Analysis of bacterial communities in rhizosphere soil of continuously cropped healthy and diseased konjac

The bacterial community and diversity in healthy and diseased konjac rhizosphere soils with different ages of continuous cropping were investigated using next-generation sequencing. The results demonstrated that the number of years of continuous cropping significantly altered soil bacterial community and diversity. Soil bacterial Shannon diversity index and Chao 1 index decreased with the increasing cropping years of konjac. After 1 year of cropping, the soil exhibited the highest bacterial relative abundance and diversity. Of the 44 bacterial genera (relative abundance ratio of genera greater than 0.3%), 14 were significantly affected by the duration of continuous cropping and plant status. With increasing continuous cropping, Alicyclobacillus decreased, while Achromobacter, Lactobacillus, Kaistobacter, Rhodoplanes increased after 3 years continuous cropping. Continuous cropping altered the structure and composition of the soil bacterial community, which led to the reduction in the beneficial bacteria and multiplication of harmful bacteria. These results will improve our understanding of soil microbial community regulation and soil health maintenance in konjac farm systems.     

Effects of long-term chlorimuron-ethyl application on the diversity and antifungal activity of soil Pseudomonas spp. in a soybean field in Northeast China

The herbicide chlorimuron-ethyl has been applied widely for weed control in farmland, especially in soybean fields in China over the past decade, but the chronic effects of this herbicide on soil microorganisms, particularly Pseudomonas spp., is not well understood. Taking a continuously cropped soybean field in the town of Fuyuan—a soybean production base of Heilongjiang Province in Northeast China—as a case study, soil samples were collected from plots having received 0-, 5-, and 10-year applications of chlorimuron-ethyl (30 g active component of chlorimuron-ethyl/ha/year) to study the abundance and diversity of Pseudomonas spp. Meanwhile, an in vitro assay was used to examine the antifungal activities of isolated Pseudomonas spp. against soil-borne pathogens (Fusarium graminearum, Fusarium oxysporum, and Rhizoctonia solani) causing soybean root rot disease. The production of siderophore, hydrogen cyanide (HCN), and lytic enzymes (cellulase, pectinase, and chitinase) by Pseudomonas spp. was also investigated. With 5- and 10- year chlorimuron-ethyl application, the numbers of soil Pseudomonas spp. decreased from 121?×?10² CFU/g dry soil in the control to 40?×?10² CFU/g dry soil and 13?×?10² CFU/g dry soil, and the Shannon index values decreased from 6.23 to 3.71 and 1.73, respectively. The numbers of antifungal Pseudomonas spp. also decreased, and the proportions of Pseudomonas spp. with antifungal activities against the different test pathogens altered. All the antifungal Pseudomonas spp. could produce siderophore and HCN but not lytic enzymes. The results suggest that long-term application of chlorimuron-ethyl in continuously cropped soybean field had negative effects on the abundance and diversity of soil Pseudomonas spp., including species with different antifungal activities against pathogens. Siderophore and HCN rather than lytic enzymes formed the antifungal metabolites of Pseudomonas spp., and the number of antifungal Pseudomonas that can produce siderophore and HCN decreased markedly under application of chlorimuron-ethyl, especially after 10-year application.     

不同土壤细菌种群结构对氯嘧磺隆胁迫的响应及降解菌系的获得

应用PCR-DGGE、DNA测序等方法,在室内驯化条件下研究了8种来源于中国不同地区土壤样品细菌种群结构对氯嘧磺隆胁迫的响应。结果表明:在氯嘧磺隆100～500mg·L-1浓度梯度下,土壤细菌群落组成有明显的更迭现象,多样性发生明显变化,驯化至10周,绝大部分细菌种群消失,样品的细菌种群结构趋于简单并呈现趋同效应;DNA测序结果表明,在驯化第10周可培养Methylophilus sp.、Beta proteobacterium、uncultured bacterium成为优势菌属,所获细菌种群出现的16个优势种群中有10个与已知的具有有机污染物降解功能和有机污染环境修复功能细菌的相似性大于97%;其中5个与嗜甲基菌16S rDNA部分序列相似性达98%以上。获得了一组对氯嘧磺隆具有降解作用的细菌菌系,可在5d内将100mg·L-1氯嘧磺隆降解67%;其主要组成为嗜甲基菌属(Methylophilus sp.)、丛毛单胞菌属(Comamonas sp.)、鞘酯杆菌属(Sphingobacterium sp.)和嗜氢菌属(Hydrogenophi-lus sp.)。     

