Estimation of Finite Population Variance Using Ratio and Product Methods of Estimation

For estimating finite population variance σ_y² of a character y under our study, estimators using auxiliary information on a character x in the form of ratio, product, ratio-type or product-type estimators have been suggested, and their comparative study with the conventional unbiased estimator s_y² of σ_y² has been made in simple random sampling with replacement. A generalized estimator representing a class of estimators for the finite populations variance, has also been studied.     

Micro Methods for Determination of 3-Ketosucrose and 3-Ketoglucose

Methods for differential determination of 3-ketosucrose and 3-ketoglucose were established. For determination of 3-ketosucrose, alkaline treatment with 0.1 N NaOH was found to be most effective. In this method, 3-ketosucrose gave a characteristic absorption spectrum with a molar extinction coefficient of 6.5 × 10³ m^?1cm^?1 at 340 mμ, while 3-ketoglucose did not show a significant absorption spectrum within a range from 300 to 400 mμ.

By mixing with 0.2 m phosphate buffer, pH 7.0, 3-ketoglucose gave a characteristic absorption spectrum with a molar extinction coefficient of 3.8 × 10³ m^?1cm^?1 at 310 mμ, while 3-ketosucrose showed little absorbance.

From the reduction rate of 2,6-dichloroindophenol with 3-ketoglucose, the ketosugar was determined. 3-Ketosucrose was not able to reduce the reagent at all.

The methods established here were not affected by fructose.     

Detection of Giardia in Environmental Waters by Immuno-PCR Amplification Methods

Genomic DNA was extracted either directly from Giardia muris cysts seeded into environmental surface waters or from cysts isolated by immunomagnetic beads (IMB). A 0.171-kbp segment of the giardin gene was PCR-amplified following “direct extraction” of Giardia DNA from seeded Cahaba river water concentrate with moderate turbidity (780 JTU's), but DNA purified from seeded Colorado river water concentrates with high turbidity (2 × 10⁵ JTUs) failed to amplify. However, if the cysts were first separated by the IMB approach from seeded Cahaba or Colorado river waters, and the DNA released by a freeze-boil Chelex?100 treatment, detection of G. muris by PCR amplification could be achieved at a sensitivity of 3 × 10⁰ or 3 × 10¹ cysts/ml, respectively. If, however, the G. muris cysts used to seed even moderately turbid river waters (780 JTUs) were formalin treated (which is conventionally used for microscopic examination), neither direct extraction nor IMB purification methods yielded amplifiable DNA. Use of immunomagnetic beads to separate Giardia cysts from complex matrices of environmental surface waters followed by DNA release and PCR amplification of the target giardin gene improved the reliability of detection of this pathogen with the required sensitivity. Received: 23 April 1997 / Accepted: 4 August 1997     

Screening Methods for Improving Rotifer Culture Quality

We studied whether the selection of rotifer B. plicatilis strain (Japanese, Russian or Australian), as well as the addition of gamma-aminobutyric acid (GABA) to the culture water, are useful in stabilising rotifer cultures. We examined the effect of a combination of the following stressors: unionized ammonia (2.4 mg l⁻¹), contamination by protozoa Euplotes sp. (10 cells ml⁻¹) and addition of methyl cellulose to increase the culture water viscosity at 15 cp. Rotifer reproductive tests and enzyme activity measurements (glucosidase) were conducted to determine the effect of the treatments. All tests were conducted at 25 °C and rotifers were fed Nannochloropsis oculata at 7 × 10⁶ cells ml⁻¹. The combined effects of the stressors caused a significant decrease in lifespan, fecundity and glucosidase activity. The effect of the stressors on reproductive characteristics and glucosidase activity could be neutralized if rotifers were treated with GABA.     

