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Microbial community analysis in the denitrification process of saline-wastewater by denaturing gradient gel electrophoresis of PCR-amplified 16S rDNA and the cultivation method.

The metallurgic wastewater generated from the processes of recovering precious metals from industrial wastes contains high concentrations of nitrogen compounds and salts. Biological nitrogen removal from this wastewater was attempted using a circulating bioreactor system equipped with an anaerobic packed bed or an anaerobic fluidized bed. The denitrification capability of the system with the anaerobic packed bed was more stable than that of the system with the anaerobic fluidized bed. The NOx removal rate of the anaerobic packed bed was as high as 97%. Microbial community analysis by denaturing gradient gel electrophoresis (DGGE) of PCR-amplified 16S ribosomal DNA (rDNA) fragments and the cultivation method revealed that the community diversity varied in accordance with wastewater composition such as the level of salinity and so on. Phylogenetic analysis suggested that the taxonomic affiliation of the dominant species in the anaerobic reactors was to the gamma-Proteobacteria including Halomonadaceae species. The PCR-DGGE method as a non-cultivation method was found to be a powerful tool for analysis of the microbial community, because the cultivation method could detect only a fraction of the microbial species present in these systems. The genetic diversity of the isolated bacteria belonging to the gamma-Proteobacteria which reduced both nitrate and nitrite in the anaerobic packed bed was higher than that of the bacteria in the anaerobic fluidized bed. This suggested that a genetically diverse microbial community stabilized the denitrifying performance in the anaerobic packed bed.

Journal Article↗

Compost microbiomes as reservoirs of cellulolytic microorganisms for cellulosic textile degradation.

Cellulosic textiles, constituting over 30% of global fibre production, are biodegradable but remain challenging to recycle at scale owing to their high crystallinity, chemical finishes, and heterogeneous waste streams. Although microorganisms drive cellulose turnover in natural ecosystems, their potential for transforming anthropogenic cellulosic waste remains largely unexplored. In this study, composting was evaluated both as a sustainable approach to textile biodegradation and a reservoir of cellulolytic microorganisms with biotechnological potential. Biodegradation assays of cotton and lyocell were integrated with shotgun metagenomics and targeted cultivation to identify microbial taxa and enzymes involved in cellulose degradation. Composting trials showed that degradation was strongly influenced by both composting system and fibre composition. Community composting achieved near-complete textile disintegration, while shredded textiles exhibited the highest degradation rates, reaching up to 97%. Shotgun metagenomic revealed a bacterial-dominated community enriched in Actinomycetota and Bacillota and characterised by an abundance of glycoside hydrolases. Culture-based screening recovered 62 microbial isolates, of which Neurospora and Aspergillus exhibited the highest cellulolytic activity (>60%). In vitro assays further showed that cotton was more readily degraded than lyocell, with several isolates achieving >70% mass loss. Metagenomic approach revealed a predominantly bacterial composting community at the sampled stage, whereas cultivation preferentially recovered fungi that, despite their low relative abundance in situ, exhibited strong cellulolytic potential. These findings highlight the potential of composting as a sustainable end-of-life strategy for cellulosic textiles and identify compost microbiomes as valuable reservoirs of cellulolytic microorganisms for the development of sustainable bioprocesses for textile waste treatment.

Cellulose↗

Microbial communities associated with skeletal tumors on Porites compressa.

Coral tumors are atypical skeletal forms found on coral reefs worldwide. Here we present an analysis of the microbial communities associated with skeletal tumors on the coral Porites compressa. Microbial growth rates on both healthy and tumorous P. compressa were decoupled from the surrounding water column. Microbial communities associated with tumorous colonies had a significantly faster growth rate than those associated with healthy P. compressa. The microbial community associated with the tumors contained more culturable Vibrio spp. and could utilize more carbon sources than the microbes associated with healthy colonies. Presence of tumors affected the composition and dynamics of the microbial population associated with the entire colony.

Animals↗

Quantitative community analysis: capillary electrophoresis techniques.

This chapter presents methodologies for RNA extraction from soils coupled with competitive reverse transcription-polymerase chain reaction and capillary electrophoresis techniques. Combined, these approaches provide new capabilities to quantify gene expression in different environments and can aid our understanding of not only community composition, but also community activity. Such information will prove important for enhancing our knowledge of how microbial communities respond to changing geochemical parameters (e.g., temperature, pH, redox conditions, substrate levels) in situ.

Electrophoresis, Capillary↗

Abundance and diversity of viruses in six Delaware soils.

