Use of length heterogeneity PCR and fatty acid methyl ester profiles to characterize microbial communities in soil

In length heterogeneity PCR (LH-PCR) a fluorescently labeled primer is used to determine the relative amounts of amplified sequences originating from different microorganisms. Labeled fragments are separated by gel electrophoresis and detected by laser-induced fluorescence with an automated gene sequencer. We used LH-PCR to evaluate the composition of the soil microbial community. Four soils, which differed in terms of soil type and/or crop management practice, were studied. Previous data for microbial biomass, nitrogen and carbon contents, and nitrogen mineralization rates suggested that the microbial characteristics of these soils were different. One site received two different treatments: no-till and conventional till perennial ryegrass. The other sites were no-till continuous grass plots at separate locations with different soil types. Community composition was characterized by assessing the natural length heterogeneity in eubacterial sequences amplified from the 5' domain of the 16S rRNA gene and by determining fatty acid methyl ester (FAME) profiles. We found that LH-PCR results were reproducible. Both methods distinguished the three sites. The most abundant bacterial community members, based on cloned LH-PCR products, were members of the beta subclass of the class Proteobacteria, the Cytophaga-Flexibacter-Bacteriodes group, and the high-G+C-content gram-positive bacterial group. Strong correlations were found between LH-PCR results and FAME results. We found that the LH-PCR method is an efficient, reliable, and highly reproducible method that should be a useful tool in future assessments of microbial community composition.     

Burning Fire-Prone Mediterranean Shrublands: Immediate Changes in Soil Microbial Community Structure and Ecosystem Functions

Wildfires subject soil microbes to extreme temperatures and modify their physical and chemical habitat. This might immediately alter their community structure and ecosystem functions. We burned a fire-prone shrubland under controlled conditions to investigate (1) the fire-induced changes in the community structure of soil archaea, bacteria and fungi by analysing 16S or 18S rRNA gene amplicons separated through denaturing gradient gel electrophoresis; (2) the physical and chemical variables determining the immediate shifts in the microbial community structure; and (3) the microbial drivers of the change in ecosystem functions related to biogeochemical cycling. Prokaryotes and eukaryotes were structured by the local environment in pre-fire soils. Fire caused a significant shift in the microbial community structure, biomass C, respiration and soil hydrolases. One-day changes in bacterial and fungal community structure correlated to the rise in total organic C and NO(3)(-)-N caused by the combustion of plant residues. In the following week, bacterial communities shifted further forced by desiccation and increasing concentrations of macronutrients. Shifts in archaeal community structure were unrelated to any of the 18 environmental variables measured. Fire-induced changes in the community structure of bacteria, rather than archaea or fungi, were correlated to the enhanced microbial biomass, CO(2) production and hydrolysis of C and P organics. This is the first report on the combined effects of fire on the three biological domains in soils. We concluded that immediately after fire the biogeochemical cycling in Mediterranean shrublands becomes less conservative through the increased microbial biomass, activity and changes in the bacterial community structure.     

A comparison of DNA profiling techniques for monitoring nutrient impact on microbial community composition during bioremediation of petroleum-contaminated soils

Amplicon length heterogeneity PCR (LH-PCR) and terminal restriction fragment length polymorphisms (TRFLP) were used to monitor the impact that nutrient amendments had on microbial community dynamics and structural diversity during bioremediation of petroleum-contaminated soils. Slurried soils contaminated with petroleum hydrocarbons were treated in airlift bench-scale bioreactors and were either amended with optimal inorganic nutrients or left unamended. Direct DNA extraction and PCR amplification of whole eubacterial community DNA were performed with universal primers that bracketed the first two or three hypervariable regions of the 16S rDNA gene sequences. The LH-PCR method profiled a more diverse microbial community than did the TRFLP method. The LH-PCR method also tracked differences between the communities due to nutrient amendments. An in silico database search for bacterial genera with amplicon lengths represented in the community fingerprints was performed. It was possible to qualitatively identify different groups in the microbial community based on the amplicon length variations. A similar "virtual" search was performed for the TRFLP fragments using the web-based TAP-TRFLP program. Cloning and sequencing of the PCR products confirmed the in silico database matches. The application of the LH-PCR method as a monitoring tool for bioremediation could greatly enhance and extend the current understanding of the microbial community dynamics during the biodegradation of environmental contaminants.     

