城市污水生物处理系统中微生物酶的活性及其分布

通过水解模式底物进行分光光度测定的方法,研究了城市污水生物处理系统中微生物酶的活性.选取亮氨酸氨基肽酶、β-葡萄糖苷酶、碱性磷酸酶和脂酶等胞外酶进行分析,结果发现4种胞外酶的平均活性在31.O～88.5 μmol/(L·h),其中76.9%～94.8%的酶活性分布在活性污泥中,说明绝大部分胞外酶是与细胞相连或固定在细胞外多聚絮体基质里.混合水样和出水水样中的DOC几乎相等,而各胞外酶与DOC不存在任何显著的相关关系.增加NO3-浓度可提高亮氨酸氨基肽酶和β-葡萄糖苷酶的活性,但对碱性磷酸酶和脂酶却有明显的抑制作用.     

Responses of a soil bacterium, Pseudomonas chlororaphis O6 to commercial metal oxide nanoparticles compared with responses to metal ions

The toxicity of commercially-available CuO and ZnO nanoparticles (NPs) to pathogenic bacteria was compared for a beneficial rhizosphere isolate, Pseudomonas chlororaphis O6. The NPs aggregated, released ions to different extents under the conditions used for bacterial exposure, and associated with bacterial cell surface. Bacterial surface charge was neutralized by NPs, dependent on pH. The CuO NPs were more toxic than the ZnO NPs. The negative surface charge on colloids of extracellular polymeric substances (EPS) was reduced by Cu ions but not by CuO NPs; the EPS protected cells from CuO NPs-toxicity. CuO NPs-toxicity was eliminated by a Cu ion chelator, suggesting that ion release was involved. Neither NPs released alkaline phosphatase from the cells’ periplasm, indicating minimal outer membrane damage. Accumulation of intracellular reactive oxygen species was correlated with CuO NPs lethality. Environmental deposition of NPs could create niches for ion release, with impacts on susceptible soil microbes.     

Chitosan/albumin/CaCO3 as mimics for membrane bioreactor fouling: genesis of structural mineralized-EPS-building blocks and cake layer compressibility

Membrane bioreactor biofouling is usually described as an extracellular matrix in which biopolymers, inorganic salts and active microbes co-exist. For that reason, biomineralization (BM) models can be useful to describe the spatial organization and environmental constraints within the referred scenario. BM arguments were utilized as background in order to (1) evaluate CaCO₃ influence on flux decline; pore blocking and cake layer properties (resistance, permeability and compressibility) in a wide range of Chitosan/Bovine serum albumin (BSA) mixtures during step-pressure runs and, (2) perform membrane autopsies in order to explore the genesis of mineralized extracellular building blocks (MEBB) during cake layer build up. Using low molecular weight chitosan (LC) and BSA, 2 L of 5 LC/BSA mixtures (0.25-1.85 ratio) were pumped to an external ultra filtration (UF) membrane (23.5 cm², hydrophobic, piezoelectric, 100 kDa as molecular weight cut-off). Eight different pressure steps (40 ± 7 to 540 ± 21 kPa) were applied. Each pressure step was held for 900 s. CaCO₃ was added to LC/BSA mixtures at 0.5, 1.5 and 3 mM in order to create MEBB during the filtration tests. Membrane autopsies were performed after the filtration tests using thermo gravimetric, scanning microscopy and specific membrane mass (mg cm⁻²) analyses. Biopolymer-CaCO₃ step-pressure filtration created compressible cake layers (with inner voids). The formation of an internal skeleton of MEBB may contribute to irreversible fouling consolidation. A hypothesis for MEBB genesis and development was set forth.     

Quantification of the interactions between Ca,Hg and extracellular polymeric substances (EPS) of sludge

The interactions between metals (Ca²⁺ and Hg²⁺) and extracellular polymeric substances (EPS) extracted from the aerobic and anaerobic sludge in wastewater treatment reactors were investigated using a combination of zeta potential measurement and 3-dimensional excitation–emission matrix (EEM) fluorescence spectroscopy with parallel factor (PARAFAC) analysis. Results show that Ca²⁺ had no substantial effects on the EEM fluorescence spectra of the EPS, but their zeta potentials increased with the increasing Ca²⁺ dosage. However, Hg²⁺ had a significant effect on the EEM fluorescence spectra of the EPS, while their zeta potentials seemed not to be affected by the dose of Hg²⁺. The interactions between Hg²⁺ and EPS were elucidated using the fluorescence quenching with PARAFAC analysis, while the interactions between Ca²⁺ and EPS were evaluated by the zeta potential technique. The binding constants for Hg²⁺ and EPS were two orders of magnitude higher than those for Ca²⁺ and EPS, suggesting that the binding mechanisms between Ca²⁺ and EPS were different from those between Hg²⁺ and EPS. The results might be useful for understanding the roles of EPS in bacterial self-protection against heavy metals and the aggregate formation mechanisms through ionic bridging interactions.     

