Analysis of BIOLOG GN Substrate Utilization Patterns by Microbial Communities

BIOLOG GN plates are increasingly used to characterize microbial communities by determining the ability of the communities to oxidize various carbon sources. Studies were done to determine whether the BIOLOG GN plate assay accurately reflects the catabolic potential of the inoculum used. To gain insight into which populations of microbial communities contribute to the BIOLOG patterns, denaturing gradient gel electrophoresis and temperature gradient gel electrophoresis (TGGE) were used to assess the diversity of ribotypes in the inocula and individual wells of BIOLOG plates following incubation. These studies were done with microbial communities from the rhizosphere of potatoes and an activated sludge reactor fed with glucose and peptone. TGGE analyses of BIOLOG wells inoculated with cell suspensions from the potato rhizosphere revealed that, compared with the inoculum, there was a decrease in the number of 16S rRNA gene fragments obtained from various wells, as well as a concomitant loss of populations that had been numerically dominant in the inoculum. The dominant fragments in TGGE gels could be assigned to the gamma subclass of the class Proteobacteria, suggesting that fast-growing bacteria adapted to high substrate concentrations were numerically dominant in the wells and may have been primarily responsible for the patterns of substrate use that were observed. Similarly, the community structure changed in wells inoculated with cells from activated sludge; one or more populations were enriched, but all dominant populations of the inoculum could be detected in at least one well. This study showed that carbon source utilization profiles obtained with BIOLOG GN plates do not necessarily reflect the functional potential of the numerically dominant members of the microbial community used as the inoculum.     

Patterns of Formation of Microbial Communities of Alluvial Soils in the Selenga Estuary

Microbial communities of alluvial soils in the facies on islands in the Selenga estuary and near Stepnoi Dvorets and Istomino Settlements were studied. The distribution of the CFU-dominant microorganisms (Bacillus and Pseudomonas + Arthrobacter) has been revealed in the studied island soils. In addition, actinomycetes Streptomyces were found among the dominants in soils of the near-terrace floodplain. The quantitative distribution of microorganisms across soil layers was demonstrated. The effect of ecological factors in formation of microbial communities on the islands and central floodplain regions was revealed.     

Spatial Patterns of Soil Microbial Communities in a Norway Spruce (Picea abies) Plantation

The phospholipid fatty acid (PLFA) profiles of soil microbial communities were determined in relation to the patterns of tree cover in a mature Norway spruce plantation. Replicate samples of the surface organic layers were taken close to the trunk, at 1 m and at 2 m (under the edge of the canopy) beneath five trees. Samples were analyzed for standard PLFAs to assess the initial composition of the microbial communities. Replicate samples were then incubated under constant or fluctuating moisture conditions for 30 d to test the hypothesis that the patterns of microbial community structure (or its physiological state) might be determined by biophysical conditions under the tree canopies. The PLFA profiles near the trunks and at 2 m were similar, but samples taken 1 m from the bases of the trees contained lower concentrations of polyunsaturated (fungal) and monounsaturated PLFAs, and higher concentrations of saturated PLFAs. These differences in PLFA profiles were maintained during laboratory incubation under a regime of drying and wetting cycles, but there was some evidence of convergence in community structure under constant moisture conditions resulting from significant increases and decreases in specific bacterial PLFA concentrations. There were no effects of either moisture treatment on fungal PLFA concentrations. It is concluded that variation in the soil biophysical environment beneath the tree canopies resulted in the differentiation of spatially defined bacterial communities that were tolerant of moisture stress. The anomaly that differences in community structure were largest at an intermediate position of 1 m between the trunk and below the canopy edge was not explained but may relate to tree root distribution.     