氯嘧磺隆对土壤微生物群落结构的影响

采用磷脂脂肪酸(PLFA)法研究了黑龙江省苇河地区不同氯嘧磺隆施药历史下土壤微生物群落结构的差异, 并测定了土壤中氯嘧磺隆的残留量.结果表明：不同施药历史下氯嘧磺隆在土壤中的残留量均很低;随着施用氯嘧磺隆年限的增加,土壤微生物的PLFA总量减少, 革兰氏阴性菌/革兰氏阳性菌（GN/GP）和真菌/细菌比降低,土壤微生物的压力指数增加.主成分分析表明,氯嘧磺隆显著改变了大豆田土壤微生物群落结构.     

Conversion from natural wetlands to paddy field alters the composition of soil bacterial communities in Sanjiang Plain,Northeast China

The Sanjiang Plain is the largest freshwater wetlands in Northeast China. In order to feed the growing population, about 84 % of the wetlands in this area have been converted to farmland, especially to paddy fields, since the 1950s. However, little is known about the influence of this conversion on soil microbial community composition. In this study, soil samples were collected from two natural wetlands dominated by plant species Carex lasiocarpa and Deyeuxia angustifolia and from a neighboring paddy field that was changed from wetland more than 10 years ago. The composition and diversity of bacterial communities in the soils were estimated by clone library analysis of nearly full length of 16S rDNA sequences. The results revealed that bacterial diversity was higher in paddy fields, and that the composition of bacterial communities differed among the three samples; the difference was more notable between the paddy field and two natural wetlands than between two natural wetlands. The distribution of clones into different bacterial phyla differed among soil samples, and the conversion from natural wetlands to paddy field increased the abundance of Proteobacteria and Firmicutes but decreased the abundance of Chloroflexi. About 63 % and 71 % of clones from two natural wetlands and 49 % of clones from the paddy field had <93 % similarity with known bacteria, suggesting that the majority of bacteria in natural wetland soils in the Sanjiang Plain are phylogenetically novel. In general, this study demonstrated that long-term conversion from natural wetlands to paddy field changes soil bacterial communities in the Sanjiang Plain.     

Response of soil nematode communities to long-term application of inorganic fertilizers in the black soil of Northeast China

The effects of inorganic fertilizers on soil nematode communities were studied in a long-term fertilization experimental field in the Black Soil Region of Northeast China, where no fertilizer (CK), N fertilizer (N), combined application of N and P (NP), combined application of N and K (NK), and combined application of N, P and K (NPK) were compared. The results showed that the total nematode abundance was not affected significantly by inorganic fertilizers in the long-term field experiment. The numbers of bacterivores increased significantly in the NP treatment compared to the CK treatment, and those of fungivores and plant-parasites were inhibited in the NPK treatment. The similarity between CK and NPK treatment and the nematode diversity were higher than in other treatments. The stability of the soil ecosystem was disturbed by the inorganic fertilizers, as indicated by the change in MI values under different treatments. The response of soil nematodes mainly depended on the types of inorganic fertilizers applied.     

