Nondestructive and three-dimensional visualization by identifying elements using synchrotron radiation microscale X-ray CT reveals microbial and cavity distributions in anaerobic granular sludge.

We developed a nondestructive three-dimensional microbial visualization method utilizing synchrotron radiation X-ray microscale computed tomography to better understand the relationship between microorganisms and their surrounding habitats. The method was tested and optimized using a mixture of axenic Escherichia coli and Comamonas testosteroni. The osmium-thiocarbohydrazide-osmium method was used to stain all the microbial cells, and gold in situ hybridization was used to detect specific phylogenetic microbial groups. The stained samples were embedded in epoxy resin for microtomographic analysis. Differences in X-ray absorbances were calculated by subtracting the pre-L3-edge images from the post-L3-edge images to visualize the osmium and gold signals. Although we successfully detected cells stained with osmium, those labeled with gold were not detected, probably because of the insufficient density of gold atoms in the microbial cells. We then applied the developed technique to anaerobic granules and visualized the distribution of microbial cells and extracellular polymeric substances. Empty spaces were highlighted to determine the cavity distribution in granules. Numerous independent cavities of different sizes were identified in the granules. The developed method can be applied to various environmental samples for deeper insights into microbial life in their habitats. IMPORTANCE: Microorganisms inhabit diverse environments and often form biofilms. One factor that affects their community structure is the surrounding physical environment. The arrangement of residential space within the formed biofilm plays a crucial role in the supply and transportation of substances, as well as the discharge of metabolites. Conventional approaches, such as scanning electron microscopy and confocal laser scanning microscopy combined with fluorescence in situ hybridization, have limitations as they provide information primarily from the biofilm surface and cross-sections. In this study, we developed a method for detecting microorganisms in biofilms using synchrotron radiation X-ray microscale computer tomography. The developed method allows nondestructive three-dimensional observation of biofilms at a single-cell resolution (voxel size of approximately 200 nm), facilitating an understanding of the relationship between microorganisms and their physical habitats.

Iron Recycle-Driven Organic Capture and Sidestream Anaerobic Membrane Bioreactor for Revolutionizing Bioenergy Generation in Municipal Wastewater Treatment.

The practicality of intensifying organic matter capture for bioenergy recovery to achieve energy-neutral municipal wastewater treatment is hindered by the lack of sustainable methods. This study developed innovative processes integrating iron recycle-driven organic capture with a sidestream anaerobic membrane bioreactor (AnMBR). Iron-assisted chemically enhanced primary treatment achieved elemental redirection with 75.2% of chemical oxygen demand (COD), 20.2% of nitrogen, and 97.4% of phosphorus captured into the sidestream process as iron-enhanced primary sludge (Fe-PS). A stable and efficient biomethanation of Fe-PS was obtained in AnMBR with a high methane yield of 224 mL/g COD. Consequently, 64.1% of the COD in Fe-PS and 48.2% of the COD in municipal wastewater were converted into bioenergy. The acidification of anaerobically digested sludge at pH = 2 achieved a high iron release efficiency of 96.1% and a sludge reduction of 29.3% in total suspended solids. Ultimately, 87.4% of iron was recycled for coagulant reuse, resulting in a theoretical 70% reduction in chemical costs. The novel system evaluation exhibited a 75.2% improvement in bioenergy recovery and an 83.3% enhancement in net energy compared to the conventional system (primary sedimentation and anaerobic digestion). This self-reliant and novel process can be applied in municipal wastewater treatment to advance energy neutrality at a lower cost.

Advanced anaerobic digestion of household food waste pretreated by in situ-produced mixed enzymes via solid-state fermentation: Feasibility and application perspectives.

Enzymatic pretreatment is an effective method which can improve the anaerobic digestion (AD) efficiency of household food waste (HFW). As an alternative to expensive commercial enzymes, mixed enzymes (MEs) produced in situ from HFW by solid-state fermentation (SSF) can greatly promote the hydrolysis rate of HFW and achieve advanced anaerobic digestion (AAD) economically sustainable. In this paper, strategies for improving the efficiency of the enzyme-production process and the abundance of MEs are briefly discussed, including SSF, fungal co-cultivation, and stepwise fermentation. The feasibility of using HFW as an applicable substrate for producing MEs (amylase, protease, and lignocellulose-degrading enzymes) and its potential advantages in HFW anaerobic digestion are comprehensively illustrated. Based on the findings, an integrated AAD process of HFW pretreated with MEs produced in situ was proposed to maximise bioenergy recovery. The mass balance results showed that the total volatile solids removal rate could reach 98.56%. Moreover, the net energy output could reach 2168.62 MJ/t HFW, which is 9.79% higher than that without in situ-produced MEs and pretreatment. Finally, perspectives for further study are presented.

