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.

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.

Propidium monoazide - polymerase chain reaction reveals viable microbial community shifts in anaerobic membrane bioreactors treating domestic sewage at low temperature.

An anaerobic membrane bioreactor (AnMBR) treated domestic sewage at 15 °C under different hydraulic retention time (HRT) conditions (6, 12, 16, and 24 h). Propidium monoazide (PMA)-PCR excluded microorganisms without intact cell membranes, focusing on the viable microbial community in anaerobic digestion. The results showed that the 6-hour HRT had poor treatment performance: low chemical oxygen demand removal efficiency (below 80%) and high mean trans-membrane pressure and flux (15 kPa and 9.4 L/(m2 h)). Comparatively, PMA-PCR combined with next-generation sequencing improved the identification of microbial changes compared to conventional 16S rRNA gene sequencing. HRT influenced microorganisms in the hydrolysis and acid-production stages, including carbohydrate-degrading bacteria such as Bifidobacterium and Prevotella 1. Remarkably, a comparison with an AnMBR at 25 °C showed Proteobacteria to be the main cause of membrane fouling in the low-temperature AnMBR, with most operational taxonomic units negatively correlated with HRT and solids retention time.

Anammox upflow hybrid reactor: Nitrogen removal performance and potential for phosphorus recovery.

Echoing to the call of recovering high-value-added chemicals from wastewater and achieving carbon-neutral operation in wastewater treatment, an anammox upflow hybrid reactor was successfully applied for nitrogen removal, and the potential for phosphorus recovery was put forward. Moreover, the spatial pattern of removal capacities, and distribution of biomass and HAP precipitates were recognized and demonstrated as height-oriented. The intensity of HAP precipitates was highly consistent with the amount of anammox biomass and the relative abundance of the Candidatus Kuenenia, indicating that HAP formation was encouraged by the anammox reaction itself and heterogeneous nucleation induced by organic matters (proteins and polysaccharides). The fixed bed also played an important role in immobilizing the anammox biomass, secreted organic matrix, and HAP precipitates. This finding also provoked the thought that in the anammox process, HAP precipitation was more achievable, effective and practicable using the fixed-carrier system.

Enhanced digestion of sludge via co-digestion with food waste in a high-solid anaerobic membrane bioreactor: Performance evaluation and microbial response.

A 15 L high-solid mesophilic AnMBR was operated for the digestion of food waste, primary sludge and excess sludge. The digestion performance was evaluated from the perspective of methane generation, permeate quality and organic reduction. Furthermore, the change in the microbial community was investigated by 16S rRNA gene analysis. The results showed that the introduction of sludge decreased the H2S levels in biogas compared with the mono-digestion of food waste and the co-digestion with food waste increased biogas and methane production compared with the mono-digestion of sludge. A substitution ratio of 25 % became a turning point of permeate composition and reaction rates. The energy recovery ratios of the mesophilic AnMBR were over 75 % based on stoichiometric analysis. In reaction kinetics analysis, hydrolysis as the first step of anaerobic digestion was found to be most influenced by the composition of the substrate. Finally, the microbial community structures were stable under tested conditions while the evolutionary relationships within the dominant phyla were observed. In the archaea community, Methanosaeta was the dominant methanogen regardless sludge ratio in the substrate.

Sodium hypochlorite induced inhibition in anaerobic digestion and possible approach to maintain methane fermentation performance.

Since sodium hypochlorite (NaClO), a commonly used chemical to deal with membrane fouling, is toxic to microorganisms, it is a major concern in the membrane cleaning process. In this study, the concentration-dependent effects of NaClO (0-9 g/L) on the biodegradation performance and microbial activity were investigated via batch experiments. The methane production (obtained approximately 140 mL) and microbial community revealed by principal coordinates analysis were almost unaffected when the NaClO concentration ranged between 0 and 3 g/L. A follow-up batch experiment was conducted and revealed that the microbial products could help protect or recover the activity of anaerobic microorganisms at a high NaClO concentration of 10 g/L. Additionally, correlation analysis was used to investigate the associations between the 15 major bacterial genera. Moreover, the microbial analysis results indicated that the top 10 operational taxonomic units most affected by NaClO were primarily coryneform and filamentous bacteria.

