大庆龙凤湿地底泥氨氧化细菌与氨氧化古菌多样性分析

为研究湿地底泥中氨氧化细菌(Ammonia-Oxidizing Bacteria,AOB)与氨氧化古菌(Ammonia-Oxidizing Archaea,AOA)的群落多样性,构建AOB与AOA的amo A基因克隆文库,同时利用限制性片段长度多态性(RFLP)技术将克隆文库阳性克隆子进行归类并测序。结果发现,从湿地底泥中均得到3个AOB与AOA的可操作分类单元(OTU)。其中,所有的AOB均属于亚硝化单胞菌属(Nitrosomonas),而AOA属于泉古菌门(Crenarcharota),与河口沉积物等环境中获得的序列具有较高同源性。     

Ammonia oxidizer abundance in paddy soil profile with different fertilizer regimes

Studies about ammonia-oxidizing bacteria (AOB) and archaea (AOA) are often focused on topsoil, but little is known about their activity and distribution in subsoil. A long-term fertilizer experiment was conducted to assess the effects of different fertilizer treatments on AOB and AOA in vertical soil profiles of paddy soil plots that received no nitrogen fertilizer control (CK), NPK chemical fertilizers (CF), organic–inorganic mixed fertilizer (OIMF) and organic fertilizer (OF). Soil properties, potential nitrification rate (PNR) and amoA gene abundance of AOB and AOA were measured and analyzed by two-way ANOVA and correlation analysis. Quantitative PCR analysis of amoA genes showed that AOA were more abundant than AOB in all the soil samples. AOB declined sharply with soil depth. Compared with CK and OF treatments, CF and OIMF treatments had higher abundance of AOB throughout the soil profiles. However, AOA tend less responsive to soil depth and fertilizers compared to AOB. This caused the AOA/AOB ratios in subsoil higher than in topsoil, and in CK and OF higher than in CF and OIMF treatments. These results suggest that AOA are more abundant and can be better adapted to nutrient-poor subsoils than AOB, and autotrophic nitrification could likely be determined by a complex suite of environmental factors in vertical profiles of the paddy soil tested.     

水稻根际和周围土壤中微生物功能基因丰度与碳、氮状况及其通量之间的关系

Rapid nitrogen（N） transformations and losses occur in the rice rhizosphere through root uptake and microbial activities. However,the relationships between rice roots and rhizosphere microbes for N utilization are still unclear. We analyzed different N forms（NH＋4,NO-3, and dissolved organic N）, microbial biomass N and C, dissolved organic C, CH4 and N2O emissions, and abundance of microbial functional genes in both rhizosphere and bulk soils after 37-d rice growth in a greenhouse pot experiment. Results showed that the dissolved organic C was significantly higher in the rhizosphere soil than in the non-rhizosphere bulk soil, but microbial biomass C showed no significant difference. The concentrations of NH＋4, dissolved organic N, and microbial biomass N in the rhizosphere soil were significantly lower than those of the bulk soil, whereas NO-3in the rhizosphere soil was comparable to that in the bulk soil. The CH4 and N2O fluxes from the rhizosphere soil were much higher than those from the bulk soil. Real-time polymerase chain reaction analysis showed that the abundance of seven selected genes, bacterial and archaeal 16 S rRNA genes, amoA genes of ammonia-oxidizing archaea and ammonia-oxidizing bacteria, nosZ gene, mcrA gene, and pmoA gene, was lower in the rhizosphere soil than in the bulk soil, which is contrary to the results of previous studies. The lower concentration of N in the rhizosphere soil indicated that the competition for N in the rhizosphere soil was very strong, thus having a negative effect on the numbers of microbes. We concluded that when N was limiting, the growth of rhizosphere microorganisms depended on their competitive abilities with rice roots for N.     