Black Band Disease Microbial Community Variation on Corals in Three Regions of the Wider Caribbean

Black band disease (BBD) is a pathogenic consortium of microorganisms that primarily affects massive framework-building scleractinian corals on reefs worldwide. There has been considerable debate concerning the microbial community composition of BBD. The aim of this study was to utilize microbial profiling to assess overall patterns of variation in the BBD bacterial community with respect to geographic location, host coral species, time, and nutrient regime. Length heterogeneity polymerase chain reaction (LH-PCR) was employed to differentiate BBD communities based on the natural variation in the sequence lengths within hypervariable domains of the 16S rRNA gene. Analysis of LH-PCR profiles of 97 BBD samples using multivariate ordination methods and analysis of similarity revealed significant clustering with respect to geographic region when comparing BBD sampled from reefs near Lee Stocking Island in the Bahamas’ Exuma Chain, the Northern Florida Keys (NFK), and St. John in the US Virgin Islands. There was much variability in BBD community composition on a regional basis, between sites in the NFK, and in terms of coral host species. The observed differences among BBD microbial community profiles were driven primarily by variation in relative abundance of 313–316-bp amplicons, which correspond to cyanobacteria and α-proteobacteria. The results obtained in this study support previous reports of intrinsic variability and complexity of the BBD microbial community but also suggest that this variability has biogeographic patterns.     

An assessment of the hypervariable domains of the 16S rRNA genes for their value in determining microbial community diversity: the paradox of traditional ecological indices

Amplicon length heterogeneity PCR (LH-PCR) was investigated for its ability to distinguish between microbial community patterns from the same soil type under different land management practices. Natural sagebrush and irrigated mouldboard-ploughed soils from Idaho were queried as to which hypervariable domains, or combinations of 16S rRNA gene domains, were the best molecular markers. Using standard ecological indices to measure richness, diversity and evenness, the combination of three domains, V1, V3 and V1+V2, or the combined V1 and V3 domains were the markers that could best distinguish the undisturbed natural sagebrush communities from the mouldboard-ploughed microbial communities. Bray-Curtis similarity and multidimensional scaling were found to be better metrics to ordinate and cluster the LH-PCR community profiling data. The use/misuse of traditional ecological indices such as diversity and evenness to study microbial community profiles will remain a major point to consider when performing metagenomic studies.     

Bacterial community changes and enrichment of Burkholderia-like bacteria induced by chlorinated benzoates in a peat-forest soil-microcosm

Bacterial community shifts in a peat-forest soil spiked with 3-chlorobenzoate (3CBA) or 2,5-dichlorobenzoate (2,5DCB) were monitored by PCR-amplification of the V6 to V8 regions of the 16S rRNA and rDNA, followed by separation of the amplicons by temperature gradient gel electrophoresis. 3CBA disappeared to non-detectable levels after 15 days by a biologically mediated process, while 2,5DCB remained at the initial concentration values. The experiments were conducted under microcosms systems. Addition of the chlorinated benzoates to the soil resulted in a rapid decrease of the microbial diversity, as judged by a time-dependent reduction in the number of amplicons detected by temperature gradient gel electrophoresis. Few amplicons specifically enriched in the spiked soils were cloned and characterised by sequence analysis. The identity of the cloned DNA and the corresponding soil amplicons was confirmed by hybridisation with a radioactively labelled V6-probe. Analysis of the 16S rDNA sequences indicated that Burkholderia-related bacteria dominated the enriched soil populations under 3CBA stress. In addition, enrichment cultures growing on 3CBA as sole C-source were obtained from the respective spiked soil, which were found to contain bacteria with identical 16S rDNA sequences as those induced by 3CBA stress in soil.     