Insights into current bio-processes and future perspectives of carbon-neutral treatment of industrial organic wastewater: A critical review.

With the rise of the concept of carbon neutrality, the current wastewater treatment process of industrial organic wastewater is moving towards the goal of energy conservation and carbon emission reduction. The advantages of anaerobic digestion (AD) processes in industrial organic wastewater treatment for bio-energy recovery, which is in line with the concept of carbon neutrality. This study summarized the significance and advantages of the state-of-the-art AD processes were reviewed in detail. The application of expanded granular sludge bed (EGSB) reactors and anaerobic membrane bioreactor (AnMBR) were particularly introduced for the effective treatment of industrial organic wastewater treatment due to its remarkable prospect of engineering application for the high-strength wastewater. This study also looks forward to the optimization of the AD processes through the enhancement strategies of micro-aeration pretreatment, acidic-alkaline pretreatment, co-digestion, and biochar addition to improve the stability of the AD system and energy recovery from of industrial organic wastewater. The integration of anaerobic ammonia oxidation (Anammox) with the AD processes for the post-treatment of nitrogenous pollutants for the industrial organic wastewater is also introduced as a feasible carbon-neutral process. The combination of AnMBR and Anammox is highly recommended as a promising carbon-neutral process for the removal of both organic and inorganic pollutants from the industrial organic wastewater for future perspective. It is also suggested that the AD processes combined with biological hydrogen production, microalgae culture, bioelectrochemical technology and other bio-processes are suitable for the low-carbon treatment of industrial organic wastewater with the concept of carbon neutrality in future.

Exploring the maximum nitrite production rate through the granular sludge-type reactor to match the needs of anammox process realizing efficient nitrogen removal.

The reliable and efficient nitrite production rate (NPR) through nitritation process is the prerequisite for the efficient running of subsequent processes, like the anammox process and the nitrite shunt. However, there has been scant research on stable and productive nitritation process in recent years. In this study, at a stable hydraulic retention time of 12.0 h and with precise and strict DO control, the upper limit of the NPR was initially investigated using a continuous-flow granular sludge reactor. The NPR of 1.69 kg/m3/d with a nitrite production efficiency of 81.97% was finally achieved, which set a record until now in similar research. The median sludge particle size of 270.0 µm confirmed the development of clearly defined granular sludge. The genus Nitrosomonas was the major ammonium oxidizing bacteria. In conclusion, this study provides valuable insights for the practical application of the effective nitritation process driving subsequent nitrogen removal processes.

Comparing the mechanisms of syntrophic volatile fatty acids oxidation and methanogenesis recovery from ammonia stress in regular and biochar-assisted anaerobic digestion: Different roads lead to the same goal.

Biochar has been recognized as a promising additive to mitigate ammonia inhibition during syntrophic methanogenesis, while the key function of biochar in this process is still in debates. This study clarified the distinct mechanisms of syntrophic volatile fatty acids -oxidizing and methanogenesis recovery from ammonia inhibition in regular and biochar-assisted anaerobic digestion. Under 5 g/L ammonia stress, adding biochar shortened the methanogenic lag time by 10.9% and dramatically accelerated the maximum methane production rate from 60.3 to 94.7 mLCH4/gVSsludge/d. A photometric analysis with a nano-WO3 probe revealed that biochar enhanced the extracellular electron transfer (EET) capacity of suspended microbes (Pearson's r = -0.98), confirming that biochar facilitated methanogenesis by boosting EET between syntrophic butyrate oxidizer and methanogens. Same linear relationship between EET capacity and methanogenic rate was not observed in the control group. Microbial community integrating functional genes prediction analysis uncovered that biochar re-shaped syntrophic partners by enriching Constridium_sensu_stricto/Syntrophomonas and Methanosarcina. The functional genes encoding Co-enzyme F420 hydrogenase and formylmethanofuran dehydrogenase were upregulated by 1.4-2.3 times, consequently enhanced the CO2-reduction methanogenesis pathway. Meanwhile, the abundances of gene encoding methylene-tetrahydrofolate transformation, a series of intermediate processes involved in acetate oxidation, in the biochar-assisted group were 28.2-63.7% higher than these in control group. Comparatively, Methanosaeta played a pivotal role driving aceticlastic methanogenesis in the control group because the abundance of gene encoding acetyl-CoA decarbonylase/synthase complex increased by 1.9 times, suggesting an aceticlastic combining H2-based syntrophic methanogenesis pathway was established in control group to resist ammonia stress. A 2nd period experiment elucidated that although depending on distinct mechanisms, the volatile fatty acid oxidizers and methanogens in both groups developed sustained and stable strategies to resist ammonia stress. These findings provided new insights to understand the distinct methanogenic recovery strategy to resist toxic stress under varied environmental conditions.

