DGGE with genomic DNA: Suitable for detection of numerically important organisms but not for identification of the most abundant organisms

Identification of all important community members as well as of the numerically dominant members of a community are key aspects of microbial community analysis of bioreactor samples. A systematic study was conducted with artificial consortia to test whether denaturing gradient gel electrophoresis (DGCE) is a reliable technique to obtain such community data under conditions where results would not be affected by differences in DNA extraction efficiency from cells. A total of 27 consortia were established by mixing DNA extracted from Escherichia coli K12, Burkholderia cepacia and Stenotrophomonas maltophilia in different proportions. Concentrations of DNA of single organisms in the consortia were either 0.04, 0.4 or 4 ng/mu l. DGGE-PCR of genomic DNA with primer sets targeted at the V3 and V6-V8 regions of the 16S rDNA failed to detect the three community members in only 7% of consortia, but provided incorrect information about dominance or co-dominance for 85% and 89% of consortia with the primer sets for the V6-V8 and V3 regions, respectively. The high failure rate in detection of dominant B. cepacia with the primers for the V6-V8 region was attributable to a single nucleoticle primer mismatch in the target sequences of both, the forward and reverse primer. Amplification bias in PCR of E. coli and S. maltophilia for the V6-V8 region and for all three organisms for the V3 region occurred due to interference of genomic DNA in PCR-DGGE, since a nested PCR approach, where PCR-DGGE was started from mixtures of 16S rRNA genes of the organisms, provided correct information about the relative abundance of original DNA in the sample. Multiple bands were not observed in pure culture amplicons produced with the V6-V8 primer pair, but pure culture V3 DGGE profiles of E. coli, S. maltophilia and B. cepacia contained 5, 3 and 3 bands, respectively. These results demonstrate DGGE was suitable for identification of all important community members in the three-membered artificial consortium, but not for identification of the dominant organisms in this small community. Multiple DGGE bands obtained for single organisms with the V3 primer pair could greatly confound interpretation of DGGE profiles. (C) 2008 Elsevier Ltd. All rights reserved.

Improved voice activity detection algorithm using wavelet and support vector machine

This paper proposes an improved voice activity detection (VAD) algorithm using wavelet and support vector machine (SVM) for European Telecommunication Standards Institution (ETS1) adaptive multi-rate (AMR) narrow-band (NB) and wide-band (WB) speech codecs. First, based on the wavelet transform, the original IIR filter bank and pitch/tone detector are implemented, respectively, via the wavelet filter bank and the wavelet-based pitch/tone detection algorithm. The wavelet filter bank can divide input speech signal into several frequency bands so that the signal power level at each sub-band can be calculated. In addition, the background noise level can be estimated in each sub-band by using the wavelet de-noising method. The wavelet filter bank is also derived to detect correlated complex signals like music. Then the proposed algorithm can apply SVM to train an optimized non-linear VAD decision rule involving the sub-band power, noise level, pitch period, tone flag, and complex signals warning flag of input speech signals. By the use of the trained SVM, the proposed VAD algorithm can produce more accurate detection results. Various experimental results carried out from the Aurora speech database with different noise conditions show that the proposed algorithm gives considerable VAD performances superior to the AMR-NB VAD Options 1 and 2, and AMR-WB VAD. (C) 2009 Elsevier Ltd. All rights reserved.

Flow injection analysis of ethyl xanthate by gas diffusion and UV detection as CS(2) for process monitoring of sulfide ore flotation

A sensitive and robust analytical method for spectrophotometric determination of ethyl xanthate, CH(3)CH(2)OCS(2)(-) at trace concentrations in pulp solutions from froth flotation process is proposed. The analytical method is based on the decomposition of ethyl xanthate. EtX(-), with 2.0 mol L(-1) HCl generating ethanol and carbon disulfide. CS(2). A gas diffusion cell assures that only the volatile compounds diffuse through a PTFE membrane towards an acceptor stream of deionized water, thus avoiding the interferences of non-volatile compounds and suspended particles. The CS(2) is selectively detected by UV absorbance at 206 nm (epsilon = 65,000 L mol(-1) cm(-1)). The measured absorbance is directly proportional to EtX(-) concentration present in the sample solutions. The Beer`s law is obeyed in a 1 x 10(-6) to 2 x 10(-4) mol L(-1) concentration range of ethyl xanthate in the pulp with an excellent correlation coefficient (r = 0.999) and a detection limit of 3.1 x 10(-7) mol L(-1), corresponding to 38 mu g L. At flow rates of 200 mu L min(-1) of the donor stream and 100 mu L min(-1) of the acceptor channel a sampling rate of 15 injections per hour could be achieved with RSD < 2.3% (n = 10, 300 mu L injections of 1 x 10(-5) mol L(-1) EtX(-)). Two practical applications demonstrate the versatility of the FIA method: (i) evaluation the free EtX(-) concentration during a laboratory study of the EtX(-) adsorption capacity on pulverized sulfide ore (pyrite) and (ii) monitoring of EtX(-) at different stages (from starting load to washing effluents) of a flotation pilot plant processing a Cu-Zn sulfide ore. (C) 2010 Elsevier By. All rights reserved.

