity and succession of Bacillus in different flooding treatments were analyzed by Detrended Canonial Correspondence Analysis(DCCA) and Canonical Correspondence Analysis(CCA).It showed that the Bacillus community structure was taken place the successive changes under different flooding times.The R-strategists organisms dominated in the early succession,while the K-strategists organisms gradually replaced them in the late succession.At the early stage of incubation(1 h~1 d),the similarity coefficient indices of predominant Bacillus were relatively high but the variety of diversity indices were low;during 1 d and 5 d,the diversity index changed more;during the later stage of incubation(20~30 d),there were the highest similarity coefficient indices and diversity indices kept invariableness.The flooding process could be divided into three different ecotopes including early-stage,mid-stage and later-stage,on the basis of DCCA.Further more,the CCA ordination among DGGE profiles and environmental variable factors demonstrated that Fe(Ⅱ) concentration and the changes of diversity and richness of Bacillus correlated with the above three ecotopes.From the phylogenetic tree,we found that the nine preponderant DGGE bands belonged to Bacillus except of the HZ-B1 band divided into Paenibacillus sp..Most of them had close relations with Bacillus sp.isolated from paddy soil fields.

OBJECTIVE:The diversity of Fe-hydrogenase based on Fe-hydrogenase gene of Clostridium was studied for exploring the biochemical mechanism of soil microbial hydrogen production and revealing the Fe-hydrogenase microbial community structure changes during the paddy soil flooding incubation. METHODS:We used denatured gradient gel electrophoresis and real-time quantitative PCR to achieve the goal. RESULTS:The band number of Fe-hydrogenase denatured gradient gel electrophoresis fingerprints indicated Fe-hydrogenase microbes structure varied significantly during the completely flooding incubation. The Principal Component Analysis (PCA) showed the highly similar communities of Fe-hydrogenase microbial was divided into three groups: 1 d and 20 d, the 5 d, 30 d and 40 d. With the growth of the flooding incubation time Fe-hydrogenase microbial community structure became relatively stable and convergence. Alpha diversity index analysis found that the richness index (R), Shannon-Weaver index (H'), Simpson index (D(S)) numerical values of 1 d and 10 d were lower compared with other points, indicated the two time points of low Fe-hydrogenase diversity, simple Fe-hydrogenase microbial community structure and successive variation of community structure during the whole flooding incubation. After sequencing 15 Fe-hydrogenase preponderant bands ( labeled by G1 - G15), the phylogenetic tree of Fe-hydrogenase showed that the preponderant bands all had high similarity with the Clostridium Fe-hydrogenase in earlier flooding incubation stage and non-Clostridium Fe-hydrogenase in the later stage. Real-time quantitative PCR results demonstrated that the Fe-hydrogenase gene copy number was 10(6) level. CONCLUSION:In the research we found 4 kinds of Clostridium Fe-hydrogenases and three kinds of non-Clostridium Fe-hydrogenases, the corresponding Fe-hydrogenase microbial community structure had successively significant variation in the early incubation stage and tend to be relatively stable and convergence in the late stage.

Microbially-mediated Fe(III) reduction is of environmental significance in wetland ecosystems such as rice fields. Despite a number of incubation experiments showing the dynamic structure and activity of microbial communities in rice paddy soils amended with different substrates, little is known regarding the succession of Fe(III)-reducing bacterial populations in non-amended, natural paddy soils upon flooding. in this study, a 30-d laboratory incubation experiment was conducted to examine the diversity, dynamic and abundance of representative Fe(III)-reducing bacterial family Geobacteraceae in anaerobic natural paddy soil slurry incubations. The Logistic model showed that the microbial Fe(III) reduction rate in paddy slurry reached the highest level (1.36 mg g(-1) d(-1)) after 3.3-d flooding, and the accumulated Fe(II) level stabilized at 8.14 mg g(-1) on day 20. Quantitative, real-time PCR assay showed that the absolute abundances of Geobacteraceae and total bacterial populations varied in similar trends. Both decreased from day 1-10 and peaked on day 20 (13.98 x 10(6) and 5.29 x 10(8) copies of 16S rDNA g(-1) dry soil, respectively), followed by large decreases on day 30 (1.94 x 10(6) and 0.62 x 10(8) copies of 16S rDNA g(-1) dry soil, respectively). The relative abundance of Geobacteraceae, i.e., the proportion of Geobacteraceae to total bacteria reached the highest level (similar to 4%) following 5-d flooding, and then slightly fluctuated at 2.6%-3.9% till the end of the experiment. Clone library construction and sequencing analysis showed that the Geobacteraceae mainly consisted of Geobacter spp. which promoted bacterial Fe(III) reduction in paddy slurries upon flooding. UniFrac principal coordinate analysis revealed the succession of Geobacteraceae species, with the highest diversity observed in the initial stage (1 h-1 d) and the dominant successional members in the late stage (20-30 d). These results indicated that Geobacteraceae species contributed to Fe(III) reduction in flood paddy soils, and that the structure of Geobacteraceae population was maintained through the common occurrence of generalized species and the succession of specialized species. (c) 2013 Elsevier Masson SAS. All rights reserved.