The importance of viruses in marine microbial ecology has been established over the past decade. Specifically, viruses influence bacterial abundance and community composition through lysis and alter bacterial genetic diversity through transduction and lysogenic conversion. By contrast, the abundance and distribution of viruses in soils are almost completely unknown. This study describes the abundance and diversity of autochthonous viruses in six Delaware soils: two agricultural soils, two coastal plain forest soils, and two piedmont forest soils. Viral abundance was measured using epifluorescence microscopy, while viral diversity was assessed from morphological data obtained through transmission electron microscopy. Extracted soil virus communities were dominated by bacteriophages that demonstrated a wide range of capsid diameters (20 nm to 160 nm) and morphologies, including filamentous forms and phages with elongated capsids. The reciprocal Simpson's index suggests that forest soils harbor more diverse assemblages of viruses, particularly in terms of morphological distribution. Repeated extractions of virus-like particles (VLPs) from soils indicated that the initial round of extraction removes approximately 70% of extractable viruses. Higher VLP abundances were observed in forest soils (1.31 x 10(9) to 4.17 x 10(9) g(-1) dry weight) than in agricultural soils (8.7 x 10(8) to 1.1 x 10(9) g(-1) dry weight). Soil VLP abundance was significantly correlated to moisture content (r = 0.988) but not to soil texture. Land use (agricultural or forested) was significantly correlated to both bacterial (r = 0.885) and viral (r = 0.812) abundances, as were soil organic matter and water content. Thus, land use is a significant factor influencing viral abundance and diversity in soils.

Agriculture↗

Analyses of microbial community within a composter operated using household garbage with special reference to the addition of soybean oil.

A commercially available composter was operated using fixed composition of garbage with or without the addition of soybean oil. The composter was operated without adding seed microorganisms or bulking materials. Microflora within the composter were analyzed by denaturing gradient gel electrophoresis (DGGE) in the case of oil addition, or by 16/18 S rRNA gene sequencing of the isolated microorganisms in the case of no oil addition. The results showed that, irrespective of the addition of oil, the bacteria identified were all gram positive, and that lactobacilli seemed to be the key microorganisms. Based on the results, suitable microflora for use in a household composter are discussed.

Journal Article↗

Testing for differentiation of microbial communities using phylogenetic methods: accounting for uncertainty of phylogenetic inference and character state mapping.

Comparative analyses of microbial communities increasingly involve the assay of 16S rRNA (or other gene) sequences from environmental DNA. Determining whether the composition of two or more communities differ in their phylogenetic composition involves testing for covariation between phylogeny and community type. This approach requires estimating the phylogenetic relationships among all sampled sequences and assessing whether the distribution of sequences among communities differs from the null expectation that sequences are randomly distributed. One method developed for implementing the phylogeny-based test of differentiation, referred to as the Phylogenetic test, relies on a single estimate of the phylogeny. However, for most data sets, many alternative phylogenetic trees provide statistically equivalent descriptions of the data. Because the actual phylogeny is unknown, phylogenetic tests of differentiation among microbial communities must account for phylogenetic uncertainty. In this article, we evaluate bootstrapping and Bayesian phylogenetic methods when implementing the Phylogenetic test using parsimony to map character states, and we investigate the effects of character mapping uncertainty by using a Bayesian approach to stochastically map character states on trees. Our approaches incorporate uncertainty into the tests of two closely related null hypotheses: (1) populations are panmictic, and (2) identical communities existed in both environments over the course of evolutionary history. We use two data sets previously implemented in tests for community differentiation: nitrite reductase genes sampled from marsh and upland soils and 16S rDNA sequences sampled from the human mouth and gut. We show that accounting for phylogenetic and mapping uncertainties can drastically affect results when implementing the Phylogenetic test. Accounting for phylogenetic and character mapping uncertainty provides a more conservative and robust test of covariation between phylogeny and environment when comparing microbial communities using DNA sequences.

Bacteria↗

[The classification and the monitoring of the state of mouth riverine and lacustrine ecosystems in lake Baikal based on the composition of local microbiocenoses and their activity].

The paper presents the results of the long-term investigation of microbial communities in the technogenically vulnerable mouth riverine and lacustrine ecosystems of Lake Baikal. The structural and functional parameters of the microbial communities were analyzed from the standpoint of developing destructive processes. The analysis showed that the total number of microorganisms (TNM), the number of saprophytic bacteria (NSB), and bacterial production (BP) were greater in the river-mouth water than in the near-mouth lake water. In the offshore direction, TNM and NSB decreased by a factor of 1.5 to 2, and BP decreased by a factor of 4 to 7. Based on TNM, NSB, and BP data, we classified the Lake Baikal rivers with respect to the degree of the impact of human activities on them. The degrading capability of the riverine microbial communities was found to be such that they degrade daily from tenths of a percent to 3.5% of the total amount of organic compounds polluted the river waters.

