bluuebunny/crossref_metadata_2025 huggingface dataset:
Comparison of Program Costs for Parent‐Only and Family‐Based Interventions for Pediatric Obesity in Medically Underserved Rural Settings
Authors: Janicke David M. , Sallinen Bethany J. , Perri Michael G. , Lutes Lesley D. , Silverstein Janet H. , Brumback Babette
Published: June 2009

ABSTRACT: Purpose:To compare the costs of parent‐only and family‐based group interventions for childhood obesity delivered through Cooperative Extension Services in rural communities.Methods:Ninety‐three overweight or obese children (aged 8 to 14 years) and their parent(s) participated in this randomized controlled trial, which included a 4‐month intervention and 6‐month follow‐up. Families were randomized to either a behavioral family‐based intervention (n = 33), a behavioral parent‐only intervention (n = 34), or a waitlist control condition (n = 26). Only program costs data for the parent‐only and family‐based programs are reported here (n = 67). Assessments were completed at baseline, post‐treatment (month 4) and follow‐up (month 10). The primary outcome measures were total program costs and cost per child for the parent‐only and family interventions.Findings:Twenty‐six families in the parent‐only intervention and 24 families in the family intervention completed all 3 assessments. As reported previously, both intervention programs led to significantly greater decreases in weight status relative to the control condition at month 10 follow‐up. There was no significant difference in weight status change between the parent‐only and family interventions. Total program costs for the parent‐only and family interventions were $13,546 and $20,928, respectively. Total cost per child for the parent‐only and family interventions were $521 and $872, respectively.Conclusions:Parent‐only interventions may be a cost‐effective alternative treatment for pediatric obesity, especially for families in medically underserved settings.
From Neighbourhood to Network: A Review of the Significance of Neighbourhood in Studies of Social Relations
Authors: Clark Andrew
Published: June 2009

This article reviews changing approaches to the idea of neighbourhood communities in social science research. It specifically considers the implications for thinking about the neighbourhood either as the settings where social relations and communal bonds are produced or as non‐geographical networks. There is a long history of territorial‐based community studies that have focused on everyday life in neighbourhood locations; most typically, working‐class urban neighbourhoods or edge‐of‐city housing estates. Research exploring networked communities in contrast considers ways in which individuals are caught up in webs of networked organisations and individuals that stretch across space and seemingly render place less relevant to community formation. The review outlines some for the challenges of these approaches to the idea of community and cautions against dismissing the significance of neighbourhood in the ways in which individuals construct communities of networks, contacts and relations with others.
Dye–Surfactant Interaction: Modulation of Photophysics of an Ionic Styryl Dye
Authors: Sahoo D. , Chakravorti S.
Published: June 2009

The interesting modulation of multibond rotation–induced intramolecular charge transfer photophysics of 2‐(4‐(dimethylamino) styryl)‐1‐methylpyridinium iodide in different micelles due to different contributions of twisted intramolecular charge transfer (TICT) and hydrogen bonding deactivation channels have been reported in this paper. 2‐(4‐(dimethylamino) styryl)‐1‐methylpyridinium iodide enters into all the micelles in different positions from the water solution due to active hydrophobic force and electrostatic field, as revealed from the shift and intensity of intramolecular charge transfer (ICT) band. The presence of mechanically trapped water with the addition of salt and inherent thermodynamic water controls the ICT emission. Analysis of spectral data before and after the addition of salt confirms the orientation of 2‐(4‐(dimethylamino) styryl)‐1‐methylpyridinium iodide in cationic and anionic micelles.
Limited Proteolysis of Luciferase as a Reporter in Nanosystem Biology: A Comparative Study
Authors: Ataei Farangis , Hosseinkhani Saman , Khajeh Khosro
Published: June 2009

Firefly luciferase is a 62 kDa protein that produces a flash of light on the oxidation of luciferin in the presence of ATP, Oxygen and Mg2+. Luciferase has a broad range of applications for analytical purposes and in vivo imaging for nanosystem biology studies. However, the enzyme is highly susceptible to proteolytic degradation that reduces its half‐life. Rate of proteolytic digestion between two members of luciferase family (Photinus pyralis and Lampyris turkestanicus) is compared. Proteolytic sensitivity of L. turkestanicus luciferase was found to be more than P. pyralis luciferase, due to higher rate of hydrolysis under identical conditions. Both luciferases showed more sensitivity to chymotrypsin than trypsin with different digestion pattern. Digestion of P. pyralis by trypsin produced some fragments which were found to be more resistant to further degradation, whereas in L. turkestanicus initial fragments subdigested by trypsin, like chymotrypsin effect on both luciferases. Furthermore, both luciferases become increasingly labile to proteolysis as the temperature increases. The rate of inactivation and the rate of degradation between luciferases were different in a specific time of incubation. Appearance of similar bands for both luciferases confirmed exposure of specific regions, in spite of structural differences.
A regional approach to plant DNA barcoding provides high species resolution of sedges (Carex and Kobresia, Cyperaceae) in the Canadian Arctic Archipelago
Authors: CLERC‐BLAIN JESSICA LE , STARR JULIAN R. , BULL ROGER D. , SAARELA JEFFERY M.
Published: June 2009

