Development of a growth-coupled selection platform for directed evolution of heme biosynthetic enzymes in Corynebacterium glutamicum.

Heme is an important tetrapyrrole compound, and has been widely applied in food and medicine industries. Although microbial production of heme has been developed with metabolic engineering strategies during the past 20 years, the production levels are relatively low due to the multistep enzymatic processes and complicated regulatory mechanisms of microbes. Previous studies mainly adopted the strategies of strengthening precursor supply and product transportation to engineer microbes for improving heme biosynthesis. Few studies focused on the engineering and screening of efficient enzymes involved in heme biosynthesis. Herein, a growth-coupled, high-throughput selection platform based on the detoxification of Zinc-protoporphyrin IX (an analogue of heme) was developed and applied to directed evolution of coproporphyrin ferrochelatase, catalyzing the insertion of metal ions into porphyrin ring to generate heme or other tetrapyrrole compounds. A mutant with 3.03-fold increase in k cat/K M was selected. Finally, growth-coupled directed evolution of another three key enzymes involved in heme biosynthesis was tested by using this selection platform. The growth-coupled selection platform developed here can be a simple and effective strategy for directed evolution of the enzymes involved in the biosynthesis of heme or other tetrapyrrole compounds.

Assessment and optimization of the oxygen based membrane biofilm reactor as a novel technology for source-diverted greywater treatment.

The oxygen based membrane biofilm (O2-MBfR) has been proved to be a novel technology in treating greywater (GW) and response surface methodology (RSM) was used to model the removal of chemical oxygen demand (COD) and total nitrogen (TN) with operation parameters COD/TN ratio, system pH and lumen air pressure (LAP). Results indicated that the all target single factors affect GW treatment efficiency, and the regression model with central composite design (CCD) showed good agreement with the experimental results with high R2 and R2 adj values (all >0.97) for all the target responses. Statistical evaluation revealed that system pH was the most significant parameter affecting COD and TN removal, followed by COD/TN ratio and LAP. The interaction between COD/TN ratio and system pH also played an important role on the GW treatment. The optimized maximum removal of COD (96.48%) and TN (133 g N/m2-day) were achieved with the COD/TN ratio 17.76 g COD/g TN, system pH 7.10 and LAP 1.00 psi. Thus, RSM combined with CCD could be used for predicting the organics and nitrogen removal during GW treatment in the O2-MBfR.

Explore the sludge stabilization process in sludge drying bed by modeling study from mesocosm experiments.

Sludge drying bed (SDB) treatment is a valuable alternative to conventional sludge treatment methods. However, changes in sludge hydrotexture during dewatering present a barrier for direct modeling of the SDB process. This study proposes a modeling strategy to simulate the sludge stabilization process in SDB treatment by separating sludge dewatering and sludge solids stabilization into independent processes. Two cell decay theories widely used by activated sludge models (ASM), death-regeneration concept and endogenous respiration theory, are compared to describe the biokinetic processes of sludge digestion. Both cell decay theories are found to adequately describe effluent total COD, NH4-N, NO3-N, and sludge layer composition, but have pronounced differences in describing effluent COD compositions. Results show that natural aeration does not maintain adequate aerobic/anoxic sludge digestion within the sludge layer to fully nitrify NH4-N released by cell decay. Results also indicate that the kinetics of sludge digestion are adaptable over time, indicating the need to adopt lumped values for biokinetic simulations. While lowered sludge dewatering rates (outflow) can increase biodegradable COD for cell metabolism, increased sludge loading rates (inflow) lead to higher effluent COD and NH4-N concentrations. Contrary to conventional judgement, this study demonstrates the merit of sludge layer formation to reduce leaching loss of biodegradable COD. Overall, the proposed modeling strategy is proven capable of simulating deposited sludge digestion processes in an SDB.

Nanometer CeO2 doped high silica ZSM-5 heterogeneous catalytic ozonation of sulfamethoxazole in water.

