Microbial diversity associated with odor modification for production of fertilizers from chicken litter

Litter from the chicken industry can present several environmental challenges, including offensive odors and runoff into waterways leading to eutrophication. An economically viable solution to the disposal of waste from chicken houses is treatment to produce a natural, granulated fertilizer that can be commercially marketed for garden and commercial use. Odor of the final product is important in consumer acceptance, and an earthy odor is desirable. By understanding and manipulating the microbial processes occurring during this process, it may be possible to modify the odors produced. Geosmin and related volatiles produced by soil actinomycetes are responsible for earthy odors, and actinomycetes are likely to be present in the composting manure. Bacterial communities at each stage of the process were analyzed by culturing studies and denaturing gradient gel electrophoresis (DGGE). The processing steps changed the culturable bacterial community, but the total community was shown by DGGE to be stable throughout the process. A local agricultural soil was analyzed in parallel as a potential source of geosmin-producing actinomycetes. This agricultural soil had higher microbial diversity than the compost at both the culturable and the molecular levels. Actinomycete bacteria were isolated and analyzed by AromaTrax, a gas chromatography-olfactometry system. This system enables the odor production of individual isolates to be monitored, allowing for rational selection of strains for augmentation experiments to improve the odor of the final fertilizer product.     

Impact of Agricultural Practices on the Zea mays L. Endophytic Community

Agricultural practices are known to alter bulk soil microbial communities, but little is known about the effect of such practices on the plant endophytic community. We assessed the influence of long-term applications (20 years) of herbicides and different fertilizer types on the endophytic community of maize plants grown in different field experiments. Nested PCR-denaturing gradient gel electrophoresis (DGGE) analyses targeting general bacteria, type I or II methanotrophs, actinomycetes, and general fungi were used to fingerprint the endophytic community in the roots of Zea mays L. Low intraplant variability (reproducible DGGE patterns) was observed for the bacterial, type I methanotroph, and fungal communities, whereas the patterns for endophytic actinomycetes exhibited high intraplant variability. No endophytic amplification product was obtained for type II methanotrophs. Cluster and stability analysis of the endophytic type I methanotroph patterns differentiated maize plants cultivated by using mineral fertilizer from plants cultivated by using organic fertilizer with a 100% success rate. In addition, lower methanotroph richness was observed for mineral-fertilized plants than for organically fertilized plants. The use of herbicides could not be traced by fingerprinting the endophytic type I methanotrophs or by evaluating any other endophytic microbial group. Our results indicate that the effect of agrochemicals is not limited to the bulk microbial community but also includes the root endophytic community. It is not clear if this effect is due to a direct effect on the root endophytic community or is due to changes in the bulk community, which are then reflected in the root endophytic community.     

Impact of agricultural practices on the Zea mays L. endophytic community

Agricultural practices are known to alter bulk soil microbial communities, but little is known about the effect of such practices on the plant endophytic community. We assessed the influence of long-term applications (20 years) of herbicides and different fertilizer types on the endophytic community of maize plants grown in different field experiments. Nested PCR-denaturing gradient gel electrophoresis (DGGE) analyses targeting general bacteria, type I or II methanotrophs, actinomycetes, and general fungi were used to fingerprint the endophytic community in the roots of Zea mays L. Low intraplant variability (reproducible DGGE patterns) was observed for the bacterial, type I methanotroph, and fungal communities, whereas the patterns for endophytic actinomycetes exhibited high intraplant variability. No endophytic amplification product was obtained for type II methanotrophs. Cluster and stability analysis of the endophytic type I methanotroph patterns differentiated maize plants cultivated by using mineral fertilizer from plants cultivated by using organic fertilizer with a 100% success rate. In addition, lower methanotroph richness was observed for mineral-fertilized plants than for organically fertilized plants. The use of herbicides could not be traced by fingerprinting the endophytic type I methanotrophs or by evaluating any other endophytic microbial group. Our results indicate that the effect of agrochemicals is not limited to the bulk microbial community but also includes the root endophytic community. It is not clear if this effect is due to a direct effect on the root endophytic community or is due to changes in the bulk community, which are then reflected in the root endophytic community.     

