Reliability for detecting composition and changes of microbial communities by T-RFLP genetic profiling

Terminal restriction fragment length polymorphism (T-RFLP) analysis is commonly used for profiling microbial communities in various environments. However, it may suffer from biases during the analytic process. This study addressed the potential of T-RFLP profiles (1) to reflect real community structures and diversities, as well as (2) to reliably detect changing components of microbial community structures. For this purpose, defined artificial communities of 30 SSU rRNA gene clones, derived from nine bacterial phyla, were used. PCR amplification efficiency was one primary bias with a maximum variability factor of 3.5 among clones. PCR downstream analyses such as enzymatic restriction and capillary electrophoresis introduced a maximum bias factor of 4 to terminal restriction fragment (T-RF) signal intensities, resulting in a total maximum bias factor of 14 in the final T-RFLP profiles. In addition, the quotient between amplification efficiency and T-RF size allowed predicting T-RF abundances in the profiles with high accuracy. Although these biases impaired detection of real community structures, the relative changes in structures and diversities were reliably reflected in the T-RFLP profiles. These data support the suitability of T-RFLP profiling for monitoring effects on microbial communities.     

Wastewater treatment by alkali bacteria and dynamics of microbial communities in two bioreactors

In this study, an alkali bacterial consortium was obtained by enrichment cultivation and was used to treat printing and dyeing wastewater (PDW, pH 11-12). The treatment effects and dynamic changes were evaluated in a biocontact oxidation reactor (BOR) and a sequencing batch reactor (SBR). During 3 months of continuous operation, the two bioreactors had similar treatment efficiencies (polyvinyl alcohol, PVA, 74.5-81.3%; COD, 73.5-77.4%; 2.15 pH decreases). Molecular biological analysis indicated that the microbial communities underwent dramatic changes during the operation, in which the SBR was superior to the BOR in retaining the alkali bacteria at the start-up stage, however, the BOR seemed to be more advantageous when the frequently changing influents were considered. The bacterial communities in BOR and SBR were diverse and included Bacteroidetes, Firmicutes, Proteobacteria, Actinobacteria and an unidentified cluster. Among these only Paracoccus sp. was successfully isolated and confirmed to have the ability to degrade PVA.     

Long-term cathode performance and the microbial communities that develop in microbial fuel cells fed different fermentation endproducts

To better understand how cathode performance and substrates affected communities that evolved in these reactors over long periods of time, microbial fuel cells were operated for more than 1 year with individual endproducts of lignocellulose fermentation (acetic acid, formic acid, lactic acid, succinic acid, or ethanol). Large variations in reactor performance were primarily due to the specific substrates, with power densities ranging from 835 ± 21 to 62 ± 1 mW/m³. Cathodes performance degraded over time, as shown by an increase in power of up to 26% when the cathode biofilm was removed, and 118% using new cathodes. Communities that developed on the anodes included exoelectrogenic families, such as Rhodobacteraceae, Geobacteraceae, and Peptococcaceae, with the Deltaproteobacteria dominating most reactors. Pelobacter propionicus was the predominant member in reactors fed acetic acid, and it was abundant in several other MFCs. These results provide valuable insights into the effects of long-term MFC operation on reactor performance.     

Change in microbial communities in acetate- and glucose-fed microbial fuel cells in the presence of light

Power densities produced by microbial fuel cells (MFCs) in natural systems are changed by exposure to light through the enrichment of photosynthetic microorganisms. When MFCs with brush anodes were exposed to light (4000 lx), power densities increased by 8–10% for glucose-fed reactors, and 34% for acetate-fed reactors. Denaturing gradient gel electrophoresis (DGGE) profiles based on the 16S rRNA gene showed that exposure to high light levels changed the microbial communities on the anodes. Based on 16S rRNA gene clone libraries of light-exposed systems the anode communities using glucose were also significantly different than those fed acetate. Dominant bacteria that are known exoelectrogens were identified in the anode biofilm, including a purple nonsulfur (PNS) photosynthetic bacterium, Rhodopseudomonas palustris, and a dissimilatory iron-reducing bacterium, Geobacter sulfurreducens. Pure culture tests confirmed that PNS photosynthetic bacteria increased power production when exposed to high light intensities (4000 lx). These results demonstrate that power production and community composition are affected by light conditions as well as electron donors in single-chamber air-cathode MFCs.     