A novel CeO2 doped high silica ZSM-5(CeO2@HSZSM-5) composite was originally fabricated via ammonia precipitation for the catalytic ozonation of sulfamethoxazole (SMX). Physicochemical properties have been investigated through electron microscope, Raman spectroscopy, X-ray photoelectron spectroscopy, etc. The prepared nanometer CeO2@HSZSM-5 had a much higher specific surface (348-395 m2/g), a finer crystallite size (8.2-33.5 nm) and superior stability. Temperature-programmed desorption and reduction analysis revealed that the formed CeO2 nanoparticles on the surface of CeO2@HSZSM-5 could improve the reducibility of surface-capping oxygen, induce more oxygen vacancies and promote oxygen migration. CeO2@HSZSM-5 exhibited excellent catalytic performance for SMX mineralization in the pH range of environmental waters. The great enhancement of CeO2@HSZSM-5 catalytic activity was ascribed to the conversion of O3 into active oxygen involved in SMX mineralization, including .OH, O2.- and 1O2. This work provides a reference for the removal of pollutants by zeolite supported Ce catalytic ozonation process in water.

Managing biofilm growth and clogging to promote sustainability in an intermittent sand filter (ISF).

The sustainability of rural sanitation includes the long-term welfare of both rural and urban societies. As a commonly used rural sanitation technology, operation of intermittent sand filters (ISF) is impacted by biofilm clogging inside the ISF. In this study ISF performance is studied at low hydraulic loading rates (HLR) to explore the interaction between biofilm growth and wastewater treatment efficiency. CW2D/HYDRUS, a simulation model which does not include media hydraulic property changes caused by biofilm growth, is utilized as a numerical control to contrast the effects of biofilm growth inside an experimental ISF. A paired experiment with simulation demonstrate that biofilm clogging comprised dominantly of heterotrophs occurred in the top layers of the ISF. Lowered HLR slows clogging development but not final clogging extent. The biofilm clogging development zone offers adequate removal of applied biodegradable COD and NH4+ - N. However, the spatial distribution of heterotrophs and biodegradable COD does not match the denitrification requirement of the resulting NO3- - N. A simultaneous nitrification and denitrification (SND) potential is manifested in the clogging development zone, but lowered HLR reduces media moisture level to a less favorable level for denitrification. Furthermore, slowed water movement under lower HLR aggravates the accumulation of NO3- - N, which can potentially result in counterproductive salt accumulation. Since biofilm growth is a natural and self-adaptive response to wastewater application, this study suggests accepting limited, managed biofilm growth and clogging in ISFs. In addition, this study calls for further research to manage biofilm growth and clogging for long-term ISF sustainability.

Effects of loading rates and plant species on sludge characteristics in earthworm assistant sludge treatment wetlands.

Sludge treatment wetlands (STWs) are widely used to treat surplus sludge in recent years. However, the effects of plant species and loading rates on sludge characteristics in earthworm assistant STWs remain unclear. In the current study, six STWs planted with two plant species (Phragmites australis, Typha angustifolia) were investigated under four loading rates (60, 80, 90 and 120 kg DS/m2/yr) regarding the influence on sludge characteristics. Furthermore, earthworms were added in three STWs to evaluate their role on sludge stabilization during resting period. Results showed that the best sludge dewatering (dry solids (DS) of 45.0%) and stabilization (volatile solids to total solids (VS/TS) of 40.5%) were determined in the P. australis STWs at the loading rate of 80 kg DS/m2/yr. Furthermore, VS/TS and Escherichia coli contents in earthworm STWs were 5.5-11.2% and 12-39% lower than that in the control without earthworm addition. Meanwhile, earthworm also decreased the nutrient contents in STWs. However, earthworms had insignificant effects on heavy metal contents in STWs. Nevertheless, the bioavailability of Cd and Cr in STWs were decreased by earthworm addition, with an acid-soluble fraction of Cd and Cr reduced by 11.2-18% and 2.5-7.5%, respectively. In conclusion, sludge characteristics can be improved by earthworm addition in P. australis STWs.

Sulfonamide antibiotics sorption by high silica ZSM-5: Effect of pH and humic monomers (vanillin and caffeic acid).

Sulfonamides (SAs) in the aquatic environment are a serious threat to human health, environmental ecology and water reuse. Natural organic matter and SAs are known to compete for adsorption sites on adsorbent. SAs adsorption by high silica ZSM-5 (HSZSM-5) affected by humic monomers remains unclear. The impact of representative humic monomers (HMs) (caffeic acid (CA) and vanillin (VNL)) on the sorption of SAs by HSZSM-5 at different pH was evaluated for the first time. Fourier transform infrared and X-ray photoelectron spectroscopy were used to obtain the group transformations. The presence of HM did not change the main adsorption mechanism of SAs on pristine and loaded HSZSM-5 (pH-dependent and hydrophobic interaction). Lewis acid-base electron interaction, π-π electron donor-acceptor (EDA), and H-bonding had a positive contribution to the adsorption of SAs by HSZSM-5 after HM coating. However, HM coating and competition reduced the adsorption capacity of HSZSM-5 towards SAs. In co-introduction system of HM and SAs, the reaction between CO in HM and N-H in SAs resulted in SAs and HM co-adsorbed on HSZSM-5. Meanwhile, CA concentration <20 mg/L was in favor of SMX adsorption. The findings give an insight into the interactions among HM and SAs in high silica zeolites adsorption system, thereby improving SAs removal in actual water.