Geobacteraceae is identified as a typical family of dissimilatory iron(Ⅲ)reducer with distribution in anaerobic environment.It plays an important role on the decomposition of organic matter,the suppression of methane production,the bioremediation of heavy metal pollution with valence fluctuation,and the transformation of phosphorus,sulfur and other nutrient elements.In this study,total DNA was extracted from six paddy soil slurries including 1 h,1 d,5 d,10 d,20 d and 30 d treatments of flooding incubation.The changing characteristics of community diversity and succession of Geobacteraceae were analyzed by using denatured gradient gel electrophoresis(DGGE)based on 16S rRNA gene(rDNA)of Geobacteraceae.The influence factors on characteristics of community diversity and succession of Geobacteraceae in different flooding treatments were analyzed by canonical correspondence analysis(CCA).It showed that the Geobacteraceae community structure was taken place the successive changes under different flooding time.The r-strategists organisms dominated in the early succession,while the k-strategists organisms gradually replaced them in the late succession.Similarity coefficient indices were the lowest between 1 h and 1 d treatment and the highest between 20 d and 30 d treatment.Richness indices and Shannon-Weiner indices were the highest in flooding 5 d treatment,and the lowest in flooding 1 h treatment.The result of PCA(principal component analysis)based on digitized DGGE patterns and the sequences showed that the changes of the community structure were stable at the end of flood incubation.The CCA ordination among DGGE profiles and environmental variable factors demonstrated that Fe(Ⅱ)concentration strongly correlated with the total variation of the bacterial community.The phylogenetic tree of preponderant DGGE bands could be divided into two groups.The major sequences could get together with the sequences of uncultured Geobacter spp.from paddy soil.The dynamic variation of community structure resulted from different flooding time will provide a theoretical basis to clarify the characteristics of community structures of iron-reducing bacteria and phylogenetic classification.

Dissimilatory iron-reducing bacteria grown in soil play an important role in bioremediation of organics and heavy metal pollution.The aim of this study is to explore the internal relationship between the iron-reducing ability and hydrogen-evolution of Clostridium,which is a typical iron-reducing bacteria.In this paper,a Clostridium strain isolated from paddy soil as the research object.Using homologous clone,a fragment(761 bp) of hydrogenase gene was obtained.Bioinformatics analysis showed that this gene fragment contained the active centers of hydrogenase,and was its major functional domain.Moreover,knockout vector(pMD-19-HTH) included tetracycline resistance gene was constructed by overlap PCR technique with the purpose of knockout hydrogenase function and lay the foundation for uncovering relationship between the iron-reducing ability and hydrogen-evolution.

The diversity changes of iron-reducing bacteria from different marine areas of Bohai sediments were analyzed by 16S rDNA-RFLP technique.The bacteria were treated by glucose enrichment culture under short-term anaerobic condition,previously.The results showed that,digesting 16S rDNA of iron-reducing bacteria by HhaⅠresulted in 9 RFLP types(OTUs);α diversity index indicated that,both the change trends of Shannon-Wiener index(H′) and Simpson index(Ds) were 14C14P6P 3C6C 3P;Margalef index(dMa) change trend was 14P6P 14C6C3C3P;species evenness index(E) followed by 14C3C14P6P6C3P.Different treatments had same dominant bacterial patterns,but the proportions were dissimilar.It was found that most of the bacteria obtained in this research belonged to Bacillus sp.and Paenibacillus sp.in family Firmicutes by alignment of partial iron-reducing bacteria 16S rDNA sequence.Compared with control treatment,the glucose enrichment culture could change the community structure of iron-reducing bacteria from different marine sediments.