Molecular biomass and MetaTaxogenomic assessment of soil microbial communities as influenced by soil DNA extraction procedure

Three soil DNA extraction procedures (homemade protocols and commercial kit) varying in their practicability were applied to contrasting soils to evaluate their efficiency in recovering: (i) soil DNA and (ii) bacterial diversity estimated by 16S rDNA pyrosequencing. Significant differences in DNA yield were systematically observed between tested procedures. For certain soils, 10 times more DNA was recovered with one protocol than with the others. About 15 000 sequences of 16S rDNA were obtained for each sample which were clustered to draw rarefaction curves. These curves, as well as the PCA ordination of community composition based on OTU clustering, did not reveal any significant difference between procedures. Nevertheless, significant differences between procedures were highlighted by the taxonomic identification of sequences obtained at the phylum to genus levels. Depending on the soil, differences in the number of genera detected ranged from 1% to 26% between the most and least efficient procedures, mainly due to a poorer capacity to recover populations belonging to Actinobacteria, Firmicutes or Crenarchaeota. This study enabled us to rank the relative efficiencies of protocols for their recovery of soil molecular microbial biomass and bacterial diversity and to help choosing an appropriate soil DNA extraction procedure adapted to novel sequencing technologies.     

Patterns and governing forces in aquatic microbial communities

In this review we survey recent publications employing molecular techniques to investigate the distribution of microbial species in aquatic environments. We analyzed the occurrence of microbial phyla in freshwater and marine habitats and observed patterns of distribution that could be explained by the adaptation of microorganisms to physical and biological parameters that vary in aquatic habitats. The gram-positive bacteria, the Verrucomicrobiales and the - and -subdivisions of the Proteobacteria are distributed throughout a range of aquatic habitats, while other phylogenetic groups appear to be adapted to more narrowly defined environmental niches such as anoxic water and sediments (-Proteobacteria) or floating aggregates (Cytophaga-Flexibacter-Bacteroides phylum). -proteobacterial sequence types have been detected throughout freshwater habitats, but these organisms are largely absent from open ocean environments. Within several of these divisions, clusters of closely related small sub unit ribosomal RNA sequence types have been detected in geographically disparate environments, suggesting that some microbial species are globally distributed. In addition to physical variables such as salinity and pH, biological variables also influence microbial community composition. This was illustrated by changes that occurred in the eukaryotic and bacterial species composition in laboratory mesocosms after a viral outburst. We conclude that physical and biological forces govern the composition of aquatic microbial communities and result in divergent evolutionary histories of the indigenous microbial species.     

Linking microbial community composition and soil processes in a California annual grassland and mixed-conifer forest

To investigate the potential role of microbial community composition in soil carbon and nitrogen cycling, we transplanted soil cores between a grassland and a conifer ecosystem in the Sierra Nevada California and measured soil process rates (N-mineralization, nitrous oxide and carbondioxide flux, nitrification potential), soil water and temperature, and microbial community parameters (PLFA and substrate utilization profiles) over a 2 year period. Our goal was to assess whether microbial community composition could be related to soil process rates independent of soil temperature and water content. We performed multiple regression analyses using microbial community parameters and soil water and temperature as X-variables and soil process rates and inorganic N concentrations as Y-variables. We found that field soil temperature had the strongest relationship with CO₂ production and soil NH₄⁺ concentration, while microbial community characteristics correlated with N₂O production, nitrification potential, gross N-mineralization, and soil NO₃^– concentration, independent of environmentalcontrollers. We observed a relationship between specific components of the microbial community (as determined by PLFA) and soil processes,particularly processes tightly linked to microbial phylogeny (e.g. nitrification). The most apparent change in microbial community composition in response to the 2 year transplant was a change in relative abundance of fungi (there was only one significant change in PLFA biomarkers for bacteria during 2 years). The relationship between microbial community composition and soil processes suggests that prediction of ecosystem response to environmental change may be improved by recognizing and accounting for changes in microbial community composition and physiological ecology.     