Removal of sulfonamide antibiotics from water by high-silica ZSM-5.

Adsorption characteristics of high-silica zeolites (HSZSM-5) for two selected sulfonamide antibiotics (SAs) (sulfamethoxazole and sulfadiazine) were investigated. The SAs were almost completely (>90%) removed from the water by HSZSM-5. Adsorption followed second-order kinetics with liquid-film diffusion as the dominant mechanism. SA adsorption capacity on high-silica zeolites was examined in terms of pH, temperature, and the presence of natural organic matter (NOM). HSZSM-5 had better adsorption performance in acidic conditions, and the apparent distribution coefficient indicated that SA0 species were the major contribution to the overall adsorption at pH of 2-10. Adsorption of SAs on HSZSM-5 was a spontaneous and exothermic physisorption process. SA removal by HSZSM-5 was a mixed mechanism through ion-exchange and hydrophobic interaction. HSZSM-5 has potential application prospects in removing SAs from wastewater.

Model-based evaluation of fouling mechanisms in powdered activated carbon/membrane bioreactor system.

Identifying the fouling degree of a membrane bioreactor (MBR) provides guidance on the determination of suitable membrane cleaning methods. There is still a lack of knowledge on the effects of powdered activated carbon (PAC) refresh ratio on the MBR fouling mechanism. Major fouling mechanisms of an MBR with constant flow rate at different PAC replenishment ratios were investigated by individual and combined mechanistic fouling models. The root mean square errors were employed to assess the prediction accuracy of the used fouling models. The combined models showed better prediction. The cake-complete model provided far better fits of the transmembrane pressure data, and provided good fits of other individual model predictions regardless of the PAC refreshment ratio. Fourier transform infrared spectroscopy, scanning electron microscopy and energy dispersive X-ray spectroscopy confirmed that the cake layer clogging was the main fouling mechanism followed by complete blockage and standard plugging. The cake-complete model may be used to predict the fouling mechanisms in PAC/MBR systems.

Purification of leachate from sludge treatment beds by subsurface flow constructed wetlands: effects of plants and hydraulic retention time.

Sludge treatment beds (STBs) have been used widely in many countries due to low energy consumption, low operating and maintenance costs, and better environmental compatibility. Penetration, evaporation, and transpiration are the main processes for sludge dewatering in STBs. However, the leachate quality from STBs usually cannot meet discharge limits. Moreover, such leachate has very low COD/N ratio, which makes it difficult to treat. In the present study, two subsurface flow (SSF) constructed wetlands (CWs) were investigated for the treatment of leachate from STBs under three different hydraulic retention time (HRT) (3 days, 4 days, 6 days), aiming for evaluating the effects of plants and HRT on treatment performance, as well as the potential of SSF CWs to treat sludge leachate with low COD/N ration. The results showed that plants play an important role in leachate treatment. The best treatment performance was achieved with HRT of 4 days. In this condition, the mean removal efficiencies of COD (chemical oxygen demand), NH4+-N, TN (total nitrogen), and TP (total phosphorus) in the planted and the unplanted CWs were 61.6% (unplanted - 3.7%), 76.6% (unplanted 43.5%), 70% (unplanted 41%), and 65.6% (unplanted 6%), respectively. Heavy metal concentrations were below the Chinese integrated wastewater discharge standard during the experimental period in the planted CW, and the removal efficiencies in the planted CW system were higher than in the unplanted CW system. In all, planted SSF CWs can be an effective approach in removing leachate from sludge treatment beds. Furthermore, considering to temperature and seasonal variation, the leachate from STBs needs to be further studied in pilot- and full-scale condition.

Effect of powdered activated carbon dosage on sludge properties and membrane bioreactor performance in a hybrid MBR-PAC system.