Application of deep learning for predicting the treatment performance of real municipal wastewater based on one-year operation of two anaerobic membrane bioreactors.

In this study, data-driven deep learning methods were applied in order to model and predict the treatment of real municipal wastewater using anaerobic membrane bioreactors (AnMBRs). Based on the one-year operating data of two AnMBRs, six parameters related to the experimental conditions (temperature of reactor, temperature of environment, temperature of influent, influent pH, influent COD, and flux) and eight parameters for wastewater treatment evaluation (effluent pH, effluent COD, COD removal efficiency, biogas composition (CH4, N2, and CO2), biogas production rate, and oxidation-reduction potential) were selected to establish the data sets. Three deep learning network structures were proposed to analyze and reproduce the relationship between the input parameters and output evaluation parameters. The statistical analysis showed that deep learning closely agrees with the AnMBR experimental results. The prediction accuracy rate of the proposed densely connected convolutional network (DenseNet) can reach up to 97.44%, and the single calculation time can be reduced to within 1 s, suggesting the high performance of AnMBR treatment prediction with deep learning methods.

Architecture of HAP-anammox granules contributed to high capacity and robustness of nitrogen removal under 7°C.

The anaerobic ammonium oxidation (anammox) process is an autotrophic nitrogen removal process with great potential as a cost-effective and highly efficient technology in the wastewater treatment field. The main challenges yet to be overcome in this new frontier technology are operating at lower temperatures and achieving a high and stable nitrogen removal efficiency. In this study, an up-flow expanded bed reactor with hydroxyapatite (HAP)-anammox granules was operated for more than 200 days at 7°C. The nitrogen loading rate (NLR) was improved from 1.0 g-N/L/d to 3.6 g-N/L/d, together with a high-level nitrogen removal efficiency of 84-92%, which is the highest to date at extremely low temperatures in a continuous experiment. Candidatus Kuenenia was revealed to be the only dominant anammox genus, with a relative abundance of 35.3-37.5%. The optimal operational temperature was around 35°C and the apparent activation energy (Ea) was calculated as 78.37 kJ/mol. The three-layers architecture and architectural evolution of HAP-anammox granules into HAP-cores and peeling biofilms with outstanding settling performance were characterized. Under 7°C, the high capacity of nitrogen removal with robust removal efficiency using HAP-anammox granules was achieved.

Methanogenic performance and microbial community during thermophilic digestion of food waste and sewage sludge in a high-solid anaerobic membrane bioreactor.

The methanogenic performance and microbial community of the thermophilic anaerobic mono-digestion and co-digestion of food waste and sewage sludge in a high-solid membrane bioreactor were investigated by a continuous experiment. The methane recovery rate of the system reached 98.0% and 89.0% when the substrate was pure food waste and 25% sewage sludge substitution, respectively. Kinetics characterization showed that hydrolysis was the rate-limiting step in both mono-digestion and co-digestion while methanogenic performance and microbial community were significantly affected by feed condition. The dominant archaea for methane generation shifted from Methanothermobacter thermophilus (72.82%) to Methanosarcina thermophila (96.25%) with sewage sludge gradually added from 0% to 100% in the substrate. The relationships between digestion performance, such as the accumulation of soluble proteins in the reactor, and functional microbial groups were also carefully analyzed. Finally, reasonable metabolic pathways for mono-digestion and co-digestion were summarized.

Important effects of temperature on treating real municipal wastewater by a submerged anaerobic membrane bioreactor: Removal efficiency, biogas, and microbial community.

