ABSTRACT Understanding processes and mechanisms governing microbial community structure and function is a central goal in microbial ecology. Previous studies disentangling the community assembly mechanisms were mainly based on taxonomic diversity but were rarely combined with species’ functional traits and interactions. Here, we showed how species’ functional traits and interactions determined microbial community structure and functions by a well-controlled laboratory experiment with nitrate-mediated sulfur oxidation systems using both culture-independent and culture-dependent technologies. The results showed that species were different in functional traits of nitrate-mediated sulfide and thiosulfate oxidation, which determined their relative abundance in the nitrate-mediated sulfur oxidation systems. Those thiosulfate-oxidizing microbes co-occurred with Thiobacillus by using intermediates (e.g., thiosulfate) secreted by Thiobacillus during sulfide oxidation process. Such metabolic dependencies exerted great effects on community functions. Metabolic dependencies between Thiobacillus and genera that oxidized thiosulfate to more sulfate (e.g., Ciceribacter) sustained high and stable oxidation activities of sulfide to sulfate. In contrast, metabolic dependencies between Thiobacillus and genera that oxidized thiosulfate to tetrathionate (e.g., Pseudoxanthomonas) slowed down the production of sulfate, indicating changes in the metabolic flow. In addition, competitions among species were mostly detrimental to the stability of community function. These results revealed that species’ functional traits and interactions were the intrinsic factors determining community structure and functions. This study advances our understanding of microbial community assembly and functions of the nitrate-mediated sulfur oxidation process from the perspectives of species’ functional traits and interactions and has important implications for designing and constructing microbiomes with expected functions. Importance Understanding the processes and mechanisms governing microbial community assembly and their linkages to ecosystem functioning has long been a core issue in microbial ecology. An in-depth insight still requires combining with analyses of species’ functional traits and microbial interactions. Our study showed how species’ functional traits and interactions determined microbial community structure and functions by a well-controlled laboratory experiment with nitrate-mediated sulfur oxidation systems using high-throughput sequencing and culture-dependent technologies. The results provided solid evidences that species’ functional traits and interactions were the intrinsic factors determining community structure and function. More importantly, our study established quantitative links between community structure and function based on species’ functional traits and interactions, which would have important implications for the design and synthesis of microbiomes with expected functions.

ABSTRACT The organic pollutant-degrading microorganisms with high cell surface hydrophobicity (CSH) are generally favorable due to the positive role of high CSH in pollutant capture and cell colonization. Sphingomonads, an important bacterial group with metabolic versatility, have significant potential for biodegradation and bioremediation of organic pollutants and generally harbor higher CSH than typical Gram-negative bacteria. However, the molecular mechanisms underlying their high CSH are still unclear. In this study, Sphingobium xenophagum C1, the most hydrophobic sphingomonad ever known, and its hydrophilic variant C2 were used to identify the genome variations responsible for the CSH difference by comparative genome and transcriptome analysis, as well as gene knockout verification. Our results indicated that the high CSH of strain C1 was largely attributed to the low copy number of the native plasmid p3, which mainly affected the transcriptional levels of outer membrane protein genes through direct and indirect means. In addition, loss of the genes on the native plasmid p5 involved in polysaccharide synthesis and secretion could increase CSH and cell surface friction. The bioinformatics analysis further revealed that some sphingomonad genomes also contained the long homologous fragments and/or key genes of p3 and p5. This study demonstrated the important role of native plasmids in regulating the CSH of sphingomonads, suggesting a novel CSH regulation strategy and evolution process. IMPORTANCE Microbial cell surface hydrophobicity (CSH) reflects nonspecific adhesion ability and affects various physiological processes, such as biofilm formation and pollutant biodegradation. Understanding the regulation mechanisms of CSH will contribute to illuminating microbial adaptation strategies and provide guidance for controlling CSH artificially to benefit humans. Sphingomonads, a common bacterial group with great xenobiotic-degrading ability, generally show higher CSH than typical Gram-negative bacteria, which plays a positive role in organic pollutant capture and cell colonization. This study verified that the variations of two native plasmids involved in synthesizing outer membrane proteins and polysaccharides greatly affected the CSH of sphingomonads. It is feasible to control their CSH by changing the plasmid copy number and sequences. Additionally, considering that plasmids are likely to evolve faster than chromosomes, the CSH of sphingomonads may evolve quickly to respond to environmental changes. Our results provide valuable insights into the CSH regulation and evolution of sphingomonads.