基于宏基因组学的酸性森林土壤氨氧化微生物群落特征研究

全球30%以上陆地面积是酸性土壤(pH5.5),而酸性土壤中氨氧化微生物群落特征研究是破译其硝化过程微生物学机理的基础。尤其随着完全硝化微生物(Complete ammonia oxidizer,comammox)的发现,亟需重新认知酸性土壤中氨氧化微生物类群。以酸性马尾松林为研究对象,综合利用荧光定量PCR(qPCR)、凝胶电泳半定量和宏基因组测序等技术研究土壤中氨氧化古菌(Ammonia-oxidizing archaea,AOA)、氨氧化细菌(Ammonia-oxidizing bacteria,AOB)和Comammox的相对丰度以及群落组成特征。研究发现AOA和AOB amoA基因丰度分别为2.61×106 copies·g~(-1)和1.45×106copies·g~(-1);而comammoxamoA基因qPCR结果存在显著的非特异性扩增,导致其丰度被高估,而经凝胶电泳半定量矫正后,约为(1.38～1.47)×106copies·g~(-1),该结果和土壤宏基因测序揭示的comammox相对丰度基本吻合。此外,宏基因组分析发现经典嗜酸group1.1a-associated仅占AOA总类群的12%,而group1.1b则占88%,尽管目前仍未有嗜酸group 1.1b AOA纯菌株的报道。AOB主要类群为Nitrosospira(约64%),而Nitrosomonas约占36%。Comammox主要类群为clade B(约64%),而clade A仅占36%且均隶属于clade A.1亚枝,这暗示clade B与已报道的嗜中性comammox clade A纯菌株有极大的生理代谢差异。总之,本研究提供了综合利用qPCR、半定量和宏基因组分析土壤氨氧化微生物群落的策略,并建议优化comammox的qPCR引物,同时本研究系统分析了酸性马尾松林土壤中氨氧化微生物的相对丰度和群落组成特征。     

不同水肥条件对农田土壤细菌生理菌群的影响

采用田间试验研究了大豆苗期、花期和鼓粒期在干旱、自然和充足三种水分及无肥、无机肥和无机肥+有机肥三种施肥方式条件下,根际士壤中氨氧化细菌、自生周氮菌和反硝化细菌三种不同生理菌群的变化.结果表明:施用无机肥条件下自然降水使氨氧化细菌数量高于充足水和干旱处理,干旱使好气性自生固氮菌和反硝化细菌数量高于自然降水和充足水,在自然降水条件不同施肥使功能性细菌发生变化,施肥促进三种功能菌的生长,其中,有机肥配施化肥的效果最为显著反硝化细菌符处理数量相近,有机肥数量稍高.     

高通量测序分析人工养殖成年林麝粪便古菌菌群多样性

本试验的目的是采用高通量测序技术研究人工养殖成年林麝粪便中古菌的结构和组成,并对比不同性别之间的差异。选取12只3岁健康的林麝依据性别分为雄性组(M组)和雌性组(F组),每组各6只。采集其新鲜粪便,提取DNA,用古菌通用引物PCR扩增古菌16S rRNA的V4~V5区,用M i Seq 300 PE测序平台对扩增产物进行高通量测序,用QIIM E等软件对测序序列进行分析统计。结果表明:在从门到属的各级分类水平上,F组与M组古菌的相对丰度差异均不显著(P0.05)。M组和F组的组内遗传距离分别为0.16±0.03和0.27±0.06,组间为0.24±0.07,样品的相似度很高。林麝粪便中的古菌可以分为3个门,优势门为阔古细菌门(Euryarchaeota);在属水平上可以分为7个已知属,优势属为甲烷短杆菌属(Methanobrevibacter),其次为热裸单胞菌属(Thermogymnomonas)。结果提示,雄性和雌性林麝粪便中古菌的结构和组成都没有显著差异,Methanobrevibacter是林麝粪便中的优势古菌属     

A lysimeter study of nitrate leaching from grazed grassland as affected by a nitrification inhibitor,dicyandiamide, and relationships with ammonia oxidizing bacteria and archaea