用PCR-DGGE研究长期施用无机肥对种稻红壤微生物群落多样性的影响

以中国科学院红壤生态试验站的发育于第四纪红粘土的种稻红壤为研究对象,采用PCR-DGGE方法研究了长期施用无机肥对土壤微生物群落多样性的影响。在种植双季稻、连续13a施用不同无机肥后,土壤中细菌、古菌、放线菌和真菌的群落结构发生了较大的变化。未种植水稻的土壤与种稻土壤间四类微生物SSUrDNADGGE带谱相似性只有33%~66%。施磷肥的处理NP、PK、NPK之间微生物群落结构相似性较高,4类微生物的SSUrDNADGGE带谱相似性高达75%~81%。施氮钾肥(NK)、不施肥(CK)处理与施磷肥处理间土壤微生物群落结构的差异较大,其四类微生物的SSUrDNADGGE带谱相似性分别为69%~77%、55%~77%。研究的目的是深入地了解土壤中微生物群落的多样性,为科学施肥、合理利用土壤、保护微生物多样性和实现农业生态系统的可持续发展提供科学依据。     

Rhizosphere soil microorganism populations and community structures of different watermelon cultivars with differing resistance to Fusarium oxysporum f. sp. niveum

Fusarium wilt is an increasingly serious disease of watermelon that reduces crop productivity. Changes in microorganism populations and bacterial and fungal community structures in rhizosphere soil of watermelon cultivars resistant or susceptible to Fusarium oxysporum f. sp. niveum were investigated using a plate culture method and PCR-DGGE analysis. Plate culture showed that populations of culturable bacteria and actinomycetes were more abundant in the rhizosphere of the resistant watermelon cultivar than the susceptible cultivar, but the fungi population had the opposite pattern. Populations of Penicillium , Fusarium , and Aspergillus were significantly lower in the resistant cultivar than the susceptible cultivar at the fruiting and uprooting stages (p?< 0.05). Pattern matching analysis generated the dendrogram of the DGGE results indicating the relatedness of the different resistant watermelon cultivars and their corresponding rhizosphere microbial communities. Further sequencing analysis of specific bands from DGGE profiles indicated that different groups of bacteria and fungi occurred in the rhizosphere of different watermelon cultivars. Our results demonstrated that plant genotype had a significant impact on soil microbial community structure, and the differences in the rhizosphere microbial community may contribute to the differences in resistance to F. oxysporum f. sp. niveum.     

微生物菌剂施用对设施茄子根际土壤养分和细菌群落多样性的影响

[背景]设施茄子连作种植和化学肥料过量施用造成土壤养分失衡、微生物多样性降低、土传病害严重等土壤质量问题,微生物制剂是改善土壤环境质量的一项重要措施。[目的]确定微生物菌剂施用对设施茄子根际土壤养分及细菌群落多样性的影响。[方法]在河北省农林科学院鹿泉大河实验园区,以含有哈茨木霉和巨大芽孢杆菌的微生物菌剂为供试菌剂,采用传统的化学分析方法测定微生物菌剂处理和对照茄子开花期、拉秧期土壤的养分含量;采用稀释涂平板方法测定土壤可培养微生物的数量;采用高通量测序技术测定土壤细菌16S rRNA基因V3-V4区,分析处理和对照土壤的微生物多样性和群落分布规律。[结果]与对照相比,微生物菌剂处理提高了茄子开花期和拉秧期的根际土壤养分含量,拉秧期微生物菌剂处理根际土壤中全氮、碱解氮、有效磷、速效钾含量分别较对照增加13.85%、21.07%、31.51%和55.94%;施用微生物菌剂能够改变土壤可培养微生物的数量和构成,微生物菌剂处理土壤中细菌、真菌、放线菌的数量均显著增加,其中细菌、放线菌在微生物总量中所占比例增加,而真菌的占比则有所降低。微生物菌剂处理能够提高土壤微生物的Shannon指数、降低...     

放射性污染对铀矿区土壤微生物群落结构的影响

为探究放射性及其伴生污染对铀矿区附近土壤微生物群落功能多样性的影响，采集退役铀矿冶周围不同辐射值的土壤为研究对象，通过测定其土壤理化性质、土壤可培养微生物计数及Biolog-ECO微孔板培养，分析土壤放射性污染程度及理化性质对土壤微生物群落功能多样性和数量的影响。结果表明，矿区放射性污染土壤中重金属含量普遍较高，尤其是铅已超出管控标准。随着辐射值的增加，土壤中可培养微生物数量显著下降，特别是放线菌和真菌的数量；微生物群落功能多样性指数、物种丰富度指数、优势度、均一性指数显著下降；对碳源利用也逐渐下降，尤其是对酚酸类和胺类碳源，利用率分别下降25.8%、29.7%。RDA分析发现放射性核素是该区域驱动土壤微生物群落代谢发生变化的主要因素。铀矿区放射性污染及伴随的重金属污染明显改变土壤微生物群落结构，抑制了土壤微生物群落的代谢活性，造成土壤微生物群落功能多样性下降。     