Microbial Communities in Pre-Columbian Coprolites

The study of coprolites from earlier cultures represents a great opportunity to study an “unaltered” composition of the intestinal microbiota. To test this, pre-Columbian coprolites from two cultures, the Huecoid and Saladoid, were evaluated for the presence of DNA, proteins and lipids by cytochemical staining, human and/or dog-specific Bacteroides spp. by PCR, as well as bacteria, fungi and archaea using Terminal Restriction Fragment analyses. DNA, proteins and lipids, and human-specific Bacteroides DNA were detected in all coprolites. Multidimensional scaling analyses resulted in spatial arrangements of microbial profiles by culture, further supported by cluster analysis and ANOSIM. Differences between the microbial communities were positively correlated with culture, and SIMPER analysis indicated 68.8% dissimilarity between the Huecoid and Saladoid. Proteobacteria, Bacteroidetes and methanogens were found in all coprolite samples. Propionebacteria, Shewanella and lactic acid bacteria dominated in the Huecoid samples, while Acidobacteria, and peptococci were dominant in Saladoid samples. Yeasts, including Candida albicans and Crypotococcus spp. were found in all samples. Basidiomycetes were the most notable fungi in Huecoid samples while Ascomycetes predominated in Saladoid samples, suggesting differences in dietary habits. Our study provides an approach for the study of the microbial communities of coprolite samples from various cultures.     

Simultaneous Amplicon Sequencing to Explore Co-Occurrence Patterns of Bacterial,Archaeal and Eukaryotic Microorganisms in Rumen Microbial Communities

Ruminants rely on a complex rumen microbial community to convert dietary plant material to energy-yielding products. Here we developed a method to simultaneously analyze the community''s bacterial and archaeal 16S rRNA genes, ciliate 18S rRNA genes and anaerobic fungal internal transcribed spacer 1 genes using 12 DNA samples derived from 11 different rumen samples from three host species (Ovis aries, Bos taurus, Cervus elephas) and multiplex 454 Titanium pyrosequencing. We show that the mixing ratio of the group-specific DNA templates before emulsion PCR is crucial to compensate for differences in amplicon length. This method, in contrast to using a non-specific universal primer pair, avoids sequencing non-targeted DNA, such as plant- or endophyte-derived rRNA genes, and allows increased or decreased levels of community structure resolution for each microbial group as needed. Communities analyzed with different primers always grouped by sample origin rather than by the primers used. However, primer choice had a greater impact on apparent archaeal community structure than on bacterial community structure, and biases for certain methanogen groups were detected. Co-occurrence analysis of microbial taxa from all three domains of life suggested strong within- and between-domain correlations between different groups of microorganisms within the rumen. The approach used to simultaneously characterize bacterial, archaeal and eukaryotic components of a microbiota should be applicable to other communities occupying diverse habitats.     

Disordered Microbial Communities in Asthmatic Airways

Background

A rich microbial environment in infancy protects against asthma [1], [2] and infections precipitate asthma exacerbations [3]. We compared the airway microbiota at three levels in adult patients with asthma, the related condition of COPD, and controls. We also studied bronchial lavage from asthmatic children and controls.

Principal Findings

We identified 5,054 16S rRNA bacterial sequences from 43 subjects, detecting >70% of species present. The bronchial tree was not sterile, and contained a mean of 2,000 bacterial genomes per cm² surface sampled. Pathogenic Proteobacteria, particularly Haemophilus spp., were much more frequent in bronchi of adult asthmatics or patients with COPD than controls. We found similar highly significant increases in Proteobacteria in asthmatic children. Conversely, Bacteroidetes, particularly Prevotella spp., were more frequent in controls than adult or child asthmatics or COPD patients.

Significance

The results show the bronchial tree to contain a characteristic microbiota, and suggest that this microbiota is disturbed in asthmatic airways.     

Cypris Habitat Selection Facilitated by Microbial Films Influences the Vertical Distribution of Subtidal Barnacle Balanus trigonus

The potential driving force(s) of the vertical distribution of subtidal barnacle Balanus trigonus Darwin were investigated using both field and laboratory experiments. Early juveniles (∼24 h old) placed in intertidal [∼0.5 m above mean low water level (MLWL)] and subtidal (∼3 m below MLWL) habitats survived equally well, indicating that the intertidal absence of B. trigonus in Hong Kong waters was not determined by differential mortality. However, enhanced attachment of cyprids in subtidal habitats indicated the importance of differential larval choice in determining their vertical distribution. In the laboratory, cyprids preferred to attach in response to subtidal microbial films, which may implicate microbial films as a primary cue in driving the adult vertical distribution. Microbial films developed in these two habitats differed in their biomass (=total organic carbon), abundance of bacteria and diatoms (determined by fluorescence microscopy), and bacterial diversity (determined by DNA fingerprinting analysis). For example, 6-day films in subtidal habitat had a significantly higher biomass than in films from intertidal habitat (P<0.05). There was no difference in the biomass of films from these two habitats in 9-day films (P>0.05); however, bacterial abundance was greater in subtidal films than in intertidal films, irrespective of the age of the film, although there was no difference in diatom abundance in films from these two habitats. Neither the abundance of bacteria and diatoms nor the biomass correlated with the attachment preferences of cyprids. This study has not provided any data to prove the existence of inductive and inhibitive (to cyprid attachment) bacterial species in subtidal and intertidal films, respectively; however, results indicate that bacterial community provided qualitative information that might explain the preferential attachment of B. trigonus cyprids in subtidal habitat.     