Microbial extracellular electron transfer (EET) is essential in many natural and engineering processes. Compared with the versatile EET pathways of Gram-negative bacteria, the EET of Gram-positive bacteria has been studied much less and is mainly limited to the flavin-mediated pathway. Here, we investigate the EET pathway of a Gram-positive filamentous bacterium Lysinibacillus varians GY32. Strain GY32 has a wide electron donor spectrum (including lactate, acetate, formate, and some amino acids) in electrode respiration. Transcriptomic, proteomic, and electrochemical analyses show that the electrode respiration of GY32 mainly depends on electron mediators, and c-type cytochromes may be involved in its respiration. Fluorescent sensor and electrochemical analyses demonstrate that strain GY32 can secrete cysteine and flavins. Cysteine added shortly after inoculation into microbial fuel cells accelerated EET, showing cysteine is a new endogenous electron mediator of Gram-positive bacteria, which provides novel information to understand the EET networks in natural environments. IMPORTANCE Extracellular electron transport (EET) is a key driving force in biogeochemical element cycles and microbial chemical-electrical-optical energy conversion on the Earth. Gram-positive bacteria are ubiquitous and even dominant in EET-enriched environments. However, attention and knowledge of their EET pathways are largely lacking. Gram-positive bacterium Lysinibacillus varians GY32 has extremely long cells (.1 mm) and conductive nanowires, promising a unique and enormous role in the microenvironments where it lives. Its capability to secrete cysteine renders it not only an EET pathway to respire and survive, but also an electrochemical strategy to connect and shape the ambient microbial community at a millimeter scale. Moreover, its incapability of using flavins as an electron mediator suggests that the common electron mediator is species-dependent. Therefore, our results are important to understanding the EET networks in natural and engineering processes.

Zero valent iron (ZVI)-microbe technology has an increasing application on the removal of organic pollution, yet the molecular mechanism of microbe respond to ZVI is still a mystery. Here, we established a successive ZVI-enhanced microbial system to remove azo dye (a typical organic pollutant) by Shewanella decolorationis S12 (S. decolorationis S12, an effective azo dye degradation bacterium) and examined the gene expression time course (10, 30, 60, and 120 min) by whole genome transcriptional analysis. The addition of ZVI to the microbial degradation system increases the rate of azo reduction from similar to 60% to over 99% in 16 h reaction, suggesting the synergistic effect of ZVI and S12 on azo dye degradation. Comparing with the treatment without ZVI, less filamentous cells were observed in ZVI treated system, and approximately 8% genes affiliated with 10 different gene expression profiles in S. decolorationis 512 were significantly changed in 120 min during the ZVI-enhanced azo reduction. Intriguingly, MarR transcriptional factor might play a vital role in regulating ZVI-enhanced azo reduction in the aspect of energy production, iron homeostasis, and detoxification. Further investigation showed that the induced [Ni-Fe] H(2)ase genes (hyaABCDEF) and azoreductase genes (mtrABC-omcA) contributed to ZVI-enhanced energy production, while the reduced iron uptake (hmuVCB and feoAB), induced sulfate assimilation (cysPTWA) and cysteine biosynthesis (cysM) related genes were essential to iron homeostasis and detoxification. This study disentangles underlying mechanisms of ZVI-enhanced organic pollution biotreatment in S. decolorationis S12.