Bacteria↗

Plants actively control nitrogen cycling: uncorking the microbial bottleneck.

Ecologists have tried to link plant species composition and ecosystem properties since the inception of the ecosystem concept in ecology. Many have observed that biological communities could feed back to, and not simply result from, soil properties. But which group of organisms, plants or microorganisms, drive those feedback systems? Recent research asserts that soil microorganisms preclude plant species feedback to soil nitrogen (N) transformations due to strong microbial control of soil N cycling. It has been well documented that litter properties influence soil N cycling. In this review, we stress that under many circumstances plant species exert a major influence over soil N cycling rates via unique N attainment strategies, thus influencing soil N availability and their own fitness. We offer two testable mechanisms by which plants impart active control on the N cycle and thereby allow for plant-litter-soil-plant feedback. Finally, we describe the characteristics of plants and ecosystems that are most likely to exhibit feedback.

Ecosystem↗

Experimental manipulations of microbial food web interactions in a humic lake: shifting biological drivers of bacterial community structure.

A previous multiyear study observed correlations between bacterioplankton community composition (BCC) and abundance and the dynamics of phytoplankton populations and bacterivorous grazers in a humic lake. These observations generated hypotheses about the importance of trophic interactions (both top-down and bottom-up) for structuring bacterial communities in this lake, which were tested using two multifactorial food web manipulation experiments that separately manipulated the intensity of grazing and the composition of the phytoplankton community. Our results, combined with field observations, suggest that a hierarchy of drivers structures bacterial communities in this lake. While other studies have noted links between aggregate measures of phytoplankton and bacterioplankton communities, we demonstrate here correlations between succession of phytoplankton assemblages and BCC as assessed by automated ribosomal intergenic spacer analysis (ARISA). We used a novel approach linking community ARISA data to phylogenetic assignments from sequence analysis of 16S rRNA gene clone libraries to examine the responses of specific bacterial phylotypes to the experimental manipulations. The synchronous dynamics of these populations suggests that primary producers may mediate BCC and diversity through labile organic matter production, which evolves in quality and quantity during phytoplankton succession. Superimposed on this resource-mediated control of BCC are brief periods of intense bacterivory that impact bacterial abundance and composition.

Animals↗

Biotic disturbance, recolonization, and early succession of bacterial assemblages in intertidal sediments.

The role of disturbance in structuring natural microbial communities has been largely unexplored. Disturbance associated with invertebrate ingestion can reduce bacterial biomass and alter metabolic activities and compositions of bacterial assemblages in marine sediments. The primary objectives of the research presented here were to test whether ingestion by a taxonomically diverse group of deposit feeders constituted a disturbance, and to determine the mechanisms by which bacterial assemblages recover following deposit-feeder ingestion. To test the question of disturbance, we compared fresh egesta vs surficial sediments with respect to bacterial assemblage structure. In emersed intertidal sediments, microbial recovery could be due to regrowth of bacterial populations surviving gut passage or to immigration from adjacent sediments. To differentiate between these modes of recolonization we used field manipulative experiments to exclude migration by isolating freshly extruded fecal coils of three deposit-feeding species from surrounding sediments. We then followed the quantitative and qualitative recovery in egesta and sediments through time using epifluorescence microscopy and PCR-DGGE analysis of 16S rDNA. Our findings indicate that (1) the degree and nature of the disturbance to bacterial assemblages from deposit feeding varies among invertebrate taxa, (2) recovery was significant but incomplete over 3 h, and (3) recolonization of biotically disturbed sediments is dominated by immigration.

Analysis of Variance↗

Minimizing decompression and warming during deep seawater collection increases abundance and activity of autochthonous bacteria and archaea.