Urea Transformation of Wetland Microbial Communities

Transformation of urea to ammonium is an important link in the nitrogen cycle in soil and water. Although microbial nitrogen transformations, such as nitrification and denitrification, are well studied in freshwater sediment and epiphytic biofilm in shallow waters, information about urea transformation in these environments is scarce. In this study, urea transformation of sedimentary, planktonic, and epiphytic microbial communities was quantified and urea transformation of epiphytic biofilms associated with three different common wetland macrophyte species is compared. The microbial communities were collected from a constructed wetland in October 2002 and urea transformation was quantified in the laboratory at in situ temperature (12°C) with the use of the ¹⁴C-urea tracer method, which measures the release of ¹⁴CO₂ as a direct result of urease activity. It was found that the urea transformation was 100 times higher in sediment (12–22 mmol urea-N m⁻² day⁻¹) compared with the epiphytic activity on the surfaces of the submerged plant Elodea canadensis (0.1–0.2 mmol urea-N m⁻² day⁻¹). The epiphytic activity of leaves of Typha latifolia was lower (0.001–0.03 mmol urea-N m⁻² day⁻¹), while urea transformation was negligible in the water column and on the submerged leaves of the emergent plant Phragmites australis. However, because this wetland was dominated by dense beds of the submerged macrophyte E. canadensis, this plant provided a large surface area for epiphytic microbial activity—in the range of 23–33 m² of plant surfaces per square meter of wetland. Thus, in the wetland system scale at the existing plant distribution and density, the submerged plant community had the potential to transform 2–7 mmol urea-N m⁻² day⁻¹ and was in the same magnitude as the urea transformation in the sediment.     

Spatial Organization of Microbial Biofilm Communities

Abstract The application of advanced microscopy and molecular and electrochemical high-resolution methods has provided insights into the structural organization and function of biofilm communities. It appears that cellular properties such as growth differentiation, chemotaxis, and cell-to-cell signaling enable biofilm communities to organize structurally in response to the external conditions and the activities of the different biofilm members. Thereby resource utilization becomes optimized, and processes which require syntrophic relationships or special micro-environments become facilitated. Received: 23 February 2000; Accepted: 8 June 2000; Online Publication: 28 August 2000     

Microbial Communities and Exopolysaccharides from Polynesian Mats

Microbial mats present in two shallow atolls of French Polynesia were characterized by high amounts of exopolysaccharides associated with cyanobacteria as the predominating species. Cyanobacteria were found in the first centimeters of the gelatinous mats, whereas deeper layers showing the occurrence of the sulfate reducers Desulfovibrio and Desulfobacter species as determined by the presence of specific biomarkers. Exopolysaccharides were extracted from these mats and partially characterized. All fractions contained both neutral sugars and uronic acids with a predominance of the former. The large diversity in monosaccharides can be interpreted as the result of exopolymer biosynthesis by either different or unidentified cyanobacterial species. Received July 25, 2000; accepted October 21, 2000     

Patterns in the Composition of Microbial Communities from a Subtropical River: Effects of Environmental,Spatial and Temporal Factors

