Persistence of pathogenic prion protein during simulated wastewater treatment processes

Transmissible spongiform encephalopathies (TSEs, prion diseases) are a class of fatal neurodegenerative diseases affecting a variety of mammalian species including humans. A misfolded form of the prion protein (PrP(TSE)) is the major, if not sole, component of the infectious agent. Prions are highly resistant to degradation and to many disinfection procedures suggesting that, if prions enter wastewater treatment systems through sewers and/or septic systems (e.g., from slaughterhouses, necropsy laboratories, rural meat processors, private game dressing) or through leachate from landfills that have received TSE-contaminated material, prions could survive conventional wastewater treatment. Here, we report the results of experiments examining the partitioning and persistence of PrPTSE during simulated wastewater treatment processes including activated and mesophilic anaerobic sludge digestion. Incubation with activated sludge did not result in significant PrPTSE degradation. PrPTSE and prion infectivity partitioned strongly to activated sludge solids and are expected to enter biosolids treatment processes. A large fraction of PrPTSE survived simulated mesophilic anaerobic sludge digestion. The small reduction in recoverable PrPTSE after 20-d anaerobic sludge digestion appeared attributable to a combination of declining extractability with time and microbial degradation. Our results suggest that if prions were to enter municipal wastewater treatment systems, most would partition to activated sludge solids, survive mesophilic anaerobic digestion, and be present in treated biosolids.     

石英砂过滤澄清食醋工艺的改进

采用石英砂配料对石英砂滤料进行改进处理,应用这种过滤器对食醋进行处理,研究了配料种类、粒径、添加量及操作压力等因素对过滤效果的影响,确定了用配料过滤食醋的最佳条件。实验结果表明:在室温,选择配料a、配料粒径范围20~25μm、添加量为m(配料):m(固体杂质)=0.32、操作压力为0.03MPa的条件下,经改进石英砂过滤,杂质去除率可达92.3%,且处理后食醋的风味未变。     

Environmentally-relevant forms of the prion protein

Scrapie and chronic wasting disease (CWD) are prion diseases of particular environmental concern as they are horizontally transmissible and can remain infectious after years in the environment. Recent evidence suggests that the N-terminus of PrPSC, the infectious conformation of the prion protein, plays an important role in the mechanism of sorption to soil particles. We hypothesize that, in a prion-infected animal carcass, a portion of the N-terminus of PrPSc could be cleaved by proteinases in the brain at ordinary temperatures. Hamster (HY transmissible mink encephalopathy-infected), transgenic mice (CWD-infected), and elk (CWD-infected) brain homogenates were incubated at 22 and 37 degrees C for up to 1 month and then analyzed by Western blot using N-terminal and middle region monoclonal anti-PrP antibodies. For all three systems, there was a very faint or undetectable N-terminal PrP signal after 35 days at both temperatures, which indicates that full-length PrPSc might be rare in the brain matter of animal carcasses. Future studies on prion-soil interactions should therefore consider N-terminal-degraded PrPSc in addition to the full-length form. Both mouse and elk CWD PrPSc demonstrated greater resistance to degradation than HY TME PrPSc. This indicates that the transgenic mouse-CWD model is a good surrogate for natural CWD prions, but that other rodent prion models might not accurately represent CWD prion fate in the environment.     

Mobility of functionalized quantum dots and a model polystyrene nanoparticle in saturated quartz sand and loamy sand

Quantum dots (QDs) are one example of engineered nanoparticles (ENPs) with demonstrated toxic effects. Yet, little is known about the behavior of QDs in the natural environment. This study assessed the transport of two commercial carboxylated QDs (CdTe and CdSe) and carboxylated polystyrene latex (nPL) as a model nanoparticle using saturated laboratory-scale columns. The influence of solution ionic strength (IS) and cation type (K(+) or Ca(2+)) on the transport potential of these ENPs was examined in two granular matrices - quartz sand and loamy sand. The retention of all three particles was generally low in the quartz sand columns within the range of studied IS (0.1-100 mM) for the monovalent salt (KCl). In contrast, the retention of the three ENPs in the quartz sand was significant in the presence of 10 mM Ca(2+). Moreover, ENP attachment efficiencies (α) were enhanced by at least 1 order of magnitude in columns packed with loamy sand (for IS between 0.1-10 mM KCl). Although all three ENPs used here are carboxylated, they differ in the type of surface coating (e.g., choice of polymers or polyelectrolytes). Regardless of the surface coatings, the three ENPs exhibit comparable mobility in the quartz sand. However, the ENPs demonstrate variable transport potential in loamy sand suggesting that differences in the binding affinities of surface-modified ENPs for specific soil constituents can play a key role in the fate of ENPs in soils.     