Molecular diversity of microbial community in acid mine drainages of Yunfu sulfide mine

Two acid mine drainage (AMD) samples were studied by a PCR-based cloning approach, which were from Yunfu sulfide mine in Guangdong province, China. A total of 15 operational taxonomic units (OTUs) were obtained from the two AMD samples. The percentage of overlapped OTUs in two AMD samples was 42.1%. Phylogenetic analysis revealed that the bacterium in the two samples fell into four putative divisions, which were Nitrospira, α-Proteobacteria, β-Proteobacteria, and γ-Proteobacteria four families. Organisms of genuses Acidithiobacillus and Gallionella, which were in γ-Proteobacteria family and β-Proteobacteria family, respectively, were dominant in two samples. The proportions of clones affiliated with Gallionella in each sample were 47.2% (G2) and 16.9% (G1). The result suggested that organisms of Gallionella were a very important composition in microbial communities of the two AMD samples we studied. In addition, the PCR amplification of archaeal 16S rDNA genes form these two AMD samples have been performed with two sets of archaea-specific primers, but no PCR product found. Zhiguo He, Shengmu Xiao, Xuehui Xie, and Hui Zhong equally contributed to this work.     

Methane emission and soil microbial communities in early rice paddy as influenced by urea-N fertilization

Plant and Soil - Moso bamboo (Phyllostachys edulis) invasions into adjacent forests are becoming increasingly common. Moso bamboo invasions affect litter quality, soil nutrients, and microbial...     

Structure and role of microbial communities in southern taiga soils

General regularities in the structure of the microbial communities of southern taiga soil ecosystems and taxonomic differences between the microbial communities of soils with different hydrothermal characteristics are discussed with reference to the main types of soils of the Central State Forest Biosphere Reserve.     

Comparison between biological and chemical treatment of wastewater containing nitrogen and phosphorus

The present work compared chemical and biological treatment methods to achieve the most efficient treatment for the reduction or elimination of phosphorus and nitrogen from mixed industrial–domestic wastewaters. Batch chemical precipitation by ferric chloride and aluminum sulfate (alum) and a continuous biological suspended growth system were investigated as well as the optimum operating conditions. Concerning chemical treatment, Alum generally achieved a higher removal efficiency percentage for the investigated pollutants compared with FeCl₃ at their optimum pH and dose, especially with chemical oxygen demand (COD). FeCl₃ treatment achieved success only with phosphorus removal, while none of the COD, 5-day biochemical oxygen demand (BOD₅), total nitrogen (TN) and N–NH₃ achieved acceptable treatment and remained above the maximum permissible limits (MPL). Thus, for such wastewaters, alum is more efficient than FeCl₃. Biological treatment exhibited higher efficiencies, particularly towards nitrogen. TN removal increased by increasing the flow rate to 30–60 l/day. N–NH₃ removal was effective at the slowest flow rate and decreased with increasing flow rate, while an opposite trend was recorded for N–NO₃. At all flow rates, phosphorus levels were below the accepted MPL for discharging into natural systems. Moreover, there was a general trend for the proposed biological treatment to achieve a high removal efficiency for BOD₅ and COD, bringing them to acceptable levels to be released into watercourses safely, especially at the slowest flow rates. Thus, integration between the proposed chemical and biological treatment is highly recommended, producing high-quality effluents acceptable by the environmental law.     