Extracellular polymeric substance induces biogenesis of vivianite under inorganic phosphate-free conditions

Vivianite is often found in reducing environments rich in iron and phosphorus from organic debris degradation or phosphorus mineral dissolution. The formation of vivianite is essential to the geochemical cycling of phosphorus and iron elements in natural environments. In this study, extracellular polymeric substances (EPS) were selected as the source of phosphorus. Microcosm experiments were conducted to test the evolution of mineralogy during the reduction of polyferric sulfate flocs (PFS) by Shewanella oneidensis MR-1 (S. oneidensis MR-1) at EPS concentrations of 0, 0.03, and 0.3 g/L. Vivianite was found to be the secondary mineral in EPS treatment when there was no phosphate in the media. The EPS DNA served as the phosphorus source and DNA-supplied phosphate could induce the formation of vivianite. EPS impedes PFS aggregation, contains redox proteins and stores electron shuttle, and thus greatly promotes the formation of minerals and enhances the reduction of Fe(III). At EPS concentration of 0, 0.03, and 0.3 g/L, the produced HCl-extractable Fe(II) was 107.9, 111.0, and 115.2 mg/L, respectively. However, when the microcosms remained unstirred, vivianite can be formed without the addition of EPS. In unstirred systems, the EPS secreted by S. oneidensis MR-1 could agglomerate at some areas, resulting in the formation of vivianite in the proximity of microbial cells. It was found that vivianite can be generated biogenetically by S. oneidensis MR-1 strain and EPS may play a key role in iron reduction and concentrating phosphorus in the oligotrophic ecosystems where quiescent conditions prevail.     

Degradation of extracellular genomic,plasmid DNA and specific antibiotic resistance genes by chlorination

Extracellular DNA structure damaged by chlorination was characterized. Integrity of extracellular ARG genetic information after chlorination was determined. Typical chlorine doses will likely effectively diminish extracellular DNA and ARGs. Plasmid DNA/ARGs were less readily broken down than genomic DNA. The Bioanalyzer methodology effectively documented damage incurred to DNA. There is a need to improve understanding of the effect of chlorine disinfection on antibiotic resistance genes (ARGs) in order to advance relevant drinking water, wastewater, and reuse treatments. However, few studies have explicitly assessed the physical effects on the DNA. Here we examined the effects of free chlorine (1–20 mg Cl₂/L) on extracellular genomic, plasmid DNA and select ARGs. Chlorination was found to decrease the fluorometric signal of extracellular genomic and plasmid DNA (ranging from 0.005 to 0.05 mg/mL) by 70%, relative to a no-chlorine control. Resulting DNA was further subject to a fragment analysis using a Bioanalyzer, indicating that chlorination resulted in fragmentation. Moreover, chlorine also effectively deactivated both chromosomal- and plasmid-borne ARGs, mecA and tetA, respectively. For concentrations >2 mg Cl₂//L × 30 min, chlorine efficiently reduced the qPCR signal when the initial concentration of ARGs was 10⁵ copies/mL or less. Notably, genomic DNA and mecA gene signals were more readily reduced by chlorine than the plasmid-borne tetA gene (by ~2 fold). Based on the results of qPCR with short (~200 bps) and long amplicons (~1200 bps), chlorination could destroy the integrity of ARGs, which likely reduces the possibility of natural transformation. Overall, our findings strongly illustrate that chlorination could be an effective method for inactivating extracellular chromosomal- and plasmid-borne DNA and ARGs.     

Relation between extracellular polymers' composition and its ability to complex Cd,Cu and Pb

Activated sludges originated from wastewater treatment plants (WWTPs) play an important role in heavy metal removal from effluents. Extracellular polymers (ECP) form a major part of the activated sludge and are heavily involved in biosorption of heavy metals. The complexation of three heavy metals (Cd, Cu and Pb) with ECP extracted from six activated sludges, originated from different WWTPs, was investigated at pH 7.