Nitrate (NO₃^?) can contribute to surface water eutrophication and is deemed harmful to human health if present at high concentrations in the drinking water. In grazed grassland, most of the NO₃^?‐N leaching occurs from animal urine‐N returns. The objective of this study was to determine the effectiveness of a nitrification inhibitor, dicyandiamide (DCD), in decreasing NO₃^? leaching in three different soils from different regions of New Zealand under two different rainfall conditions (1260 mm and 2145 mm p.a.), and explore the relationships between NO₃^?‐N leaching loss and ammonia oxidizing bacteria (AOB) and ammonia oxidizing archaea (AOA). The DCD nitrification inhibitor was found to be highly effective in decreasing NO₃^?‐N leaching losses from all three soils under both rainfall conditions. Total NO₃^?‐N leaching losses from the urine patch areas were decreased from 67.7–457.0 kg NO₃^?‐N/ha to 29.7–257.4 kg NO₃^?‐N/ha by the DCD treatment, giving an average decrease of 59%. The total NO₃^?‐N leaching losses were not significantly affected by the two different rainfall treatments. The total NO₃^?‐N leaching loss was significantly related to the amoA gene copy numbers of the AOB DNA and to nitrification rate in the soil but not to that of the AOA. These results suggest that the DCD nitrification inhibitor is highly effective in decreasing NO₃^? leaching under these different soil and rainfall conditions and that the amount of NO₃^?‐N leached is mainly related to the growth of the AOB population in the nitrogen rich urine patch soils of grazed grassland.     

农田土壤好氧氨氧化和甲烷氧化交互作用机制的研究进展

从农田土壤生态系统中硝化和甲烷氧化的研究意义、氨氧化和甲烷氧化的功能微生物演替规律，以及二者交互作用机制三个方面综述了现阶段取得的主要研究成果，并进一步阐述了土壤碳氮元素生物地球化学循环机制研究的科学问题和面临的挑战。未来研究应充分利用学科交叉，结合宏观结果和土壤微观动态过程，揭示土壤中不可培养微生物代谢能力及其在土壤生物地球化学循环中的重要作用，逐步实现对土壤生态过程的预测和调控。     

长期施肥对水稻土剖面氨氧化古菌和细菌丰度及组成的影响

长期施肥影响稻田土壤理化性质和硝化微生物群落,但长期施肥对稻田不同土层氨氧化古菌(AOA)和氨氧化细菌(AOB)群落结构的影响尚不明确.以湖南宁乡稻田不同施肥制度长期定位试验为平台,选取不施肥(CK)、施秸秆有机肥(ST)、有机-无机肥配施(OM)和施全量化肥(NPK)4个处理,采用实时荧光定量PCR和Illumina...     

红壤稻田不同生育期土壤氨氧化微生物群落结构

以福建省红壤稻田土壤为对象,通过提取土壤总DNA,利用特异引物进行PCR(聚合酶链反应)扩增和DGGE(变性梯度凝胶电泳)并结合DNA克隆测序,研究了水稻生长过程中稻田土壤氨氧化细菌和氨氧化古菌群落结构的变化。结果显示:稻田土壤具有丰富的氨氧化细菌和氨氧化古菌资源。水稻生长过程中土壤氨氧化细菌群落组成较为稳定,只表现出水稻生长前期(苗期、分蘖期)和中后期(孕穗期、成熟期)间存在一定差异。而土壤氨氧化古菌群落组成变化较大,在水稻生长的苗期、分蘖期、孕穗期和成熟期4个时期间均存在一定差异。在水稻生长过程中,土壤氨氧化细菌群落多样性指数无显著性变化,但氨氧化古菌群落多样性指数随水稻生长明显提高,孕穗期后才达到平稳。水稻生长前期土壤硝化势也具有显著上升趋势,孕穗期时达到最高,而后有所下降。土壤硝化势与氨氧化古菌群落多样性指数具有显著正相关性,与氨氧化细菌没有相关性。研究表明,氨氧化古菌对红壤稻田土壤硝化作用的影响程度较大,证实了氨氧化微生物尤其是氨氧化古菌在稻田土壤微生物组成及其生态系统功能中的重要性。