Spatial variation in deposition rate coefficients of an adhesion-deficient bacterial strain in quartz sand

The transport of bacterial strain DA001 was examined in packed quartz sand under a variety of environmentally relevant ionic strength and flow conditions. Under all conditions, the retained bacterial concentrations decreased with distance from the column inlet at a rate that was faster than loglinear, indicating that the deposition rate coefficient decreased with increasing transport distance. The hyperexponential retained profile contrasted againstthe nonmonotonic retained profiles that had been previously observed for this same bacterial strain in glass bead porous media, demonstrating that the form of deviation from log-linear behavior is highly sensitive to system conditions. The deposition rate constants in quartz sand were orders of magnitude below those expected from filtration theory, even in the absence of electrostatic energy barriers. The degree of hyperexponential deviation of the retained profiles from loglinear behavior did not decrease with increasing ionic strength in quartz sand. These observations demonstrate thatthe observed low adhesion and deviation from log-linear behavior was not driven by electrostatic repulsion. Measurements of the interaction forces between DA001 cells and the silicon nitride tip of an atomic force microscope (AFM) showed that the bacterium possesses surface polymers with an average equilibrium length of 59.8 nm. AFM adhesion force measurements revealed low adhesion affinities between silicon nitride and DA001 polymers with approximately 95% of adhesion forces having magnitudes < 0.8 nN. Steric repulsion due to surface polymers was apparently responsible for the low adhesion to silicon nitride, indicating that steric interactions from extracellular polymers controlled DA001 adhesion deficiency and deviation from log-linear behavior on quartz sand.     

High-resolution measurement of pore saturation and colloid removal efficiency in quartz sand using fluorescence imaging

Colloid deposition in unsaturated, nonuniform porous media is poorly explained by current models and difficult to measure using breakthrough curves and retained mass profiles. We present new methods which enable time-lapse fluorescence imaging to quantify variations in pore saturation, theta, and colloid deposition in 2D, nonuniform unsaturated flow fields. Calibration experiments revealed direct proportionality between fluorescence F and theta in 20/30 mesh quartz sand. Analysis of breakthrough data in fluorescence images allows quantification of the mean mobile concentration, mean deposition rate, and hence the colloid removal efficiency eta directly from data at the pixel-scale throughoutthe flow field. We imaged carboxylate-modified latex microspheres from a point source in saturated flow and unsaturated flow across a capillary fringe at 10(-3), 10(-2), and 10(-1) M NaCl. Total numbers of colloids deposited and values of eta increased with ionic strength. We modeled the observed variations in eta with theta to estimate the partitioning of colloid deposition between air-water and solid-water interfaces. In the broad saturation range 0.2 < theta < 1, our results suggest that only at the lowest ionic strength, where deposition at solid-water interfaces was strongly unfavorable, did colloid deposition associated with air-water interfaces significantly influence the total colloid removal.     

Field and laboratory investigations of inactivation of viruses (PRD1 and MS2) attached to iron oxide-coated quartz sand

Field and laboratory experiments were conducted to investigate inactivation of viruses attached to mineral surfaces. In a natural gradient transport field experiment, bacteriophage PRD1, radiolabeled with 32P, was injected into a ferric oxyhydroxide-coated sand aquifer with bromide and linear alkylbenzene sulfonates. In a zone of the aquifer contaminated by secondary sewage infiltration, small fractions of infective and 32P-labeled PRD1 broke through with the bromide tracer,followed bythe slow release of 84% of the 32P activity and only 0.011% of the infective PRD1. In the laboratory experiments, the inactivation of PRD1, labeled with 35S (protein capsid), and MS2, dual radiolabeled with 35S (protein capsid) and 32P (nucleic acid), was monitored in the presence of groundwater and sediment from the contaminated zone of the field site. Release of infective viruses decreased at a much faster rate than release of the radiolabels, indicating that attached viruses were undergoing surface inactivation. Disparities between 32P and 35S release suggest that the inactivated viruses were released in a disintegrated state. Comparison of estimated solution and surface inactivation rates indicates solution inactivation is approximately 3 times as fast as surface inactivation. The actual rate of surface inactivation may be substantially underestimated owing to slow release of inactivated viruses.     