Anammox-Mediated Hydroxyapatite Granules: Physicochemical Properties, 3D Hierarchy, and Biofilm Thickness.

Biomineralization inspired the development of simultaneous biological transformations and chemical precipitation for simultaneous nitrogen removal and phosphorus recovery from wastewater, which could compensate for the incapacity of phosphorus management in the new biological route of anaerobic ammonium oxidation (anammox). In this study, we strengthened anammox-mediated biomineralization by long-term feeding of concentrated N, P, and Ca substrates, and a self-assembled matrix of anammox bacteria and hydroxyapatite (HAP) was fabricated in a granular shape, defined as HAP-anammox granules. HAP was identified as the dominant mineral using elemental analysis, X-ray diffraction, and Raman spectroscopy. The intensive precipitation of HAP resulted in a higher inorganic fraction and substantially improved settleability of anammox biomass, which facilitated HAP precipitation by acting as nucleation and metabolically elevated pH. By using X-ray microcomputed tomography, we visually represented the hybrid texture of interwoven HAP pellets and biomass, the core-shell layered architecture of different-sized HAP-anammox granules, and their homogeneously regulated thickness of the outer biofilm (from 118 to 635 µm). This unique architecture endows HAP-anammox granules with outstanding settleability, active biofilm, and tightly bonded biofilm with the carrier, which may explain the excellent performance of these HAP-anammox granules under various challenging operational conditions in previous studies.

Improved Properties and Enhancement Strategies of Hydroxyapatite-Based Functional Granular Sludge for a High-Rate Partial Nitritation/Anammox System.

Retaining sufficient anammox bacteria (AnAOB) while keeping the anammox-based process stable is the focus of the study of anammox technology, especially in a one-stage partial nitritation/anammox (PNA) process. The use of hydroxyapatite (HAP) granules in an anammox-based process is innovative for its potential to improve the nitrogen removal rate and achieve simultaneous removal of phosphorus. In this study, the HAP-based granular sludge was employed using enhancement strategies for an excellent nitrogen removal performance in a one-stage PNA process. Compared to those of other granular sludge PNA systems, a remarkable sludge volume index of 7.8 mL/g and an extremely high mixed liquor volatile suspended solids of 15 g/L were achieved under a low hydraulic retention time of 2 h. Consequently, an unprecedented nitrogen removal rate as high as 4.8 kg N/m3/d at 25 °C was obtained under a nitrogen loading rate of 6 kg N/m3/d. After a long-term operation of 870 days, the enhancement strategies underlying the superior performance of the granular sludge were identified. These findings clearly demonstrate that the enhancement strategies are crucial for the superior operating performance of the PNA process, and they can promote the application of the anammox-based process.

Anaerobic membrane bioreactor for carbon-neutral treatment of industrial wastewater containing N, N-dimethylformamide: Evaluation of electricity, bio-energy production and carbon emission.