A Gram stain-positive, non-spore-forming, non-motile and rod-shaped actinomycete, strain 5221(T), was isolated from the sediment of a river collected at Ronggui in the Pearl River Delta, PR China. Phylogenetic analysis based on 16S rRNA gene sequences revealed that the strain formed a distinct lineage within the genus Brevibacterium and had the highest sequence similarity to Brevibacterium pityocampae Tp12(T) (96.7%), followed by Brevibacterium daeguense 2C6-41(T) (96.5 %), Brevibacterium samyangense SST-8(T) (96.0 %) and Brevibacterium ravenspurgense 20(T) (95.9%). The results of chemotaxonomic analyses, including detecting anteiso-C-15:0, anteiso-C-17:0, and C-16:0 as the major cellular fatty acids, diphosphatidylglycerol, phosphatidylglycerol and three phosphoglycolipids as the polar lipids, MK-8(H-2) as the major menaquinone, and a DNA G+C content of 72.4 mol%, supported that strain 5221(T) is a member of the genus Brevibacterium. Furthermore, low sequence similarities of 16S rRNA gene sequences, differences in fatty acid compositions and differential physiological characteristics such as enzyme activity and carbon sources utilization ability distinguished the isolate from its close relatives. Therefore, strain 5221(T) represents a novel species of the genus Brevibacterium, for which the name Brevibacterium rongguiense sp. nov. is proposed, with the type strain 5221(T) (=GDMCC 1.1766(T)=KACC 21700(T)).

Bacteria promoters along with operators are crucial elements in the control of gene expression in microbes in response to environmental stress changes. A genome-wide promoter DNA regulatory library is in demand to be developed for a microbe reporter method to monitor the existence of any given environmental stress substance. In this study, we utilized Escherichia coli ( E. coli ) as a model system for the preparation of both cell lysates and genomic DNA fragments. Through enriching protein-bound DNA fragments to construct luciferase reporter libraries, we found that, of 280 clones collected and sequenced, 131 clones contained either the promoter-35 and -10 conservative sequences and/or an operator transcription factor binding sites (TFBS) region. To demonstrate the functionality of the identified clones, five of 131 clones containing LexA binding sequence have been demonstrated to be induced in response to mitomycin C treatment. To evaluate our libraries as a functional screening library, 80 randomly picked up clones were cultured and treated with and without MMC, where two clones were shown to have greater than twofold induction. In addition, two arsenite-responsive clones were identified from 90 clones, one having the well-known ArsR and another having the osmotically inducible lipoprotein (OsmE1). The newly discovered osmE1 has been quantitatively validated to be induced by arsenite treatment with real-time PCR in a dose response and time course manner. This enriching protein-bound DNA luciferase reporter libraries and functional screening facilitate the identification of stress-responsive transcriptional factors in microbes. We developed functional libraries containing E. coli genomic-wide protein-bound DNA as enhancers/operators to regulate downstream luciferase in response to stress.

All well-established cell size homeostasis paradigms are based on the researches of rod bacteria like B. subtilis and E. coli, suggesting a constant division time (timer model), division size (sizer model) or added size (adder model) before division. However, Lysinibacillus varians, a new species with regular filament-to-rod cell cycle, is inconsistent with existing models. In this study, the cell size parameters of the type strain GY32, were investigated by combing multiple microscopy techniques and single-cell approach. Our results showed that the filaments of strain GY32 were unicellular cells with multiple nucleoids. The division time of GY32 cells was variable and their daughter cells produced by asymmetric binary fission had different birth sizes, which were proportional to their elongation rates, resulting in high heterogeneity among the sister cells. Furthermore, the added size from birth to division was significantly shorter than birth size (P < 0.01) and decreased along generations. The results above revealed that the asymmetric division site and varied cell size parameters resulted in filament-to-rod cell cycle of L. varians, and cell size homeostasis could be a more complex and dynamic process than previously assumed. These findings would be helpful in elucidating the open questions in cell division and cell size heterogeneity.