甜菜红在双蛋白纤维上的吸附性能

采用甜菜红色素对双蛋白纤维进行染色,探讨染色温度、时间、pH值、氯化钠和染料用量对双蛋白纤维的染色性能的影响,分析甜菜红色素对双蛋白纤维的染色热力学,测试染色织物的色牢度。甜菜红色素对双蛋白纤维的优化染色工艺为:甜菜红1.5%,pH=2~3,染色温度30℃,染色时间90 min;甜菜红色素在双蛋白纤维上的吸附符合Langmuir吸附模型。     

Effects of phosphate on the transport of Escherichia coli O157:H7 in saturated quartz sand

Consumption of groundwater contaminated with E. coli O157:H7 has led to several waterborne disease outbreaks over the past decade. A thorough understanding of the transport of E. coli O157:H7 within the soil-groundwater system is critical to the protection of public health. Although phosphate is ubiquitous in the natural environment, the influence of phosphate on the transport of E. coli O157:H7 in the groundwater system remains unknown. In this research, we performed column transport experiments to evaluate the effect of phosphate on the transport of E. coli O157:H7 cells within saturated sand. The pH of the solutions was maintained at 7.2, the ionic strength varied from 10 to 100 mM, and the phosphate concentration ranged from 0 to 1 mM. Our results show that (1) phosphate could enhance the transport of E. coli O157:H7 cells under both ionic strength conditions; (2) E. coli O157:H7 displayed lower retention in sand under higher ionic strength conditions; (3) increased phosphate in the mobile aqueous phase led to the release of previously immobilized E. coli O157:H7 cells. The response of E. coli O157:H7 cells to variations in phosphate concentrations and ionic strength conditions are explained using the extended DLVO (XDLVO) theory and the steric repulsion caused by extracellular macromolecules. In summary, our results suggest that phosphate could widen the spread of E. coli O157:H7 cells, and potentially other types of bacterial cells, within the soil-groundwater system.     

Airborne respirable silica near a sand and gravel facility in central California: XRD and elemental analysis to distinguish source and background quartz

Despite the potential toxicity of respirable quartz to humans, little is known about the transport of airborne quartz from sources to receptors and how to distinguish anthropogenically generated quartz from natural background in a receptor sample. Airborne quartz emissions near a sand and gravel facility were determined using PM10 and PM2.5 filter samples collected at four downwind sites (D1: 22 m, D2: 62 m, D3: 259 m, and D4: 745 m from the facility) as well as one upwind site (U1: 1495 m) during summer sampling. X-ray diffraction was used to determine quartz concentration and elemental composition was analyzed using PIXE, XRF, PESA, and HIPS techniques. Elemental composition of the PM samples was used to determine the X-ray mass absorption coefficients that are essential for accurate quartz quantification by XRD. Elemental composition was found to be a useful tool to distinguish source and background crystalline silica. Both PM10 and PM2.5 samples collected at the D1, D2, and D3 sites contained more Si, Al, and Fe and less H, Na, and S, compared to those at the U1 site, whereas site D4 sample compositions were similar to those at the U1 site. A composite variable, SOIL/(H+Na+S), where SOIL = 2.20Al + 2.49Si + 1.63Ca + 1.94Ti + 2.42Fe, was used to distinguish source materials from background. Average dry season quartz concentrations in replicate PM10 samples were 4.6 +/- 0.9) microg m(-3) at U1, 60.6 (+/- 5.4) microg m(3) at D1, 62.4 (+/- 3.6) at D2, 32.6 (+/- 2.1) microg m(-3) at D3, and 9.41 (+/- 0.9) microg m(-3) at D4. The mass fraction of quartz was the highest at the D1 site and decreased with increasing distance from the facility. The mass of PM2.5 samples was too low to determine quartz concentrations. These results identify the facility as the main source of quartz and other silicate minerals downwind of the plant and that the air quality of the most remote sampling site, located approximately 750 m downwind, was still impacted by the facility's activity.     