The feasibility of anaerobic membrane bioreactor (AnMBR) for the treatment of N, N-dimethylformamide (DMF)-containing wastewater was theoretically compared with the conventional activated sludge (CAS) process in this study. The electricity consumption and expenditure, bio-energy production and CO2 emission were investigated using the operational results of a lab-scale AnMBR operated in a long-term operation. The AnMBR was capable of producing bio-methane from wastewater and generated 3.45 kWh/m3 of electricity as recovered bio-energy while the CAS just generated 1.17 kWh/m3 of electricity from the post-treatment of excessive sludge disposal. The large quantity of bio-methane recovered by the AnMBR can also be sold as sustainable bioresource for the use of household natural gas with a theoretical profit gain of 29,821 US$/year, while that of the CAS was unprofitable. The AnMBR was also demonstrated to significantly reduce the carbon emission by obtaining a theoretical negative CO2 production of -2.34 kg CO2/m3 with the recycle of bio-energy while that for the CAS was 4.50 kg CO2/m3. The results of this study demonstrate that the AnMBR process has promising potential for the carbon-neutral treatment of high-strength DMF-containing wastewater in the future.

Increasing the organic loading rate of household food waste anaerobic digestion by landfill leachate addition: Performance and mechanism.

A high amount of easily degradable organics and the absence of trace metals (TMs) in household food waste (HFW) lowered the stability and efficiency of anaerobic digestion (AD) of HFW. Leachate addition to the AD of HFW can provide ammonia nitrogen and TMs to address the accumulation of volatile fatty acids and the lack of TMs. To study the effect of leachate addition on increasing organic loading rate (OLR), both mono-digestion of HFW and AD of HFW with leachate addition were evaluated using two continuously stirred tank reactors. The OLR of the mono-digestion reactor only reached 2.5 g COD/L/d. However, with the addition of ammonia nitrogen and TMs, the OLR of the failed mono-digestion reactor increased by 2 and 3.5 g COD/L/d, respectively. The specific methanogenic activity increased by 94.4% and the hydrolysis efficiency increased by 135%. Finally, the OLR of mono-digestion of HFW reached 8 g COD/L/d, with a hydraulic retention time (HRT) of 8 days and methane production rate of 2.4 L/L/d. In the leachate addition reactor, the OLR reached 15 g COD/L/d, while the HRT and methane production were 7 days and 3.4 L/L/d, respectively. This study demonstrates that leachate addition substantially improves the AD efficiency of HFW. The two main mechanisms of increasing the OLR of an AD reactor are the buffer capacity of ammonia nitrogen and the stimulation of methanogen by TMs from leachate.

EDTA-enhanced alkaline anaerobic fermentation of landfill leachate-derived waste activated sludge for short-chain fatty acids production: Metals chelation and EPSs destruction.

Alkaline anaerobic fermentation (AAF) of waste activated sludge (WAS) has been demonstrated to be promising for short-chain fatty acids (SCFAs) recovery. However, high-strength metals and EPSs in the landfill leachate-derived WAS (LL-WAS) would stabilize its structure, suppressing AAF performance. To improve sludge solubilization and SCFAs production, AAF was coupled with EDTA addition for LL-WAS treatment. The results show that sludge solubilization at AAF-EDTA was promoted by 62.8% than AAF, releasing 21.8% more soluble COD. The maximal SCFAs production of 477.4 mg COD/g VSS was thus achieved, i.e., 1.21 and 6.13 times those at AAF and the control, respectively. SCFAs composition was also improved with more acetic and propionic acids (80.8% versus 64.3%). Metals bridging EPSs were chelated by EDTA, which significantly dissolved metals from sludge matrix (e.g., 23.28 times higher soluble Ca than AAF). EPSs tightly bound with microbial cells were thus destructed (e.g., 4.72 times more protein release than alkaline treatment), causing an easier sludge disruption and subsequently a higher SCFAs production by hydroxide ions. These findings suggest an effective EDTA-supported AAF for metals and EPSs-rich WAS to recover carbon source.

Microbial characteristics in anaerobic membrane bioreactor treating domestic sewage: Effects of HRT and process performance.