Microbial communities in wetlands of the Athabasca oil sands: genetic and metabolic characterization

Naphthenic acids are a complex family of naturally occurring cyclic and acyclic carboxylic acids that are present in the acidic fraction of petroleum. Naphthenic acids are acutely toxic to aquatic organisms. Previous studies showed that wetland sediments exposed to oil sands process water containing naphthenic acids had higher rates of naphthenic acid degradation in vitro compared with unexposed wetlands. In this study we compare the microbial community structures in sediments from wetlands exposed to different amounts of oil sands process water using BIOLOG, phospholipid fatty acid analysis and denaturing gradient gel electrophoresis of total bacterial DNA. Community profiles were compared using cluster analysis. BIOLOG profiles were primarily influenced by seasonal trends rather than naphthenic acids content. In contrast, phospholipid fatty acid analysis comparisons clustered communities that had higher levels of residual oil, although this association was not strong. In contrast, cluster diagrams produced from the denaturing gradient gel electrophoresis data clearly separated bacterial communities according to naphthenic acids concentrations, indicating that naphthenic acids content was a major influence on the composition of the bacterial community. In addition, denaturing gradient gel electrophoresis profiles indicated that naphthenic acids-exposed bacterial communities were homogeneous on a scale of meters, whereas unexposed (off-site) wetlands were less homogeneous.     

Field and microcosm experiments to evaluate the effects of agricultural Cu treatment on the density and genetic structure of microbial communities in two different soils

The effects of Cu amendment on indigenous soil microorganisms were investigated in two soils, a calcareous silty clay (Ep) and a sandy soil (Au), by means of a 1-year field experiment and a two-month microcosm incubation. Cu was added as 'Bordeaux mixture' [CuSO(4), Ca(OH)(2)] at the standard rate used in viticulture (B1=16 kg Cu kg(-1) soil) and at a higher level of contamination (B3=48 kg Cu ha(-1) soil). More extractable Cu was observed in sandy soil (Au) than in silty soil (Ep). Furthermore, total Cu and Cu-EDTA declined with time in Au soil, whereas they remained stable in Ep soil. Quantitative modifications of the microflora were assessed by C-biomass measurements and qualitative modifications were assessed by the characterization of the genetic structure of bacterial and fungal communities from DNA directly extracted from the soil, using B- and F-ARISA (bacterial and fungal automated ribosomal intergenic spacer analysis). In the field study, no significant modifications were observed in C-biomass whereas microcosm incubation showed a decrease in B3 contamination only. ARISA fingerprinting showed slight but significant modifications of bacterial and fungal communities in field and microcosm incubation. These modifications were transient in all cases, suggesting a short-term effect of Cu stress. Microcosm experiments detected the microbial community modifications with greater precision in the short-term, while field experiments showed that the biological effects of Cu contamination may be overcome or hidden by pedo-climatic variations.     

用微型生物群落评价常德市水系的研究

本文应用PFU法对常德水系四季微型生物群落的结构和功能进行了分析。根据MacArthur-Wilson平衡模型提出了3个功能参数,测定了微型生物的2个结构参数。同步进行了16个站四季的水质分析,提出化学综合污染指数(P)。经统计学处理,P和微型生物群落的HI、S_(eq)、G、T_(90％)等指数均呈显著性相关。物种多样性(d)也在一定程度上反映出城市水体的不同污染程度。再次证明PFU微型生物监测方法的科学性,为生物监测提供了快速、经济、正确的新方法。     

Diversity and assembly patterns of activated sludge microbial communities: A review

Understanding diversity and assembly patterns of microbial communities in activated sludge (AS) is pivotal for addressing fundamental ecological questions and wastewater treatment engineering. Recent applications of molecular methods especially high-throughput sequencing (HTS) have led to the explosion of information about AS community diversity, including the identification of uncultured taxa, and characterization of low-abundance but environmentally important populations such as antibiotic resistant bacteria and pathogens. Those progresses have facilitated the leverage of ecological theories in describing AS community assembly. The lognormal species abundance curve has been applied to estimate AS microbial richness. Taxa-area and taxa-time relationships (TAR and TTR) have been observed for AS microbial communities. Core AS microbial communities have been identified. Meanwhile, the roles of both deterministic and stochastic processes in shaping AS community structures have been examined. Nonetheless, it remains challenging to define tempo-spatial scales for reliable identification of community turnover, and find tight links between AS microbial structure and wastewater treatment plant (WWTP) functions. To solve those issues, we expect that future research will focus on identifying active functional populations in AS using omics- methods integrated with stable-isotope probing (SIP) with the development of bioinformatics tools. Developing mathematic models to understand AS community structures and utilize information on AS community to predict the performance of WWTPs will also be vital for advancing knowledge of AS microbial ecology and environmental engineering.     

