Dynamic changes of microbial community diversity in a photohydrogen producing reactor monitored by PCR-DGGE

A PCR-DGGE (denaturing gradient gel electrophoresis of polymerase chain reaction) protocol was used for monitoring the dynamic changes in the microbial population during photohydrogen production. Total DNA was extracted directly from the mixed bacterial community in the reactor and subjected to PCR with V3-16S rDNA and pufM gene primers, and the amplifications were then analyzed by DGGE. The DGGE patterns demonstrated the dynamics of community structure and the shift of microbial diversity, which correspond...     

Denaturing gradient gel electrophoresis fingerprinting of soil bacteria in the vicinity of the Chinese Great Wall Station, King George Island, Antarctica

Bacterial diversity was investigated in soil samples collected from 13 sites around the Great Wall Station, Fildes Peninsula, King George Island, Antarctica, using denaturing gradient gel electrophoresis (DGGE) of 16S rRNA genes. The classes α-, β-, and γ-Proteobacteria, as well as the phylum Actinobacteria, were found to be the dominant bacteria in the soils around the Great Wall Station. Although the selected samples were not contaminated by oil, a relationship between soil parameters, microbial biodiversity, and human impact was still seen. Sample sites in human impacted areas showed lower bacterial biodiversity (average H′= 2.65) when compared to nonimpacted sites (average H′= 3.05). There was no statistically significant correlation between soil bacterial diversity and total organic carbon (TOC), total nitrogen, or total phosphorus contents of the soil. Canonical correlation analysis showed that TOC content was the most important factor determining bacterial community profiles among the measured soil parameters. In conclusion, microbial biodiversity and community characteristics within relatively small scales (1.5 km) were determined as a function of local environment parameters and anthropogenic impact.     

Temporal and spatial changes of microbial community in an industrial effluent receiving area in Hangzhou Bay

Anthropogenic activities usually contaminate water environments, and have led to the eutrophication of many estuaries and shifts in microbial communities. In this study, the temporal and spatial changes of the microbial community in an industrial effluent receiving area in Hangzhou Bay were investigated by 454 pyrosequencing. The bacterial community showed higher richness and biodiversity than the archaeal community in all sediments. Proteobacteria dominated in the bacterial communities of all the samples; Marine_Group_Ⅰ and Methanomicrobia were the two dominant archaeal classes in the effluent receiving area. PCoA and AMOVA revealed strong seasonal but minor spatial changes in both bacterial and archaeal communities in the sediments. The seasonal changes of the bacterial community were less significant than those of the archaeal community, which mainly consisted of fluctuations in abundance of a large proportion of longstanding species rather than the appearance and disappearance of major archaeal species. Temperature was found to positively correlate with the dominant bacteria, Betaproteobacteria, and negatively correlate with the dominant archaea,Marine_Group_Ⅰ; and might be the primary driving force for the seasonal variation of the microbial community.     

Monitoring impact of mefenacet treatment on soil microbial communities by PCR-DGGE fingerprinting and conventional testing procedures

The effect of acetanilide herbicide mefenacet on soil microbial communities was studied using paddy soil samples with different short-term treatments. The culturable bacteria （plate counts）, dehydrogenase activity and changes in community structure （denaturing gradient gel electrophoresis （DGGE） analysis） were used for biological community assessments. Mefenacet was a significant stimulus to cultural aerobic bacteria and dehydrogenase activity while Sphingobacterium multivorum Y1, a bacterium efficiently degrading the mefenacet, only induced the increasing colony-forming unit （CFU） of bacteria but little effect on dehydrogenase activity during the whole experiment. The degree of similarity between the 16S rDNA profiles of the communities was quantified by numerically analyzing the DGGE band patterns. Similarity dendrograms showed that the microbial community structures of the mefenacet-treated and non-treated soils were not significantly different. But supplement of S. multivorum Y1 could increase the diversity of the microbial community in the mefenacet-polluted paddy soil. This work is a new attempt to apply the S. multivorum Y1 for remediation of the mefenacet-polluted environments.     

