Fluorecent staining for study of extracellular polymeric substances in membrane biofouling layers

Membrane biofouling by microbial products adversely impacts the feasibility of adopting membrane bioreactors (MBRs) for treating wastewater. The fouling layer structure determines the pressure drop across the fouling layer. Three-dimensional distributions of nucleic acids, proteins, alpha-D-GLUCOPYRANOSE POLYSACCHARIDES, AND B-D-glucopyranose polysaccharides in the fouling layer formed on a mixed cellulose ester membrane were generated utilizing a quadruple staining protocol combined with confocal laser scanning microscopy (CLSM). For the first time, this study constructed a three-dimensional volumetric grids model representing the fouling layer structure on the basis of a series of CLSM images. Quantitative structural information about the fouling layer was extracted from the CLSM images.     

Influence of extracellular polymeric substances on Pseudomonas aeruginosa transport and deposition profiles in porous media

The impact of cell surface extracellular polymeric substances (EPS) on bacterial transport and retention profiles was investigated in saturated columns packed with glass beads. Three genetically well-defined isogenic Pseudomonas aeruginosa strains with different EPS secretion capability and EPS composition were used to systematically examine their deposition behavior over a range of solution chemistry. The presence of EPS on nonmucoid strain PA01 and mucoid strain PD0300 significantly increased bacterial adhesion over the EPS deficient PA01 psl pel mutant strain despite their similar surface charge as indicated by the zeta potential measurements. Retained bacterial profiles show the deposition rate coefficients with various shapes and degrees of deviation from those expected from the classic filtration theory. Non-monotonic deviations from the log-linear deposition pattern with the majority of the bacteria retained downgradient of the column inlet were observed when bacterial cells were encapsuled by EPS under both high and low ionic strength conditions. In contrast, the EPS-deficient strain exhibited monotonic deviation from theory only under low ionic strength conditions. The results demonstrate that the non-monotonic deviation from filtration theory observed in this study was driven by steric interactions between extracellular polymers and glass beads. Analysis of the retained polysaccharides (carbohydrates and uronic acids) and protein profiles suggests that bacterial re-entrainment and re-entrapment may have contributed to the downgradient movement of the maximum retained bacteria. The detachment of bacteria may leave behind various constituents of EPS as their "footprints," which can be a valuable tool for tracking the trajectory of bacterial transport.     

酒酒球菌(Oenococcus oeni)产胞外聚合物培养基的优化及其抗冷冻干燥性能评价

为了研究酒酒球菌(Oenococcus oeni)产胞外聚合物(extracellular polymeric substances,EPS)在冷冻干燥过程中的保护性能,通过优化培养基提高EPS产量,并通过EPS对细菌冷冻干燥存活率的影响以及观察冷冻干燥过程中酒酒球菌内部及外表面的变化,评价其抗冷冻干燥的性能。正交试验优化可得,在初始pH值4. 8、蛋白胨含量20 g/L、葡萄糖含量15 g/L的条件下,能产119. 7 mg/100 mL酒酒球菌EPS。EPS作为冷冻干燥保护剂时细菌冷冻干燥后的存活率为70. 97%,高于其他常规保护剂,扫描电子显微镜(scanning electron microscope,SEM)和透射电子显微镜(transmission electron microscope,TEM)的图像表明添加冷冻干燥保护剂的实验组细胞形态更加完整,说明EPS有明显的冷冻干燥保护作用。该研究初步探究了酒酒球菌EPS冷冻干燥保护的机制,并为EPS在冷冻干燥保护剂中的应用提供理论基础。     

Removal of Pseudomonas putida biofilm and associated extracellular polymeric substances from stainless steel by alkali cleaning