A Comparison of Different Estimation Methods for Fungicide EC50 and EC95 Values

EC₅₀ and EC₉₅ (the effective concentrations to cause inhibitions by 50 and 95%, respectively) are commonly used to express fungicide potency. Different methods are currently employed to calculate EC₅₀ and EC₉₅ values. In this study, EC₅₀ and EC₉₅ values for fungicide epoxiconazole against 34 isolates of Sclerotinia sclerotiorum were calculated with seven different methods. Results showed that for both EC₅₀ and EC₉₅ calculations, there was no significant difference among three statistical programs IBM spss ^®, GraphPad Prism^® and dps ^® (P ≥ 0.066). Methods linear log (linear regression of mycelial growth inhibition vs. logarithmic concentration) and interpolation log (linear interpolation from inhibition and logarithmic concentration data) were not significantly different (P ≥ 0.058) from IBM spss in EC₅₀ calculations. These results indicate that among the seven methods, the three statistical programs IBM spss , GraphPad Prism, dps and linear log method are appropriate for EC₅₀ calculations. But for EC₉₅ calculations, only the three statistical programs are recommended, and GraphPad Prism is likely to give a little higher values than spss and dps .     

Methods of Estimation of Visceral Fat: Advantages of Ultrasonography

Objective: To compare methods for the assessment of visceral fat with computed tomography (CT) and establish cutoffs to define visceral obesity based on such alternative methods. Research Methods and Procedures: One hundred women (50.4 ± 7.7 years; BMI 39.2 ± 5.4 kg/m²) underwent anthropometric evaluation, bioelectrical impedance, DXA, abdominal ultrasonography (US), and CT scan. Results: Waist circumference, waist‐to‐hip ratio (WHR), and US‐determined visceral fat values showed the best correlation coefficients with visceral fat determined by CT (r = 0.55, 0.54, and 0.71, respectively; p < 0.01). Fat mass determined by DXA was inversely correlated with visceral‐to‐subcutaneous‐fat ratio (r = ?0.47, p < 0.01). Bioimpedance‐determined fat mass and skinfolds were correlated with only subcutaneous abdominal fat quantified by CT. Linear regression indicated US visceral‐fat distance and WHR as the main predictors of CT‐determined visceral fat (adjusted r² = 0.51, p < 0.01). A waist measurement of 107 cm (82.7% specificity, 60.6% sensitivity) and WHR of 0.97 (78.8% specificity, 63.8% sensitivity) were chosen as discriminator values corresponding with visceral obesity diagnosed by CT. A value of 6.90 cm for visceral fat US‐determined diagnosed visceral obesity with a specificity of 82.8%, a sensitivity of 69.2%, and a diagnostic concordance of 74% with CT. Discussion: US seemed to be the best alternative method for the assessment of intra‐abdominal fat in obese women. Its diagnostic value could be optimized by an anthropometric measurement. Prospective studies are needed to establish CT and US cutoffs for defining visceral‐fat levels related to elevated cardiovascular risk.     

Comparison of Methods to Assess Body Composition Changes during a Period of Weight Loss

Objective: To assess the accuracy of body composition measurements by air displacement plethysmography and bioelectrical impedance analysis (BIA) compared with DXA during weight loss. Research Methods and Procedures: Fifty‐six healthy but overweight participants, 34 women and 22 men (age, 52 ± 8.6 years; weight, 92.2 ± 11.6 kg; BMI, 33.3 ± 2.9 kg/m²) were studied in an outpatient setting before and after 6 months of weight loss (weight loss, 5.6 ± 5.5 kg). Subjects were excluded if they had initiated a new drug therapy within 30 days of randomization, were in a weight loss program, or took a weight loss drug within 90 days of randomization. Subjects were randomly assigned either to a self‐help program, consisting of two 20‐minute sessions with a nutritionist and provision of printed materials and other self‐help resources, or to attendance at meetings of a commercial program (Weight Watchers). Body composition was examined by each of the methods before and after weight loss. Results: BIA (42.4 ± 5.8%) underestimated percentage fat, whereas the BodPod (Siri = 51.7 ± 6.9%; Brozek = 48.5 ± 6.5%) overestimated percentage fat compared with DXA (46.1 ± 7.9%) before weight loss. Correlation coefficients for detecting changes in body composition between DXA and the other methods were relatively high, with Brozek Δfat mass (FM; r² = 0.63), Siri FM (r² = 0.65), tetrapolar BIA percentage fat (r² = 0.57), and Tanita FM (r² = 0.61) being the highest. Discussion: In conclusion, all of the methods were relatively accurate for assessing body composition compared with DXA, although there were biases. Furthermore, each of the methods was sensitive enough to detect changes with weight loss.     