Effects of Cd（Ⅱ） and Cu（Ⅱ） on microbial characteristics in 2-chlorophenol-degradation anaerobic bioreactors

The effects of Cd^2＋ and Cu^2＋ at 300 mg/L on anaerobic microbial communities that degrade 2-cholorophenol （2-CP） were examined. Based on the polymerase chain reaction （PCR） of 16S rDNA, bacterial community diversity and archaeal community structure were analyzed with denaturing gradient gel electrophoresis （DGGE） and cloning, respectively. Degradation capabilities of the anaerobic microbial community were drastically abated and the degradation efficiency of 2-CP was reduced to 60% after shock by Cu^2＋ and Cd^2＋, respectively. The bacterial community structure was disturbed and the biodiversity was reduced after shock by Cu^2＋ and Cd^2＋ for 3 d. Some new metal-resistant microbes which could cope with the new condition appeared. The sequence analysis showed that there existed common Archaea species in control sludge and systems when treated with Cu^2＋ and Cd^2＋, such as Methanothrix soehngenii, Methanosaeta concilii, uncultured euryarchaeote, and so on. Both the abundance and diversity of archaeal species were altered with addition of Cd^2＋ and Cu^2＋ at high concentration. Although the abundance of the predominant archaeal species decreased with Cd^2＋ and Cu^2＋ addition for 3 d, they recovered to some extent after 10 d. The diversity of archaeal species was remarkably reduced after recovery for 10 d and the shift in archaeal composition seemed to be irreversible. The 2-CP-degradation anaerobic system was more sensitive to Cu^2＋ than Cd^2＋.     

Performance and microbial community of a membrane bioreactor system——Treating wastewater from ethanol fermentation of food waste

In this study, a lab-scale biological anaerobic/anaerobic/anoxic/membrane bioreactor(A_-~3MBR) was designed to treat wastewater from the ethanol fermentation of food waste,a promising way for the disposal of food waste and reclamation of resources. The 454 pyrosequencing technique was used to investigate the composition of the microbial community in the treatment system. The system yielded a stable effluent concentration of chemical oxygen demand(202 ± 23 mg/L), total nitrogen(62.1 ± 7.1 mg/L), ammonia(0.3 ±0.13 mg/L) and total phosphorus(8.3 ± 0.9 mg/L), and the reactors played different roles in specific pollutant removal. The exploration of the microbial community in the system revealed that:(1) the microbial diversity of anaerobic reactors A_1 and A_2, in which organic pollutants were massively degraded, was much higher than that in anoxic A_3 and aerobic MBR;(2) although the community composition in each reactor was quite different, bacteria assigned to the classes Clostridia, Bacteroidia, and Synergistia were important and common microorganisms for organic pollutant degradation in the anaerobic units, and bacteria from Alphaproteobacteria and Betaproteobacteria were the dominant microbial population in A_3 and MBR;(3) the taxon identification indicated that Arcobacter in the anaerobic reactors and Thauera in the anoxic reactor were two representative genera in the biological process. Our results proved that the biological A_-~3MBR process is an alternative technique for treating wastewater from food waste.     

A novel acidophile community populating waste ore deposits at an acid mine drainage site

Waste ore samples （pH 3.0） were collected at an acid mine drainage （AMD） site in Anhui, China. The present acidophillc microbial community in the waste ore was studied with 16S rRNA gene clone library and denaturing gradient gel electrophoresis （DGGE）. Eighteen different clones were identified and affiliated withActinobacteria, low G ＋ C Gram-positives, Thermomicrobia, Acidobacteria, Proteobacteria, candidate division TM7, and Planctomycetes. Phylogenetic analysis of 16S rRNA gene sequences revealed a diversity of acidophiles in the samples that were mostly novel. It is unexpected that the moderately thermophilic acidophiles were abundant in the acidic ecosystem and may play a great role in the generation of AMD. The result of DGGE was consistent with that of clone library analysis. These findings help in the better understanding of the generation mechanism of AMD and in developing a more efficient method to control AMD.     