Microbes are key components of aquatic ecosystems and play crucial roles in global biogeochemical cycles. However, the spatiotemporal dynamics of planktonic microbial community composition in riverine ecosystems are still poorly understood. In this study, we used denaturing gradient gel electrophoresis of PCR-amplified 16S and 18S rRNA gene fragments and multivariate statistical methods to explore the spatiotemporal patterns and driving factors of planktonic bacterial and microbial eukaryotic communities in the subtropical Jiulong River, southeast China. Both bacterial and microbial eukaryotic communities varied significantly in time and were spatially structured according to upper stream, middle-lower stream and estuary. Among all the environmental factors measured, water temperature, conductivity, PO₄-P and TN/TP were best related to the spatiotemporal distribution of bacterial community, while water temperature, conductivity, NO_x-N and transparency were closest related to the variation of eukaryotic community. Variation partitioning, based on partial RDA, revealed that environmental factors played the most important roles in structuring the microbial assemblages by explaining 11.3% of bacterial variation and 17.5% of eukaryotic variation. However, pure spatial factors (6.5% for bacteria and 9.6% for eukaryotes) and temporal factors (3.3% for bacteria and 5.5% for eukaryotes) also explained some variation in microbial distribution, thus inherent spatial and temporal variation of microbial assemblages should be considered when assessing the impact of environmental factors on microbial communities.     

Habitat Use of Fish Communities in A Virginia Stream System

Fish habitat use during summer was examined at micro- and meso-levels, to determine species associations in the upper Roanoke River watershed, Virginia. Based on multivariate-mesohabitat analysis and examination of mean microhabitat use, seven habitat-use guilds were apparent. These included four rheophilic (fast-riffle, riffle/run, fast-generalist, and shallow-rheophilic) and three limnophilic guilds (pool/run, open-pool, and pool-cover) that were reasonably robust across two river segments and two years. Although simple-hydraulic, bottom-topographic, and turbulence variables all segregated fish habitat-use guilds, turbulence variables were redundant with simple-hydraulic variables, substratum use by limnophilic fishes was related to availability, and only one guild consistently selected high cover levels. At the family level, suckers, darters, and especially minnows were notable for occupying several habitat-use guilds, because of species differences in habitat preferences. Such formulation of guilds can simplify habitat-impact analyses in biodiverse, warmwater streams, via focus on habitat needs of guilds rather than on individual species.     

Functional Patterns of Microbial Communities of Rhizospheric Soils Across the Development Stages of a Young Mangrove in French Guiana

The functional patterns of microbial communities (microbial respiration, enzyme activities, functional diversity) and the relevant physico-chemical characteristics of rhizospheric soils were studied during the process of mudflat colonization by mangrove. The study site is a fringe mangrove stand located in Montabo Bay at Cayenne (French Guiana). It is characterized by different vegetation development stages dominated by an assemblage of Avicennia germinans and Laguncularia racemosa. Rhizospheric and surface soils were collected from three stations based on successional stages of mangrove colonization: pioneer (P), coppice (C), and young forest (F). The microbial functional patterns showed significant progressive shifts along the mangrove vegetation profile. The P stages, those most influenced by tide currents, were macroscopically characterized by hydro-sedimentary instability and micro-phytobenthic colonization of mudflat. This stage, characterized by low total organic carbon (TOC) content and quality, showed the lowest extracellular enzymatic activities and the highest functional metabolic diversities. TOC quality analyses by ¹³C CPMAS NMR provided evidence of progressive TOC enrichment and an increasing imprint of aboveground vegetation on C quality as succession occurs. These differences in the origin, amount, and quality of soil organic matter (SOM) of older stages exerted both a quantitative and qualitative control over microbial functional responses. This indicated the enhancement of aboveground–belowground functional linkages, leading to the expression of high decomposition activities and a functional loss and specialization of rhizospheric microbial communities.     

Generation of Electricity and Analysis of Microbial Communities in Wheat Straw Biomass-Powered Microbial Fuel Cells