An improved insight into the effect of powdered activated carbon (PAC) on membrane fouling is crucial to the MBR performance. Sludge key property, soluble microbial products (SMP) and transmembrane pressure (TMP) were monitored. The membrane fouling rate in the MBRs was also analyzed based on TMP profile and resistance-in-series model. PAC reduced the membrane filtration resistance and significantly decreased the fouling rate. The sludge filterability was improved by extending the filtration time by almost twofold. PAC affected the SMP release and protein/polysaccharide (carbohydrate) was in a lower ratio. Fourier transform infrared (FTIR) analysis indicated that PAC decreased the impact of organic carbon, and reduced the proteins' and polysaccharides' absorption and deposition on the membrane surface and in the pores. The degree of reversible and irreversible fouling was related to the PAC content added into the MBRs. At the optimum dosage of 2 g/L, the results signified the PAC potential as a mitigation strategy of membrane fouling.

Identification of fouling mechanisms in MBRs at constant flowrate: model applications and SEM-EDX characterizations.

A fundamental understanding of fouling mechanisms is critical to improving filtration operations. The performance of four parallel membrane bioreactors (MBRs) with different sludge retention times (SRTs) was monitored during long-term constant flowrate filtration. The characteristics of the membrane and transmembrane pressure (TMP) profiles obtained were studied to demonstrate fouling mechanisms. Both classical blocking models and their combined models were evaluated. The intermediate model provided very good agreement with all the TMP data. However, the combined cake-intermediate and intermediate-standard models were more effective in the description of the experimental data. Contributions analysis indicated that the cake, intermediate and standard blocking models were the dominant fouling mechanisms. Scanning electron microscopy and energy dispersive X-ray (SEM-EDX) imaging showed that cake blocking by organic matter and standard blocking by inorganic matter made the main contributions to membrane fouling. The combined cake-intermediate and intermediate-standard models may be applicable to systems where these two models are consistent with the experimentally observed fouling mechanisms in an MBR.

Fouling modeling of the mixed liquor in MBR by the individual and combined models.

To predict the membrane fouling phenomena in the membrane filtration operation, the individual models derived from Darcy's law and the corresponding combined models were employed to investigate the kind of models that provided better fits. The filtration of the mixed liquid from membrane bioreactors with different sludge retention time (SRT) at a constant pressure was carried out. The variation of applied pressure had significantly effect on the kind of the individual model provided better fit for the data at longer SRT and less effect for the data at shorter SRT, though it had less effect on the kind of the combined model that provided better fit. The kind of model that provided better fit did not change when the concentration of the diluted mixed liquor was at a certain range, even though the dilution ratio would lead to the variation of the prediction results. The cake-standard model and the complete-standard model provided good fits at different pressure and at different dilution ratio, respectively. The cake-standard and complete-standard models may be applicable to systems where these models are consistent with the experimentally observed fouling mechanisms.

Fundamental characteristics study of anion-exchange PVDF-SiO(2) membranes.

A new type of poly(vinylidene fluoride)(PVDF)-SiO(2) hybrid anion-exchange membrane was prepared by blending method. The anion-exchange groups were introduced by the reaction of epoxy groups with trimethylamine (TMA). Contact angle between water and the membrane surface was measured to characterize the hydrophilicity change of the membrane surface. The effects of nano-sized SiO(2) particles in the membrane-forming materials on the membrane mechanical properties and conductivity were also investigated. The experimental results indicated that PVDF-SiO(2) anion-exchange membranes exhibited better water content, ion-exchange capacity, conductivity and mechanic properties, and so may find potential applications in alkaline membrane fuel cells and water treatment processes.

Comparative performance between intermittently cyclic activated sludge-membrane bioreactor and anoxic/aerobic-membrane bioreactor.

A process of intermittently cyclic activated sludge-membrane bioreactor (ICAS-MBR) was developed to enhance the performance of biological phosphorus removal (EBPR), which was constructed under sequencing anoxic/anaerobic/aerobic condition. The performance between ICAS-MBR and conventional anoxic/aerobic-membrane bioreactor (A/O-MBR) in terms of phosphorus removal, nitrification performance and microbial aspects were investigated in this study. The experimental results indicated that the phosphorus removal efficiency in ICAS-MBR process increased from 65% to 83% when compared with A/O-MBR. It was also found that the COD removal efficiencies of the two processes were over 94%, and NH(4)(+)-N and TN average removal efficiencies were 96% and 69% in ICAS-MBR and 96% and 78% in A/O-MBR, respectively. Furthermore, micrographs observation obtained confirmed the succession and diversification of microorganisms in the two systems followed a similar regularity.