A submerged anaerobic membrane bioreactor (SAnMBR) was used in the treatment of real municipal wastewater at operation temperatures ranging from 15 °C to 25 °C and hydraulic retention time (HRT) of 6 h. The treatment process was evaluated in terms of organic removal efficiency, biogas production, sludge growth and membrane filtration. During long-term operation, the SAnMBR achieved chemical oxygen demand removal efficiencies of about 90% with a low sludge yield (0.12-0.19 g-VSS/g-CODrem) at 20-25 °C. Approximately 1.82-2.27 kWh/d of electric energy was generated during the wastewater treatment process at 20-25 °C, 0.67 kWh/d was generated at 15 °C. The microbial community analysis results showed that microbial community was dominated by aceticlastic methanogens, coupled by hydrogenotrophic methanogens and a very small quantity of methylotrophic methanogens. It was also shown that the stabilization of the microbial community could be attributed to the carbohydrate-protein degrading bacteria and the carbohydrate degrading bacteria.

One-year operation of a 20-L submerged anaerobic membrane bioreactor for real domestic wastewater treatment at room temperature: Pursuing the optimal HRT and sustainable flux.

A 20 L hollow-fiber submerged anaerobic membrane bioreactor (SAnMBR) was used to treat real domestic wastewater at 25 °C with hydraulic retention times (HRTs) ranging from 4 to 12 h. The process performance was evaluated by organic removal efficiency, biogas production, sludge yield, and filtration behaviors during one-year's operation. For HRTs ranging between 6 and 12 h, the AnMBR showed good organic removal efficiency with chemical oxygen demand (COD) and biochemical oxygen demand (BOD) removal efficiencies of about 89% and 93%, respectively. The biogas yield was 0.26 L-gas/g-CODfed, with approximately 80% methane content, and the sludge yield was 0.07-0.11 g-VSS/g-CODrem. While at an HRT of 4 h, with the higher wastewater treatment capacity and organic loading rate (OLR), the biogas production was lower (0.17 L-gas/g-CODfed), and the sludge production was higher (0.22 g-VSS/g-CODrem). The organic removal performance (COD 84% and BOD 89%) at HRT of 4 h was acceptable due to the effective separation effect of the membrane filtration process. According to COD balance analysis, the low biogas yield and high sludge yield at HRT of 4 h were due to insufficient biodegradation under an OLR of 2.05 g-COD/L-reactor/d. Theoretical calculations based on Henry's law indicate that the ideal methane content in the biogas should be 82-85% when the operational temperature was 25 °C. To achieve a high flux and sustainable AnMBR operation, the impact of mixed liquor suspended solid (MLSS) and gas sparging velocity (GSV) on the filtration performance was analyzed. The critical flux increased with increase in the GSV from 24.2 to 174.3 m/h, but decreased with increase in the MLSS concentration from 8.2 to 20.2 g/L. Therefore, decreasing fouling rate to 0.8-1.2 kPa/d by efficiently controlling GSV and MLSS, sustainable operation could be achieved at a flux of 0.34 m/d.

Temperature-phased anaerobic co-digestion of food waste and paper waste with and without recirculation: Biogas production and microbial structure.

Two temperature-phased anaerobic digestion (TPAD) systems (55 °C in the first reactor and 35 °C in the second reactor) with and without recirculation were operated in parallel for the co-digestion of food waste and paper waste. A long-term experiment was carried out for these two systems with the paper waste ratios elevated from 0 to 50%. The removal efficiencies of COD, TS, VS, carbohydrate and protein in the recirculated TPAD system were higher than those of the non-recirculated system. The successful acclimation of thermophilic cellulose-degrading bacteria in the first reactor (RT1), partly due to recirculation, ensured the effective degradation of cellulose when the paper waste ratio was higher than 40%, resulting in the production of large amounts of hydrogen in reactor RT1. In the absence of recirculation, the main substance produced in the first reactor of the non-recirculated system (T1) was lactic acid. This gradually led to over-acidification and a low degradation efficiency and no methane or hydrogen was produced in T1. Recirculation helped to establish a stable bacterial community capable of producing bio-hydrogen in reactor RT1. The relatively low pH of 5.5 in the RT1 inhibited the activity of hydrogenotrophic archaea without consuming hydrogen, facilitating high hydrogen production levels.