Comparative Evaluation of Quantitative Polymerase Chain Reaction Methods for Routine Enumeration of Specific Bacterial DNA in Aquatic Samples

Summary Three quantitative polymerase chain reaction (PCR) methods, the internal standard method (IS-PCR), competitive PCR (cPCR) and most probable number-PCR (MPN-PCR), were compared in terms of their ability to quantify specific bacterial DNA in environmental samples. Serially diluted Pseudomonas putida BH, the target bacterium, was inoculated into sterilized potassium phosphate buffer (PPB), river water and activated sludge, total DNA was extracted, and the number of pheB genes carried by P. putida BH in each sample was enumerated. IS-PCR and cPCR could not quantify the pheB gene at low concentrations (1.0 × 10³ copies ml^-1 in all samples and 1.0 × 10⁴ copies ml^--1 in some samples) and tended to give overestimations because of differences in amplification efficiencies between pheB gene and the internal standard/competitor in a reaction tube. Although reproducibility of MPN-PCR was slightly lower than that of the other two methods, MPN-PCR was the most sensitive, enabling us to quantify the pheB gene at 1.0 × 10³ copies ml^--1, and it had a good correlation with the inoculum size of P. putida BH. These results suggest that MPN-PCR is the best suited for routine microbial monitoring in natural environmental samples because of the simple handling, the ease of modification as occasion demands and the wide detection range, especially at low cell densities of the target microbe.     

Prediction of Side Chain Orientations in Proteins by Statistical Machine Learning Methods

Abstract

We develop ways to predict the side chain orientations of residues within a protein structure by using several different statistical machine learning methods. Here side chain orientation of a given residue i is measured by an angle Ω_i between the vector pointing from the center of the protein structure to the C^α _i atom and the vector pointing from the C^α _i atom to the center of its side chain atoms. To predict the Ω_i angles, we construct statistical models by using several different methods such as general linear regression, a regression tree and bagging, a neural network, and a support vector machine. The root mean square errors for the different models range only from 36.67 to 37.60 degrees and the correlation coefficients are all between 30% and 34%. The performances of different models in the test set are, thus, quite similar, and show the relative predictive power of these models to be significant in comparison with random side chain orientations.     

Comparison of Detection Methods for Legionella Species in Environmental Water by Colony Isolation,Fluorescent Antibody Staining,and Polymerase Chain Reaction

Three detection methods for Legionella species in water samples from cooling towers and a river were examined. Direct counting of bacteria stained with fluorescent antibody (FA) for L. pneumophila (serogroups 1 to 6) could detect the cell of 10⁴ to 10⁶ cell/100 ml in all 14 samples, while colony counting method detected 10 to 10³ CFU/100 ml only in 8 samples from cooling towers. Polymerase chain reaction (PCR) assay with primers to amplify 16S ribosomal DNA sequence of most Legionella species (LEG primer) detected legionellae in 13 samples, while species-specific primers for L. pneumophila detected the DNAs from 3 samples. In laboratory examination, LEG primers could amplify DNAs of 29 species of genus Legionella with high sensitivity, even from 1 cell of L. pneumophila GIFU 9134. The PCR assay with LEG primers was specific and sensitive methods to be satisfied the survey of legionellae. Thus, PCR assay is a suitable method to detect and monitor Legionella species in an environment.     

Impact of Chiral Bioanalytical Methods on the Bioequivalence of Ibuprofen Products Containing Ibuprofen Lysinate and Ibuprofen Base

The purpose was to assess the impact of the use of a chiral bioanalytical method on the conclusions of a bioequivalence study that compared two ibuprofen suspensions with different rates of absorption. A comparison of the conclusion of bioequivalence between a chiral method and an achiral approach was made. Plasma concentrations of R‐ibuprofen and S‐ibuprofen were determined using a chiral bioanalytical method; bioequivalence was tested for R‐ibuprofen and for S‐ibuprofen separately and for the sum of both enantiomers as an approach for an achiral bioanalytical method. The 90% confidence interval (90% CI) that would have been obtained with an achiral bioanalytical method (90% CI: C_max: 117.69–134.46; AUC₀^t: 104.75–114.45) would have precluded the conclusion of bioequivalence. This conclusion cannot be generalized to the active enantiomer (90% CI: C_max: 103.36–118.38; AUC₀^t: 96.52–103.12), for which bioequivalence can be concluded, and/or the distomer (90% CI: C_max: 132.97–151.33; AUC₀^t: 115.91–135.77) for which a larger difference was observed. Chiral bioanalytical methods should be required when 1) the enantiomers exhibit different pharmacodynamics and 2) the exposure (AUC or C_max) ratio of enantiomers is modified by a difference in the rate of absorption. Furthermore, the bioequivalence conclusion should be based on all enantiomers, since the distomer(s) might not be completely inert, in contrast to what is required in the current regulatory guidelines. In those cases where it is unknown if the ratio between enantiomers is modified by changing the rate of absorption, chiral bioanalytical methods should be employed unless enantiomers exhibit the same pharmacodynamics. Chirality 28:429–433, 2016. © 2016 Wiley Periodicals, Inc.     