ECP in the study were shown to be mainly composed of proteins, humic acids, uronic acids and polysaccharides along with smaller amounts of lipids and nucleic acids. IR spectra confirmed the presence of the functional groups usually found in the ECP and the content in each fraction was determined using colorimetric methods. The determination of surface charge was carried out on each ECP sample and allowed two pK_a values characteristic of two distinctive functional groups to be determined. At the pH used in the study, the value of these constants indicates that only one functional group is under protonated form.

A polarographic method was used to determine the complexation parameters (number of binding sites and complexation constant) of ECP solutions towards metals. The following orders were established for the number of binding sites: Cu > Pb  Cd and for the stability of the ECP–metal complex: Cd > Pb  Cu.

A matrix of correlation between the composition of the polymers and the complexation parameters showed that the number of binding sites and the complexation constant were strongly linked to proteins, polysaccharides and humic substances content.     

Nitrate removal and extracellular polymeric substances of autohydrogenotrophic bacteria under various pH and hydrogen flow rates

In recent years there has been an increasing interest in the use of autohydrogenotrophic bacteria to treat nitrate from wastewater. However, our knowledge about the characteristics of extracellular polymeric substances (EPS) releasing by these activities is not yet very advanced. This study aimed to investigate the change in EPS compositions under various pH values and hydrogen flow rates, taking into consideration nitrogen removal. Results showed that pH 7.5 and a hydrogen flow rate of 90 mL/min were the optimal operating conditions, resulting in 100% nitrogen removal after 6 hr of operation. Soluble and bound polysaccharides decreased, while bound proteins increased with increasing pH. Polysaccharides increased with increasing hydrogen flow rate. No significant change of bound proteins was observed at various hydrogen flow rates.     

Purification and molecular weight distribution of a key exopolysaccharide component of Bacillus megaterium TF10

Extracellular polymeric substances (EPS) are organic metabolic compounds excreted by microorganisms. They largely impact microbial aggregate structures and functions. Extracellular polysaccharides (EP) in EPS are responsible for the formation of microbial aggregates. In this work, we successfully separated and characterized EP from EPS of the bacterium Bacillus megaterium TF10. Extraction of EP from EPS was optimized using Sevag''s reagent. Chemical characteristics, functional groups, and molecular weight (MW) distribu-tion of EP were compared with the harvested EPS and soluble microbial products (SMP). We found that the polymers of lower MW and free proteins were successfully removed by Sevag''s reagent. The higher MW components of EPS were predominantly polysaccharides, while the polymers of lower MW tended to secrete to the supernatant and were described as SMP. A part of the proteins in the EP was polysaccharide-bonded. Our results can be further used in elucidating the complex flocculation mechanisms in which EP play a major role.     

Insights into sludge granulation during anaerobic treatment of high-strength leachate via a full-scale IC reactor with external circulation system

In this study, a full-scale internal circulation(IC) reactor coupled with an external circulation system was developed to treat high-strength leachate from a municipal solid waste(MSW)incineration plant, in which anaerobic sludge granulation was intensively investigated. Results showed that the IC reactor achieved excellent treatment performance under high organic loading rates(OLR) of 21.06–25.16 kg chemical oxygen demand(COD)/(m3? day). The COD removal efficiency and biogas yield respectively reached 89.4%–93.4% and 0.42–0.50 m3/kg COD.The formation of extracellular polymeric substances(EPS) was closely associated with sludge granulation. Protein was the dominant component in sludge EPS, and its content was remarkably increased from 21.6 to 99.7 mg/g Volatile Suspended Solid(VSS) during the reactor operation. The sludge Zeta potential and hydrophobicity positively correlated with the protein/polysaccharide ratio in EPS, and they were respectively increased from-26.2 m V and 30.35% to-10.6 m V and 78.67%, which was beneficial to microbial aggregation. Three-dimensional fluorescence spectroscopy(3 D-EEM) and Fourier transform infrared spectroscopy(FT-IR)analysis further indicated the importance of protein-like EPS substances in the sludge granulation. Moreover, it was also found that the secondary structures of EPS proteins varied during the reactor operation.