Bacterial anaerobic respiration using an extracellular electron acceptor plays a predominant role in global biogeochemical cycles. However, the mechanisms of bacterial adaptation to the toxic organic pollutant as the extracellular electron acceptor during anaerobic respiration are not clear, which limits our ability to optimize the strategies for the bioremediation of a contaminated environment. Here, we report the physiological characteristics and the global gene expression of an ecologically successful bacterium, Shewanella decolorationis S12, when using a typical toxic organic pollutant, amaranth, as the extracellular electron acceptor. Our results revealed that filamentous shift (the cells stretched to fiber-like shapes as long as 18 μm) occurred under amaranth stress. Persistent stress led to a higher filamentous cell rate and decolorization ability in subcultural cells compared to parental strains. In addition, the expression of genes involved in cell division, the chemotaxis system, energy conservation, damage repair, and material transport in filamentous cells was significantly stimulated. The detailed roles of some genes with significantly elevated expressions in filamentous cells, such as the outer membrane porin genes ompA and ompW, the cytochrome c genes arpC and arpD, the global regulatory factor gene rpoS, and the methyl-accepting chemotaxis proteins genes SHD_2793 and SHD_0015, were identified by site-directed mutagenesis. Finally, a conceptual model was proposed to help deepen our insights into both the bacterial survival strategy when toxic organics were present and the mechanisms by which these toxic organics were biodegraded as the extracellular electron acceptors.IMPORTANCE Keeping toxic organic pollutants (TOPs) in tolerable levels is a huge challenge for bacteria in extremely unfavorable environments since TOPs could serve as energy substitutes but also as survival stresses when they are beyond some thresholds. This study focused on the underlying adaptive mechanisms of ecologically successful bacterium Shewanella decolorationis S12 when exposed to amaranth, a typical toxic organic pollutant, as the extracellular electron acceptor. Our results suggest that filamentous shift is a flexible and valid way to solve the dilemma between the energy resource and toxic stress. Filamentous cells regulate gene expression to enhance their degradation and detoxification capabilities, resulting in a strong viability. These novel adaptive responses to TOPs are believed to be an evolutionary achievement to succeed in harsh habitats and thus have great potential to be applied to environment engineering or synthetic biology if we could picture every unknown node in this pathway.

Electronic waste (e-waste) has caused a severe worldwide pollution problem. Despite increasing isolation of degradative microorganisms from e-waste contaminated environments, the mechanisms underlying their adaptive evolution in such habitats remain unclear. Sphingomonads generally have xenobiotic-degrading ability and may play important roles in bioremediation. Sphingobium hydrophobicum C1T, characterized with superior cell surface hydrophobicity, was recently isolated from e-waste contaminated river sediment. To dissect the mechanisms driving its adaptive evolution, we evaluated its stress resistance, sequenced its genome and performed comparative genomic analysis with 19 other Sphingobium strains. Strain C1T can feed on several kinds of e-waste-derived xenobiotics, exhibits a great resistance to heavy metals and possesses a high colonization ability. It harbors abundant genes involved in environmental adaptation, some of which are intrinsic prior to experiencing e-waste contamination. The extensive genomic variations between strain C1T and other Sphingobium strains, numerous C1T-unique genes, massive mobile elements and frequent genome rearrangements reflect a high genome plasticity. Positive selection, gene duplication, and especially horizontal gene transfer drive the adaptive evolution of strain C1T. Moreover, presence of type IV secretion systems may allow strain C1T to be a source of beneficial genes for surrounding microorganisms. This study provides new insights into the adaptive evolution of sphingomonads, and potentially guides bioremediation strategies.

BACKGROUND:A transcriptional reporter is the key component in bacterial biosensors which are employed to monitor the induction or repression of a reporter gene corresponding to environmental change. Interaction of a transcription factor with its consensus sequence generated by using a position weight matrix (PWM) model is crucial for its sensitivity of the reporter. However, recent studies suggest that PWM model based on independent contribution of individual consensus base pairs to protein interaction is often insufficient to explain complex regulation, such as the effect of nonconsensus sequences on the protein-DNA binding affinity. In the present study, we employed a simpler prokaryotic arsenic repressor (ArsR) regulation system to access the protein-DNA recognition. Contribution of nonconsensus base pairs within ArsR binding sequences toward ArsR-DNA binding and arsenic-mediated transcriptional induction was studied.RESULTS:We constructed a series of arsenic responsive reporters, each comprising two copies of the ArsR binding sequences from different resources. We found that high arsenic-mediated induction specifically requires the binding sequence from Escherichia coli to be placed at the first binding sequence; however, no such preference was observed for the second binding sequence, which could be from Acidithiobacillus ferrooxidans, plasmid R773, Synechococcus, or a core binding sequence of arsR. By creating a series of reporters differed at the nonconsensus base pairs of the second binding sequence, we observed that some constructs bound weakly while others strongly to ArsR. Most interestingly, although a number of these reporters showed similar binding affinity to ArsR, their arsenic-dependent induction differed significantly.CONCLUSIONS:The results indicated that nonconsensus base pairs could have profound influence on protein binding and may also modulate post-binding function. These findings provide new insights into the complex regulation of gene expression and facilitate the development of transcriptional reporter-based biosensors.

