Mar 2011, Volume 5 Issue 1

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    Zhili HE, Joy D. VAN NOSTRAND, Ye DENG, Jizhong ZHOU

    Functional gene arrays (FGAs) are a special type of microarrays containing probes for key genes involved in microbial functional processes, such as biogeochemical cycling of carbon, nitrogen, sulfur, phosphorus, and metals, biodegradation of environmental contaminants, energy processing, and stress responses. GeoChips are considered as the most comprehensive FGAs. Experimentally established probe design criteria and a computational pipeline integrating sequence retrieval, probe design and verification, array construction, data analysis, and automatic update are used to develop the GeoChip technology. GeoChip has been systematically evaluated and demonstrated to be a powerful tool for rapid, specific, sensitive, and quantitative analysis of microbial communities in a high-throughput manner. Several generations of GeoChip have been developed and applied to investigate the functional diversity, composition, structure, function, and dynamics of a variety of microbial communities from different habitats, such as water, soil, marine, bioreactor, human microbiome, and extreme ecosystems. GeoChip is able to address fundamental questions related to global change, bioenergy, bioremediation, agricultural operation, land use, human health, environmental restoration, and ecological theories and to link the microbial community structure to environmental factors and ecosystem functioning.

    Husen ZHANG

    New high-throughput technologies continue to emerge for studying complex microbial communities. In particular, massively parallel pyrosequencing enables very high numbers of sequences, providing a more complete view of community structures and a more accurate inference of the functions than has been possible just a few years ago. In parallel, quantitative real-time polymerase chain reaction (QPCR) allows quantitative monitoring of specific community members over time, space, or different environmental conditions. In this review, the principles of these two methods and their complementary applications in studying microbial ecology in bioenvironmental systems are discussed. The parallel sequencing of amplicon libraries and using barcodes to differentiate multiple samples in a pyrosequencing run are explained. The best procedures and chemistries for QPCR amplifications are also described and advantages of applying automation to increase accuracy are addressed. Three examples in which pyrosequencing and QPCR were used together to define and quantify members of microbial communities are provided: in the human large intestine, in a methanogenic digester whose sludge was made more bioavailable by a high-voltage pretreatment, and on the biofilm anode of a microbial electrolytic cell. The key findings in these systems and how both methods were used in concert to achieve those findings are highlighted.

    Juntaek LIM, Seung Gu SHIN, Seungyong LEE, Seokhwan HWANG

    Real-time quantitative polymerase chain reaction (qPCR) has gained popularity as a technique to detect and quantify a specific group of target microorganisms from various environmental samples including soil, water, sediments, and sludge. Although qPCR is a very useful technique for nucleic acid quantification, accurately quantifying the target microbial group strongly depends on the quality of the primer and probe used. Many aspects of conducting qPCR assays have become increasingly routine and automated; however, one of the most important aspects, designing and selecting primer and probe sets, is often a somewhat arcane process. In many cases, failed or non-specific amplification can be attributed to improperly designed primer-probe sets. This paper is intended to provide guidelines and general principles for designing group-specific primers and probes for qPCR assays. We demonstrate the effectiveness of these guidelines by reviewing the use of qPCR to study anaerobic processes and biologic nutrient removal processes. qPCR assays using group-specific primers and probes designed with this method, have been used to successfully quantify 16S ribosomal Ribonucleic Acid (16S rRNA) gene copy numbers from target methanogenic and ammonia- oxidizing bacteria in various laboratory- and full-scale biologic processes. Researchers with a good command of primer and probe design can use qPCR as a valuable tool to study biodiversity and to develop more efficient control strategies for biologic processes.

    Qingmei YAN, Xuxiang ZHANG, Tong ZHANG, Herbert H P FANG

    Activated sludge was monthly sampled from a saline sewage treatment plant of Hong Kong (China) during June 2007 to May 2008 to analyze the microbial community shift along with environmental variations using denaturing gradient gel electrophoresis of polymerase chain reaction amplified 16S rDNA fragments. Environmental changes resulted into a seasonal microbial community shift characterized by alterations in species number and abundance in the sewage treatment plant. Correspondence analysis and cluster analysis on community structure profile showed that the 12 monthly samples fell into four groups, which is in accordance with season changing in Hong Kong. Canonical correspondence analysis revealed that PO43--P and NH4+-N posed more significant effects on community structure than total phosphorus and total nitrogen, respectively. Compared with sludge retention time, influent total phosphorus had an inverse effect on the community structure shift, and chemical oxygen demand and NH4+-N showed a similar effects. Results of this study may contribute to the development of new knowledge involving the microbial community shift in sewage treatment plants.