Impacts of produced water origin on bacterial community structures of activated sludge

The purpose of this study was to reveal how activated sludge communities respond to influent quality and indigenous communities by treating two produced waters from different origins in a batch reactor in succession. The community shift and compositions were investigated using Polymerase Chain Reaction–denaturing gradient gel electrophoresis(PCR–DGGE) and further16 S ribosomal DNA(r DNA) clone library analysis. The abundance of targeted genes for polycyclic aromatic hydrocarbon(PAH) degradation, nah Ac/phn Ac and C12O/C23 O, was tracked to define the metabolic ability of the in situ microbial community by Most Probable Number(MPN) PCR. The biosystem performed almost the same for treatment of both produced waters in terms of removals of chemical oxygen demand(COD) and PAHs. Sludge communities were closely associated with the respective influent bacterial communities(similarity 60%), while one sludge clone library was dominated by the Betaproteobacteria(38%) and Bacteriodetes(30%)and the other was dominated by Gammaproteobacteria(52%). This suggested that different influent and water quality have an effect on sludge community compositions. In addition, the existence of catabolic genes in sludge was consistent with the potential for degradation of PAHs in the treatment of both produced waters.     

Bacterial diversity in soils around a lead and zinc mine

Five samples of soil collected from a lead and zinc mine were used to assess the effect of combined contamination of heavy metals on soil bacterial communities using a polyphasic approach including characterization of isolates by culture method, community level catabolic profiling in BIOLOG GN microplates, and genetic community fingerprinting by denaturing gradient gel electrophoresis of 16S rDNA fragments amplified by PCR from community DNA （PCR-DGGE）. The structure of the bacterial community was affected to a certain extent by heavy metals. The PCR-DGGE analysis of 16S rRNA genes showed that there were significant differences in the structure of the microbial community among the soil samples, which were related to the contamination levels. The number of bacteria and the number of denaturing gradient gel electrophoresis （DGGE） bands in the soils increased with increasing distance from the lead and zinc mine tailing, whereas the concentration of lead （Pb） and cadmium （Cd） was decreased. Heavily polluted soils could be characterized by a community that differs from those of lightly polluted soils in richness and structure of dominating bacterial populations. The clustering analysis of the DGGE profiles showed that the bacteria in all the five samples of soil belonged to three clusters. The data from the BIOLOG analysis also showed the same result. This study showed that heavy metal contamination decreased both the biomass and diversity of the bacterial community in soil.     

Assessing the impact of fungicide enostroburin application on bacterial community in wheat phyllosphere

Fungicides have been used extensively for controlling fungal pathogens of plants. However, little is known regarding the effects that fungicides upon the indigenous bacterial communities within the plant phyllosphere. The aims of this study were to assess the impact of fungicide enostroburin upon bacterial communities in wheat phyllosphere. Culture-independent methodologies of 16S rDNA clone library and 16S rDNA directed polymerase chain reaction with denaturing gradient gel electrophoresis (PCR-DGGE) were used for monitoring the change of bacterial community. The 16S rDNA clone library and PCR-DGGE analysis both confirmed the microbial community of wheat plant phyllosphere were predominantly of the γ-Proteobacteria phyla. Results from PCR-DGGE analysis indicated a significant change in bacterial community structure within the phyllosphere following fungicide enostroburin application. Bands sequenced within control cultures were predominantly of Pseudomonas genus, but those bands sequenced in the treated samples were predominantly strains of Pantoea genus and Pseudomonas genus. Of interest was the appearance of two DGGE bands following fungicide treatment, one of which had sequence similarities (98%) to Pantoea sp. which might be a competitor of plant pathogens. This study revealed the wheat phyllosphere bacterial community composition and a shift in the bacterial community following fungicide enostroburin application.     