Two anaerobic membrane bioreactors (AnMBRs) equipped with different membrane pore size (0.4 or 0.05 µm) were operated at 25˚C and fed with domestic wastewater. The hydraulic retention time (HRT) of the reactors was shortened. The microbial communities of the two AnMBRs were investigated by 16S rRNA gene amplicon sequencing to see the effects of HRT. The predominant Archaea was an aceticlastic methanogen Methanosaeta. The composition of hydrogenotrophic methanogens changed with the HRTs: the population of Methanobacterium was higher for longer HRTs, whereas the population of unclassified Methanoregulaceae was higher for shorter HRTs. The Anaerolineae, Bacteroidia and Clostridia bacteria were dominant in both of the reactors, with a combined relative abundance of over 55%. The relative abundance of Anaerolineae was proportional to the biogas production performance. The change in the population of hydrogenotrophic methanogens or Anaerolineae can be used as an indicator for process monitoring. The sum of the relative abundance of Anaerolineae and Clostridia fluctuated slightly with changes in the HRT in both AnMBRs when the reactor was stably operated. The co-occurrence analysis revealed the relative abundance of the operational taxonomic units belonging to Anaerolineae and Clostridia was functionally equivalent during the treatment of real domestic sewage. A principal coordination analysis revealed that the changes in the microbial community in each reactor were consistent with the change of HRT. In addition, both the HRT and the stability of the process are important factors for maintaining microbial community structures.

Sequence-Specific Capture of Oligonucleotide Probes (SCOPE): a Simple and Rapid Microbial rRNA Quantification Method Using a Molecular Weight Cutoff Membrane.

A method named sequence-specific capture of oligonucleotide probes (SCOPE) was developed for quantification of microbial rRNA molecules in a multiplex manner. In this method, a molecular weight cutoff membrane (MWCOM) was used for the separation of fluorescence-labeled oligonucleotide probes hybridized with rRNA from free unhybridized probes. To demonstrate proof of concept, probes targeting bacteria or archaea at different taxonomic levels were prepared and were hybridized with rRNAs. The hybridization stringency was controlled by adjusting reaction temperature and urea concentration in the mixture. Then, the mixture was filtered through the MWCOM. The rRNA and hybridized probes collected on the MWCOM were recovered and quantified using a spectrophotometer and fluorospectrometer, respectively. The method showed high accuracy in detecting specific microbial rRNA in a defined nucleic acid mixture. Furthermore, the method was capable of simultaneous detection and quantification of multiple target rRNAs in a sample with sensitivity up to a single-base mismatch. The SCOPE method was tested and benchmarked against reverse transcription-quantitative PCR (RT-qPCR) for the quantification of Bacteria, Archaea, and some key methanogens in anaerobic sludge samples. It was observed that the SCOPE method produced more reliable and coherent results. Thus, the SCOPE method allows simple and rapid detection and quantification of target microbial rRNAs for environmental microbial population analysis without any need for enzymatic reactions. IMPORTANCE Microorganisms play integral roles in the Earth's ecosystem. Microbial populations and their activities significantly affect the global nutrient cycles. Quantification of key microorganisms provides important information that is required to understand their roles in the environment. Sequence-based analysis of microbial population is a powerful tool, but it provides information only on relative abundance of microorganisms. Hence, the development of a simpler and quick method for the quantification of microorganisms is necessary. To address the shortcomings of a variety of molecular methods reported so far, we developed a simple, rapid, accurate, and multiplexed microbial rRNA quantification method to evaluate the abundance of specific microbial populations in complex ecosystems. This method demonstrated high specificity, reproducibility, and applicability to such samples. The method is useful for quantitative detection of particular microbial members in the environment.

Uncovering Viable Microbiome in Anaerobic Sludge Digesters by Propidium Monoazide (PMA)-PCR.

Use of anaerobic sludge digester is a common practice around the world for solids digestion and methane generation from municipal sewage sludge. Understanding microbial community structure is vital to get better insight into the anaerobic digestion process and to gain better process control. However, selective analysis of viable microorganisms is limited by DNA-based assays. In this study, propidium monoazide (PMA)-PCR with 16S rRNA gene sequencing analysis was used to distinguish live and dead microorganisms based on cell membrane integrity. Microbial community structures of PMA-treated and PMA-untreated anaerobic digester sludge samples were compared. Quantitative PCR revealed that 5-30% of the rRNA genes were derived from inactive or dead cells in anaerobic sludge digesters. This caused a significant decrease in the numbers of operational taxonomic units and Chao1 and Shannon indices compared with that of the PMA-untreated sludge. Microbial community analysis showed that majority of the viable microbiome consisted of Euryarchaeota, Bacteroidetes, Deltaproteobacteria, Chloroflexi, Firmicutes, WWE1, Spirochaetes, Synergistetes, and Caldiserica. On the other hand, after the PMA treatment, numbers of Alphaproteobacteria and Betaproteobacteria declined. These were considered residual microbial members. The network analysis also revealed a relationship among the OTUs belonging to WWE1 and Bacteroidales. PMA-PCR-based 16S rRNA gene sequencing analysis is an effective tool for uncovering viable microbiome in complex environmental samples.