Response of soil nematode communities to long-term application of inorganic fertilizers in the black soil of Northeast China

The effects of inorganic fertilizers on soil nematode communities were studied in a long-term fertilization experimental field in the Black Soil Region of Northeast China,where no fertilizer (CK),N fertilizer (N),combined application of N and P (NP),combined application of N and K (NK),and combined application of N,P and K (NPK) were compared.The results showed that the total nematode abundance was not affected significantly by inorganic fertilizers in the long-term field experiment.The numbers of bacterivores increased significantly in the NP treatment compared to the CK treatment,and those of fungivores and plant-parasites were inhibited in the NPK treatment.The similarity between CK and NPK treatment and the nematode diversity were higher than in other treatments.The stability of the soil ecosystem was disturbed by the inorganic fertilizers,as indicated by the change in MI values under different treatments.The response of soil nematodes mainly depended on the types of inorganic fertilizers applied.     

Simazine application inhibits nitrification and changes the ammonia-oxidizing bacterial communities in a fertilized agricultural soil

s-Triazine herbicides are widely used for weed control, and are persistent in soils. Nitrification is an essential process in the global nitrogen cycle in soil, and involves ammonia-oxidizing Bacteria (AOB) and ammonia-oxidizing Archaea (AOA). In this study, we evaluated the effect of the s-triazine herbicide simazine on the nitrification and on the structure of ammonia-oxidizing microbial communities in a fertilized agricultural soil. The effect of simazine on AOB and AOA were studied by PCR-amplification of amoA genes of nitrifying Bacteria and Archaea in soil microcosms and denaturing gradient gel electrophoresis (DGGE) analyses. Simazine [50?μg g(-1) dry weight soil (d.w.s)] completely inhibited the nitrification processes in the fertilized agricultural soil. The inhibition by simazine of ammonia oxidation observed was similar to the reduction of ammonia oxidation by the nitrification inhibitor acetylene. The application of simazine-affected AOB community DGGE patterns in the agricultural soil amended with ammonium, whereas no significant changes in the AOA community were observed. The DGGE analyses strongly suggest that simazine inhibited Nitrosobacteria and specifically Nitrosospira species. In conclusion, our results suggest that the s-triazine herbicide not only inhibits the target susceptible plants but also inhibits the ammonia oxidation and the AOB in fertilized soils.     

根际效应对大豆田土壤线虫群落组成及多样性的影响

根际作为重要的环境界面是植物与环境之间物质能量交换的场所,关于根际效应的研究已成为土壤生态学的新兴热点领域,然而有关大豆根际效应对土壤动物多样性影响的研究报道并不多见。在三江平原选择连续耕作15a的大豆田,对大豆根际区与非根际区土壤线虫群落结构组成进行了对比分析。结果表明:大豆根际区土壤线虫总数、辛普森多样性指数(Dom)显著高于非根际区,根际区的物种数(S)、物种丰富度指数(SR)显著低于非根际区。说明大豆根际效应增加土壤线虫的丰度,但降低了线虫群落结构的复杂性。大豆根际区植物寄生线虫(PP)、食真菌线虫(FF)和食细菌线虫(BF)数量显著高于非根际区,而PP类群的比例在根际区却显著低于非根际区。这一研究结果表明食微线虫(FF和BF)类群在大豆根际区的比例增加更显著。食真菌与食细菌线虫数量比值(F/B)指示大豆根际区细菌生物量相对高于真菌生物量。研究结果丰富了农田土壤线虫多样性的研究内容,并为我国东北大豆田线虫病害的防治及定制相应的农业管理措施提供参考。     

长期施用不同无机肥对旱地红壤线虫群落的影响

红壤生态系统中土壤生物群落对于维持土壤功能的正常发挥具有重要作用.本研究基于持续25年的红壤旱地化肥定位试验,研究不同无机肥组合,包括氮磷钾(NPK)、氮磷钾补充石膏(NPKCaS)、氮磷(NP)、氮钾(NK)和磷钾(PK)对花生生长季土壤线虫群落的影响.结果表明： 土壤线虫总数、营养类群以及各生态指数在处理间差异显著(P<0.01).线虫总数由高到低的顺序为PK>NPKCaS>NPK>NP>NK.除5月外,NPK、NP、NK处理的线虫总数均显著低于NPKCaS和PK处理.NPKCaS处理的优势类群为食细菌线虫,平均丰度为42.1%,其他处理均以植食性线虫为优势营养类群,其平均丰度为38%~65%.NPKCaS处理线虫群落较高的成熟度指数、瓦斯乐斯卡指数和结构指数说明土壤食物网结构较为成熟和稳定,同时表明氮磷钾补充石膏通过缓解土壤酸化创建了良好的土壤健康状况.仅施氮钾的处理则相反.本研究证实了施用石膏和磷肥是改善红壤质量的有效措施,土壤线虫群落分析能较好地反映不同无机肥对红壤旱地生态系统的影响.
     