不同施肥制度土壤微生物量碳氮变化及细菌群落16S rDNA V3 片段PCR产物的DGGE分析

应用化学分析和变性梯度凝胶电泳（DGGE）技术分离PCR扩增的16S rDNA的方法,研究了不同施肥制度对土壤微生物量碳、氮变化及微生物多样性的影响。结果表明,连续15a长期试验下,土壤微生物量碳（SMB-C）和微生物量氮（SMB-N）的含量大小均为长期撂荒（CK0）土壤高于农田土壤,而在农田土壤中,长期施肥的处理（NPK、NPKM、NPKSt和NPKF）高于长期不施肥处理（CK）,不同的种植制度中,长期复种轮作（NPKF）高于长期复种连作（NPK）;各处理的SMB-C/SOC（土壤有机碳）和SMB-N/TN（全氮）的比值的变化趋势与SMB-C和SMB-N变化一致;从PCR-DGGE分析,长期氮磷钾化肥配施有机肥（NPKM）处理的微生物量碳、氮的含量最高,微生物丰度最高,细菌物种最多,其次为长期撂荒（CK0）,CK处理细菌物种最少。UPGMC聚类分析表明NPK和NPKF处理细菌的群落结构相似,CK和CK0处理细菌的群落结构相似,而NPKM和NPKSt处理细菌的群落结构相似。     

Characterization of the rhizosphere microbial community from different Arabidopsis thaliana genotypes using phospholipid fatty acids (PLFA) analysis

Total and culturable rhizosphere microbial communities structure from three different genotypes of Arabidopsis thaliana growing on three different substrates was studied with phospholipid fatty acid analysis (PLFA) and multivariate statistical analyses: correspondence analysis (CA) and distance based redundancy analyses (db-RDA). In addition, microbial biomass from different groups (total bacteria, Gram+, Gram? and fungi) was calculated from biomarkers PLFA peak area, both from total and culturable microbial community. db-RDA analysis showed significant differences between soils but not between plant genotypes for culturable microbial community structure. Total microbial community was significantly different between soils, and also between plant lines in each soil. Biomass of different bacterial groups showed significant higher values in soil two rhizosphere irrespective of the plant line. In addition, significant differences between plant lines were also found for microbial biomass of different bacterial groups both in total and culturable microbial community. Throughout the work we have demonstrated that PLFA analysis has been able to show a different behaviour of total microbial community with regard to the culturable fraction analyzed in this work under the influence of plant roots. Microbial biomass of different microbial groups calculated with PLFA biomarkers was a suitable tool to detect differences between soils irrespective of the plant line, and differences in the same soil between plant lines. According to this data, a previous study should be carried out before GMPs are used in field conditions to evaluate the potential alterations that may take place on rhizosphere microbial communities structure which may further affect soil productivity. In conclusion, based on data presented in this work, GMPs alter rhizosphere microbial communities structure and this effect is different depending on the soil. Furthermore, total microbial community is affected to a greater extent than the culturable fraction analyzed.     

Effects of agronomical measures on the microbial diversity of soils as related to the suppression of soil-borne plant pathogens

The diversity of soil microbial communities can be key to the capacity of soils tosuppress soil-borne plant diseases. As agricultural practice, as well as directedagronomical measures, are known to be able to affect soil microbial diversity, it isplausible that the soil microflora can be geared towards a greater suppressivity ofsoil-borne diseases as a result of the selection of suitable soil management regimes.In the context of a programme aimed at investigating the microbial diversity of soilsunder different agricultural regimes, including permanent grassland versus arableland under agricultural rotation, we assessed how soil microbial diversity is affectedin relation to the suppression of the soil-borne potato pathogen Rhizoctoniasolani AG3. The diversity in the microbial communities over about a growingseason was described by using cultivation-based – plating on different media – and cultivation-independent – soil DNA-based PCR followed by denaturing gradient gel electrophoresis (DGGE) community fingerprinting – methods. The results showed great diversity in the soil microbiota at both the culturable and cultivation-independent detection levels. Using cultivation methods, various differences between treatments with respect to sizes of bacterial and fungal populations were detected, with highest population sizes generally found in rhizospheres. In addition, the evenness of eco-physiologically differing bacterial types was higher in grassland than in arable land under rotation. At the cultivation-independent level, clear differences in the diversities of several microbial groups between permanent grassland and arable land under rotation were apparent. Bio-assays that assessed the growth of R. solani AG3 hyphae through soil indicated a greater growth suppression in grassland than in arable land soils. Similarly, an experiment performed in the glasshouse showed clear differences in both microbial diversities and suppressiveness of R. solani growth in soil, depending on the presence of either maizeor oats as the crop. The significance of these findings for designing soil managementstrategies is discussed.     