Alkali (NaOH)-based compounds are commonly used in the food industry to clean food contact surfaces. However, little information is available on the ability of alkali and alkali-based cleaning compounds to remove extracellular polymeric substances (EPS) produced by biofilm bacteria. The objectives of this study were to determine the temperature and NaOH concentration necessary to remove biofilm EPS from stainless steel under turbulent flow conditions (clean-in-place simulation) and to determine the ability of a commercial alkaline cleaner to remove biofilm EPS from stainless steel when applied under static conditions without heat. Biofilms were produced by growing Pseudomonas putida on stainless steel for 72 h at 25 degrees C in a 1:10 dilution of Trypticase soy broth. The biofilms were treated using NaOH at concentrations of 1.28 to 6.0% and temperatures ranging from 66 to 70 degrees C. Other biofilms were treated with commercial alkaline cleaner at 25 or 4 degrees C for 1 to 30 min. Removal of EPS was determined by direct microscopic observation of samples stained with fluorescent-labeled peanut agglutinin lectin. Treatment with 1.2% NaOH at 66 degrees C for 3 min was insufficient to remove biofilm EPS. A minimum of 2.5% NaOH at 66 degrees C and 2.0% NaOH at 68 degrees C for 3 min were both effective for EPS removal. Commercial alkaline cleaner removed over 99% of biofilm EPS within 1 min at 4 and 25 degrees C under static conditions. Selection of appropriated cleaning agent formulation and use at recommended concentrations and temperatures is critical for removal of biofilm EPS from stainless steel.     

Aggregation and dissolution of silver nanoparticles in natural surface water

This study investigated aggregation and silver release of silver nanoparticles suspended in natural water in the absence and presence of artificial sun light. The influence of the capping layer was investigated using uncoated particles and particles coated with citrate or Tween 80. The experiments were conducted over 15 days in batch mode using a river water matrix. Silver release was monitored over this time while the aggregation state and morphological changes of the silver nanoparticles were tracked using dynamic light scattering and transmission electron microscopy. Results indicate sterically dispersed particles coated with Tween released silver quicker than did bare- and citrate-coated particles, which rapidly aggregated. A dissolved silver concentration of 40 μg/L was reached after just 6 h in the Tween-coated particle systems, accounting for ca. 3% of the total silver. Similar levels of dissolved silver were reached in the uncoated and citrate-coated systems at the end of the 15 days. Silver release was not significantly impacted by the artificial sun light; however, the light (and citrate) imparted significant morphological changes to the particles. Their impact was masked by aggregation, which seemed to be the controlling process in this study.     

Grape and wine polymeric polyphenols: Their importance in enology

Phenolic compounds are important constituents of red wine, contributing to its sensory properties and antioxidant activity. Owing to the diversity and structural complexity, study of these compounds was mainly limited, during the last three decades, on their low-molecular-mass compounds or simple phenolic compounds. Only in recent years, much attention has been paid to highly polymerized polyphenols in grape and red wines. The reason for this is largely due to the development of analytical techniques, especially those of HPLC-ESI-MS, permitting the structural characterization of highly polymerized polyphenols. Furthermore, the knowledge on the biological properties of polymeric polyphenols of red wine is very limited. Grape polyphenols mainly consist of proanthocyanidins (oligomers and polymers) and anthocyanins, and low amount of other phenolics. Red wine polyphenols include both grape polyphenols and new phenolic products formed from them during winemaking process. This leads to a great diversity of new polyphenols and makes wine polyphenol composition more complex. The present paper summarizes the advances in the research of polymeric polyphenols in grape and red wine and their important role in Enology. Scientific results indicate that polymeric polyphenols, as the major polyphenols in grape and red wine, play a major role in red wine sensory properties, color stability and antioxidant activities.     

Contribution of extracellular polymeric substances from Shewanella sp. HRCR-1 biofilms to U(VI) immobilization

The goal of this study was to quantify the contribution of extracellular polymeric substances (EPS) to U(VI) immobilization by Shewanella sp. HRCR-1. Through comparison of U(VI) immobilization using cells with bound EPS (bEPS) and cells with minimal EPS, we show that (i) bEPS from Shewanella sp. HRCR-1 biofilms contribute significantly to U(VI) immobilization, especially at low initial U(VI) concentrations, through both sorption and reduction; (ii) bEPS can be considered a functional extension of the cells for U(VI) immobilization and they likely play more important roles at lower initial U(VI) concentrations; and (iii) the U(VI) reduction efficiency is dependent upon the initial U(VI) concentration and decreases at lower concentrations. To quantify the relative contributions of sorption and reduction to U(VI) immobilization by EPS fractions, we isolated loosely associated EPS (laEPS) and bEPS from Shewanella sp. HRCR-1 biofilms grown in a hollow fiber membrane biofilm reactor and tested their reactivity with U(VI). We found that, when reduced, the isolated cell-free EPS fractions could reduce U(VI). Polysaccharides in the EPS likely contributed to U(VI) sorption and dominated the reactivity of laEPS, while redox active components (e.g., outer membrane c-type cytochromes), especially in bEPS, possibly facilitated U(VI) reduction.     