Previous research on barcoding sedges (Carex) suggested that basic searches within a global barcoding database would probably not resolve more than 60% of the world’s some 2000 species. In this study, we take an alternative approach and explore the performance of plant DNA barcoding in the Carex lineage from an explicitly regional perspective. We characterize the utility of a subset of the proposed protein‐coding and noncoding plastid barcoding regions (matK, rpoB, rpoC1, rbcL, atpF‐atpH, psbK‐psbI) for distinguishing species of Carex and Kobresia in the Canadian Arctic Archipelago, a clearly defined eco‐geographical region representing 1% of the Earth’s landmass. Our results show that matK resolves the greatest number of species of any single‐locus (95%), and when combined in a two‐locus barcode, it provides 100% species resolution in all but one combination (matK + atpFH) during unweighted pair‐group method with arithmetic mean averages (UPGMA) analyses. Noncoding regions were equally or more variable than matK, but as single markers they resolve substantially fewer taxa than matK alone. When difficulties with sequencing and alignment due to microstructural variation in noncoding regions are also considered, our results support other studies in suggesting that protein‐coding regions are more practical as barcoding markers. Plastid DNA barcodes are an effective identification tool for species of Carex and Kobresia in the Canadian Arctic Archipelago, a region where the number of co‐existing closely related species is limited. We suggest that if a regional approach to plant DNA barcoding was applied on a global scale, it could provide a solution to the generally poor species resolution seen in previous barcoding studies.
Nucleolar size in choroidal and ciliary body melanomas and corresponding hepatic metastases
Authors: Al‐Jamal Rana’a T. , Toivonen Päivi , Kivelä Tero
Published: June 2009

Purpose:  This study aimed to investigate the relationship between hepatic metastasis and the mean diameter of the 10 largest nucleoli (MLN) in uveal melanoma.

Methods:  A cross‐sectional histopathological analysis of 37 metastases (13 surgical or needle biopsies, 24 autopsies) and corresponding primary choroidal and ciliary body melanomas was conducted, using statistical tests appropriate for paired data. The largest nucleoli were measured from digital photographs of silver‐stained sections along a 5‐mm‐wide linear field. Confounders considered were presence of epithelioid cells and microvascular density (MVD), counted as the number of discrete elements labelled by monoclonal antibody QBEND/10 to the CD34 epitope.

Results:  Hepatic metastases had more frequent epithelioid cells (p = 0.0047) and a higher MVD (median difference, 7.5 counts/0.313 mm2 more; p = 0.044) than their corresponding primary tumours. Hepatic metastases, especially in autopsy specimens rather than surgical biopsies, tended to have a smaller MLN (median 3.6 μm) than the corresponding primary tumour (median difference, 0.55 μm; p = 0.066). The MLN in hepatic metastases was not associated with presence of epithelioid cells and MVD. Overall survival after diagnosis of metastasis was comparable whether hepatic metastases had a large or small MLN (p = 0.95), whereas a high MVD tended to be associated with shorter survival (p = 0.096) among the 13 patients with known survival.

Conclusions:  The results suggest that MLN is not a useful marker for assessing prognosis after diagnosis of hepatic metastasis from uveal melanoma.
The inhibitory effect of different concentrations of topical bevacizumab on corneal neovascularization
Authors: Habot‐Wilner Zohar , Barequet Irina S. , Ivanir Yair , Moisseiev Joseph , Rosner Mordechai
Published: June 2009

Acta Ophthalmol. 2010: 88: 862–867

Purpose:  This study aimed to evaluate the effects of different concentrations of topically administered bevacizumab (Avastin) on experimental corneal neovascularization (NV) in rats.

Methods:  Corneal NV was induced by chemical cauterization with silver nitrate sticks applied to the centre of the corneas of 37 Wistar rats. The rats were then randomized to four topical treatment groups: group 1 (n = 10) received 4 mg/ml bevacizumab; group 2 (n = 9) received 2 mg/ml bevacizumab; group 3 (n = 10) received 1 mg/ml bevacizumab, and group 4 (n = 8) represented a control group and received saline. All drops were initiated immediately after cauterization and applied twice per day for 7 days. Corneal NV was assessed 8 days after cauterization in a masked fashion, both qualitatively by clinical evaluation and quantitatively by blood vessel count in photographs of histological sections.