Identification of Genes for Resistance to Bean Common Mosaic Virus and Bean Common Mosaic Necrosis Virus in Snap Bean (Phaseolus vulgaris L.) Breeding Lines Using Conventional and Molecular Methods

Bean common mosaic virus (BCMV) and Bean common mosaic necrosis virus (BCMNV) are among the biggest threats for snap bean production in Bulgaria due to their seed, aphid and mechanical transmission. Old valuable Bulgarian snap bean varieties are being neglected, because of the high percentage of virus‐infected seeds. Breeding resistant cultivars is the best way to solve the problem. The genetic control towards both viruses is assured by one dominant I gene and a number of recessive (bc‐u, bc‐1, bc‐1², bc‐2, bc‐2² and bc‐3) genes. Our aim was to identify resistance gene combinations in advanced F8 breeding lines, derived from two crosses (A‐8‐40‐7‐2‐1 × IVT 7214) and (Zaria × RH 26D), by the application of conventional and molecular approaches. Four methods were applied for the characterization of their resistance gene makeup: (i) leaf‐abscission infection test designed to identify I gene by direct inoculation with NL3 strain of BCMNV; (ii) intact‐plant infection test with strain NY15 of BCMV to separate immune genotypes, possessing bc‐ubc‐1², bc‐ubc‐2^2,bc‐ubc‐2bc‐3, I, Ibc‐1², Ibc‐2² or Ibc‐3; (iii) PCR analysis with the following markers: SCAR – SW13 (for I gene), SBD5 (for bc‐1²), ROC11 (for bc‐3) and CAPS – eIFE4 (for bc‐3); and 4) high‐temperature (more than 30°C) infection test with NL3 of BCMNV to provoke systemic necrosis in I, Ibc‐1, Ibc‐1², Ibc‐1²bc‐2² or Ibc‐3. The four methods applied worked properly and complemented each other. Valuable gene combination (Ibc‐3) was established in seven breeding lines with immune reaction to BCMNV. They will be included in the snap bean breeding programme for virus resistance.     

Green Chromatographic Methods and in Vitro Pharmacological Analysis of Varronia curassavica Leaf Derivatives

Varronia curassavica displays anti-inflammatory, antiulcerogenic, and antioxidant activities. Herein, we employed new UHPLC –UV green chromatographic methods for the analysis of in vitro antioxidant and anti-inflammatory activities of V. curassavica and its embryotoxicity in Zebrafish. Cordialin A, brickellin, and artemetin were purified from the ethanol (EtOH) extract of V. Curassavica leaves and identified using spectrometric techniques. In line with Green Analytical Chemistry principles, the proposed UHPLC methods involve the use of ethanol as organic modifier with low mobile phase consumption, and without sample pretreatment (OLE-UHPLC-UV). The application of the Agree and HPLC-EAT tools for greenness assessment yielded this pattern: HPLC-UV (reference)<UHPLC-UV<OLE-UHPLC-UV. Zebrafish assay results showed that 70 % EtOH extract of V. Curassavica leaves exhibited lower toxicity compared to 100 % EtOH extract, with LC₅₀ of 164.3 and 122.9 μg/mL, respectively, in 24 h post fertilization. Some embryos exhibited malformation phenotypes in the heart, somites, and eyes, mainly in higher extract concentrations. Extracts and brickellin exhibited higher antioxidant activity in the DPPH⋅ assay, while brickellin+artemetin displayed higher antioxidant activity compared to the extracts and isolated flavones in the O₂⋅⁻ and HOCl/OCl⁻ scavenging assays. Cordialin A and brickellin exhibited low COX-1, COX-2, and phospholipase A₂ inhibition.     