Anaerobic electrochemical corrosion of mild steel in the presence of extracellular polymeric substances produced by a culture enriched in sulfate-reducing bacteria

The corrosion of mild steel in a seawater medium containing extracellular polymeric substances (EPS) produced by sulfate-reducing bacteria (SRB) was studied by electrochemical experiments and atomic force microscopy (AFM). Under anaerobic conditions, the corrosion of mild steel increased up to 5-fold in the presence of a 1% (w/w) EPS solution but in the absence of SRB. The enhanced corrosion is mainly due to the oxidizing power of EPS with a reduction potential of E1/2 at -0.54 V (saturated calomel electrode), which is 0.4 V above that of hydrogen reduction. The electrochemical reduction of EPS provides a couple to iron oxidation, as demonstrated by H-shaped cell experiments in which the steel sample and EPS are not in physical contact but are ionically connected via the solution and electronically connected through an external wire. Fourier transformation infrared spectroscopy and X-ray photoelectron spectroscopy showed that EPS derived from SRB are comprised of 60% proteins, 37% polysaccharides, and 3% hydrocarbons. The XPS results showed that, upon corrosion, polysaccharide components were mostly converted to hydrocarbons.     

Lack of influence of extracellular polymeric substances (EPS) level on hydroxyl radical mediated disinfection of Escherichia coli

Photolysis of nitrate, a prevalent constituent in agriculturally impacted waters, may influence pathogen attenuation in such systems through production of hydroxyl radical ((?)OH). This study focuses on the efficacy of (?)OH generated during nitrate photolysis in promoting E. coli die-off as a function of extracellular polymeric substances (EPS) coverage. EPS levels of four E. coli isolates were systematically altered through a sonication extraction method and photochemical batch experiments with a solar simulator examined isolate viability loss as a function of time in nitrate solutions. E. coli viability loss over time exhibited two regimes: an initial induction time, t(s), with little decay was followed by rapid exponential decay characterized by a first-order disinfection rate constant, k. Increasing steady-state (?)OH concentrations enhanced E. coli viability loss, increasing values of k and decreasing t(s) values, both of which were quantified with a multitarget bacterial disinfection model. Notably, at a given steady-state (?)OH concentration, values of t(s) and k were independent of EPS levels, nor did they vary among the different E. coli strains considered. Results herein show that while (?)OH generated via nitrate photolysis enhances rates of disinfection in surface water, the mechanism by which (?)OH kills E. coli is relatively insensitive to common bacterial variables.     

Identification of key constituents and structure of the extracellular polymeric substances excreted by Bacillus megaterium TF10 for their flocculation capacity

Extracellular polymeric substances (EPS), a complex high-molecular-weight mixture of polymers excreted by microorganisms and produced from cell lysis, may have a high bioflocculation activity. In this work, the EPS excreted from Bacillus megaterium TF10, which was isolated from a soil sample, were systematically characterized to give insights into the relationship between their specific constituents and structure with their flocculation capacity. The results of microscopic observation, zeta potential, and TF10 EPS structure analysis show that the bridging mechanism was mainly responsible for the flocculation of the TF10. The constituents with a large molecular weight (1037-2521 kDA) and functional groups had contributed to the flocculation. GC-MS and NMR analyses demonstrate that the polysaccharides had long chain composed of rhamnose as well as glucose and galactose with uronic acids, acetyl amino sugars, and proteins as the side chains. The proteins in TF10 had no flocculation ability because of their special secondary structure and molecular weight diffusion characters. The EPS from Bacillus megaterium TF10 were found to exhibit a high flocculation activity, and the polysaccharides in EPS, which have the structure of the long backbone with active side chains, were identified as the active constituents for the high flocculation activity.     