Comparative study of microbial community structure in di erent filter media of constructed wetland

Comparisons of microbial community structure, in eight filter media of zeolites, anthracite, shale, vermiculite, ceramic filter media, gravel, steel slag and bio-ceramic, were undertaken by analyzing the phospholipid fatty acid (PLFA) composition. A total of 20 fatty acids in the range of C11 to C20 were determined but only 13 PLFAs were detected in steel slag. They consist of saturated fatty acids, branched fatty acids, monounsaturated fatty acids, and polyunsaturated fatty acids. The variation of fatty acids was revealed in the relative proportions of these fatty acids in di erent media. The aerobic prokaryotes were the predominant group in all media. The PLFA composition showed significant di erences among the eight di erent media by Tukey’s honestly test. It was found that steel slag was significantly di erent in the microbial community as compared to other filter media, probably due to its alkaline e uent. Steel slag alone is probably not a good choice of substratum in constructed wetlands. The principle components analysis (PCA) showed that zeolites, bio-ceramic, shale and vermiculite had a similar microbial community structure while steel slag and ceramic filter media were distinct from other media.     

Community analysis of ammonia and nitrite oxidizers in start-up of aerobic granular sludge reactor

A lab-scale sequencing batch reactor (SBR) was setup and the aerobic granular sludge was successfully incubated by using anaerobic granular sludge as seed sludge. Nitrogen was partially removed by simultaneous nitrification and denitrification (SND) via nitrite with free ammonia (FA) of about 10 mg/L. The denaturing gradient gel electrophoresis (DGGE) method was used to investigate community structure of α-Proteobacteria, β-Proteobacteria, ammonia oxidizing bacteria (AOB), and Nitrospira populations during start-up. The population sizes of bacteria, AOB and Nitrospira were examined using real-time PCR method. The analysis of community structure and Shannon index showed that stable structure of AOB population was obtained at the day 35, while the communities of α-Proteobacteria, β-Proteobacteria, and Nitrospira became stable after day 45. At stable stage, the average cell densities were 1.1×1012, 2.2×1010 and 1.0×1010 cells/L for bacteria, AOB and Nitrospira, respectively. The relationship between characteristics of nitrifying bacteria community and nitrogenous substrate utilization constant was discussed by calculating Pearson correlation. Certain correlation seemed to exist between population size, biodiversity, and degradation constant. And the influence of population size might be greater than that of biodiversity.     

Analysis of key microbial community during the start-up of anaerobic ammonium oxidation process with paddy soil as inoculated sludge

A sequencing batch reactor(SBR)-anaerobic ammonium oxidation(anammox) system was started up with the paddy soil as inoculated sludge. The key microbial community structure in the system along with the enrichment time was investigated by using molecular biology methods(e.g., high-throughput 16 S r RNA gene sequencing and quantitative PCR). Meanwhile,the influent and effluent water quality was continuously monitored during the whole start-up stage. The results showed that the microbial diversity decreased as the operation time initially and increased afterwards, and the microbial niches in the system were redistributed. The anammox bacterial community structure in the SBR-anammox system shifted during the enrichment, the most dominant anammox bacteria were Candidatus Jettenia. The maximum biomass of anammox bacteria achieved 1.68 × 10~9 copies/g dry sludge during the enrichment period, and the highest removal rate of TN achieved around 75%.     

Comparing results of cultured and uncultured biological methods used in biological phosphorus removal

Increasing attention has been paid to phosphate-accumulating organisms(PAOs)for their important role in biological phosphorus removal.In this study,microbial communities of PAOs cultivated under different carbon sources(sewage,glucose,and sodium acetate) were investigated and compared through culture-dependent and culture-independent methods,respectively.The results obtained using denaturing gradient gel electrophoresis(DGGE)of polymerase chain reaction-amplified 16S rDNA fragments revealed that the diversity of bacteria in a sewage-fed reactor(1#)was much higher than in a glucose-fed one(2#)and a sodium acetate-fed one(3#);there were common PAOs in three reactors fed by different carbon sources.Five strains were separated from three systems by using a phosphate- rich medium;they were from common bacteria isolated and three isolates could not be found in DGGE profile at all.Two isolates had good phosphorus removal ability.When the microbial diversity was studied,the molecular biological method was better than the culture-dependent one.When phosphorus removal characteristics were investigated,culture-dependent approach was more effective. Thus a combination of two methods is necessary to have a comprehensive view of PAOs.     