    Rongchang WANG, Xinmin ZHAN, Yalei ZHANG, Jianfu ZHAO

    Nitrogen removal performance and nitrifying population dynamics were investigated in a redox stratified membrane biofilm reactor (RSMBR) under oxygen limited condition to treat ammonium-rich wastewater. When the NH4+-N loading rate increased from 11.1±1.0 to 37.2±3.2 gNH4+-N·m-2·d-1, the nitrogen removal in the RSMBR system increased from 18.0±9.6 mgN·d-1 to 128.9±61.7 mgN·d-1. Shortcut nitrogen removal was achieved with nitrite accumulation of about 22.3±5.3 mgNO2--N·L-1. Confocal micrographs showed the stratified distributions of nitrifiers and denitrifiers in the membrane aerated biofilms (MABs) at day 120, i.e., ammonia and nitrite oxidizing bacteria (AOB and NOB) were dominant in the region adjacent to the membrane, while heterotrophic bacteria propagated at the top of the biofilm. Real-time qPCR results showed that the abundance of amoA gene was two orders of magnitude higher than the abundance of nxrA gene in the MABs. However, the nxrA gene was always detected during the operation time, which indicates the difficulty of complete washout of NOB in MABs. The growth of heterotrophic bacteria compromised the dominance of nitrifiers in biofilm communities, but it enhanced the denitrification performance of the RSMBR system. Applying a high ammonia loading together with oxygen limitation was found to be an effective way to start nitrite accumulation in MABs, but other approaches were needed to sustain or improve the extent of nitritation in nitrogen conversion in MABs.

    Deming ZHAO, Jie CHENG, Michael R. HOFFMANN

    Aqueous solutions of phenol were oxidized by hydrogen peroxide assisted by microwave (MW) irradiation. A simple kinetic model for the overall degradation of phenol in the presence of excess H2O2 is proposed in which the degradation rate of phenol is expressed as a linear function of the concentrations of phenol and H2O2. A detailed parametric study showed that the degradation rate of phenol increased with increasing [H2O2] until saturation was observed. Phenol degradation followed apparent zero-order kinetics under MW radiation or H2O2 oxidation. However, after 90 min of irradiation, the observed kinetics shifted to pseudo first order. The overall reaction rates were significantly enhanced in the combined MW/H2O2 system, mainly because microwave could accelerate H2O2 to generate hydroxyl radical (·OH) and other reactive oxygen intermediates. The observed synergetic effects of the MW/H2O2 process resulted in an increased in the net reaction rate by a factor of 5.75. When hydrogen peroxide is present in a large stoichiometric excess, the time required to achieve complete mineralization is reduced significantly.

    Yanhui ZHAN, Jianwei LIN, Yanling QIU, Naiyun GAO, Zhiliang ZHU

    Surfactant-modified natural zeolites (SMNZ) with different coverage types were prepared by loading hexadecyltrimethyl ammonium bromide (HTAB) onto the surface of a natural zeolite. The adsorption behavior of humic acid (HA) on SMNZ was investigated. Results indicate that the adsorbent SMNZ exhibited a higher affinity toward HA than the natural zeolite. HA removal efficiency by SMNZ increased with HTAB loading. Coexisting Ca2+ in solution favored HA adsorption onto SMNZ. Adsorption capacity decreased with an increasing solution pH. For typical SMNZ with bilayer HTAB coverage, HA adsorption process is well described by a pseudo-second-order kinetic model. The experimental isotherm data fitted well with the Langmuir model. Calculated maximum HA adsorption capacities for SMNZ with bilayer HTAB coverage at pH 5.5 and 7.5 were 63 and 41 mg·g-1, respectively. E2/E3 (absorbance at 250 nm to that at 365 nm) and E4/E6 (absorbance at 465 nm to that at 665 nm) ratios of the residual HA in solution were lower than that of the original HA solution. This indicates that the HA fractions with high polar functional groups, low molecular weight (MW), and aromaticity had a stronger tendency for adsorption onto SMNZ with bilayer HTAB coverage. Results show that HTAB-modified natural zeolite is a promising adsorbent for removal of HA from aqueous solution.