Ammonia-oxidizing archaea are likely the most abundant ammonia-oxidizing microbes in natural environment and they also play an important role in nitrification.In order to improve nitrogen use efficiency and explicate the indicating function of ammonia-oxidizing archaea(AOA) on changes of soil quality in the Loess Plateau,AOA community structure diversity was studied.The soil samples used in this research derived from Changwu Agro-ecological Experimental Station on the Loess Plateau,Chinese Academy of Sciences,which had received 23 years continuous fertilization treatments,include CK(control,without fertilizers),LD(unplanted,without fertilizers),N(nitrogen input),P(phosphorus input) and NP(combination of nitrogen and phosphorus fertilizers).The soil AOA community structure diversity was analyzed by restriction fragment length polymorphism(PCR-RFLP) and DNA sequence.Positive clones collected randomly from clone libraries were digested by Rsa I and Msp I,respectively.According to the statistics of diversity index,there were 25,18,29,20 and 30 restriction endonuclease types(OTUs),respectively.The α diversity indices indicated that there was a pronounced difference among five fertilizer treatments.The OTUs were the highest in both P treatment and CK treatment,while the lowest in NP treatment.The rescaled distance matrix tree indicated that the different fertilization had weak convergence of AOA community types with the CK treatment soil.Phylogenetic tree of amoA gene amino acid sequences analysis showed all AOA sequences fell within cluster S and cluster M,but the proportions were different.These results indicated that long-term fertilization resulted in change of AOA community diversity;however,different fertilizer alkaline soil had no significant impact on the species composition of dominant AOA.

OBJECTIVE:The dynamic characteristics of community structure and relative abundance of Geobacteraceae were investigated to understand their response to microbial iron (III) reducing in flooded paddy soil. METHODS:The paddy soil was incubated anaerobically and the amount of Fe(II) was determined during the flooding incubation. We retrieved Geobacteraceae sequences from clone libraries constructed for different time points (1 h and day 1, 5, 10, 20 and 30) after flooding of the paddy soil. The diversity and community structure were analyzed by using RFLP method, and the relative abundance of Geobacteraceae was detected by real-time PCR. RESULTS:Microbial reduction of iron (III) changed greatly in early time and was stable after incubated for 20 d in paddy soil. The largest iron reduction potential was 10.16 mg/g with a Vmax of 1.064 mg/(g x d) at the time of 4.84 d whereas this process achieved plateau after 20 days flooding. Diversity of Geobacteraceae, given by alpha indices, fluctuated during the flooding incubation. Two peaks of diversity were observed in treatments of 5 d and 20 d respectively, while significant low diversity appeared in samples of 10 d and 30 d. Beta indices described the differences between community structures of Geobacteraceae and hence reflected the variation of the flooding situation over time. In all samples, 10 RFLP-based preponderant types were found, which fell into clade 1 and clade 2 of Geobacteraceae. The relative abundance of Geobacteraceae was the lowest in 1 d (1.20% ) and the highest in 20 d (4.54%). CONCLUSION:The dynamic variation of Geobacteraceae diversity, community structure and abundance are closely related to microbial iron (III) reducing in flooding paddy soil.

Cultivation-independent techniques like PCR-amplified restriction fragment length polymorphism (PCR-RFLP) of 16S rRNA genes and real-time PCR were applied to assess the abundance, diversity and phylogenetic composition of Anaeromyxobacter communities over time in flooded, unplanted paddy soil slurries. Six Anaeromyxobacter communities were sampled from anoxic slurries at 1 h, and 1, 5, 10, 20 and 30 days while the Fe (II) concentrations were measured also. Bacterial Genomic DNA was extracted and PCR-amplified to obtain 16S rDNA fragments of Anaeromyxobacter which were cloned to construct 6 16s rDNA libraries. Eventually 10 major Anaeromyxobacter types were identified by RFLP fingerprintings. Results showed that the optimal increasing phase of Fe (II) was from 1h to nearly 10 days, being correspond with the growth phase of the abundance of Anaeromyxobacter. The highest diversity appeared in slurry at 30 days and the lowest was found at 30 days. Jackknife Environment Clusters by UniFrac showed that phylogenetic compositions of Anaeromyxobacter communities in slurries at 10 and 20 days were the most similar. By evolutionary distance analysis, our 10 major Anaeromyxobacter types were diverged into Group 1 and 2 in phylogenetic tree, while Group 1 was the exclusive collection of clones from our experiment. Major type P1 was present in all slurries abundantly and P9 only existed in slurry at 5 days. The abundance of Anaeromyxobacter spp., calculated as its proportion of 16S rDNA copies to the value of total Bacteria, was from 0.242% at 1 h to 5.135% at 10 days. We demonstrated that flooding time led to successional dynamics of major types and variable abundance of Anaeromyxobacter community. Flooding time also influenced the diversity of Anaeromyxobacter community on some extent. Canonical correspondence analysis (CCA) revealed that Anaeromyxobacteria spp. abundance had interrelation with Fe (II) content and the influenced the distribution of the slurries in the Biplot. Moreover, our study provides valuable information for the further isolation of Anaeromyxobacter strains from paddies.