Electricity generation from wheat straw hydrolysate and the microbial ecology of electricity-producing microbial communities developed in two-chamber microbial fuel cells (MFCs) were investigated. The power density reached 123 mW/m² with an initial hydrolysate concentration of 1,000 mg chemical oxygen demand (COD)/liter, while coulombic efficiencies ranged from 37.1 to 15.5%, corresponding to the initial hydrolysate concentrations of 250 to 2,000 mg COD/liter. The suspended bacteria found were different from the bacteria immobilized in the biofilm, and they played different roles in electricity generation from the hydrolysate. The bacteria in the biofilm were consortia with sequences similar to those of Bacteroidetes (40% of sequences), Alphaproteobacteria (20%), Bacillus (20%), Deltaproteobacteria (10%), and Gammaproteobacteria (10%), while the suspended consortia were predominately Bacillus (22.2%). The results of this study can contribute to improving understanding of and optimizing electricity generation in microbial fuel cells.Wheat straw is one of the most abundant renewable resources. According to the Food and Agriculture Organization of the United Nations, approximately 1.9 × 10⁹ tons of wheat straw annually are produced worldwide, accompanied by 6.2 × 10⁸ tons of wheat production. Wheat straw is composed of 35 to 45% cellulose and 20 to 30% hemicelluloses with a relatively low lignin content (<20%) (42). The hemicellulose fraction of the straw is easily hydrolyzed to its constituent sugars by a hydrothermal treatment process, forming a carbohydrate-enriched liquid hydrolysate (46). Chemical and biological approaches to sustainable energy production from the liquefied hydrolysates to energy carriers, such as methane, ethanol, and H₂, have been developed. However, many of these approaches encounter technical and economical hurdles (10, 12, 15, 16). An alternative strategy is direct conversion of wheat straw biomass to electrical energy in microbial fuel cells (MFCs).MFCs are bioelectrochemical reactors in which microorganisms mediate the direct conversion of chemical energy stored in organic matter or bulk biomass into electrical energy (12, 15, 16, 40). Various substrates, such as simple carbohydrates, low-molecular-weight organic acids, starch, amino acids, chitin, cellulose, domestic wastewater, food-processing wastewater, recycled paper wastewater, and marine sediment organic matter, have been successfully utilized for power generation in MFCs (16-18, 27, 30, 33). To understand the microbial constraints on various fuel-powered MFCs, microbial communities have been characterized by several groups. Microbial communities from various systems are very different and often diverse, ranging from well-known metal- and anode-reducing bacteria to unknown exoelectrogens (1, 20, 21). It has been found that parameters such as the substrates used as fuels and the inocula used for starting up the MFCs can influence the anode bacterial communities in an MFC, which subsequently influence the efficiency of the MFCs (3, 14, 22, 38, 44). Different pure substrates, such as acetate, glucose, and lactate, were used as fuel to compare the microbial communities that developed in the MFCs. Regardless of the different substrates, all anode communities contained sequences closely affiliated with Geobacter sulfurreducens (>99% similarity) and an uncultured bacterium clone belonging to the family Bacteroidaceae (99% similarity). Firmicutes were only found in glucose-fed MFCs (20). Microbial-community analyses of MFCs powered with complex substrates have also been performed by several researchers, and their results were very diverse. The microbial community in starch wastewater-powered MFC was dominated by unidentified bacteria (35.9%), followed by Betaproteobacteria (25.0%), Alphaproteobacteria (20.1%), and the Cytophaga/Flexibacter/Bacteroides group (19.0%) (21). The anode-attached consortia in a cellulose-powered MFC were related to Clostridium spp., while Comamonas spp. were abundant in the suspended consortia (13). Although many studies have reported the microbial compositions of MFCs, it is still unclear which microbial communities develop as a function of the external parameters.Wheat straw biomass constitutes a large source for bioenergy production and shows promising prospects for electricity generation in MFCs. Therefore, wheat straw biomass was used to study the microbial communities that develop during the operation of an MFC in order to better understand the microbial electrochemical roles and potentially improve MFC performance.The objectives of this study were to (i) test wheat straw hydrolysate as a potential fuel in an MFC for electricity generation and (ii) study the microbial composition and evolution of electricity-producing communities in a two-chamber MFC system. Phylogenetic-diversity analysis of the enriched consortia was conducted to verify the presence of hydrolytic and respiratory anaerobes that could couple hydrolysate oxidation with proton reduction in the anode chamber. This is the first report of exploiting microbial communities for direct conversion of wheat straw hydrolysate to electrical energy in an MFC.     

Syntrophy and Interspecies Electron Transfer in Methanogenic Microbial Communities

Microbiology - Anaerobic digestion of municipal and other organic waste is a microbial process for conversion of complex organic substances to biogas (a renewable energy source) comprising a...