A review of diversity-stability relationship of soil microbial community: What do we not know?

The impact of decreased biodiversity on ecosystem stability, or the diversity-stability (D-S) relationship, is one of the major concerns of modern ecological studies. Studies on the D-S relationship for soil microbial communities began in 2000 when the fumigation method was developed to generate different levels of soil microbial biodiversity. The studies used various measures and levels of biodiversity, and covered several functional parameters. Due to the lack of general concepts and reliable approaches to define microbial species, studies on the D-S relationship of soil microbial communities concentrate on genetic diversity and functional diversity more than species diversity. Contradictory results were observed in various studies on D-S relationship with possible factors affecting or even changing the directions of the D-S relationship including: (1) the methods of stability measurement, (2) the techniques in microbial diversity measurement, (3) the measures and levels of diversity, (4) the type and strength of disturbance, (5) the traits of functions, and (6) the hidden treatments stemming from diversity manipulation. We argue that future studies should take diversity, species composition and interaction, and soil environmental conditions holistically into account in D-S studies to develop modeling to predict soil functional stability. We also suggest that studies should be carried out on a wider range of disturbance types and functional parameters, and efforts be shifted towards long-term field studies.     

Characterization of depth-related microbial communities in lake sediment by denaturing gradient gel electrophoresis of amplified 16S rRNA fragments

The characterization of microbial communities of different depth sediment samples was examined by a culture-independent method and compared with physicochemical parameters, those are organic matter （OM）, total nitrogen （TN）, total phosphorus （TP）, pH and redox potential （Eh）. Total genomic DNA was extracted from samples derived from different depths. After they were amplified with the GC-341 f/907r primer sets of partial bacterial 16S rRNA genes, the products were separated by denaturing gradient gel electrophoresis （DGGE）. The profile of DGGE fingerprints of different depth sediment samples revealed that the community structure remained relatively stable along the entire 45 cm sediment core, however, principal-component analysis of DGGE patterns revealed that at greater sediment depths, successional shifts in community structure were evident. The principle coordinates analysis suggested that the bacterial communities along the sediment core could be separated into two groups, which were located 0-20 cm and 21-45 cm, respectively. The sequencing dominant bands demonstrated that the major phylogenetic groups identified by DGGE belonged to Bacillus, Bacterium, Brevibacillus, Exiguobacterium, γ-Proteobacterium, Acinetobacter sp. and some uncultured or unidentified bacteria. The results indicated the existence of highly diverse bacterial community in the lake sediment core.     

Effect of seed sludge on characteristics and microbial community of aerobic granular sludge

Aerobic granular sludge was cultivated by using different kinds of seed sludge in sequencing batch airlift reactor.The influence of seed sludge on physical and chemical properties of granular sludge was studied;the microbial community structure was probed by using scanning electron microscope and polymerase chain reaction-denaturing gradient gel electrophoresis(PCR-DGGE).The results showed that seed sludge played an important role on the formation of aerobic granules.Seed sludge taken from beer wastewater treatment plant(inoculum A) was more suitable for cultivating aerobic granules than that of sludge from municipal wastewater treatment plant(inoculum B).Cultivated with inoculum A, large amount of mature granules formed after 35 days operation, its SVI reached 32.75 mL/g, and SOUR of granular sludge was beyond 1.10 mg/(g·min).By contrast, it needed 56 days obtaining mature granules using inoculum B.DGGE profiles indicated that the dominant microbial species in mature granules were 18 and 11 OTU when inoculum A and B were respectively employed as seed sludge.The sequencing results suggested that dominant species in mature granules cultivated by inoculum A were Paracoccus sp., Devosia hwasunensi, Pseudoxanthomonas sp., while the dominant species were Lactococcus raffinolactis and Pseudomonas sp.in granules developed from inoculum B.     