Evaluating Methods for Counting Cryptic Carnivores

ABSTRACT Numerous techniques have been proposed to estimate carnivore abundance and density, but few have been validated against populations of known size. We used a density estimate established by intensive monitoring of a population of radiotagged leopards (Panthera pardus) with a detection probability of 1.0 to evaluate efficacy of track counts and camera-trap surveys as population estimators. We calculated densities from track counts using 2 methods and compared performance of 10 methods for calculating the effectively sampled area for camera-trapping data. Compared to our reference density (7.33 ± 0.44 leopards/100 km²), camera-trapping generally produced more accurate but less precise estimates than did track counts. The most accurate result (6.97 ± 1.88 leopards/100 km²) came from camera-trap data with a sampled area buffered by a boundary strip representing the mean maximum distance moved by leopards outside the survey area (MMDMOSA) established by telemetry. However, contrary to recent suggestions, the traditional method of using half the mean maximum distance moved from photographic recaptures did not result in gross overestimates of population density (6.56 ± 1.92 leopards/100 km²) but rather displayed the next best performance after MMDMOSA. The only track-count method comparable to reference density employed a capture-recapture framework applied to data when individuals were identified from their tracks (6.45 ± 1.43 leopards/100 km²) but the underlying assumptions of this technique limit more widespread application. Our results demonstrate that if applied correctly, camera-trap surveys represent the best balance of rigor and cost-effectiveness for estimating abundance and density of cryptic carnivore species that can be identified individually.     

Methods and milliliter scale devices for high-throughput bioprocess design

Based on electromagnetic simulations as well as on computational fluid dynamics simulations gas-inducing impellers and their magnetic inductive drive were optimized for stirred-tank reactors on a 10 ml-scale arranged in a bioreaction block with 48 bioreactors. High impeller speeds of up to 4,000 rpm were achieved at very small electrical power inputs (63 W with 48 bioreactors). The maxima of local energy dissipation in the reaction medium were estimated to be up to 50 W L⁻¹ at 2,800 rpm. Total power input and local energy dissipation are thus well comparable to standard stirred-tank bioreactors. A prototype fluorescence reader for 8 bioreactors with immobilized fluorometric sensor spots was applied for online measurement of dissolved oxygen concentration making use of the phase detection method. A self-optimizing scheduling software was developed for parallel control of 48 bioreactors with a liquid-handling system for automation of titration and sampling. It was shown on the examples of simple parallel batch cultivations of Escherichia coli with different media compositions that high cell densities of up to 16.5 g L⁻¹ dry cell mass can be achieved without pH-control within 5 h with a high parallel reproducibility (standard deviation<3.5%, n=48) due to the high oxygen transfer capability of the gas-inducing stirred-tank bioreactors.     

Projection Methods for the Analysis of Complex Motions in Macromolecules

Abstract

In studies of macromolecular dynamics it is often desirable to analyze complex motions in terms of a small number of coordinates. Only for simple types of motion, e.g., rigid-body motions, these coordinates can be easily constructed from the Cartesian atomic coordinates. This article presents an approach that is applicable to infinitesimal or approximately infinitesimal motions, e.g., Cartesian velocities, normal modes, or atomic fluctuations. The basic idea is to characterize the subspace of interesting motions by a set of (possibly linearly dependent) vectors describing elementary displacements, and then project the dynamics onto this subspace. Often the elementary displacements can be found by physical intuition. The restriction to small displacements facilitates the study of complicated coupled motions and permits the construction of collective-motion subspaces that do not correspond to any set of generalized coordinates.

As an example for this technique, we analyze the low-frequency normal modes of proteins up to ≈ 20 THz (600 cm^?1) in order to see what kinds of motions occupy which frequency range. This kind of analysis is useful for the interpretation of spectroscopic measurements on proteins, e.g., inelastic neutron scattering experiments.     

Quadrupole Relaxation of the 7Li+ Ion in Dilute Aqueous Solution Determined by Experimental and Theoretical Methods

A combination of molecular dynamics simulations (MD), ab initio selfconsistent field (SCF) calculations and nuclear magnetic resonance relaxation time experiments (NMR) is a powerful battery of techniques to investigate the molecular origins of the nuclear quadrupole relaxation mechanism for ⁷Li⁺ ions in dilute aqueous solution.     