Microbial diversity of topographical gradient profiles in Fushan forest soils of Taiwan

To evaluate the microbial diversity of Fushan forest soils, the variation of soil properties, microbial populations, and soil DNA with soil depth in three sites of different altitude were analyzed. Microbial population, moisture content, total organic carbon (C_org), and total nitrogen (N_tot) decreased with increasing soil depth. The valley site had the lowest microbial populations among the three tested sites due to the low organic matter content. Bacterial population was the highest among the microbial populations. The ratios of cellulolytic microbes to the total bacteria in organic layers were high, implying their roles in the carbon cycle. The microbial biomass carbon (C_mic) and nitrogen (N_mic) contents ranged from 130.5 to 564.1 μg g⁻¹ and from 16.7 to 95.4 μg g⁻¹, respectively. The valley had the lowest C_mic and N_mic. The organic layer had the highest C_mic and N_mic and decreased with soil depth. Analysis using denaturing gradient gel electrophoresis (DGGE) of polymerase chain reaction (PCR) amplicons of 16S rDNA showed that the bacterial diversity of the three sites were very similar to each other in the major bands, and the variation was in the minor bands. However, the patterns in PCR-DGGE profile through gradient horizons were different, indicating the prevalence of specific microbes at different horizons. These results suggest that the microbial diversity in the deeper horizons is not simply the diluted analogs of the surface soils and that some microbes dominate only in the deeper horizons. Topography influenced the quantity and diversity of microbial populations.     

水田改果园后土壤性质的变化及其特征

近年来,水田改作经济林地,在我国南方地区非常普遍。为深入了解这一转变对土壤质量的影响,以浙江省典型水稻土(青粉泥田)及其改果园不同年限的系列表层土壤(0—15 cm)为研究对象,应用磷脂脂肪酸生物标记等方法,研究了水田改果园后土壤理化性质和微生物群落结构等性质的变化以及它们之间的关系。结果表明,水田改果园后,土壤中大于0.25 mm水稳定性团聚体、盐基饱和度、p H值、有机质、全氮和碱解氮等随着改果园年限的延长而显著降低(P0.05)。土壤微生物生物量碳氮、微生物商和土壤呼吸强度随改果园年限增加而显著下降(P0.01)。土壤微生物群落结构也发生明显变化:磷脂脂肪酸总量显著降低(P0.01),微生物种类减少,原生动物在土壤微生物中所占比例增加,革兰氏阴性细菌与革兰氏阳性细菌比值降低(P0.01),好氧细菌/厌氧细菌和甲烷氧化菌/细菌增加(P0.01),表征养分胁迫的环丙基脂肪酸/前体物和异式脂肪酸/反异支链脂肪酸显著增加(P0.01)。冗余分析表明,土壤含水率、有机质和碱解氮是决定水田和果园土壤微生物群落结构差异的最重要因子(P0.01);改果园后,土壤微生物群落结构发生了阶段性变化,不同利用方式对微生物群落结构的影响程度要大于同一利用方式耕作不同年限对微生物群落结构的影响。研究表明,水田改果园后土壤理化性质以及生物学性质发生退化,土壤质量下降;而水田中微生物数量和种类都比较丰富,因而认为水田是土壤(地)可持续利用的一种有效方式。     

Dynamics of bacterial community exposed to hydrocarbons and oleophilic fertilizer in high-Arctic intertidal beach

Exposure of pristine microbial environments to hydrocarbon contamination stimulates growth of the initially small fraction of indigenous hydrocarbon-degrading bacteria. Custom-made oleophilic fertilizers have been demonstrated to promote oil bioremediation by boosting this proliferation. In the present study, the temporal dynamics of the bacterial community structure and the individual influences of hydrocarbons and an oleophilic fertilizer in shaping the community structure was explored during a 78 days bioremediation experiment in a high-Arctic intertidal beach environment. A combination of cultivation-independent 16S rRNA gene length-heterogeneity polymerase chain reaction (LH-PCR) profiling and identification of hydrocarbon-degrading isolates based on partial 16S rRNA gene sequences was employed. LH-PCR community profiles in the fertilizer alone and fertilized kerosene plots were largely indistinguishable throughout the experimental period, while kerosene alone plots showed a markedly different composition of dominant groups. This pointed to the fertilizer as the more decisive factor in shaping the community structure. Most prominent LH-PCR fragments which emerged after kerosene or fertilizer addition could be provisionally assigned to bacterial taxa through coinciding LH-PCR fragment lengths with hydrocarbon-degrading isolates obtained from the same type of experimental units. However, a few quantitatively significant LH-PCR groups had no counterparts among the cultivated bacteria. One of these was affiliated to a hitherto unspeciated subgroup within the Alkanindiges/Acinetobacter clade of Moraxellaceae by a 16S rRNA gene cloning approach.     