The key component in bacteria-based biosensors is a transcriptional reporter employed to monitor induction or repression of a reporter gene corresponding to environmental change. In this study, we made a series of reporters in order to achieve highly sensitive detection of arsenite. From these reporters, two biosensors were developed by transformation of Escherichia coli DH5α with pLHPars9 and pLLPars9, consisting of either a high or low copy number plasmid, along with common elements of ArsR-luciferase fusion and addition of two binding sequences, one each from E. coli and Acidithiobacillus ferrooxidans chromosome, in front of the R773 ArsR operon. Both of them were highly sensitive to arsenite, with a low detection limit of 0.04 μM arsenite (~ 5 μg/L). They showed a wide dynamic range of detection up to 50 μM using high copy number pLHPars9 and 100 μM using low copy number pLLPars9. Significantly, they differ in metal specificity, pLLPars9 more specific to arsenite, while pLHPars9 to both arsenite and antimonite. The only difference between pLHPars9 and pLLPars9 is their copy numbers of plasmid and corresponding ratios of ArsR to its binding promoter/operator sequence. Therefore, we propose a working model in which DNA bound-ArsR is different from its free form in metal specificity.

Aquatic sediment contamination by refractory organic pollutants is an urgent environmental issue in the world.Microorganism is an important driver in the remediation of contaminated aquatic sediments.Due to the flexibilities of microorganism in respiration and metabolism,microorganism can use a variety of contaminants as electron donors or acceptors in anaerobic respiration and construct a metabolic network with the biotic and abiotic factors in its surrounding environment coupling the degradation and transformation of refractory organic pollutants.In this paper,we review recent research progress in microbial anaerobic respiration,electron transfer network,and their remediation mechanisms for aquatic sediments contaminated by refractory organic pollutants.The prospects and challenges in developing the theories and technologies for microbial remediation of contaminated aquatic sediments are also discussed.

An increasing number of industrial, agricultural and commercial chemicals in the aquatic environment lead to various deleterious effects on organisms, which is becoming a serious global health concern. In this study, the Ames test and SOS/umu test were conducted to investigate the potential genotoxicity and mutagenicity caused by organic extracts from drinking water sources. Organic content of source water was extracted with XAD-2 resin column and organic solvents. Four doses of the extract equivalent to 0.25, 0.5, 1 and 2L of source water were tested for toxicity. All the water samples were collected from six different locations in Guangdong province. The results of the Ames test and SOS/umu test showed that all the organic extracts from the water samples could induce different levels of DNA damage and mutagenic potentials at the dose of 2 L in the absence of S9 mix, which demonstrated the existence of genotoxicity and mutagenicity. Additionally, we found that Salmonella typhimurium strain TA98 was more sensitive for the mutagen. Correlation analysis between genotoxicity, Organochlorine Pesticides (OCPs) and Polycyclic Aromatic Hydrocarbons (PAHs) showed that most individual OCPs were frame shift toxicants in drinking water sources, and there was no correlation with total OCPs and PAHs.

Two facultative chemolithotrophic, nitrate-reducing thiosulfate-oxidizing strains, F43bT and F21, were isolated from the sulfide-rich anoxic sediment of an urban creek in Pearl River Delta, China. Both strains were Gram-negative, facultatively anaerobic, non-spore-forming and rod-shaped with a flagellum. Phylogenetic analyses of 16S rRNA genes and the thrC, recA, glnII and atpD housekeeping genes revealed that the type strain shared high sequence similarities to Ciceribacter lividus MSSRFBL1T, with 98.8, 90.9, 94.8, 95.4 and 96.1 % identity, respectively. In addition, the major isoprenoid quinone (ubiquinone Q-10) and the DNA G+C content (66.0 mol%) of the type strain were similar to those of Ciceribacter lividus MSSRFBL1T. These results strongly support the classification of strains F43bT and F21 into the genus Ciceribacter. However, these strains diverged markedly from strain MSSRFBL1T with respect to several physiological and biochemical properties such as their semi-translucent colonies and nitrate-reducing and simultaneous thiosulfate-oxidizing respiration. Furthermore, the predominant fatty acids of strain F43bT were summed feature 2 (C18 : 1ω9t and/or C18 : 1ω9c and/or C18 : 1ω11t), C14 : 0 3-OH, C18 : 0 and C16 : 0, and its polar lipids were diphosphatidylglycerol, phosphatidylethanolamine, phosphatidymonomethylethanolamine and an unidentified glycolipid, which represented another two significant differences from strain MSSRFBL1T. Importantly, the DNA-DNA relatedness between strain F43bT and MSSRFBL1T was only 47.7 %. Based on the aforementioned polyphasic taxonomic results, the two isolates are suggested to represent a novel species of the genus Ciceribacter, for which the name Ciceribacterthiooxidans sp. nov. is proposed; the type strain is F43bT (=CCTCC AB 2016062T=KCTC 52231T).