Influence of temperature and soil water content on bacterial, archaeal and denitrifying microbial communities in drained fen grassland soil microcosms

In this study, microcosms were used to investigate the influence of temperature (4 and 28 degrees C) and water content (45% and 90% WHC) on microbial communities and activities in carbon-rich fen soil. Bacterial, archaeal and denitrifier community composition was assessed during incubation of microcosms for 12 weeks using terminal restriction fragment length polymorphism (T-RFLP) profiling of 16S rRNA and nitrous oxide reductase (nosZ) genes. In addition, microbial and denitrifier abundance, potential denitrification activity and production of greenhouse gases were measured. No detectable changes were observed in prokaryote or denitrifier abundance. In general, cumulatively after 12 weeks more carbon was respired at the higher temperature (3.7 mg CO(2) g(-1) soil), irrespective of the water content, whereas nitrous oxide production was greater under wet conditions (98-336 microg N(2)O g(-1) soil). After an initial lag phase, methane emissions (963 microg CH(4) g(-1) soil) were observed only under warm and wet conditions. T-RFLP analyses of bacterial 16S rRNA and nosZ genes revealed small or undetectable community changes in response to temperature and water content, suggesting that bacterial and denitrifying microbial communities are stable and do not respond significantly to seasonal changes in soil conditions. In contrast, archaeal microbial community structure was more dynamic and was strongly influenced by temperature.     

喀斯特土壤固氮微生物群落与植被、土壤的关系

固氮微生物作为土壤中重要的功能微生物群之一,其与地上植物群落、土壤环境之间的关系尚不清楚。在桂西北的环江县、都安县和大化县选取喀斯特典型植被类型(草丛、灌丛、次生林)建立样方,通过植被调查、测定土壤理化性质和构建克隆文库的方法,研究了土壤固氮微生物群落的结构与组成,分析了固氮微生物群落与植物群落、土壤理化性质之间的关系。结果表明:研究区典型植被类型土壤中的优势固氮微生物为慢生根瘤菌。Mantel相关性分析表明植物群落与固氮微生物群落显著相关(r=0.6116,P=0.011);结合PCo A分析和Venn图可看出,植物群落组成与结构越相似,土壤固氮微生物群落结构与组成也越相似。CCA分析前两轴的解释率之和仅为22.72%,其中总氮、有效态钾、有效态钙对固氮微生物群落的影响显著,这表明本研究涵盖的土壤理化性质指标并不能完全解释固氮微生物群落的变异,需要补充更多的土壤数据进行更深入的研究。由此可见,在喀斯特生态恢复过程中,不仅要关注地上植被群落的恢复与重建,同时也应重视地下功能微生物群落的恢复与重建。     

Responses of bacterial and fungal communities to an elevation gradient in a subtropical montane forest of China

Bacteria and fungi are ecologically important contributors to various functioning of forest ecosystems. In this study, we examined simultaneously the bacterial and fungal distributions in response to elevation changes of a forest. By using clone library analysis from genomic DNA extracted from forest humic clay soils, the composition and diversity of bacterial and fungal communities were determined across an elevation gradient from low via medium to high, in a subtropical forest in the Mountain Lushan, China. Our results showed that soil water content and nutrient availability, specifically total carbon, differed significantly with elevation changes. Although the soil acidity did not differ significantly among the three sites, low pH (around 4) could be an important selection factor selecting for acidophilic Acidobacteria and Alphaproteobacteria, which were the most abundant bacterial clones. As the majority of the fungi recovered, both Basidiomycota and Ascomycota, and their relative abundance were most closely associated with the total carbon. Based on the Shannon–Weaver diversity index and ∫-libshuff analysis, the soil at medium elevation contained the highest diversity of bacteria compared with those at high and low elevations. However, it is difficult to predict overall fungal diversity along elevation. The extreme high soil moisture content which may lead to the formation of anaerobic microhabitats in the forest soils potentially reduces the overall bacterial and fungal diversity.     