The deep ocean hosts autochthonous pressure-adapted microorganisms that are unique to this environment, as well as allochthonous pressure-sensitive members transported from shallow depths by vertical advection and particle-sinking. However, conventional sampling instruments decompress and warm deep-sea samples during retrieval, potentially altering microbial properties when studied ex situ. Here, we assess this potential sampling bias by comparing seawater microbial communities collected with or without measures aimed at minimizing pressure and temperature effects. When compared to samples collected under pressurized conditions, conventional sampling (using Niskin bottles) was found to affect prokaryotic cells retrieved by reducing their total numbers, diminishing protein synthesis activity (>10%), and also causing overall shifts in the community composition. The most significant compositional change was a >20% decrease in metagenomic archaeal representation (TACK-group/Thaumarchaeota/Nitrososphaerota). Deep-sea bacterial groups had mixed responses to preserving pressure during retrieval, with some groups exhibiting higher representation when samples were maintained pressurized (e.g. members of the family Pelagibacteraceae, unclassified Thiotricales, Thioglobaceae, and Chitinophagaceae), whereas others increased their representation when decompressed (e.g. Burkholderiaceae, Comamonadaceae, and Oxalobacteraceae). This study reveals the existence of bias introduced by the complete decompression of samples retrieved with traditional instrumentation, as well as a decrease in overall bacterial activity when samples are completely decompressed during retrieval. Additionally, incubations lasting for >24 h were shown to transform the original prokaryotic community composition. Precautions addressing these effects are necessary to enhance the reliability of ex situ measurements and improve our understanding of deep-sea microbial ecology and biogeochemistry.

Seawater↗

Succession of aquatic microbial communities as a result of the water quality variations in continuous water.

The changes of structural and functional parameters of aquatic microbial communities in continuous water on campus of Tsinghua University, China are investigated, by polyurethane foam unit (PFU) method. The measured compositions of the communities include alga, protozoa, and some metazoa (such as rotifers). The measured indicators of water quality include water temperature, pH value, dissolved oxygen (DO), potassium permanganate index (COD(Mn)), total nitrogen (TN), total phosphorus (TP) and chlorophyll-a (Chla). The trophic level, expressed by the trophic level indices (TLIc), is assessed with analytic hierarchy process and principal component analysis (AHP-PCA) method. The changing trends of the structural and functional parameters of aquatic microbial communities, such as Margalef index of diversity (D), Shannon-weaver index of diversity (H), Heterotropy index (HI), number of species when the colonization gets equilibrium (S(eq)), colonizing speed constant (G) and time spent when 90 percent of S(eq) colonized in PFU (T(80%)), are also analyzed. The experimental results showed the succession of aquatic microbial communities along the water flow is consistent with the water quality changes, so the parameters of microbial community can reflect the changes of water quality from the ecological view.

Animals↗

Community structure and diversity of biofilms from a beer bottling plant as revealed using 16S rRNA gene clone libraries.

The microbial composition of biofilms from a beer bottling plant was analyzed by a cultivation independent analysis of the 16S rRNA genes. Clone libraries were differentiated by amplified 16S rRNA gene restriction analysis and representative clones from each group were sequenced. The diversity of the clone libraries was comparable with the diversity found for environmental samples. No evidences for the presence of strictly anaerobic taxa or important beer spoilers were found, indicating that biofilms developed for more than 6 months at the plant formed no appropriate habitat for those microorganisms. The genus Methylobacterium was one of the dominating groups of the clone libraries. The size of this population was assessed by fluorescence in situ hybridization and fatty acid analysis. In addition, considerable numbers of clones were assigned to uncultivated organisms.

Bacteria↗

Genomic analysis of bacteriophage PhiJL001: insights into its interaction with a sponge-associated alpha-proteobacterium.

Bacteriophage PhiJL001 infects a novel marine bacterium in the alpha subclass of the Proteobacteria isolated from the marine sponge Ircinia strobilina. PhiJL001 is a siphovirus and forms turbid plaques on its host. The genome sequence of PhiJL001 was determined in order to better understand the interaction between the marine phage and its sponge-associated host bacterium. The complete genome sequence of PhiJL001 comprised 63,469 bp with an overall G+C content of 62%. The genome has 91 predicted open reading frames (ORFs), and 17 ORFs have been assigned putative functions. PhiJL001 appears to be a temperate phage, and the integrase gene was identified in the genome. DNA hybridization analysis showed that the PhiJL001 genome does not integrate into the host chromosome under the conditions tested. DNA hybridization experiments therefore suggested that PhiJL001 has some pseudolysogenic characteristics. The genome of PhiJL001 contains many putative genes involved in phage DNA replication (e.g., helicase, DNA polymerase, and thymidylate synthase genes) and also contains a putative integrase gene associated with the lysogenic cycle. Phylogeny based on DNA polymerase gene sequences indicates that PhiJL001 is related to a group of siphoviruses that infect mycobacteria. Designation of PhiJL001 as a siphovirus is consistent with the morphology of the phage visualized by transmission electron microscopy. The unique marine phage-host system described here provides a model system for studying the role of phages in sponge microbial communities.

Alphaproteobacteria↗

Effects on the structure of Arctic ecosystems in the short- and long-term perspectives.