【Objective】In order to improve the efficiency of nitrogen utilization and explicate the function of ammonia-oxidizing archaea(AOA) under the changes of soil quality in the Loess Plateau,the community structure diversity and abundance of AOA were studied.【Method】 The influence of long-term fertilization treatments including CK,M,NM,PM and NPM on soil AOA community structure diversity and amoA gene copy numbers were analyzed by restriction fragment length polymorphism(PCR-RFLP) and real-time PCR.【Result】From the clone libraries of the different fertilization treatments,there were 25,18,29,20 and 30 restriction endonuclease types,respectively.The α diversity indices indicated that there was a pronounced difference among five fertilizer treatments.The OTUs was the highest in NPM treatment and the lowest diversity in M treatment.The rescaled distance matrix tree indicated that the different fertilization had the largest convergence coefficient of AOA community types with the CK treatment soil,so the different fertilization led to significant changes of AOA communities.The amoA gene copy numbers of AOA changes were different among the treatments,whereas the highest copy numbers were detected in the NPM treatment,and had a pronounced difference with other fertilizer treatments.All preponderant sequences of AOA fell within soils/fresh water sediments based on phylogenetic tree of amoA gene amino acid sequences analysis.【Conclusion】Long-term fertilization resulted in changes of AOA community diversity and abundance.

Soil microbial community is often affected by type and level of fertilization. To analyze the potential impacts of different nitrogen forms on bacterial community in Earth-cumuli-Orthic Anthrosols in northwest China, short-term indoor thermostatic incubations including treatments of excessive amide-nitrogen(T1), nitrate-nitrogen(T2) and ammonium-nitrogen(T3) were performed and their responding bacterial 16S rDNA libraries were then constructed and analyzed by PCR-RFLP, as well as controls without fertilization(CK1) and with normal fertilization(CK2). According to the fingerprints generated by restrict cleaving with endonuclease Hha I and Rsa I and agarose gel electrophoresis, we obtained 17, 25, 130, 119 and 187 OTUs(Operational Taxonomic Units) from the established libraries of treatments T1, T2, T3 and controls of CK1 and CK2, respectively. Results showed that normal fertilization triggered the appearance of the most OTUs while excessive utilization of amide-nitrogen and nitrate-nitrogen reduced the OTUs to approximately 9.09% and 13.36% of that of normal fertilization, whereas over usage of ammonium-nitrogen generated a similar OTU numbers as the control without fertilization. Estimation by α indices gave a decreasing order of both diversity(indices of Shannon-wiener and Simpson,H' and Ds) and richness(index of Margalef, dMa) of all incubations as CK2>T3>CK1>T2>T1, indicating that normal fertilization was beneficial to the bacterial diversity in our study while excessive amide-nitrogen treatment decreased it and ammonium-nitrogen overusing maintained the bacterial diversity on a moderate level. In addition, we observed the presence of dominant pattern in all the incubations except for control with normal fertilization. As shown by phylogenetic analysis of evolutionary distance, dominant clones from T1 entirely fell into the radiation of uncultured bacteria, while the distributions of Pseudomona sp. Stenotrophomonas sp, uncultured Gemmatimonadetes sp., and uncultured soil bacterium were found in T2 and clones from T3 were closely affiliated to uncultured acidobacterium sp. Stenotrophomonas sp. and Bacillus sp.The variation of bacterial diversity, driven by different fertilizing manners of nitrogen, was proofed in our present study in molecular approach.