The Dongjiang River plays an important role in southern China, as a source for irrigation and potable water of Hongkong and the other parts of the Pearl River Delta (PRD). The water quality index (WQI) was calculated to assess the spatial and temporal variability and identify the classification of water quality in the river. In order to simplify the procedure and reduce the analytical costs of the water quality evaluation, a modified WQI (defined as WQImin) was introduced based on Principal Component Analysis (PCA) and correlations analyses of the water parameters detected in dry and wet seasons during 2011–2012. Compared with the previous index, similar spatial changing trend and classification of the water quality were obtained by WQImin, which was composed of pH, temperature, total suspended solid, NH4+-N, and NO3−-N. The results showed an excellent water quality in the tributary site near the reservoir, a good water quality in the upstream of the river, and medium water quality in the downstream of the river, which suggested that the urban wastewater originated from increasing population size and industry development in the downstream mainly led to the deterioration of water quality along the river. Moreover, WQImin could more adequately reflect the seasonal changes of water quality which was slightly worse in dry season than wet season. Our results also suggest that continuous monitoring should be conducted to prevent pollution from industry and anthropogenic activities.

Terrestrial plant uptake of PBDEs from contaminated soils has been widely reported recently. In this study the fate of deca-BDE within a plant/PBDEs/aquatic environment system was investigated through simulated pot experiments. Accumulations of the total PBDEs and deca-BDE were observed in tissues of seven test aquatic plant species, namely Phragmites australis, Cyperus papyrus, Alternanthera philoxeroides, Colocasia esculenta, Scirpus validus, Acorus calamus and Oryza sativa. In all seven plants, O. sativa leads the uptake and accumulation both in the total PBDEs (444.8 ng g(-1)) and deca-BDE (368.0 ng g(-1)) in roots. Among the six common phytoremediation aquatic plants, A. calamus leads the uptake (236.2 ng g(-1)), and P. australis leads the translocation (Cshoot/Croot = 0.35), while A. philoxeroides (43.4%) and P. australis (80.0%) lead in the metabolism efficiencies in the root and shoot, respectively. The detection of seventeen lesser brominated PBDE congeners provided the debromination evidence, and the specific PBDEs profiles in test plant species indicated there is no common metabolic pattern. Furthermore, a relative high proportion of lesser brominated PBDE congeners in shoots suggested the possible metabolic difference between roots and shoots. Finally, a noticeable percentage of penta- and octa-BDE derived from deca-BDE also hint the ecological risk in deca-BDE use. This comparative research on the aquatic plants provide a broad vision on the understanding of plant/PBDEs/aquatic environment interaction system, and may be applied to remediate PBDEs in contaminated waters and sediments.

Electron acceptor redox potential (EARP) was presumed to be a determining factor for microbial metabolism in many natural and engineered processes. However, little is known about the potentially global effects of EARP on bacteria. In this study, we compared the physiological and transcriptomic properties of Shewanella decolorationis S12 respiring with different EARPs in microbial electrochemical systems to avoid the effects caused by the other physicochemical properties of real electron acceptor. Results showed that the metabolic activities of strain S12 were nonlinear responses to EARP. The tricarboxylic acid cycle for central carbon metabolism was down-regulated while glyoxylate shunt was up-regulated at 0.8 V compared to 0.2 and −0.2 V, which suggested that EARP is an important but not the only determinant for metabolic pathways of strain S12. Moreover, few cytochrome c genes were differentially expressed at different EARPs. The energy intensive flagella assembly and assimilatory sulfur metabolism pathways were significantly enriched at 0.8 V, which suggested strain S12 had stronger electrokinesis behavior and oxidative stress-response at high EARP. This study provides the first global information of EARP regulations on microbial metabolism, which will be helpful for understanding microorganism respiration.