Soil microbiome responses to the short‐term effects of Amazonian deforestation

Slash‐and‐burn clearing of forest typically results in increase in soil nutrient availability. However, the impact of these nutrients on the soil microbiome is not known. Using next generation sequencing of 16S rRNA gene and shotgun metagenomic DNA, we compared the structure and the potential functions of bacterial community in forest soils to deforested soils in the Amazon region and related the differences to soil chemical factors. Deforestation decreased soil organic matter content and factors linked to soil acidity and raised soil pH, base saturation and exchangeable bases. Concomitant to expected changes in soil chemical factors, we observed an increase in the alpha diversity of the bacterial microbiota and relative abundances of putative copiotrophic bacteria such as Actinomycetales and a decrease in the relative abundances of bacterial taxa such as Chlamydiae, Planctomycetes and Verrucomicrobia in the deforested soils. We did not observe an increase in genes related to microbial nutrient metabolism in deforested soils. However, we did observe changes in community functions such as increases in DNA repair, protein processing, modification, degradation and folding functions, and these functions might reflect adaptation to changes in soil characteristics due to forest clear‐cutting and burning. In addition, there were changes in the composition of the bacterial groups associated with metabolism‐related functions. Co‐occurrence microbial network analysis identified distinct phylogenetic patterns for forest and deforested soils and suggested relationships between Planctomycetes and aluminium content, and Actinobacteria and nitrogen sources in Amazon soils. The results support taxonomic and functional adaptations in the soil bacterial community following deforestation. We hypothesize that these microbial adaptations may serve as a buffer to drastic changes in soil fertility after slash‐and‐burning deforestation in the Amazon region.     

集约经营对毛竹林土壤固氮细菌群落结构和丰度的影响

为揭示集约经营对毛竹林土壤固氮细菌群落特征的影响,采用变性梯度凝胶电泳(DGGE)和荧光定量PCR技术,分析集约经营0(CK)、10、15、20、25年毛竹林土壤固氮菌群落结构和丰度的变化规律,并探讨了影响土壤固氮菌群落的主要环境因素.结果表明： 毛竹林集约经营导致土壤pH下降而速效养分积累;集约经营初期(10年)和后期(25年)土壤固氮细菌的群落结构与对照相似,而中期的15年和20年则与对照差异较大.固氮菌多样性指数和丰度均呈现先减少后增加的趋势,经营15年时达到最小值;土壤固氮细菌表现出对集约经营干扰的抵抗和恢复反应.冗余分析表明,土壤速效钾、水解氮、硝态氮和铵态氮的含量与固氮菌群落结构的变化有较强的相关性,表明集约经营措施导致了土壤固氮细菌短期的变化,但长期而言,不会对土壤固氮细菌产生不良影响.     

长期重金属胁迫对农田土壤微生物生物量、活性和种群的影响

调查了沈阳张士灌区长期污水灌溉造成的原位农田土壤重金属污染状况,从土壤微生物生物量、微生物活性和微生物种群数量的角度评价了长期重金属污染对农田土壤生态系统的影响.结果表明,张士灌区土壤存在严重的Cd污染,土壤Cd含量达1.75～3.89 mg·kg ^-1,部分区域还伴有Cu、Zn复合污染.在目前污染程度下,土壤微生物生物量碳(C_mic)、微生物商(qM)、土壤脱氢酶活性以及自生固氮菌数量随土壤重金属含量增加呈下降趋势,代谢商(qCO₂)随土壤重金属含量增加显著升高,而底物诱导呼吸强度(SIR)、纤维素酶活性以及细菌、放线菌和真菌数量无明显变化.相关性分析表明,土壤Cd含量变化是影响微生物参数变化的主要因素,在微生物参数中微生物商和代谢商对重金属污染最敏感.     