Application of two anaerobic membrane bioreactors with different pore size membranes for municipal wastewater treatment.

Pore size is one of the most important properties in the successful operation of membrane-based bioprocesses for the treatment of municipal wastewater. The characteristics of two anaerobic membrane bioreactors (AnMBRs), one with a hollow fiber membrane of 0.4 µm pore size (AnMBR1), and the other with a membrane of 0.05 µm pore size (AnMBR2) were investigated for the treatment of real municipal wastewater at room temperature (25 °C) under varied hydraulic retention times (HRTs). Process performance was evaluated in terms of organic removal efficiency, biogas production and membrane filtration behaviours during a long-term continuous operation. Both AnMBRs showed good organic removal performance with COD and BOD removal efficiencies of around 89% and 93%, respectively. High energy recovery potential was achieved, with the biogas yield ranging between 0.20 and 0.26 L-gas/g-CODrem and a methane content of approximately 75%. The differences in the membrane filtration behaviours in the two AnMBRs included different permeate flux and total filtration resistance (Rt). In the AnMBR with a 0.4 µm pore size membrane, an average Rt of 1.08 × 10^12 m-1 was obtained even when the permeate flux was a high 0.274 m/day, while a higher average Rt of 1.51 × 10^12 m-1 was observed in the AnMBR with 0.05 µm pore size membrane even when the flux was a low 0.148 m/day. The off-line membrane cleaning strategy used for AnMBR1 indicated that the membrane restoration efficiency was 90.2%.

Response of morphology and microbial community structure of granules to influent COD/SO42 - ratios in an upflow anaerobic sludge blanket (UASB) reactor treating starch wastewater.

Biochemical properties of granules are of vital importance to UASB performance. This study characterized the granules cultivated at different COD/SO42- ratios to elucidate the influence of sulfidogenesis on starch wastewater (1000 mg-COD L-1) biodegradation kinetics and process stability. Suitable sulfate addition enriched granular microecosystems and stimulated the secretion of extracellular substances, facilitating cells cohersion and sludge aggregation. The percentage of granules larger than 2.8 mm increased from <10.0% to 58.8-69.4% with decreasing COD/SO42- ratio from 10 to 2. Starch-fed granules tended to grow flagella-like filaments on the surface. The filaments overwhelmed by hydrophilic biopolymers had high affinity for biogas-bubbles and water-molecules aggravating granule floatation and washout. 16 s rRNA gene analysis revealed that decreasing COD/SO42- ratio shifted Syntrophobacterales to Desulfovibrio, which co-worked with Methanosaeta while suppressing Methanobacterium thereby altering starch bioconversion routes. Decrease in Syntrophobacterales caused propionate accumulation and slight process upset.

Sulfidogenesis process to strengthen re-granulation for biodegradation of methanolic wastewater and microorganisms evolution in an UASB reactor.

A lab-scale methanolic wastewater-fed (3000 mg COD L-1) UASB reactor was operated for 235 days to evaluate the influence of the sulfidogenesis process on metabolic routes, the re-granulation of dispersed granules and long-term process performance. Various sulfidogenesis scenarios were created by stepwise decreasing the influent COD/SO42- ratio from 20 to 0.5 at a fixed organic loading rate (OLR) of 12 g COD L-1 d-1. It was shown that the conversion of methanol to methane was stable at a wide COD/SO42- range of ≥2, attaining high biogas production rate of 3.78 ± 0.32 L L-1 d-1 with efficient concurrent removal of the total COD (96.5 ± 4.4%) and sulfate (56.3 ± 13.0%). The methane content in biogas remained relatively stable at 81.5 ± 1.6% for all COD/SO42- ratios tested. The particle size of the granules was shown to clearly increase as the COD/SO42- ratios decreased. A slight linear decline was noted in the number of electrons utilized by methane producing archaea (MPA) (from 98.5 ± 0.5% to 80.0 ± 2.4%), whereas consumption by sulfate reducing bacteria (SRB) increased (from 1.5 ± 0.5% to 20.0 ± 2.4%) with the decreasing COD/SO42- ratio. According to the results of activity tests and microbial community analysis, the conversion of methanol to methane at a low COD/SO42- ratio, except from Methanomethylovorans sp., depends not only on low levels of acetoclastic and hydrogenotrophic methanogens, but also on incomplete oxidizer SRB species (e.g. Desulfovibrio sp.) that utilize H2-CO2 with acetate to mineralize the methanol. This serves to diversify the metabolic pathway of methanol. Further analysis through scanning electron microscopy (SEM) revealed that a lower COD/SO42- ratio favored the sulfidogenesis process and diversified the microbial community inside the reactor. The benefical sulfidogenesis process subsequently invoked the formation of a sufficient, rigid [-Fe-EPS-]n network (EPS: extracellular polymeric substances), binding and immobilizing the sludge, and resulting in the re-granulation of the dispersed granules.