Species individualistic responses to warming and increased UV-B radiation are moderated by the responses of neighbors within communities, and trophic interactions within ecosystems. All of these responses lead to changes in ecosystem structure. Experimental manipulation of environmental factors expected to change at high latitudes showed that summer warming of tundra vegetation has generally led to smaller changes than fertilizer addition. Some of the factors manipulated have strong effects on the structure of Arctic ecosystems but the effects vary regionally, with the greatest response of plant and invertebrate communities being observed at the coldest locations. Arctic invertebrate communities are very likely to respond rapidly to warming whereas microbial biomass and nutrient stocks are more stable. Experimentally enhanced UV-B radiation altered the community composition of gram-negative bacteria and fungi, but not that of plants. Increased plant productivity due to warmer summers may dominate food-web dynamics. Trophic interactions of tundra and sub-Arctic forest plant-based food webs are centered on a few dominant animal species which often have cyclic population fluctuations that lead to extremely high peak abundances in some years. Population cycles of small rodents and insect defoliators such as the autumn moth affect the structure and diversity of tundra and forest-tundra vegetation and the viability of a number of specialist predators and parasites. Ice crusting in warmer winters is likely to reduce the accessibility of plant food to lemmings, while deep snow may protect them from snow-surface predators. In Fennoscandia, there is evidence already for a pronounced shift in small rodent community structure and dynamics that have resulted in a decline of predators that specialize in feeding on small rodents. Climate is also likely to alter the role of insect pests in the birch forest system: warmer winters may increase survival of eggs and expand the range of the insects. Insects that harass reindeer in the summer are also likely to become more widespread, abundant and active during warmer summers while refuges for reindeer/caribou on glaciers and late snow patches will probably disappear.

Animals↗

Desulfotomaculum and Methanobacterium spp. dominate a 4- to 5-kilometer-deep fault.

Alkaline, sulfidic, 54 to 60 degrees C, 4 to 53 million-year-old meteoric water emanating from a borehole intersecting quartzite-hosted fractures >3.3 km beneath the surface supported a microbial community dominated by a bacterial species affiliated with Desulfotomaculum spp. and an archaeal species related to Methanobacterium spp. The geochemical homogeneity over the 650-m length of the borehole, the lack of dividing cells, and the absence of these microorganisms in mine service water support an indigenous origin for the microbial community. The coexistence of these two microorganisms is consistent with a limiting flux of inorganic carbon and SO4(2-) in the presence of high pH, high concentrations of H2 and CH4, and minimal free energy for autotrophic methanogenesis. Sulfide isotopic compositions were highly enriched, consistent with microbial SO4(2-) reduction under hydrologic isolation. An analogous microbial couple and similar abiogenic gas chemistry have been reported recently for hydrothermal carbonate vents of the Lost City near the Mid-Atlantic Ridge (D. S. Kelly et al., Science 307:1428-1434, 2005), suggesting that these features may be common to deep subsurface habitats (continental and marine) bearing this geochemical signature. The geochemical setting and microbial communities described here are notably different from microbial ecosystems reported for shallower continental subsurface environments.

Desulfotomaculum↗

A statistical procedure for the analysis of microbial communities based on phenotypic properties of isolates.

A novel statistical procedure for the analysis of microbial communities based on phenotypic properties of randomly collected isolates is presented and discussed. The procedure allows the representation of the microbial communities as a set of ellipses in a bidimensional graph. This representation is obtained by the following steps: (a) measurement of a set of binary phenotypic properties for n isolates belonging to k samples, each representing a different community; (b) repeated sampling by bootstrapping of the m samples, thus obtaining, for each community, i subsamples of j isolates; (c) calculation of the frequency of positive results for each test for each subsample; (d) calculation of the matrix of Euclidean distances between the k x i frequency vectors; (e) use of multidimensional scaling (MDS) to obtain a representation in two dimensions of the distance relationships between the frequency vectors; (f) plotting of the 95% confidence ellipses for the i frequency vectors of each of the k communities. By using both simple, synthetic microbial communities, and samples of lactic acid bacteria isolated from natural microbial communities (sourdoughs, compressed yeast, fermented sausages), it was demonstrated that the position and shape of the ellipses are clearly related to the composition of the community, while the relationship between the size of the ellipses and the phenotypical diversity of the community is less straightforward: while communities with very different diversity (measured with the Functional Evenness index and the mean taxonomic distance) had ellipses that were very different in size, there was no strict proportionality between the size of the ellipse and the diversity of the community. Nevertheless, the representation of microbial communities obtained by bootstrapping and multidimensional scaling appears to be superior to the more usual representation based on tabulation of the frequencies of isolates belonging to different clusters.

Animals↗