Stability and removal of water soluble CdTe quantum dots in water

Commercial use of quantum dots (GDs) will lead to their entry into aquatic environments. This study examines the characteristics and stability of CdTe QDs with thioglycolate capping ligands in water as well as their removal by alum salts. The capping ligands of QDs are a key factor in determining their fate in water. Protonated thioglycolate capping ligands cause QDs to aggregate. The stability of QDs depends more on their ionic composition in water than on the ionic strength. In KCl solution, QDs remain stable even under 0.15 M ionic strength. Relatively low concentrations (< or = 2 meq/L) of divalent (Mg2+ and Ca2+) or trivalent (Al3+) cations, however, can induce aggregation. The proposed mechanism for this phenomenon is that multivalent metal cations (or their hydrated species) react with capping ligands to form complexes that bridge QDs or neutralize their surface charges. Because the complexation of hydrated Al3+ with capping ligands inhibits the formation of Al(OH)3 precipitates, alum dosages higher than the A3+ solubility are required to form settleable flocs and remove QDs from nanopure water by sedimentation. Divalent cations (Mg2+ and Ca2+) in tap water induce the formation of settleable QD flocs such that 70-80% of the QGDs by mass settle out.     

Multiple fluorescence labeling and two dimensional FTIR-13C NMR heterospectral correlation spectroscopy to characterize extracellular polymeric substances in biofilms produced during composting

Knowledge on the structure and function of extracellular polymeric substances (EPS) in biofilms is essential for understanding biodegradation processes. Herein, a novel method based on multiple fluorescence labeling and two-dimensional (2D) FTIR-(13)C NMR heterospectral correlation spectroscopy was developed to gain insight on the composition, architecture, and function of EPS in biofilms during composting. Compared to other environmental biofilms, biofilms in the thermophilic (>55 °C) and cooling (mature) stage of composting have distinct characteristics. The results of multiple fluorescence labeling demonstrated that biofilms were distributed in clusters during the thermophilic stage (day 14), and dead cells were detected. In the mature stage (day 26), the biofilm formed a continuous layer with a thickness of approximately 20-100 μm around the compost, and recolonization of cells at the surface of the compost was easily observed. Through 2D FTIR-(13)C NMR correlation heterospectral spectroscopy, the following trend in the ease of the degradation of organic compounds was observed: heteropolysaccharides > cellulose > amide I in proteins. And proteins and cellulose showed significantly more degradation than heteropolysaccharides. In summary, the combination of multiple fluorescence labeling and 2D correlation spectroscopy is a promising approach for the characterization of EPS in biofilms.     

基于量子点的传感技术在农药残留检测中的应用

食品中农药的残留日益威胁人类健康,而基于量子点发展起来的传感技术在农药残留的检测中有着不可比拟的优势。文中阐述了近几年来几种主要的传感技术在农药残留中的应用,并探讨了这些传感技术在农药残留检测中面临的挑战。     

海绵及其共生微生物的生态多样性与重要性

海绵是底栖生物群落中的重要成员,广泛分布于温带、热带及极地地区,其丰度、多样性及分布规律在不同地区有明显差异。海绵可能影响其它底栖生物及生态系统特征,因而在海洋生态系统中有多种功能作用。海绵不仅在海洋生态系统中起重要作用,而且能提供大量新颖的生物活性产物。大量研究显示多种微生物能寄生在海绵体内,在微生物与其寄主海绵之间可能存在互惠共生关系。文中对海绵的重要生态作用、生物多样性、与微生物的共生关系和作为新天然产物的资源进行了讨论。     

巯基乙酸稳定的CdTe发光量子点的合成与表征

以巯基乙酸为稳定剂,合成了水溶性的具有各种荧光发射波长的CdTe纳米粒子体系,其光谱范围为537～577nm.TEM,XRD和红外线光谱结果表明,该纳米粒子分散均匀,尺寸分布较窄,立方晶相,晶格中有少量的S参与配位,并且TGA的巯基和CdTe纳米粒子表面的镉离子发生了配位作用,TGA稳定的CdTe纳米粒子在碱性条件下稳定,溶血实验表明,TGA稳定的CdTe量子点具有一定的生物相容性,这些量子点可有效地应用于生物体系中.     