Application of biocathode in microbial fuel cells: cell performance and microbial community

Instead of the utilization of artificial redox mediators or other catalysts, a biocathode has been applied in a two-chamber microbial fuel cell in this study, and the cell performance and microbial community were analyzed. After a 2-month startup, the microorganisms of each compartment in microbial fuel cell were well developed, and the output of microbial fuel cell increased and became stable gradually, in terms of electricity generation. At 20 ml/min flow rate of the cathodic influent, the maximum power density reached 19.53 W/m3, while the corresponding current and cell voltage were 15.36 mA and 223 mV at an external resistor of 14.9 Omega, respectively. With the development of microorganisms in both compartments, the internal resistance decreased from initial 40.2 to 14.0 Omega, too. Microbial community analysis demonstrated that five major groups of the clones were categorized among those 26 clone types derived from the cathode microorganisms. Betaproteobacteria was the most abundant division with 50.0% (37 of 74) of the sequenced clones in the cathode compartment, followed by 21.6% (16 of 74) Bacteroidetes, 9.5% (7 of 74) Alphaproteobacteria, 8.1% (6 of 74) Chlorobi, 4.1% (3 of 74) Deltaproteobacteria, 4.1% (3 of 74) Actinobacteria, and 2.6% (2 of 74) Gammaproteobacteria.     

Water pollution control by aquatic vegetation of treatment wetlands

Supplying polluted river water to nature reserves in The Netherlands often leads to eutrophication of the reserve. The eutrophication can be caused directly by the high nutrient input (external eutrophication) or indirectly by altering nutrient availability due to changes in nutrient desorption or mineralization. This paper investigates the potential of a ditch system that is tested for its potential to improve the water quality of polluted river water prior to supplying to the wet meadow reserve De Meije in The Netherlands. Concentrations of the macro-ions chloride, sulphate, calcium and bicarbonate in the polluted river water were much higher than original background values, measured in the reserve. During transport of the river water through the ditch system, no decline was observed in the concentrations of these macro-ions. The phosphorus concentration, however, decreased along the flow path and was significantly negatively correlated with the distance from the inlet point. High phosphorus removal occurred in a stretch of the ditch system where submerged and free floating species such as Fontinalis antipyretica and Lemna trisulca were dominant. The N: P ratio of F. antipyretica was especially low (N : P < 5) at sampling stations where high phosphorus concentrations were measured. The high N: P ratio indicated a luxury consumption of phosphorus. With decreasing phosphorus concentrations, the N: P ratio of F. antipyretica increased to a maximum of N: P = 25. The nutrient budget of the ditch system showed that supply of river water was the main input of phosphorus (12 kg P) whereas the main inputs of nitrogen of the ditch system were atmospheric deposition (66 kg N) and leaching from the wet meadows (44 kg N). For both nutrients, harvesting the aquatic vegetation in September was the main removal mechanism from the ditch system with 92 kg of nitrogen (80% of the annual input N) and 14 kg of phosphorus (95% of the annual P input) removed. It was concluded that the ditch system with aquatic vegetation could successfully remove nutrients from polluted river water. The concentrations of macro-ions, however, are not influenced by the ditch systems and internal eutrophication due to changes in adsorption or mineralization may still occur.     

Balance sheet of soil phosphorus and potassium as influenced by intensive cropping and fertilizer use

Summary The balance sheet of phosphorus and potassium were worked out from a long term manure and fertilizer experiment conducted for eight years and is still continuing at Ranchi Agricultural College, Kanke, Ranchi, India. Increasing levels of fertilizer combination with organic manure and lime give the highest yield as well as removed the highest amount of phosphorus and potassium from the soil and gave the positive gain of soil phosphorus and potassium in intensive cropping. The highest gains of 59.0 and 278.0 kg/ha of phosphorus and potassium respectively were recorded in 150% NPK application.