New insights into the cultivation of N, N-dimethylformamide-degrading methanogenic consortium: A long-term investigation on the variation of prokaryotic community inoculated with activated sludge.

The cultivation of the N, N-dimethylformamide (DMF)-degrading methanogenic consortium is considered difficult. In this study, an up-flow anaerobic sludge blanket (UASB) was inoculated with activated sludge in order to culture the DMF-degrading anaerobic sludge under a constant DMF concentration of approximately 2000 mg L-1. While the UASB realized a nearly 100% degradation of DMF and a high methane production of 1.03 L d-1 for the first two months, both the removal efficiency and methane production continued to decrease until the end. The characterization of the prokaryotic community reveals that those DMF-hydrolyzing bacteria (DHB) originating from the activated sludge were responsible for the effective degradation of DMF. However, even when fed with a constant concentration of DMF, the DHB kept decreasing all the time while methane-producing archaea were rapidly cultivated. The variation of prokaryotic community suggests that the DHB could not proliferate anaerobically without utilizing the intermediate products from the hydrolysis of DMF, resulting in an unstable DMF-degrading consortium. The cultivation of DHB under the anaerobic condition of the UASB was therefore difficult. The reason it was not possible to culture a DMF-degrading methanogenic consortium in this study is that the DHB are denitrifying bacteria which require nitrate for their cell growth under the anaerobic condition. The solution to maintain the abundance of these DHB is to add doses of nitrate into the system. Nitrate is likely to help these DHB recapture intermediates from methanogens, enabling them to perform a heterotrophic denitrification by using a small proportion of DMF as the carbon source while simultaneously maintaining the cell growth of DHB.

Effect of treated sewage characteristics on duckweed biomass production and microbial communities.

Duckweed biomass production in a duckweed pond fed with three differently treated sewage (i.e. sewage treated by primary sedimentation (PS); conventional activated sludge process (CAS); and downflow hanging sponge process (DHS)) was evaluated in order to assess the effects of water quality on biomass yield. Higher and stable biomass production was observed when the duckweed pond was fed with PS or DHS-effluent than with CAS-effluent, evidently due to the difference in nutrient loads. Availability of nutrients, especially phosphorus, affected the biomass production rate: higher the nutrient, faster the production. Microbial community analysis revealed that the members of Rhizobiales were likely to contribute to stable and high biomass growth. From the results of the study, a sewage treatment system consisting of a primary sedimentation followed by a duckweed pond and a tertiary treatment unit can be proposed to maximize biomass production without compromising treatment objectives. Size and operational parameters of the duckweed pond should be determined primarily based on the nutrient availability in the influent water to maximize duckweed growth.

Optimizing biomethane production of mesophilic chicken manure and sheep manure digestion: Mono-digestion and co-digestion kinetic investigation, autofluorescence analysis and microbial community assessment.

Optimization of mesophilic methane production from Chicken manure (CM) and Sheep manure (SM) at total solid (TS) of 8% and 1.6% were obtained by sequence tests in mono-digestion. However, the positive synergy of co-digestion with an optimum CM/SM of 2.5 (310 mLCH4/gVSadded) resulted in a high hydrolytic capacity and methane production. The modified Gompertz model (R2 > 0.98) and modified Aiba model (R2 > 0.88) illustrated co-digestion significantly improved the methane generation rate with strong ammonia tolerance. Dissolved Organic Matter (DOM) variation in response to the metabolic rate of microbial community illustrated that the SMP-like and protein-like components half-split by EEM-PARAFAC were significantly negative corresponded to bio-methane production. Moreover, the canonical correlation analysis (CCA) resulted a significant difference between the substrate and DOM composition. Potential functional metabolic illustrated statistically significance difference between mono and co-digestion, however, Methanosaeta and Syntrophobacter predominated the syntrophic methanogenesis. The constructed complex metabolic cooperation caused the co-digestion stable and high efficiency.