Studies on the kinetics of ascorbate oxidation at a ruthenium oxide hexacyanoferrate modified electrode towards the detection at microenvironments

The electrocatalytic oxidation of ascorbate on a ruthenium oxide hexacyanoferrate (RuOHCF) glassy carbon (GC) modified electrode was investigated at pH 6.9 by using rotating disc electrode (RDE) voltammetry. The influence of the systematic variation of rotation rate, film thickness, ascorbate concentration and the electrode potential indicated that the rate of cross-chemical reaction between Ru(III) centres immobilized into the film and ascorbate controls the overall process. The kinetic regime may be classified as a Sk `` mechanism and the second order rate constant for the surface electrocatalytic reaction was found to be 1.56 x 10(-3) mol(-1) L-1 s(-1) cm. A carbon fibre microelectrode modified with the RuOHCF film was successfully used as an amperometric sensor to monitor the ascorbate diffusion in a simulated microenvironment experiment. (C) 2008 Elsevier B.V. All rights reserved.

Environmental formaldehyde analysis by active diffusive sampling with a bundle of polypropylene porous capillaries followed by capillary zone electrophoretic separation and contactless conductivity detection

Compared to other volatile carbonylic compounds present in outdoor air, formaldehyde (CH2O) is the most toxic, deserving more attention in terms of indoor and outdoor air quality legislation and control. The analytical determination of CH2O in air still presents challenges due to the low-level concentration (in the sub-ppb range) and its variation with sampling site and time. Of the many available analytical methods for carbonylic compounds, the most widespread one is the time consuming collection in cartridges impregnated with 2,4-dinitrophenylhydrazine followed by the analysis of the formed hydrazones by HPLC. The present work proposes the use of polypropylene hollow porous capillary fibers to achieve efficient CH2O collection. The Oxyphan (R) fiber (designed for blood oxygenation) was chosen for this purpose because it presents good mechanical resistance, high density of very fine pores and high ratio of collection area to volume of the acceptor fluid in the tube, all favorable for the development of air sampling apparatus. The collector device consists of a Teflon pipe inside of which a bundle of polypropylene microporous capillary membranes was introduced. While the acceptor passes at a low flow rate through the capillaries, the sampled air circulates around the fibers, impelled by a low flow membrane pump (of the type used for aquariums ventilation). The coupling of this sampling technique with the selective and quantitative determination of CH2O, in the form of hydroxymethanesulfonate (HMS) after derivatization with HSO3-, by capillary electrophoresis with capacitively coupled contactless conductivity detection (CE-(CD)-D-4) enabled the development of a complete analytical protocol for the CH2O evaluation in air. (C) 2008 Published by Elsevier B.V.

Evaluation of Red Colour Segmentation Algorithms in Traffic Signs Detection

Colour segmentation is the most commonly used method in road signs detection. Road sign contains several basic colours such as red, yellow, blue and white which depends on countries.The objective of this thesis is to do an evaluation of the four colour segmentation algorithms. Dynamic Threshold Algorithm, A Modification of de la Escalera’s Algorithm, the Fuzzy Colour Segmentation Algorithm and Shadow and Highlight Invariant Algorithm. The processing time and segmentation success rate as criteria are used to compare the performance of the four algorithms. And red colour is selected as the target colour to complete the comparison. All the testing images are selected from the Traffic Signs Database of Dalarna University [1] randomly according to the category. These road sign images are taken from a digital camera mounted in a moving car in Sweden.Experiments show that the Fuzzy Colour Segmentation Algorithm and Shadow and Highlight Invariant Algorithm are more accurate and stable to detect red colour of road signs. And the method could also be used in other colours analysis research. The yellow colour which is chosen to evaluate the performance of the four algorithms can reference Master Thesis of Yumei Liu.