Soil bacterial communities of different natural forest types in Northeast China

Background and aims

The types of natural forests have long been suggested to shape below-ground microbial communities in forest ecosystem. However, detailed information on the impressionable bacterial groups and the potential mechanisms of these influences are still missing. The present study aims to deepen the current understanding on the soil microbial communities under four typical forest types in Northeast Asia, and to reveal the environmental factors driving the abundance, diversity and composition of soil bacterial communities.

Methods

Four forest types from Changbai Nature Reserve, representing mixed conifer-broadleaf forest and its natural secondary forest, evergreen coniferous forest, and deciduous coniferous forest were selected for this study. Namely, Broadleaf-Korean pine mixed forest (BLKP), secondary Poplar-Birch forest (PB), Spruce-Fir forest (SF), and Larch forest (LA), respectively. Soil bacterial community was analyzed using bar-coded pyrosequencing. Nonmetric multidimensional scaling (NMDS) was used to illustrate the clustering of different samples based on both Bray-Curtis distances and UniFrac distances. The relationship between environmental variables and the overall community structure was analyzed using the Mantel test.

Results

The two mixed conifer-broadleaf forests (BLKP and PB) displayed higher total soil nutrients (organic carbon, nitrogen, and phosphorus) and soil pH, but a lower C/N ratio as compared to the two coniferous forests (SF and LA). The mixed conifer-broadleaf forests had higher alpha-diversity and had distinct bacterial communities from the coniferous forests. Soil texture and pH were found as the principle factors for shaping soil bacterial diversity and community composition. The two mixed conifer-broadleaf forests were associated with higher proportion of Acidobacteria, Verrucomicrobia, Bacteroidetes, and Chloroflexi. While the SF and LA forests were dominated by Proteobacteria and Gemmatimonadetes.

Conclusions

Different natural forest type each selects for distinct microbial communities beneath them, with mixed conifer-broadleaf forests being associated with the low-activity bacterial groups, and the coniferous forests being dominated by the so-called high-activity members. The differentiation of soil bacterial communities in natural forests are presumably mediated by the differentiation in terms of soil properties, and could be partially explained by the copiotroph/oligotroph ecological classification model and non-random co-occurrence patterns.     

连作马铃薯根际土壤真菌种群结构及其生物效应

连作障碍已成为马铃薯产业发展的主要限制因素,为了探明马铃薯连作障碍的机理,减轻连作障碍对产量的影响,采用大田试验与PCR-DGGE分子指纹图谱技术相结合的方法,研究马铃薯连作对根际土壤真菌种群结构的影响及其生物效应.结果表明： 随着连作年限的增加,根际土壤中真菌DGGE图谱的条带数量增加,连作1～5年处理的操作分类单元(OTU)分别比对照(轮作)增加了38.5%、38.5%、30.8%、46.2%和76.9%,说明马铃薯连作使根际土壤中真菌优势种群的个体数明显增多.随着连作年限的增加,各处理间真菌种群结构的相似性越来越低.通过真菌DGGE条带的克隆测序比对发现,随着连作年限的增加,马铃薯根际土壤土传病害病原菌尖孢镰刀菌和茄病镰刀菌的数量明显增加,而球毛壳菌作为一种生防菌,连作5年时数量明显减少.连作使根际土壤中病原真菌种群过渡成为优势种群,根际微生态环境恶化,从而作用于根系,使根系活力和吸收面积下降,最终导致块茎产量大幅度-下降.     