设施土壤生态环境特点与调控

论述了设施土壤生态环境的特性主要是土壤养分富集 ,土壤板结 ;土壤酸化、次生盐渍化和连作障碍严重 ;土壤产生有害气体 ;土壤微生物区系变化很大。这些特点主要是盲目大量施肥 ,土壤长期连续利用 ,土壤环境密闭 ,单一化栽培 ,用肥种类不合理及某些化学物质积累造成的。利用生物、农业耕作、地上环境和工程措施调控相结合 ,是调控设施土壤生态环境的有效方法 ,土壤生物学的应用将是改善设施土壤生态环境新的研究方向。     

微生物产生的土腥味化合物及其清除方法

自然界中存在许多具有土腥味的化合物,以Geosmin和MIB为代表,它们的嗅阈值极低,即便含量极微,也能嗅到浓烈的土腥味、土臭味、泥土味和霉味。这些土腥味挥发性物质主要是放线菌和藻类的次生代谢产物,不同生物具有不同的合成途径。它们极易影响饮用水的品质,也给渔业和食品业带来极大危害。通过物理、化学和生物处理等方法,可以控制或清除土腥味化合物,其中微生物降解法是最有应用前景的净化方法。综述了土腥味化合物的来源、生物合成途径、分析检测技术及净化方法。     

Cultivation-dependent and -independent approaches for determining bacterial diversity in heavy-metal-contaminated soil

In recent years, culture-independent methods have been used in preference to traditional isolation techniques for microbial community analysis. However, it is questionable whether uncultured organisms from a given sample are important for determining the impact of anthropogenic stress on indigenous communities. To investigate this, soil samples were taken from a site with patchy metal contamination, and the bacterial community structure was assessed with a variety of approaches. There were small differences in microscopic epifluorescence bacterial counts. Denaturing gradient gel electrophoresis (DGGE) profiles of 16S rRNA gene fragments (16S-DGGE) amplified directly from soil samples were highly similar. A clone library generated from the most contaminated sample revealed a diverse bacterial community, which showed similarities to pristine soil communities from other studies. However, the proportion of bacteria from the soil samples that were culturable on standard plate-counting media varied between 0.08 and 2.2%, and these values correlated negatively with metal concentrations. The culturable communities from each sample were compared by 16S-DGGE of plate washes and by fatty acid profiling of individual isolates. Each approach indicated that there were considerable differences between the compositions of the culturable communities from each sample. DGGE bands from both culture-based and culture-independent approaches were sequenced and compared. These data indicated that metal contamination did not have a significant effect on the total genetic diversity present but affected physiological status, so that the number of bacteria capable of responding to laboratory culture and their taxonomic distribution were altered. Thus, it appears that plate counts may be a more appropriate method for determining the effect of heavy metals on soil bacteria than culture-independent approaches.     

Cultivation-Dependent and -Independent Approaches for Determining Bacterial Diversity in Heavy-Metal-Contaminated Soil