Influence of prion protein genotypes on milk production traits in Spanish Churra sheep

The aim of this work was to analyze the possible relationships between milk production traits and prion protein genotypes in Spanish Churra sheep. For this purpose, 2 analyses were carried out. First, an association study was performed of the prion protein genotypes of 12,533 Churra ewes and their milk yield, protein percentage, fat percentage, and somatic cell score as phenotypes, followed by a quantitative trait loci screening on the chromosome where the prion protein gene was located in this population. The latter analysis was carried out using 8 genetic markers (7 microsatellites and the prion protein genotypes) spanning ovine chromosome 13 using a daughter design. Regarding genotype frequencies, the most frequent allele was ARQ (75.90%), which linked with a high susceptibility to scrapie, followed by the resistant haplotype, ARR (18.16%). The frequency of the most susceptible allele, VRQ, was around 1%. No evidence of association or linkage between prion protein genotypes and milk traits has been detected in Churra sheep. These results indicate that increasing the ARR frequency in Churra population will not have an adverse effect on selection for milk traits included in the breeding objectives. However, the low allele frequencies for ARR should be considered in the initial stages to prevent possible bottlenecks in future genetic progress.     

Transport of strontium and cesium in simulated hanford tank waste leachate through quartz sand under saturated and unsaturated flow

We investigated the effects of water saturation and secondary precipitate formation on Sr and Cs transport through quartz sand columns under saturated and unsaturated flow. Column experiments were conducted at effective water saturation ranging from 0.2 to 1.0 under steady-state flow using either 0.1 M NaNO(3) or simulated tank waste leachate (STWL; 1 M NaNO(3) and 1 M NaOH) mimicking Hanford (Washington, USA) tank waste. In 0.1 M NaNO(3) columns, Sr transported like a conservative tracer, whereas Cs was retarded relative to Sr. The transport of Sr and Cs in the 0.1 M NaNO(3) columns under all water saturations could be described with the equilibrium convection-dispersion equation (CDE). In STWL columns, Sr mobility was significantly reduced compared to the 0.1 M NaNO(3) column, because Sr was incorporated into or sorbed to neo-formed secondary precipitates. Strontium sequestration by precipitates was confirmed by additional batch and electron micrograph analyses. In contrast(,) the transport of Cs was less affected by the STWL; retardation of Cs in STWL columns was similar to that found in 0.1 M NaNO(3) columns. Analysis of STWL column data revealed that both Sr and Cs breakthrough curves showed nonideal behavior that suggest nonequilibrium conditions, although nonlinear geochemical behavior cannot be ruled out.     

Highly dense protein layers confirmed by atomic force microscopy and quartz crystal microbalance

Protein adsorption on a gold surface is investigated by comparing the results of quartz crystal microbalance method and atomic force microscopy. The adsorption of streptavidin on functional gold surfaces is directly monitored by a quartz crystal microbalance, and confirmed by atomic force microscopy. For this investigation, a modified gold substrate is fabricated to obtain a topographic image of streptavidin molecules. Both methods show a correlation in terms of the highly dense protein single-layer formation, and the modified gold electrode shows a slightly denser protein layer formation because of the difference in substrate geometry as compared with that of a mica surface.     

Adsorption of transgenic insecticidal Cry1Ab protein to SiO2. 1. Forces driving adsorption