Insights into In Vitro Binding of Parecoxib to Human Serum Albumin by Spectroscopic Methods

Herein, we report the effect of parecoxib on the structure and function of human serum albumin (HSA) by using fluorescence, circular dichroism (CD), Fourier transforms infrared (FTIR), three‐dimensional (3D) fluorescence spectroscopy, and molecular docking techniques. The Stern–Volmer quenching constants K_SV and the corresponding thermodynamic parameters ΔH, ΔG, and ΔS have been estimated by the fluorescence quenching method. The results indicated that parecoxib binds spontaneously with HSA through van der Waals forces and hydrogen bonds with binding constant of 3.45 × 10⁴ M^?1 at 298 K. It can be seen from far‐UV CD spectra that the α‐helical network of HSA is disrupted and its content decreases from 60.5% to 49.6% at drug:protein = 10:1. Protein tertiary structural alterations induced by parecoxib were also confirmed by FTIR and 3D fluorescence spectroscopy. The molecular docking study indicated that parecoxib is embedded into the hydrophobic pocket of HSA.     

Efficacy of Conventional Endoscopic Disinfection and Sterilization Methods Against Helicobacter pylori Contamination

Background. Iatrogenic transmission of Helicobacter pylori via contaminated endoscopic devices is well documented. Despite the prevalence of this infectious agent, few controlled studies have investigated the major factors that impact on reprocessing of endoscopes contaminated with H. pylori. Materials and Methods. An endoscope (Pentax) was contaminated with 10⁸ cfu/ml of H. pylori in 5% bovine calf serum as standardized inoculum. The endoscope then was passed through one of eight arms (five repetitions per arm = 40 total runs), as follows: 1, recovery control (no cleaning or disinfection); 2, manual cleaning alone; 3–5, manual precleaning followed by either 10-, 20-, or 45-minute exposure to 2% glutaraldehyde and ethanol (ETOH) drying; 6, manual cleaning followed by automated reprocessing by STERIS System; 7 and 8, automated reprocessing by STERIS with and without active peracetic acid sterilant (wash-off control). Suction-biopsy channels and air-water channels were harvested for microbiological culture. Results. Control runs recovered more than 1 × 10⁶ cfu per site, confirming the viability of the test organism and the adequacy of the biological burden for challenge. When instruments underwent manual cleaning alone (without subsequent disinfection), test organisms remained in 40% of runs at the air-water site. Manual cleaning followed by 10-, 20-, or 45-minute glutaraldehyde exposure and ETOH drying removed all test organisms from all sites in all runs (i.e., 100% disinfection). The automated STERIS system with or without active peracetic acid sterilant also removed all test organisms from all sites in all runs, as did manual cleaning followed by STERIS use. Conclusion. Manual cleaning alone does not effectively remove H. pylori from an endoscope. Current joint association recommendations for minimal disinfection (manual cleaning followed by at least 20 minutes of immersion in glutaraldehyde and ETOH drying) are effective in preventing cross-transmission of H. pylori. Reprocessing using the automated STERIS system according to manufacturer's recommendations also is highly effective in sterilizing endoscopes contaminated with H. pylori.     

Modelling of Structural and Vibrational Properties of Poly(p-Phenylene) and Polypyrrole Using Molecular Orbital Methods

Abstract

This paper concentrates on two very important conducting polymers poly(p-phenylene) and polypyrrole. Detailed atomistic molecular models have been developed with the help of ab initio and semi-empirical quantum mechanical calculations using the Cerius² and WinMOPAC (version 6.0) programs.

Their optimised geometry had been calculated and compared with experimental X-ray diffraction data. The simulated and experimental vibrational spectra of biphenyl as well as isolated pyrrole monomers and oligomers from n = 1 and 2, where n is the number of structural repeat units used, have been computed using the ab initio 3–21G basis set. The results obtained are compared with experimental data for the case of biphenyl and for oligomers with n = 2 to 5 for both neutral benzenoid and quinonoid oligopyrroles, from semi-empirical predictions obtained by AM1 and PM3. The trends in the computed harmonic force fields, vibrational frequencies and intensities are monitored as a function of the chain length. The data are analyzed in conjunction with the trends in computed equilibrium geometries.