Plankton community composition in the Three Gorges Reservoir Region revealed by PCR-DGGE and its relationships with environmental factors

To explore the relationships between community composition and the environment in a reservoir ecosystem, plankton communities from the Three Gorges Reservoir Region were studied by PCR-denaturing gradient gel electrophoresis fingerprinting. Bacterial and eukaryotic operational taxonomic units （OTUs）, generated by DGGE analysis of the PCR-amplified 16S and 18S rRNA genes, were used as surrogates for the dominant ＂biodiversity units＂. OTU composition among the sites was heterogeneous; 46.7% of the total bacterial OTUs （45） and 64.1% of the eukaryotic OTUs （39） were identified in less than half of the sampling sites. Unweighted pair group method with arithmetic averages （UPGMA） clustering of the OTUs suggested that the plankton communities in the Xiangxi Rive sites were not always significantly different from those from the Yangtze River sites, despite clear differences in their environmental characterizations. Canonical correspondence analysis （CCA） was applied to further investigate the relationships between OTU composition and the environmental factors. The first two CCA ordination axes suggested that the bacterial community composition was primarily correlated with the variables of NO3^--N, dissolved oxygen （DO）, and SiO3^2--Si, whereas, the eukaryotic community was mainly correlated with the concentrations of DO, PO4^3--P, and SiO3^2--Si.     

Nutrient depletion is the main limiting factor in the crude oil bioaugmentation process

The biodegradation was considered as the prime mechanism of crude oil degradation. To validate the efficacy and survival of the crude oil-degrading strain in a bioremediation process, the enhanced green fluorescent protein gene (egfp) was introduced into Acinetobacter sp. HC8–3S. In this study, an oil-contaminated sediment microcosm was conducted to investigate the temporal dynamics of the physicochemical characterization and microbial community in response to bacterium amendment. The introduced strains were able to survive, flourish and degrade crude oil quickly in the early stage of the bioremediation. However, the high abundance cannot be maintained due to the ammonium (NH₄⁺-N) and phosphorus (PO₄^3?-P) contents decreased rapidly after 15 days of remediation. The sediment microbial community changed considerably and reached relatively stable after nutrient depletion. Therefore, the addition of crude oil and degrading cells did not show a long-time impact on the original microbial communities, and sufficient nitrogen and phosphorus nutrients ensures the survive and activity of degrader. Our studies expand the understanding of the crude oil degradative processes, which will help to develop more rational bioremediation strategies.     

Effects of tourmaline on nitrogen removal performance and biofilm structures in the sequencing batch biofilm reactor

The effects of tourmaline on nitrogen removal performance and biofilm structures were comparatively investigated in two identical laboratory-scale sequencing batch biofilm reactors(SBBRs)(denoted SBBR1 and SBBR2) at different nitrogen loading rates(NLRs) varying from(0.24 ± 0.01) to(1.26 ± 0.02) g N/(L·day). SBBR1 was operated in parallel with SBBR2, but SBBR1 was filled with polyurethane foam loaded tourmaline(TPU) carriers and another(SBBR2) filled with polyurethane foam(PU) carriers. Results obtained from this study showed that the excellent and stable performance of SBBR1 was obtained. Ammonia nitrogen removal and total nitrogen removal were higher in SBBR1 than that in SBBR2 with increase of NLR. At an NLR of(0.24 ± 0.01) g N/(L·day), the majority of the spherical and elliptical bacteria were surrounded by the extracellular polymeric substance(EPS) and bacillus or filamentous bacteria in two SBBRs biofilms. When NLR increased to(1.26 ± 0.02) g N/(L·day), the clusters were more obvious in the SBBR1 biofilm than that in the SBBR2 biofilm. Bacteria in SBBR1 were inclined to synthesis more EPS, and the formed EPS could protect the bacteria from free ammonia(FA) under extreme condition NLR(1.26 ± 0.02) g N/(L·day). The results of polymerase chain reaction-denaturing gradient gel electrophoresis analysis showed that the microbial community similarity in SBBR2 decreased more obviously than that in SBBR1 with the increase of NLR, which the microbial community in SBBR1 was relatively stable.