五大连池火山土壤细菌多样性及其群落结构

火山熔岩生境孕育了独特的土壤微生物群落。为了解火山生态系统土壤细菌群落多样性和群落结构及其关键影响因子,选择五大连池新、老期火山为研究样点,非火山为对照,基于高通量测序方法,分析不同采样点土壤细菌群落结构和多样性,结合土壤理化指标,进一步分析影响火山生态系统土壤细菌群落多样性的环境因子。结果表明：细菌操作分类单元（OTUs）、Ace指数、Chao1指数和Simpson指数变化趋势一致,表现为非火山 > 新期火山 > 老期火山。三个样点土壤的共有OUTs数量为713个,各自特有的OTUs数量不尽相同。三个样点土壤中检测到共有细菌16个类群,其中变形菌门、酸杆菌门、放线菌门和绿弯菌门为优势菌群,老期火山土壤中酸杆菌门、疣微菌门、Rokubacteria相对丰度最大,而Patescibacteria相对丰度最小。三个样点的土壤细菌群落具有明显的空间关系,相似性差异较大,但不符合随地理距离的增加而降低的模型。土壤理化性质测定结果标明：老期火山土壤pH、有机质、全氮、全磷、铵态氮和硝态氮显著高于新期火山和非火山,新期火山土壤含水量和速效磷显著低于老期火山和非火山。喷发时间和火成岩基质等特性会导致不同火山土壤理化性质的差异,进而影响土壤细菌多样性和群落结构。Pearson相关性分析表明：土壤pH显著影响细菌的多样性指数。冗余分析（RDA）结果表明：土壤氮含量、pH和有机质是影响火山森林生态系统土壤细菌群落结构的主要因子。     

Short-term effects of wildfire on microbial biomass and abundance in black pine plantation soils in Turkey

Measurement of soil microbial biomass and abundance offers a means of assessing the response of all microbial populations to changes in the soil environment after a fire. We examined the effects of wildfire on microbial biomass C and N, and abundance of bacteria and fungi 2 months after a fire in a pine plantation. Soil organic carbon (C_org), total nitrogen (N_tot), and electrical conductivity (EC) increased following the fire. In terms of microbial abundance, the overall results showed that burned forest soils had the most bacteria and fungi. Microbial biomass C and N from soil in the burned forest were not significantly different from their unburned forest counterparts. However, microbial indices indicated that fire affects soil microbial community structure by modifying the environmental conditions. The results also suggested that low-intensity fire promotes microorganism functional activity and improves the chemical characteristics of soils under humid climatic conditions.     

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.     

An assessment of soil bacterial community structure and physicochemistry in two microtopographic locations of a palustrine forested wetland

We studied redoximorphic features, field indicators and bacterial communities of soils in hummocks and hollows of a palustrine forested wetland in Virginia. We hypothesized that presence of hydric soils, soil physicochemistry and soil bacterial community structure would differ between hummocks and hollows. We fingerprinted soils collected from different microtopographic locations using Length Heterogeneity Polymerase Chain Reaction (LH-PCR) to study their bacterial community structures. Two hummocks had silty/sandy loam soils with mean chroma values of > 4, showing no indication of ‘hydric soils’ (i.e., wetland soils). Two hollows, however, had clay loam soils with mean chroma values of 2 with gleying and redox concentrations observed, indicative of seasonally inundated wetlands. The soils of hollows also had higher organic matter content and soil moisture compared to the soils of hummocks (P < 0.05). Multidimensional scaling (MDS) and Analysis of similarity (ANOSIM) of the fingerprints revealed differences in soil microbial community structures between hummocks and hollows (Global R = 0.30, P < 0.01). The diversity measures of the fingerprints (Shannon’s H′) were also different by microtopography with higher diversity in hollows relative to hummocks (P < 0.05). LH-PCR proves to be a useful tool in examining bacterial community composition of wetland soils in this study. However, cloning and sequencing of specific community LH-PCR profiles of interest is necessary to fully characterize the community down to genus/species level. With species identities we should be able to not only better explain differences observed in the community profiles, but study their relations to hydrologic and/or physicochemical conditions of wetlands.     

Responses of Bacterial Communities to Simulated Climate Changes in Alpine Meadow Soil of the Qinghai-Tibet Plateau

