Evaluation of DNA isolation procedures for detecting and quantifying environmental Legionella by real-time quantitative PCR

Six methods, QiAamp DNA Mini Kit (Q), Q with Sepharose 4B gel column (Q/G), Q with low melting point agarose (Q/L), freeze-thaw/phenol-chloroform lysis (FT-PC), FT-PC/G, and FT-PC/L, were evaluated for their ability to isolate DNA of sufficient quality to quantify Legionella using qPCR. Samples of mixing Legionella pneumophila (ATCC33152) and humic acid (HA, 0-126.8 mg/l) were treated by the six methods. Q, Q/G, Q/L, FT-PC/G, and FT-PC/L removed HA from 1.9-126.8 to <1 mg/l determined by A260 with a spectrophotometer. Q obtained the highest DNA yield, followed by Q/G. Dilution (10- to 100-fold) of DNA arising from extraction using Q, Q/G, FT-PC, or FT-PC/G prevented qPCR inhibition. The highest recovery of cells was found in DNA extracted by Q and diluted 100-fold, and followed by Q/G. The applicability of Q and Q/G with dilution was further validated with cooling tower waters. Q or Q/G with 10-fold dilution increased L. pneumophila detection, whereas 100-fold dilution obtained the highest cell concentrations. Similar results were found for Legionella spp. except that both 10- and 100-fold dilutions increased cell concentrations. Thus, Q with 10-fold dilution is suggested to detect and quantify Legionella spp. and detect L. pneumophila. For L. pneumophila-positive samples, 100-fold diluted DNA must be re-analyzed to accurately quantify L. pneumophila.     

Dual-layer hollow fibers for gas separation processes produced by quadruple spinning

A novel quadruple spinneret to produce dual-layer hollow fiber membranes by simultaneous spinning of two polymer solutions, using the dual precipitation bath technique is proposed. Hollow fibers aimed at gas separation processes were prepared in extrusion system specifically designed and built for this purpose. A polyurethane polymer was selected as the selective layer (outer-layer), while polyethersulfone was defined as the support (inner-layer). Activated carbon powder was added into the PU solution for further improvement of the transport properties. The hollow fibers showed good adhesion between the polymer layers and a defect-free selective layer. Representative results include a CO₂/N₂ selectivity of 43.     

Environmental fate of endocrine-disrupting dimethyl phthalate esters (DMPE) under sulfate-reducing condition

Dimethyl phthalate esters (DMPE) can easily be released into the environment from plastic products. As endocrine disruptors, DMPE mimic estrogenic activities in animals and humans. The metabolites of DMPE are suspected to cause even more serious health problems. Among the common sterilization techniques adopted in the study of DMPE degradation, the average loss of the parent DMPE compounds after autoclaving was as high as 21.26%. In contrast, the loss after 0.2 microm filtration was significantly lower at 2.28%. It is suggested that filtration should be used over autoclaving for sterilizing DMPE. The environmental fate of DMPE under sulfate-reducing condition was simulated and studied in microcosm system. It was observed that dimethyl phthalate (DMP), dimethyl isophthalate (DMI) and dimethyl terephthalate (DMT) could not be mineralized over an extended period of 6 months, but with the transformation to the respective monomethyl phthalate and/or phthalic acid. The dominant species of microorganisms utilizing individual DMPE isomer as the sole carbon source were isolated and identified as facultative anaerobe Thauera sp., Xanthobacter sp. and Agrobacterium sp. for DMP, DMI and DMT, respectively. This study illustrates that the detrimental DMPE and their natural metabolites may accumulate in the sulfate-reducing environment. Accordingly, proper surveillance program should be devised to monitor both the parent compounds and degradation intermediates of DMPE in order to protect the aquatic ecosystem and human health.     

Attacker-defender models and road network vulnerability

The reliability of road networks depends directly on their vulnerability to disruptive incidents, ranging in severity from minor disruptions to terrorist attacks. This paper presents a game theoretic approach to the analysis of road network vulnerability. The approach posits predefined disruption, attack or failure scenarios and then considers how to use the road network so as to minimize the maximum expected loss in the event of one of these scenarios coming to fruition. A mixed route strategy is adopted, meaning that the use of the road network is determined by the worst scenario probabilities. This is equivalent to risk-averse route choice. A solution algorithm suitable for use with standard traffic assignment software is presented, thereby enabling the use of electronic road navigation networks. A variant of this algorithm suitable for risk-averse assignment is developed. A numerical example relating to the central London road network is presented. The results highlight points of vulnerability in the road network. Applications of this form of network vulnerability analysis together with improved solution methods are discussed.     

Assessing the cost of groundwater pollution: the case of diffuse agricultural pollution in the Upper Rhine valley aquifer.

This paper presents an assessment of the costs of diffuse groundwater pollution by nitrates and pesticides for the industrial and the drinking water sectors in the Upper Rhine valley, France. Pollution costs which occurred between 1988 and 2002 are described and assessed using the avoidance cost method. Geo-statistical methods (kriging) are then used to construct three scenarios of nitrate concentration evolution. The economic consequences of each scenario are then assessed. The estimates obtained are compared with the results of a contingent valuation study carried out in the same study area ten years earlier.     

An Integrated Segmentation and Analysis Approach for QCT of the Knee to Determine Subchondral Bone Mineral Density and Texture

We have developed a new integrated approach for quantitative computed tomography of the knee in order to quantify bone mineral density (BMD) and subchondral bone structure. The present framework consists of image acquisition and reconstruction, 3-D segmentation, determination of anatomic coordinate systems, and reproducible positioning of analysis volumes of interest (VOI). Novel segmentation algorithms were developed to identify growth plates of the tibia and femur and the joint space with high reproducibility. Five different VOIs with varying distance to the articular surface are defined in the epiphysis. Each VOI is further subdivided into a medial and a lateral part. In each VOI, BMD is determined. In addition, a texture analysis is performed on a high-resolution computed tomography (CT) reconstruction of the same CT scan in order to quantify subchondral bone structure. Local and global homogeneity, as well as local and global anisotropy were measured in all VOIs. Overall short-term precision of the technique was evaluated using double measurements of 20 osteoarthritic cadaveric human knees. Precision errors for volume were about 2-3% in the femur and 3-5% in the tibia. Precision errors for BMD were about 1-2% lower. Homogeneity parameters showed precision errors up to about 2% and anisotropy parameters up to about 4%.     

Abundant and stable char residues in soils: implications for soil fertility and carbon sequestration

Large-scale soil application of biochar may enhance soil fertility, increasing crop production for the growing human population, while also sequestering atmospheric carbon. But reaching these beneficial outcomes requires an understanding of the relationships among biochar's structure, stability, and contribution to soil fertility. Using quantitative (13)C nuclear magnetic resonance (NMR) spectroscopy, we show that Terra Preta soils (fertile anthropogenic dark earths in Amazonia that were enriched with char >800 years ago) consist predominantly of char residues composed of ～6 fused aromatic rings substituted by COO(-) groups that significantly increase the soils' cation-exchange capacity and thus the retention of plant nutrients. We also show that highly productive, grassland-derived soils in the U.S. (Mollisols) contain char (generated by presettlement fires) that is structurally comparable to char in the Terra Preta soils and much more abundant than previously thought (～40-50% of organic C). Our findings indicate that these oxidized char residues represent a particularly stable, abundant, and fertility-enhancing form of soil organic matter.