Hand Gesture Detection & Recognition System

The project introduces an application using computer vision for Hand gesture recognition. A camera records a live video stream, from which a snapshot is taken with the help of interface. The system is trained for each type of count hand gestures (one, two, three, four, and five) at least once. After that a test gesture is given to it and the system tries to recognize it.A research was carried out on a number of algorithms that could best differentiate a hand gesture. It was found that the diagonal sum algorithm gave the highest accuracy rate. In the preprocessing phase, a self-developed algorithm removes the background of each training gesture. After that the image is converted into a binary image and the sums of all diagonal elements of the picture are taken. This sum helps us in differentiating and classifying different hand gestures.Previous systems have used data gloves or markers for input in the system. I have no such constraints for using the system. The user can give hand gestures in view of the camera naturally. A completely robust hand gesture recognition system is still under heavy research and development; the implemented system serves as an extendible foundation for future work.

Modeling Of District Metered Areas With Relatively High Leakage Rate. The Case Study Of Kalipoli’s DMA

The article reviews the modelling of District Metered Areas (DMAs) with relatively high leakage rate. As a generally recognised approach in modelling of leakage does not exist, modelling of leakage by enginners and other researchers usually takes place by dividing the whole leakage rate evenly to all available nodes of the model. In this article, a new methodology is proposed to determine the nodal leakage by using a hydraulic model. The proposed methodology takes into consideration the IWA water balance methodology, the Minimum Night Flow (MNF) analysis, the number of connections related to each node and the marerial of pipes. In addition, the model is illustrated by a real case study, as it was applied in Kalipoli’s DMA. Results show that the proposed model gives reliable results.

An improved and fast UHPLC-PDA methodology for determination of L-ascorbic and dehydroascorbic acids in fruits and vegetables: evaluation of degradation rate during storage

This study provides a versatile validated method to determine the total vitamin C content, as the sum of the contents of L-ascorbic acid (L-AA) and dehydroascorbic acid (DHAA), in several fruits and vegetables and its degradability with storage time. Seven horticultural crops from two different origins were analyzed using an ultrahigh-performance liquid chromatographic–photodiode array (UHPLC-PDA) system, equipped with a new trifunctional high strength silica (100% silica particle) analytical column (100 mm×2.1 mm, 1.7 μm particle size) using 0.1% (v/v) formic acid as mobile phase, in isocratic mode. This new stationary phase, specially designed for polar compounds, overcomes the problems normally encountered in HPLC and is suitable for the analysis of large batches of samples without L-AA degradation. In addition, it proves to be an excellent alternative to conventional C18 columns for the determination of L-AA in fruits and vegetables. The method was fully validated in terms of linearity, detection (LOD) and quantification (LOQ) limits, accuracy, and inter/intraday precision. Validation experiments revealed very good recovery rate of 96.6±4.4% for L-AA and 103.1±4.8 % for total vitamin C, good linearity with r2-values >0.999 within the established concentration range, excellent repeatability (0.5%), and reproducibility (1.6%) values. The LOD of the method was 22 ng/mL whereas the LOQ was 67 ng/mL. It was possible to demonstrate that L-AA and DHAA concentrations in the different horticulture products varied oppositely with time of storage not always affecting the total amount of vitamin C during shelf-life. Locally produced fruits have higher concentrations of vitamin C, compared with imported ones, but vegetables showed the opposite trend. Moreover, this UHPLC-PDA methodology proves to be an improved, simple, and fast approach for determining the total content of vitamin C in various food commodities, with high sensitivity, selectivity, and resolving power within 3 min of run analysis.

Analysis of Factor VIII polymorphic markers as a means for carrier detection in Brazilian families with haemophilia A

Haemophilia A is an X-linked, recessively inherited bleeding disorder of varying severity, which results from the deficiency of procoagulant factor VIII f(8). Linkage diagnosis using polymorphic markers in the f8 gene is widely used to detect carriers. The objective of this study was to verify the informativeness of three polymorphic markers in the Brazilian population, to evaluate the usefulness of such markers in carrier detection procedures. Sixty-three unrelated healthy volunteers and 10 haemophilic families were studied. Two microsatellite repeats and one HindIII RFLP markers were used. Carrier and non-carrier status could be determined in 80% of females investigated. Intron 13 markers presented the highest heterozygosity rate (79%) followed by intron 22 (68%) and intron 19 (57%). When all three markers were used together, linkage analysis informativeness increased significantly. We conclude that these markers are suitable for carrier detection in the Brazilian population and we recommend their use in combination to maximize diagnostic efficiency.