亚热带米槠天然林土壤氨氧化微生物对模拟氮沉降的响应

我国亚热带地区是全球氮沉降的热点区域。氮沉降会影响氨氧化微生物的丰度和群落结构,进而改变土壤微生物驱动的养分循环。目前对新近发现的完全氨氧化菌认识不足,极大地制约了对森林土壤氨氧化微生物响应氮沉降的整体认识。本研究以福建省三明市辛口镇格氏栲自然保护区长期模拟氮沉降处理土壤为研究对象,利用实时定量PCR方法,研究氨氧化微生物(包括氨氧化细菌AOB、氨氧化古菌AOA和完全氨氧化菌comammox Nitrospira),尤其是完全氨氧化菌的amoA基因丰度。模拟氮沉降处理包括:不添加N(CK)、低氮(添加40 kg N·hm^-2·a^-1,LN)和高氮(添加80 kg N·hm^-2·a^-1,HN)。结果表明: 8年的氮添加降低了土壤pH值和有机碳含量,提高了土壤硝态氮含量。供试土壤的AOB丰度低于检测限,无法获得目的片段。高氮处理显著提高了AOA丰度,但对完全氨氧化菌clade A和clade B丰度无显著影响。两种氮添加处理均降低了完全氨氧化菌/AOA值,表明氮添加降低了完全氨氧化菌在亚热带森林土壤氨氧化微生物类群中的相对竞争力。针对完全氨氧化菌clade A和clade B的扩增都存在非特异性产物,表明针对森林土壤的高特异性和覆盖度设计引物的必要性。Clade A和clade B丰度与总氮和铵态氮含量呈显著正相关,clade B丰度还与有机碳含量呈显著正相关。总之,模拟氮沉降提高了AOA在亚热带米槠天然林土壤硝化过程中的相对重要性,这些发现可为该地区应对全球变化和氮沉降的风险评估提供理论依据。     

Characterization of the bacterial and archaeal communities in rice field soils subjected to long-term fertilization practices

The bacterial and archaeal communities in rice field soils subjected to different fertilization regimes for 57 years were investigated in two different seasons, a non-planted, drained season (April) and a rice-growing, flooded season (August), by performing soil dehydrogenase assay, real-time PCR assay and pyrosequencing analysis. All fertilization regimes increased the soil dehydrogenase activity while the abundances of bacteria and archaea increased in the plots receiving inorganic fertilizers plus compost and not in those receiving inorganic fertilizers only. Rice-growing and flooding decreased the soil dehydrogenase activity while they increased the bacterial diversity in rice field soils. The bacterial communities were dominated by Chloroflexi, Proteobacteria, and Actinobacteria and the archaeal communities by Crenarchaeota at the phylum level. In principal coordinates analysis based on the weighted Fast UniFrac metric, the bacterial and archaeal communities were separated primarily by season, and generally distributed along with soil pH, the variation of which had been caused by long-term fertilization. Variations in the relative abundance according to the season or soil pH were observed for many bacterial and archaeal groups. In conclusion, the microbial activity, prokaryotic abundance and diversity, and prokaryotic community structure in the rice field soils were changed by season and long-term fertilization.     

黄瓜连作土壤酚酸类物质积累对土壤微生物和酶活性的影响

伴随连作年限的增加,日光温室黄瓜连作土壤中酚酸类物质(对羟基苯甲酸、阿魏酸、苯甲酸)明显积累,连作5～9年的土壤酚酸类物质含量显著高于连作1～3年的土壤.伴随外源酚酸类物质处理浓度的增加,黄瓜根区土壤中细菌、放线菌和微生物总量以及N生理群均呈先升后降趋势,在80 μg·g^-1处理浓度下细菌、放线菌数量最多,处理浓度在120μg·g^-1以下土壤真菌数量(包括尖孢镰刀菌、疫霉)急剧增长;多酚氧化酶、过氧化氢酶、蔗糖酶、脲酶和蛋白酶活性也同样呈先升后降趋势,但其峰值对应的浓度不同.