    Shufang WU, Pute WU, Hao FENG, G. P. Merkley

    An evaluation of the interactions between vegetation, overland and soil erosion can provide valuable insight for the conservation of soil and water. An experiment was conducted to study water infiltration, runoff generation process, rate of sediment erosion, and hydrodynamic characteristics of overland flow from a sloping hillside with different draw-off discharges from alfalfa and control plots with 20° slope. The effect of alfalfa on runoff and sediment transport reduction was quantitatively analyzed. Alfalfa was discussed for its ability to reduce the overland flow scouring force or change the runoff movement. Compared to the bare-soil plots, alfalfa plots generated a 1.77 times increase in infiltration rate. Furthermore, the down-slope water infiltration rate for the bare soil plots was higher than in the up-slope, while the opposite was found in the alfalfa plots. In addition, alfalfa had a significant effect on runoff and sediment yield. In comparison to the control, the runoff coefficient and sediment transportation rate decreased by 28.3% and 78.4% in the grass slope, respectively. The runoff generated from the alfalfa and bare-soil plots had similar trends with an initial increase and subsequent leveling to a steady-state rate. The transport of sediment reduced with time as a consequence of the depletion of loose surface materials. The maximum sediment concentration was recorded within the first few minutes of each event. The alfalfa plots had subcritical flow while the bare-soil plots had supercritical flow, which indicate that the capability of the alfalfa slope for resisting soil erosion and sediment movement was greater than for bare soil plots. Moreover, the flow resistance coefficient and roughness coefficient for the alfalfa plots were both higher than for the bare-soil plots, which indicate that overland flow in alfalfa plots had retarded and was blocked, and the flow energy along the runoff path had gradually dissipated. Finally, the ability to erode and transport sediment had decreased.

    Yansong HOU, Wei LIANG, Liping ZHANG, Shuiping CHENG, Feng HE, Zhenbin WU

    The study of community composition of algae is essential for understanding the structure and dynamics of the aquatic ecosystem and for evaluating the eutrophic level of the water body. A high-performance liquid chromatographic (HPLC) method based on a reverse-phase C18 nonpolar column was developed for the main algal taxa, which includes cyanophytes, bacillariophytes, euglenophytes, dinophytes, and chlorophytes. Based on the elution order using HPLC, 19 pigments were identified, and they were chlorophyllide a, 19′-butanoyloxyfucoxanthin, chlorophyll c1 + c2, phephorbides a, peridinin, methyl-chlorophyllide a, fucoxanthin, neoxanthin, violaxanthin, myxoxanthophyll, diadinoxanthin, diatoxanthin, lutein, zeaxanthin, chlorophyll b allomer, chlorophyll b, chlorophyll a allomer, chlorophyll a, and β,β-carotene. A comparison study of cell microscopic counts and accessory pigment analysis indicated that HPLC analysis could be a useful tool for monitoring phytoplankton communities and their abundance.

    Xiaohui WANG, Xianghua WEN, Hengjing YAN, Kun DING, Man HU

    To determine whether the functional stability of nitrification was correlated to a stable community structure of ammonia oxidizing bacteria (AOB) in a full-scale wastewater treatment plant, the AOB community dynamics in a wastewater treatment system was monitored over one year. The community dynamics were investigated using specific PCR followed by terminal restriction fragment length polymorphism (T-RFLP) analysis of the amoA gene. The T-RFLP results indicated that during the period of nitrification stability, the AOB community structure in the full-scale wastewater treatment system was relatively stable, and the average change rate every 15 d of the system was 6.6%±5.8%. The phylogenetic analysis of the cloned amoA gene showed clearly that the dominant AOB in the system was Nitrosomonas spp. The results of this study indicated that throughout the study period, the AOB community structure was relatively stable in the full-scale wastewater treatment system with functional stability of nitrification.

    Baohua TANG, Lingyan ZHU, Qixing ZHOU

    The joint toxicity of Penta-BDE (Pe-BDE) and heavy metals including cadmium and copper on Daphnia magna (D. magna) was evaluated on the basis of determining the 48 h survival, antioxidative enzyme responses, and lipid peroxidation. The response was classified as additive, greater than additive, or less than additive by comparing the measured “toxic units, TU” with one. Based on the survival of D. magna, less-than-additive interactions were found in most of mixtures treatments. This may be attributed to the different toxicity mechanism between Pe-BDE and metals. Cu and Cd played a greater role in toxicity than what Pe-BDE did. As for the superoxide dismutase (SOD) and catalase (CAT) activity, most response was less than additive. For the glutathione S-transferases (GST) activity, most of the greater-than-additive responses were found in the Cu plus Pe-BDE treatments, but the additive responses occurred in Cd plus Pe-BDE treatments and binary metal treatments. For lipid peroxide levels, which were measured as malondialdehyde (MDA) levels, less-than-additive response occurred in the 50% Cd plus 50% Cu and ternary mixture treatments. Results suggested that Pe-BDE, Cd, and Cu could induce different patterns of antioxidant enzyme responses, such as antioxidant/prooxidant responses, depending on their capability to produce reactive oxygen species and antioxidant enzymes to detoxify them.