Pyroelectric composite film for X-ray intensity detection

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Evalution of methods for the detection of MRSA in patients of Botucatu Medical School Hospital, São Paulo, Brazil

Oxacillin is the main drug of choice for the treatment of S. aureus infections. However, S. aureus resistance to oxacillin has become a major problem in the recent decades. The study aimed assess the rates of oxacillin resistance in S. aureus samples obtained at the Botucatu Medical School Hospital, UNESP, and to compare phenotypic techniques for the detection of MRSA against the gold standard method (mecA gene detection) in these samples. A total of 102 samples, previously isolated between 2002 and 2006, and kept at the Culture Collection of the Department of Microbiology and Immunology, in the Botucatu Biosciences Institute, UNESP, were included. Oxacillin resistance was assessed by oxacillin and cefoxitin disk diffusion and agar dilution tests, screening tests using Mueller-Hinton agar with 6 mu g/mL of oxacillin and 4% NaCl, E-test, and mecA gene detection. of the samples analyzed, 46 (45.1%) were mecA-positive. Oxacillin disk sensitivity and specificity were 86.9% and 91.1%, respectively. Cefoxitin disk sensitivity and specificity were respectively 91.3% and 91.1%. The screening test with the cefoxitin disk showed almost the same level of sensitivity (91.3%) and specificity (91.1%). With E-test strips, sensitivity was higher (97.8%) and specificity was comparable to that found with the other methods (91.1%). Ninety-three percent of the samples produced beta-lactamase and five of them were mecA-negative. There was a gradual increase in the number of oxacillin-resistant S. aureus samples between 2002 and 2004. However, from 2004 to 2006, the number of resistant samples dropped from 55% of MRSA in 2004, to 45% in 2005 and 34.6% in 2006. The data obtained reveal that, among phenotypic methods, the E-test yielded the best results, with higher sensitivity levels when compared to the other methods. The decreased resistance rate observed over the most recent years may be explained by the rational use of antimicrobial agents associated with good practices in the control of hospital infection, or may be related to the diminished use of oxacillin as a treatment option.

Effect of Ionic Liquid on the Determination of Aromatic Amines as Contaminants in Hair Dyes by Liquid Chromatography Coupled to Electrochemical Detection

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Sequential injection analysis system with spectrophotometric detection for determination of norfloxacin and ciprofloxacin in pharmaceutical formulations

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Analysis of aromatic amines in surface waters receiving wastewater from a textile industry by liquid chromatographic with electrochemical detection

A high performance liquid chromatography ( HPLC) method with electrochemical detection (ED) was developed for the determination of benzidine, 3,3-dimethylbenzidine, o-toluidine and 3,3-dichlorobenzidine in the wastewater of the textile industry. The aromatic amines were eluted on a reversed phase column Shimadzu Shimpack C-18 using acetonitrile + ammonium acetate (1 x 10(-4) mol L-1) at a ratio 46: 54 v/v as mobile phase, pumped at a flow rate of 1.0 mL min(-1). The electrochemical oxidation of the aromatic amines exhibits well-defined peaks at a potential range of +0.45 to +0.78 V on a glassy carbon electrode. Optimum working potentials for amperometric detection were from 0.70 V to +1.0 V vs. Ag/AgCl. Analytical curves for all the aromatic amines studied using the best experimental conditions present linear relationship from 1 x 10(-8) mol L-1 to 1.5 x 10(-5) mol L-1, r = 0.99965, n = 15. Detection limits of 4.5 nM (benzidine), 1.94 nM (o-toluidine), 7.69 nM (3,3-dimethylbenzidine), and 5.15 nM (3,3-dichlorobenzidine) were achieved, respectively. The detection limits were around 10 times lower than that verified for HPLC with ultra violet detection. The applicability of the method was demonstrated by the determination of benzidine in wastewater from the textile industry dealing with an azo dye processing plant.