Electro-Fenton oxidation of coking wastewater: optimization using the combination of central composite design and convex optimization method.

The electro-Fenton treatment of coking wastewater was evaluated experimentally in a batch electrochemical reactor. Based on central composite design coupled with response surface methodology, a regression quadratic equation was developed to model the total organic carbon (TOC) removal efficiency. This model was further proved to accurately predict the optimization of process variables by means of analysis of variance. With the aid of the convex optimization method, which is a global optimization method, the optimal parameters were determined as current density of 30.9 mA/cm2, Fe2+ concentration of 0.35 mg/L, and pH of 4.05. Under the optimized conditions, the corresponding TOC removal efficiency was up to 73.8%. The maximum TOC removal efficiency achieved can be further confirmed by the results of gas chromatography-mass spectrum analysis.

Effect of influent COD/SO4(2-) ratios on biodegradation behaviors of starch wastewater in an upflow anaerobic sludge blanket (UASB) reactor.

A lab-scale upflow anaerobic sludge blanket (UASB) has been run for 250days to investigate the influence of influent COD/SO4(2-) ratios on the biodegradation behavior of starch wastewater and process performance. Stepwise decreasing COD/SO4(2-) ratio enhanced sulfidogenesis, complicating starch degradation routes and improving process stability. The reactor exhibited satisfactory performance at a wide COD/SO4(2-) range ⩾2, attaining stable biogas production of 1.15-1.17LL(-1)d(-1) with efficient simultaneous removal of total COD (73.5-80.3%) and sulfate (82.6±6.4%). Adding sulfate favored sulfidogenesis process and diversified microbial community, invoking hydrolysis-acidification of starch and propionate degradation and subsequent acetoclastic methanogenesis; whereas excessively enhanced sulfidogenesis (COD/SO4(2-) ratios <2) would suppress methanogenesis through electrons competition and sulfide inhibition, deteriorating methane conversion. This research in-depth elucidated the role of sulfidogenesis in bioenergy recovery and sulfate removal, advancing the applications of UASB technology in water industry from basic science.

Operation performance and granule characterization of upflow anaerobic sludge blanket (UASB) reactor treating wastewater with starch as the sole carbon source.

Long-term performance of a lab-scale UASB reactor treating starch wastewater was investigated under different hydraulic retention times (HRT). Successful start-up could be achieved after 15days' operation. The optimal HRT was 6h with organic loading rate (OLR) 4g COD/Ld at COD concentration 1000mg/L, attaining 81.1-98.7% total COD removal with methane production rate of 0.33L CH4/g CODremoved. Specific methane activity tests demonstrated that methane formation via H2-CO2 and acetate were the principal degradation pathways. Vertical characterizations revealed that main reactions including starch hydrolysis, acidification and methanogenesis occurred at the lower part of reactor ("main reaction zone"); comparatively, at the up converting acetate into methane predominated ("substrate-shortage zone"). Further reducing HRT to 3h caused volatile fatty acids accumulation, sludge floating and performance deterioration. Sludge floating was ascribed to the excess polysaccharides in extracellular polymeric substances (EPS). More efforts are required to overcome sludge floating-related issues.