Submerged anaerobic membrane bioreactor (SAnMBR) performance on sewage treatment: removal efficiencies, biogas production and membrane fouling.

A submerged anaerobic membrane reactor (SAnMBR) was employed for comprehensive evaluation of sewage treatment at 25 °C and its performance in removal efficiency, biogas production and membrane fouling. Average 89% methanogenic degradation efficiency as well as 90%, 94% and 96% removal of total chemical oxygen demand (TCOD), biochemical oxygen demand (BOD) and nonionic surfactant were obtained, while nitrogen and phosphorus were only subjected to small removals. Results suggest that SAnMBRs can effectively decouple organic degradation and nutrients disposal, and reserve all the nitrogen and phosphorus in the effluent for further possible recovery. Small biomass yields of 0.11 g mixed liquor volatile suspended solids (MLVSS)/gCOD were achieved, coupled to excellent methane production efficiencies of 0.338 NLCH4/gCOD, making SAnMBR an attractive technology characterized by low excess sludge production and high bioenergy recovery. Batch tests revealed the SAnMBR appeared to have the potential to bear a high food-to-microorganism ratio (F/M) of 1.54 gCOD/gMLVSS without any inhibition effect, and maximum methane production rate occurred at F/M 0.7 gCOD/gMLVSS. Pore blocking dominated the membrane fouling behaviour at a relative long hydraulic retention time (HRT), i.e. >12 hours, while cake layer dominated significantly at shorter HRTs, i.e. <8 hours.

On the effect of Fe(III) on proliferation of Microcystis aeruginosa at high nitrate and low chlorophyll condition.

The impact of Fe concentrations on the growth of Microcystisaeruginosa in aquatic systems under high nitrate and low chlorophyll conditions was studied. The responses of cell density, total and cell chlorophyll-a intracellular Fe content and organic elemental composition of M. aeruginosa to different concentration gradients of Fe(III) in the solutions were analysed. The results showed that the proliferation speeds of M. aeruginosa were: (1) decelerated when the Fe(III) concentration was lower than 50µg/L in the solutions, (2) promoted and positively related to the increase of Fe(III) concentration from 100 to 500µg/L in the solutions over the experimental period, and (3) promoted in the early stage but decelerated in later stages by excess adsorption of Fe by cells when the Fe(III) concentration was higher than 500µg/L in the solutions. The maximum cell density, total and cell chlorophyll-a were all observed at 500µg Fe(III)/L concentration. The organic elemental composition of M. aeruginosa was also affected by the concentration of Fe(III) in the solutions, and the molecular formula of M. aeruginosa should be expressed as C7-7.5H14O0.8-1.3N3.5-5 according to the functions for different Fe(III) concentrations. Cell carbon and oxygen content appeared to increase slightly, while cell nitrogen content appeared to decrease as Fe(III) concentrations increased from 100 to 500µg/L in the solutions. This was attributed to the competition of photosynthesis and nitrogen adsorption under varying cell Fe content.

Use of real-time PCR with propidium monoazide for enumeration of viable Escherichia coli in anaerobic digestion.

A combination of propidium monoazide (PMA) with real-time quantitative polymerase chain reaction (PMA-qPCR) was optimized to enumerate only viable Escherichia coli in anaerobic digestion processes. Repeating the PMA treatment twice and a final concentration of 100 µM resulted in an effective exclusion of DNA from heat-treated E. coli cells. In three anaerobic digestion processes, real-time PCR, PMA-qPCR, and the most probable number method (MPN) were used to estimate the numbers of total, viable, and culturable E. coli cells, respectively. Culturable concentrations of fecal coliforms were also measured by the membrane filter method. For thermophilic digestion, the reductions in total and viable E. coli cells from the digester influent to the effluent were significantly lower than those in culturable cells and fecal coliforms by two to four orders of magnitude. For mesophilic digestion, the differences in the reductions in E. coli and fecal coliforms counts were less than two orders of magnitude. Based on the measurements of viable E. coli determined by the PMA-qPCR method, the microbial quality of digester effluents was discussed for agricultural application, and pasteurization after anaerobic digestion was suggested for the destruction of viable pathogens.