In recent years, culture-independent methods have been used in preference to traditional isolation techniques for microbial community analysis. However, it is questionable whether uncultured organisms from a given sample are important for determining the impact of anthropogenic stress on indigenous communities. To investigate this, soil samples were taken from a site with patchy metal contamination, and the bacterial community structure was assessed with a variety of approaches. There were small differences in microscopic epifluorescence bacterial counts. Denaturing gradient gel electrophoresis (DGGE) profiles of 16S rRNA gene fragments (16S-DGGE) amplified directly from soil samples were highly similar. A clone library generated from the most contaminated sample revealed a diverse bacterial community, which showed similarities to pristine soil communities from other studies. However, the proportion of bacteria from the soil samples that were culturable on standard plate-counting media varied between 0.08 and 2.2%, and these values correlated negatively with metal concentrations. The culturable communities from each sample were compared by 16S-DGGE of plate washes and by fatty acid profiling of individual isolates. Each approach indicated that there were considerable differences between the compositions of the culturable communities from each sample. DGGE bands from both culture-based and culture-independent approaches were sequenced and compared. These data indicated that metal contamination did not have a significant effect on the total genetic diversity present but affected physiological status, so that the number of bacteria capable of responding to laboratory culture and their taxonomic distribution were altered. Thus, it appears that plate counts may be a more appropriate method for determining the effect of heavy metals on soil bacteria than culture-independent approaches.     

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.     

Effects of fertilization on bacterial community structure and function in a black soil of Dehui region estimated by Biolog and PCR-DGGE methods

Soil microbial community structure and function are commonly used as indicators for soil quality and fertility. In this paper, the bacterial community structure and function in a black soil of Dehui region influenced by fertilization were investigated by Biolog and PCR-DGGE (polymerase chain reaction-denaturing gradient gel electrophoresis) methods. Biolog examination showed that substrate richness and catabolic diversities of bacterial communities were the highest in the treatment of farm yard manure, and the lowest in the chemical fertilizer treatment. DGGE fingerprint showed that the majority of bands were similar among all treatments, suggesting that microbial communities with those bands were stable, and not influenced by fertilization. In general, chemical fertilizer decreased the diversity of soil bacterial communities. The PCA (principal component analysis) plots of Biolog and DGGE revealed that the structure and function of bacterial communities were similar in the non-fertilized control and the treatment of farm yard manure alone, which inferred that the application of farm yard manure increased the quantity of soil microbes but had less effect on the changes of community structure. The catabolic function was similar, but the composition structure differed between the treatments of chemical fertilizer alone and combined application of farm yard manure with chemical fertilizer. These results suggest that the use of chemical fertilizer mainly decreased the catabolic activity of the fast growth bacteria or eutrophic bacteria.     

番茄根际微生物种群动态变化及多样性

采用盆栽试验的方法对番茄根际主要微生物种群在不同生育期的动态变化进行了跟踪研究.结果表明,在番茄整个生育期内,可培养细菌数量在初花期和初果期时最多;放线菌数量从苗期到末期逐渐减少;真菌数量逐渐增多.番茄对细菌根际效应明显.DGGE图谱显示不同生育期番茄根际均具有较高的细菌多样性.根际细菌种类和数量在初花期发生较为显著的变化,初果期根际群落多样性指数(H)和物种丰度(S)值都达到最高,微生物最丰富,是筛选拮抗菌的较好时期.     

Decline in topsoil microbial quotient, fungal abundance and C utilization efficiency of rice paddies under heavy metal pollution across South China

Agricultural soils have been increasingly subject to heavy metal pollution worldwide. However, the impacts on soil microbial community structure and activity of field soils have been not yet well characterized. Topsoil samples were collected from heavy metal polluted (PS) and their background (BGS) fields of rice paddies in four sites across South China in 2009. Changes with metal pollution relative to the BGS in the size and community structure of soil microorganisms were examined with multiple microbiological assays of biomass carbon (MBC) and nitrogen (MBN) measurement, plate counting of culturable colonies and phospholipids fatty acids (PLFAs) analysis along with denaturing gradient gel electrophoresis (DGGE) profile of 16S rRNA and 18S rRNA gene and real-time PCR assay. In addition, a 7-day lab incubation under constantly 25°C was conducted to further track the changes in metabolic activity. While the decrease under metal pollution in MBC and MBN, as well as in culturable population size, total PLFA contents and DGGE band numbers of bacteria were not significantly and consistently seen, a significant reduction was indeed observed under metal pollution in microbial quotient, in culturable fungal population size and in ratio of fungal to bacterial PLFAs consistently across the sites by an extent ranging from 6% to 74%. Moreover, a consistently significant increase in metabolic quotient was observed by up to 68% under pollution across the sites. These observations supported a shift of microbial community with decline in its abundance, decrease in fungal proportion and thus in C utilization efficiency under pollution in the soils. In addition, ratios of microbial quotient, of fungal to bacterial and qCO(2) are proved better indicative of heavy metal impacts on microbial community structure and activity. The potential effects of these changes on C cycling and CO(2) production in the polluted rice paddies deserve further field studies.     