The soil microbial community plays an important role in terrestrial carbon and nitrogen cycling. However, microbial responses to climate warming or cooling remain poorly understood, limiting our ability to predict the consequences of future climate changes. To address this issue, it is critical to identify microbes sensitive to climate change and key driving factors shifting microbial communities. In this study, alpine soil transplant experiments were conducted downward or upward along an elevation gradient between 3,200 and 3,800 m in the Qinghai-Tibet plateau to simulate climate warming or cooling. After a 2-year soil transplant experiment, soil bacterial communities were analyzed by pyrosequencing of 16S rRNA gene amplicons. The results showed that the transplanted soil bacterial communities became more similar to those in their destination sites and more different from those in their “home” sites. Warming led to increases in the relative abundances in Alphaproteobacteria, Gammaproteobacteria, and Actinobacteria and decreases in Acidobacteria, Betaproteobacteria, and Deltaproteobacteria, while cooling had opposite effects on bacterial communities (symmetric response). Soil temperature and plant biomass contributed significantly to shaping the bacterial community structure. Overall, climate warming or cooling shifted the soil bacterial community structure mainly through species sorting, and such a shift might correlate to important biogeochemical processes such as greenhouse gas emissions. This study provides new insights into our understanding of soil bacterial community responses to climate warming and cooling.     

黄土丘陵沟壑区不同植被恢复格局下土壤微生物群落结构

针对典型黄土丘陵沟壑区陕西延安羊圈沟小流域坡面上单一刺槐林、单一撂荒草地以及林草搭配的草地-林地-草地及林地-草地-林地4种不同植被格局,利用磷脂脂肪酸(phospholipid fatty acid,PLFA)谱图分析法对土壤微生物群落结构进行监测研究,旨在揭示坡面上不同的植被恢复格局对土壤微生物群落结构的影响。研究发现4种不同植被格局下,2种林草搭配的植被格局磷脂脂肪酸的结构比较相似,与单一植被格局相比,表层土壤中表征真菌的特征脂肪酸所占的比例有所提高。主成分分析显示4种植被格局0—10 cm土壤微生物群落结构存在差异,差异主要存在于2种林草搭配的植被格局与2种单一的植被格局之间,其中草地-林地-草地的植被格局与刺槐林和撂荒草地之间土壤微生物群落结构的差异均达到了显著水平。不同微生物菌群的量在4种植被格局土壤间显著性差异主要存在于表层土壤中的细菌菌群和革兰氏阳性菌,革兰氏阴性菌和真菌在4种植被格局土壤之间无显著差异。总之,4种不同植被恢复格局的土壤微生物群落结构存在差异且差异主要存在于表层土壤,坡面上人工林的种植及林草搭配的恢复模式较直接撂荒更有利于提高微生物菌群的生物量。     

Molecular identification of diazotroph microbial consortia in mountain soil

Nitrogen fixing microbial consortia from soil samples taken from five altitudinal vegetation zones (alpine, subalpine, coniferous, beech, Maleia flood plain) of Parang Massif, Romania, were isolated and identified. Molecular characterisation of nitrogen fixing consortia was carried out by PCR and nested PCR with 7 primer sets specific to nifH genes. All nifH genes are specific to nitrogen fixation and are found within phylogenetically related organisms which have the nitrogenase enzyme complex. These molecular studies allowed the assessment of nifH gene diversity within these nitrogen fixing microbial consortia from different type of soils. At high altitude, a consortium of nitrogen fixing bacteria dominated by Azotobacter chroococcum and Azospirillum brasilense was found. Clostridium, Rhizobiales, Herbaspirillum, Frankia species were also found in different rations depending on the altitudinal vegetation zone.     

PCR-DGGE Comparison of Bacterial Community Structure in Fresh and Archived Soils Sampled along a Chihuahuan Desert Elevational Gradient

The polymerase chain reaction coupled with denaturing gradient gel electrophoresis (PCR-DGGE) has been used widely to determine species richness and structure of microbial communities in a variety of environments. Researchers commonly archive soil samples after routine chemical or microbial analyses, and applying PCR-DGGE technology to these historical samples offers evaluation of long-term patterns in bacterial species richness and community structure that was not available with previous technology. However, use of PCR-DGGE to analyze microbial communities of archived soils has been largely unexplored. To evaluate the stability of DGGE patterns in archived soils in comparison with fresh soils, fresh and archived soils from five sites along an elevational gradient in the Chihuahuan Desert were compared using PCR-DGGE of 16S rDNA. DNA from all archived samples was extracted reliably, but DNA in archived soils collected from a closed-canopy oak forest site could not be amplified. DNA extraction yields were lower for most archived soils, but minimal changes in bacterial species richness and structure due to archiving were noted in bacterial community profiles from four sites. Use of archived soils to determine long-term changes in bacterial community structure via PCR-DGGE appears to be a viable option for addressing microbial community dynamics for particular ecosystems or landscapes.