Results:  On clinical evaluation, groups 1 and 2 showed significantly less NV compared with the saline‐treated control group (p = 0.006 and p = 0.024, respectively). Histopathological evaluation showed that only group 1 differed significantly from controls (5% significance level) and normal corneal epithelium was seen in all groups.

Conclusions:  Topically administered bevacizumab at a concentration of 4 mg/ml significantly reduces corneal NV according to both clinical and histopathological evaluations; lower concentrations were less effective on both parameters. No corneal epitheliopathy was found using these concentrations.
A morphometric analysis of the Bonatea speciosa complex (Orchidaceae) and its implications for species boundaries
Authors: Ponsie Mariaan E. , Johnson Steven D. , Edwards Trevor J.
Published: June 2009

Species boundaries in rapidly evolving lineages are often blurred, necessitating analysis of the full range of variation within a complex in order to identify traits that can be used to diagnose taxa. The delimitation of Bonatea speciosa (L. f.) Willd. has been controversial, as most, but not all, recent authors have included within the concept of this species several additional taxa that were formerly recognised at the species level, including Bonateaboltonii (Harv.) Bolus and Bonateaantennifera Rolfe. Morphological variation across the Bonateaspeciosa complex (Orchidaceae) was explored using principal component and cluster analysis. Scatter plots were used to identify characters that could potentially be used to diagnose taxa. These data support the recognition of three distinct species, B.speciosa, B.antennifera and B.boltonii, instead of a single entity. A taxonomic account of southern African taxa within the B. speciosa complex is presented.
Bio-Medico-Mechanical Behavior of Natural Artery Blood Vessel Under Constant and Variable Internal Pulsatile Pressure Flow Test In Vitro
Authors: Yokobori A. T. , Maeyama T. , Ohkuma T. , Yokobori T. , Ohuchi H. , Nara S. , Sasaki H. , Kasai M.
Published: June 2009

Studies have been carried out on the bio-medico-mechanical behavior in vitro of natural blood vessel (dogs) under constant and variable internal pulsatile pressure flow. The apparatus designed by us well simulated the arterial system. The studies were made for the case of pressure amplitude kept as constant, of the two-step-multi-duplicated pulsatile pressure and of the fluctuating pressure. For the case of the fluctuating pressure, the strength of the artery becomes considerably lower than those under constant amplitude and two-step-multi-duplicated pulsatile pressure. SEM observations of the inner walls of the artery shows that collagen fibers are more elongated under fluctuating pulsatile pressure flow. In conclusion, in order to avoid the mechanical deterioration of the artery strength, it is useful to keep the pulsatile blood pressure at constant amplitude. Even for the case of the blood pressure fluctuation, it is necessary to manage to keep the blood pressure as near a regular wave as possible, the total number of repeated pulse being equal.
Assessment of Fiber Strength in a Urinary Bladder by Using Experimental Pressure Volume Curves: An Analytical Method
Authors: To¨zeren A.
Published: June 2009

In the present study, an analytical method is developed to deduce the constitutive equations of fibers embedded in a thick shell from the time-variant pressure volume curves obtained by experimental procedures. It is assumed that the spherical shell under consideration is composed of a fiber reinforced material and undergoes radial deflection, modeling the behavior of some biological shells such as urinary bladder. The fiber stress is expressed as a function of fiber strain, rate of strain and the degree of biochemical activation. The function form is chosen such that equations of mechanical equilibrium can be integrated analytically to yield chamber pressure as a function of chamber volume, time rate of change of volume and activation. Arbitrary coefficients appearing in the fiber stress-equation are also present in the resultant time-variant pressure-volume relation. These coefficients can be determined by curve-fitting commonly used clinical data such as cystometry measurements.
Adaptive Thermal Modeling: A Concept for Measurement of Local Blood Perfusion in Heated Tissues
Authors: Arkin H. , Chen M. M. , Holmes K. R.
Published: June 2009

A new Adaptive Thermal Modeling (ATM) method for the measurement of local tissue blood perfusion rate is introduced. The method is based on a two-phase numerical technique. The first phase includes a fast, finite difference scheme for solution of the transient temperature field. The second phase involves iterative corrections of the perfusion until the modeled temperatures coincide with those measured by the temperature sensors. The results obtained from computer generated “data”, as well as from laboratory experiments demonstrate the potential capability of the ATM method to continuously measure local perfusion rates in heated tissues. Rigorous analysis of the technique is planned for the near future so that it can be applied to in vivo measurements of local tissue blood perfusions.
A Method for the Identification of In-Vivo Segmental Stiffness Properties of the Spine
Authors: Vanderby R. , Daniele M. , Patwardhan A. , Bunch W.
Published: June 2009