Genetically modified Bt crops express insecticidal Cry proteins (Bt toxins) that may enter agricultural soils. A mechanistic understanding of Cry protein adsorption to soils is critical for risk assessment, as this process governs Cry protein fate and bioavailability. We used quartz crystal microbalance and optical waveguide lightmode spectroscopy to elucidate the driving forces of the adsorption of monomeric Cry1Ab to negatively charged quartz (SiO(2)) and positively charged poly-L-lysine (PLL) at pH 5-8 and constant ionic strength of 50 mM (NaCl). Bovine serum albumin and hen egg white lysozyme were used as reference proteins because of their known adsorption behavior. Electrostatics governed Cry1Ab adsorption; as pH increased above the isoelectric point of Cry1Ab, the initial rate and the extent of adsorption decreased on SiO(2) and increased on PLL. Reversible adsorption to SiO(2) suggested weak Cry1Ab-SiO(2) electrostatic interactions and no irreversible conformational changes of Cry1Ab at the surface. High conformational stability of Cry1Ab was further supported by supply rate-independent extent of adsorption of Cry1Ab to apolar gold. Some evidence is presented that the nonuniform surface charge distribution of Cry1Ab resulted in patch-controlled electrostatic attraction with sorbents that carried the same net charge as Cry1Ab. A more detailed discussion of this mechanism is given in a companion paper.     

Adsorption of transgenic insecticidal Cry1Ab protein to SiO2. 2. Patch-controlled electrostatic attraction

Adsorption governs the fate of Cry proteins from genetically modified Bt crops in soils. The effect of ionic strength (I) on the adsorption of Cry1Ab (isoelectric point IEP(Cry1Ab) ≈ 6) to negatively charged quartz (SiO(2)) and positively charged poly-L-lysine (PLL) was investigated at pH 5 to 8, using quartz crystal microbalance with dissipation monitoring and optical waveguide lightmode spectroscopy. Cry1Ab adsorbed via positively and negatively charged surface patches to SiO(2) and PLL, respectively. This patch controlled electrostatic attraction (PCEA) explains the observed increase in Cry1Ab adsorption to sorbents that carried the same net charge as the protein (SiO(2) at pH > IEP(Cry1Ab) and PLL at pH < IEP(Cry1Ab)) with decreasing I. In contrast, the adsorption of two reference proteins, BSA and HEWL, with different adsorption mechanism, were little affected by similar changes of I. Consistent with PCEA, Cry1Ab desorption from SiO(2) at pH > IEP(Cry1Ab) increased with increasing I and pH. Weak Cry1Ab-SiO(2) PCEA above pH 7 resulted in reversible, concentration dependent adsorption. Solution depletion experiments showed that PCEA also governed Cry1Ab adsorption to SiO(2) particles at environmentally relevant concentrations (a few ng mL(-1)). These results imply that models describing Cry1Ab adsorption to charged surfaces in soils need to account for the nonuniform surface charge distribution of the protein.     

Adsorption of transgenic insecticidal Cry1Ab protein to silica particles. Effects on transport and bioactivity

Bt crops are genetically modified to be resistant against insect pests by expressing insecticidal Cry proteins. The processes governing the fate and bioavailability of the expressed transgenic Cry proteins in soils are poorly understood. We studied adsorption of Cry1Ab to negatively charged silica (SiO(2)) particles, a major soil constituent and a model for negatively charged mineral surfaces, at pH 5 to 10 and ionic strengths I = 10 mM to 250 mM, both in solution depletion and saturated column transport experiments. Cry1Ab-SiO(2) interactions were dominated by patch-controlled electrostatic attraction (PCEA), as evident from increasing Cry1Ab attraction to SiO(2) with decreasing I at pH at which both Cry1Ab and SiO(2) were net negatively charged. Experimental and modeling evidence is provided that the surface heterogeneity of SiO(2) particles modulated PCEA, leading to a fraction of adsorption sites with slow Cry1Ab desorption kinetics. Desorption rates from these sites increased upon increasing the solution pH. In toxicity bioassays, we demonstrated that Cry1Ab retained insecticidal activity when adsorbed to SiO(2), suggesting high protein conformational stability during adsorption-desorption cycles. Models predicting Cry1A protein adsorption in soils therefore need to account for combined effects of the nonuniform protein surface charge distribution and of sorbent surface heterogeneity.     