    John E COULTER

    Through human history, wealth has been measured in grain, gold, and, now, dollars. Though counterfeiting of coins and notes goes back a long way, it is only with electronic financial accounting in a global economy tainted by toxic loans and imaginary funds that there is an urgency to search for a realistic objective way to monitor and regulate what we are doing to our Earth and ourselves. Various schemes using analysis of utility functions, oil equivalents, entropy, energy, and other units have been tried and, while helping to understand some basic processes and flows, have always been swamped by the machinations of financiers and the attention big sums of money attract. Now, the concept of exergy, pioneered in Eastern Europe in the 1950s, is being researched, developed, and applied, especially in China, driven by the desperation to measure the reality beyond the twin specters of global financial and environmental crises. A rough inventory of the matter in the biosphere at the coordinate details of an angstrom and an appreciation of how humans harness and manipulate electromagnetic forces can be enlightening as to what is and is not sustainable. Without that understanding, any financial estimate and proposed stimulus packages or IMF reform will be wildly wrong and may even be headed in the wrong direction.

    Zhaoxing HAN, Zhenyao SHEN, Yongwei GONG, Qian HONG

    Emission trading is one of the most effective alternatives to controlling water pollution. Water environmental functional zone (WEFZ) is used to determine the water quality standard and identify the zone boundary for each river or reach. In this study, a new emission trading scheme was addressed based on WEFZ, accounting for both the temporal dimension and water quality control. A temporal factor of emission trading was proposed based on variations in the environmental capacity within a year by dividing the year into three periods, including high, normal, and low periods of environmental capacity. During each period, emission trading was implemented exclusively. A water quality-control scheme was suggested based on the water quality requirement in the water functional zone, in which the water quality at the downstream boundary of the zone was required to meet the water standard following auto-purification in the stream. Two methods of calculating water quality control are addressed for point-source pollution and non-point-source pollution. The calculated temporal dimension and water quality control were located in Dongxi River of the Daning Watershed in the Three Gorges Watershed. The high period was during June, July, and August, the normal period was during April, May, September, and October, and the low period was during January, February, March, November, and December. The results from the water quality calculation demonstrated that the discharge of point-source and non-point-source pollutions led to an excess of common contaminants at the downstream boundary of WEFZ. The temporal and spatial factors above should be incorporated into the emission trading scheme based on WEFZ.

    Tonggang SHEN, Hanchang SHI, Huiming SHI, He JING, Huilei XIONG

    To improve the efficiency of nitrogen removal with lower energy consumption, the study of feedforward control was carried out on a pilot-scale anaerobic-anoxic-oxic (AAO) plant for the treatment of municipal wastewater. The effluent qualities of the pilot plant under different control strategies were investigated. The results indicated that the change of external recycle was not a suitable approach to regulate the sludge concentration of plug-flow reactors; adjusting the aeration valve and dissolved oxygen set-point according to ammonia load could overcome the impact of influent fluctuation; and the denitrification potential could be estimated based on the transit time of anoxic zone and the relative content of carbon resource entering the anoxic zone. Simple feedforward control strategies for aeration and internal recycle were subsequently proposed and validated. The nitrogen removal was successfully improved in the pilot plant. The effluent total nitrogen had decreased by 29.9% and was steadily controlled below 15 mg·L-1. Furthermore, approximately 38% of the energy for aeration had been saved.

    Sheng CHANG, Jianzheng LI, Feng LIU

    An anaerobic contact reactor (ACR) system comprising a continuous flow stirred tank reactor (CSTR) with settler to decouple the hydraulic retention time (HRT) from solids retention time (SRT) was developed for fermentative hydrogen production from diluted molasses by mixed microbial cultures. The ACR was operated at various volumetric loading rates (VLRs) of 20–44 kgCOD·m-3·d-1 with constant HRT of 6 h under mesophilic conditions of 35°C. The SRT was maintained at about 46–50 h in the system. At the initial VLR of 20 kgCOD·m-3·d-1, the hydrogen production rate dropped from 22.6 to 1.58 L·d-1 as the hydrogen was consumed by the hydrogentrophic methanogen. After increasing the VLR to 28 kgCOD·m-3·d-1 and discharging the sludge for 6 consecutive times, the hydrogentrophic methanogens were eliminated, and the hydrogen content reached 36.4%. As the VLR was increased to 44 kgCOD·m-3·d-1, the hydrogen production rate and hydrogen yield increased to 42.1 L·d-1 and 1.40 mol H2·molglucose-consumed-1, respectively. The results showed that a stable ethanol-type fermentation that favored hydrogen production in the reactor was thus established with the sludge loading rate (SLR) of 2.0–2.5 kgCOD·kgMLVSS-1·d-1. It was found that the ethanol increased more than other liquid fermentation products, and the ethanol/acetic acid (mol/mol) ratio increased from 1.27 to 2.45 when the VLR increased from 28 to 44 kgCOD·m-3·d-1, whereas the hydrogen composition decreased from 40.4% to 36.4%. The results suggested that the anaerobic contact reactor was a promising bioprocess for fermentative hydrogen production.