Extraction of high-quality microbial DNA from contaminated environmental samples is an essential step in microbial ecological study. Based on previously published methods for soil and sediment samples, a modified pretreatment method was developed for extracting microbial DNA from heavily contaminated river sediment samples via selection of optimal pretreatment parameters (i.e., reagent solution, reaction duration, and temperature). The pretreatment procedure involves washing the river sediment sample for three times with a solution containing 0.1 mol·L−1 ethylene diamine tetraacetic acid (EDTA), 0.1 mol·L−1 Tris (pH 8.0), 1.5 mol·L−1 NaCl, 0.1 mol·L−1 NaH2PO4, and Na2HPO4 at 65°C with 180 r·min−1 for 15 min to remove humic materials and heavy metals prior to the employment of standard DNA extraction procedures. We compared the results of standard procedure DNA extraction following pretreatment, without pretreatment, and with using a commercial PowerSoil™ DNA Isolation Kit. The results indicated that the pretreatment significantly improved the DNA quality based on DNA yield, DNA fragment length, and determination of prokaryotic diversity. Prokaryotic diversity exhibited in the DNA with the pretreatment was also considerably higher than that extracted with the PowerSoil™ DNA Isolation Kit only. The pretreatment method worked well even with a small amount of sediment sample (0.25 g or even lower). The method provides a novel, simple, cost-effective tool for DNA extraction for microbial community analysis in environmental monitoring and remediation processes.

A pilot-scale multilayer rapid infiltration system (MRIS) for domestic wastewater treatment was established and efficient removal of ammonia and chemical oxygen demand (COD) was achieved in this study. The microbial community composition and abundance of ammonia oxidizers were investigated. Efficient biofilms of ammonia oxidizers in the stationary phase (packing material) was formed successfully in the MRIS without special inoculation. DGGE and phylogenetic analyses revealed that proteobacteria dominated in the MRIS. Relative abundance of ammonia-oxidizing archaea (AOA) and ammonia-oxidizing bacteria (AOB) showed contrary tendency. In the flowing phase (water effluent), AOA diversity was significantly correlated with the concentration of dissolve oxygen (DO), NO3-N and NH3-N. AOB abundance was significantly correlated with the concentration of DO and chemical oxygen demand (COD). NH3-N and COD were identified as the key factors to shape AOB community structure, while no variable significantly correlated with that of AOA. AOA might play an important role in the MRIS. This study could reveal key environmental factors affecting the community composition and abundance of ammonia oxidizers in the MRIS.

A high number of aromatic compounds that have been released into aquatic ecosystems have accumulated in sediment because of their low solubility and high hydrophobicity, causing significant hazards to the environment and human health. Since nitrate is an essential nitrogen component and a more thermodynamically favorable electron acceptor for anaerobic respiration, nitrate-based bioremediation has been applied to aromatic-contaminated sediments. However, few studies have focused on the response of aromatic-degrading microbial communities to nitrate addition in anaerobic sediments. Here we hypothesized that high nitrate inputs would stimulate aromatic-degrading microbial communities and their associated degrading processes, thus increasing the bioremediation efficiency in aromatic compound-contaminated sediments. We analyzed the changes of key aromatic-degrading genes in the sediment samples from a field-scale site for in situ bioremediation of an aromatic-contaminated creek in the Pearl River Delta before and after nitrate injection using a functional gene array. Our results showed that the genes involved in the degradation of several kinds of aromatic compounds were significantly enriched after nitrate injection, especially those encoding enzymes for central catabolic pathways of aromatic compound degradation, and most of the enriched genes were derived from nitrate-reducing microorganisms, possibly accelerating bioremediation of aromatic-contaminated sediments. The sediment nitrate concentration was found to be the predominant factor shaping the aromatic-degrading microbial communities. This study provides new insights into our understanding of the influences of nitrate addition on aromatic-degrading microbial communities in sediments.