Inhibition of growth of pathogenic bacteria in raw milk by legume protein esters

Protein isolates from soybean and chickpea, as well as their methylated esters, were tested for their inhibitory action against the propagation of pathogenic bacteria in raw milk during its storage either at room temperature or under refrigeration. Raw milk was inoculated with a mixed culture of Listeria monocytogenes Scott A and Salmonella enterica serovar Enteritidis strain PT4 at ca. 2 log CFU ml?1. Aerobic plate count, coliform count, and presumptive E. coli in raw milk treated with esterified legume proteins were inhibited by 2 to 3 log relative to a control after 6 to 8 days of storage at 4°C. At room temperature, bacterial populations (aerobic plate count, coliform count, and presumptive E. coli) in raw milk treated with esterified legume proteins were inhibited by ca. 1.5 to 1.6 log relative to the control after 12 h. Supplementation of raw milk with esterified soybean protein could significantly inhibit the counts of the two inoculated pathogens (L. monocytogenes Scott A and Salmonella Enteritidis PT4), which were initially inoculated at ca. 2 log CFU ml?1, by ca. 2.4 log and 1.6 log CFU ml?1, respectively, on day 8 of storage under cold conditions. Corresponding reductions amounting to 2.7 and 1.8 log CFU ml?1 were observed after 12 h of storage at room temperature. Supplementation of raw milk with esterified soybean protein (0.5%) reduced the maximum level of titratable acidity to 0.21 and maintained the pH level at 6.4 after 8 days of storage under cold conditions as compared with 4 days for untreated raw milk. Similar results were observed when raw milk was stored at room temperature for 10 h.     

阳离子羽毛蛋白助剂对棉纤维吸附机理的研究

研究了阳离子羽毛蛋白助剂WLS-20在棉织物上的吸附性能.探究了WLS-20助剂在棉织物上的吸附动力学性能、热力学性能以及吸附牢度.结果表明,WLS-20在棉织物上吸附速率快;WLS-20的最大吸收波长是195 nm,此波长下WLS-20的浓度与吸光度成良好的线性关系;WLS-20的最佳吸附温度为40℃;准二级动力学方程能很好地描述WLS-20在棉织物上的吸附过程;WLS-20在棉织物上的吸附属于朗格缪尔(Langmuir)型,为化学定位吸附.碳酸钠作为催化剂,有利于WLS-20与染料和织物发生交联反应,使得助剂在织物上的吸附量显著提升,吸附牢度显著提高.     

Analysis of associations between the prion protein genotypes and production traits in East Friesian milk sheep

The objective of this study was to analyze associations between ovine prion protein genotypes and production traits in East Friesian milk sheep. Production traits included the type traits scores for muscle mass, wool quality, and type; the reproduction traits age at first lambing, first lambing interval, second lambing interval, and total number of lambs born; the milk performance traits; milk, fat, and protein yields; fat and protein contents; and somatic cell scores. Prion protein genotypes were available for 658 East Friesian milk sheep. Linear animal models were used for the analysis of the prion protein genotype effects. The scores of the genotyped sheep for muscle mass, type, wool quality, and fat yield were significantly superior to those of the nongenotyped animals. An explanation for this might be that breeders seek to minimize genotyping costs by preselecting animals that do not meet the top breeding requirements. No significant associations were found between the prion protein genotypes and milk performance, type, or reproduction traits.     

Adhesion kinetics of viable Cryptosporidium parvum oocysts to quartz surfaces

The transport and deposition (adhesion) kinetics of viable Cryptosporidium parvum oocysts onto ultrapure quartz surfaces in a radial stagnation point flow system were investigated. Utilizing an optical microscope and an image-capturing device enabled real time observation of oocyst deposition behavior onto the quartz surface in solutions containing either monovalent (KCl) or divalent (CaCl2) salts. Results showed a significantly lower oocyst deposition rate in the presence of a monovalent salt compared to a divalent salt. With a monovalent salt, oocyst deposition rates and corresponding attachment efficiencies were relatively low, even at high KCl concentrations where Derjaguin-Landau-Verwey-Overbeek (DLVO) theory predicts the absence of an electrostatic energy barrier. On the other hand, in the presence of a divalent salt, oocyst deposition rates increased continuously as the salt concentration was increased over the entire range of ionic strengths investigated. The unusually low deposition rate in a monovalent salt solution is attributed to "electrosteric" repulsion between the Cryptosporidium oocyst and the quartz surface, most likely due to proteins on the oocyst surface that extend into the solution. It is further proposed that specific binding of calcium ions to the oocyst surface functional groups results in charge neutralization and conformational changes of surface proteins that significantly reduce electrosteric repulsion.