Relative susceptibility and transcriptional response of nitrogen cycling bacteria to quantum dots

Little is known about the potential impacts of accidental or incidental releases of manufactured nanomaterials to microbial ecosystem services (e.g., nutrient cycling). Here, quantum dots (QDs) coated with cationic polyethylenimine (PEI) were more toxic to pure cultures of nitrogen-cycling bacteria than QDs coated with anionic polymaleic anhydride-alt-1-octadecene (PMAO). Nitrifying bacteria (i.e., Nitrosomonas europaea) were much more susceptible than nitrogen fixing (i.e., Azotobacter vinelandii, Rhizobium etli, and Azospirillum lipoferum) and denitrifying bacteria (i.e., Pseudomonas stutzeri). Antibacterial activity was mainly exerted by the QDs rather than by their organic coating or their released QD components (e.g., Cd and Zn), which under the near-neutral pH tested (to minimize QD weathering) were released into the bacterial growth media at lower levels than their minimum inhibitory concentrations. Sublethal exposure to QDs stimulated the expression of genes associated with nitrogen cycling. QD-PEI (10 nM) induced three types of nitrogenase genes (nif, anf, and vnf) in A. vinelandii, and one ammonia monooxygenase gene (amoA) in N. europaea was up-regulated upon exposure to 1 nM QD-PEI. We previously reported up-regulation of denitrification genes in P. stutzeri exposed to low concentrations of QD-PEI. (1) Whether this surprising stimulation of nitrogen cycling activities reflects the need to generate more energy to overcome toxicity (in the case of nitrification or denitrification) or to synthesize organic nitrogen to repair or replace damaged proteins (in the case of nitrogen fixation) remains to be determined.     

Effects of aging and pH on dissolution kinetics and stability of chloropyromorphite

The objectives of this research were to understand the effect of aging time on chloropyromorphite stability by dissolution, to examine physical and chemical alterations of the pyromorphite samples, and to model the kinetic data collected from the dissolution experiments. The results of this investigation indicate that chloropyromorphite formation is kinetically rapid and that its dissolution in acid is thermodynamically stable, ideal conditions for Pb immobilization that has emerged as a potential remediation strategy. In terms of aging prior to dissolution, X-ray absorption fine structure (XAFS) and X-ray diffraction (XRD) spectroscopies were unable to distinguish fundamental differences in progressively aged samples; however, high-resolution thermogravimetric analysis (HRTGA) did demonstrate thatthe thermostability of the chloropyromorphite material increased with increasing residence time. The stirred-flow and batch dissolution studies suggest that the aging process ceased within 24 h and that the dissolution rate of the 1-day aged sample was not significantly different than the 1-year aged specimen. The amount of Pb released peaked at 21% (1-h sample, stirred-flow, pH 2.0) and was as low as 0.17% (1-year sample, batch method, pH 6.0). Postdissolution analyses of chloropyromorphite with XAFS, XRD, and HRTGA revealed no detectable chemical alterations of the pyromorphite samples signifying only release of dissolved Pb to solution and no formation of secondary products during dissolution.     

量子点标记荧光免疫法检测花生中黄曲霉毒素B_1

建立了基于量子点标记二抗的间接竞争荧光免疫吸附测定方法（indirect competitive fluorescence-linked immunosorbent assay,cFLISA）,并研究检测花生中黄曲霉毒素B1的可行性。采用谷胱甘肽为稳定剂,在水相中直接合成碲化镉（CdTe）量子点,并利用1-（3-二甲氨基丙基）-3-乙基碳二亚胺盐酸盐（EDC）与兔抗鼠二抗进行共价偶联,以黄曲霉毒素B1的单克隆抗体建立cFLISA方法。结果表明,该方法的灵敏度和最低检测限值分别为0.023ng/mL和0.001ng/mL,与传统的有机染料FITC-二抗法比较,灵敏度提高了30倍,花生样品加标0.1、0.05和0.025ng/g,回收率范围在88%～116%之间,变异系数均小于10%。建立的cFLISA方法可以较好的检测花生中黄曲霉毒素B1,并为其它真菌毒素的检测提供参考。