长期施肥对新疆灰漠土土壤微生物群落结构与功能多样性的影响

以20a新疆国家灰漠土土壤肥力与肥料效益长期定位试验为平台,采用常规培养法,结合Biolog技术对可培养微生物、生理菌群数量和碳源利用进行测定分析,研究撂荒(CK0)、耕作不施肥(CK)、不同化肥(N、NK、NP、PK、NPK)、化肥配施低量高量有机肥(NPKM1和NPKM2)和秸秆还田(NPKS)等10种处理土壤微生物特征,揭示长期施肥对土壤微生物群落结构与功能多样性的影响。结果表明:(1)可培养微生物:与CK处理相比,CK0处理显著提高了细菌、放线菌和真菌的数量(P0.05),NPKS处理微生物数量则显著降低(P0.05);不同化肥处理的细菌(除PK处理外)、放线菌(除PK和N处理外)数量也有所增加,增幅在8.14%—135.70%和15.30%—44.78%之间;真菌数量(除NK处理外)则有一定幅度的降低;NPKM1和NPKM2处理,微生物数量最高,细菌分别增加了162.20%和173.75%,放线菌增加了34.39%和39.37%,真菌增加了63.33%和488.33%;(2)生理菌群:与CK0相比,CK处理显著提高了自生固氮菌和亚硝化细菌数量(P0.05),显著降低了氨化细菌和纤维素分解菌数量(P0.05);与CK相比,NPKM1和NPKM2处理显著提高土壤中与氮素转化有关的生理菌群数量(P0.05),不同化肥处理和NPKS处理的影响不相同,NPK处理显著高于其余处理(P0.05);(3)微生物碳源利用:微生物活性表现为NK、NPKM1、NPKM2N、NPK、CKPK、NPKSCK0、NP;CK0处理3个多样性指数以及NPKM1、NPKM2和NK处理Shannon(H)指数最高,其余施肥处理差异不显著;糖类、氨基酸类、羧酸类和胺类是微生物利用的主要碳源。(4)聚类分析表明,除NP处理外,施氮处理土壤有较为相似的碳源利用,细菌和真菌与养分之间有较好的相关性,可培养微生物和生理菌群与微生物碳源利用的相关性较差。因此,长期不同施肥对新疆灰漠土土壤微生物群落结构和功能多样性产生了显著的影响,长期耕作不施肥降低了土壤微生物群落结构和功能多样性,不同化肥配合施用对微生物群落的影响不同,NPK及NPK配施有机肥可提高土壤微生物多样性。     

Denaturing Gradient Gel Electrophoresis Can Rapidly Display the Bacterial Diversity Contained in 16S rDNA Clone Libraries

Two different strategies for molecular analysis of bacterial diversity, 16S rDNA cloning and denaturing gradient gel electrophoresis (DGGE), were combined into a single protocol that took advantage of the best attributes of each: the ability of cloning to package DNA sequence information and the ability of DGGE to display a community profile. In this combined protocol, polymerase chain reaction products from environmental DNA were cloned, and then DGGE was used to screen the clone libraries. Both individual clones and pools of randomly selected clones were analyzed by DGGE, and these migration patterns were compared to the conventional DGGE profile produced directly from environmental DNA. For two simple bacterial communities (biofilm from a humics-fed laboratory reactor and planktonic bacteria filtered from an urban freshwater pond), pools of 35–50 clones produced DGGE profiles that contained most of the bands visible in the conventional DGGE profiles, indicating that the clone pools were adequate for identifying the dominant genotypes. However, DGGE profiles of two different pools of 50 clones from a lawn soil clone library were distinctly different from each other and from the conventional DGGE profile, indicating that this small number of clones poorly represented the bacterial diversity in soil. Individual clones with the same apparent DGGE mobility as prominent bands in the humics reactor community profiles were sequenced from the clone plasmid DNA rather than from bands excised from the gel. Because a longer fragment was cloned (∼1500 bp) than was actually analyzed in DGGE (∼350 bp), far more sequence information was available using this approach that could have been recovered from an excised gel band. This clone/DGGE protocol permitted rapid analysis of the microbial diversity in the two moderately complex systems, but was limited in its ability to represent the diversity in the soil microbial community. Nonetheless, clone/DGGE is a promising strategy for fractionating diverse microbial communities into manageable subsets consisting of small pools of clones.