A numerical algorithm is used to estimate in-vivo segmental stiffness properties of individual spine segments based upon existing load-displacement data. A static nonlinear finite element model stimulates a pathological spine and corrective instrumentation system. A systematic procedure for establishing the model’s stiffness parameters is described, in the form of a nonlinear constrained optimal design problem. The numerical method is demonstrated using as an example a case of adolescent idiopathic scoliosis requiring corrective surgery.
An Apparatus to Study the Response of Cultured Endothelium to Shear Stress
Authors: Viggers R. F. , Wechezak A. R. , Sauvage L. R.
Published: June 2009

An apparatus which has been developed to study the response of cultured endothelial cells to a wide range of shear stress levels is described. Controlled laminar flow through a rectangular tube was used to generate fluid shear stress over a cell-lined coverslip comprising part of one wall of the tube. A finite element method was used to calculate shear stresses corresponding to cell position on the coverslip. Validity of the finite element analysis was demonstrated first by its ability to generate correctly velocity profiles and wall shear stresses for laminar flow in the entrance region between infinitely wide parallel plates (two-dimensional flow). The computer analysis also correctly predicted values for pressure difference between two points in the test region of the apparatus for the range of flow rates used in these experiments. These predictions thus supported the use of such an analysis for three-dimensional flow. This apparatus has been used in a series of experiments to confirm its utility for testing applications. In these studies, endothelial cells were exposed to shear stresses of 60 and 128 dynes/cm2. After 12 hr at 60 dynes/cm2, cells became aligned with their longitudinal axes parallel to the direction of flow. In contrast, cells exposed to 128 dynes/cm2 required 36 hr to achieve a similar reorientation. Interestingly, after 6 hr at 128 dynes/cm2, specimens passed through an intermediate phase in which cells were aligned perpendicular to flow direction. Because of its ease and use and the provided documentation of wall shear stress, this flow chamber should prove to be a valuable tool in endothelial research related to atherosclerosis.
A Flow System for the Study of Shear Forces Upon Cultured Endothelial Cells
Authors: Koslow A. R. , Stromberg R. R. , Friedman L. I. , Lutz R. J. , Hilbert S. L. , Schuster P.
Published: June 2009

A parallel plate chamber in a flow system has been designed to study the effects of fluid shear stresses on cells. The system was applied to the study of cultured endothelial cells grown on cover slips which were accommodated in recessed wells in the base plate. Dye injection studies in the chamber indicated laminar flow over the cells. Shear rates measured over the cover slips by an electrochemical technique were found to be linear with flow rate. Laser doppler anemometry showed parabolic profiles between the plates. Endothelial cells subjected to flow showed a correlation between the time required for orientation and the magnitude of the shear stress.
Mass Transfer to Fluids Flowing Through Rotating Nonaligned Straight Tubes
Authors: Berman J. , Mockros L. F.
Published: June 2009

Relatively inefficient heat/mass transfer is characteristic of tubular devices if the Reynolds number is low. One method of improving the heat/mass transfer efficiency of such devices is by inducing transverse laminar secondary circulations that are superimposed on the primary flow field; the resulting transverse velocity components lead to fluid mixing and hence augmented mass transfer in the tube lumen. The present work is a theoretical and experimental investigation of the enhanced transport in rotating, nonaligned, straight tubes, a method of transport enhancement that utilizes Coriolis acceleration to create transverse fluid mixing. This technique couples the transport advantages of coiled tubes with the design advantages of straight tubes. The overall mass balance equation is numerically solved for transfer into fluids flowing steadily through rotating nonaligned straight tubes. This solution, for small Coriolis disturbances, incorporates a third order perturbation solution for the primary and secondary flow fields. For sufficiently small Coriolis disturbances the bulk concentration increase is found to be uniquely determined by the value of a single similarity parameter. As the Coriolis disturbance is increased, however, two additional parameters are required to accurately characterize the mass transfer. In general, increasing the Coriolis accelerations results in an increase in mass transfer. There are solution regimes, however, in which increasing this acceleration can lead to a decrease in mass transfer efficiency. This interesting phenomena, which has important design implications, appears to result from velocity-weighting effects on the exiting sample. Experiments, involving the measurement of oxygen transferred into water and blood, produced data that agree with the theoretical predictions.
Design of a Portable Artificial Heart Drive System Based on Efficiency Analysis
Authors: Kitamura T.
Published: June 2009

This paper discusses a computer simulation of a pneumatic portable piston-type artificial heart drive system with a linear d-c-motor. The purpose of the design is to obtain an artificial heart drive system with high efficiency and small dimensions to enhance portability. The design employs two factors contributing the total efficiency of the drive system. First, the dimensions of the pneumatic actuator were optimized under a cost function of the total efficiency. Second, the motor performance was studied in terms of efficiency. More than 50 percent of the input energy of the actuator with practical loads is consumed in the armature circuit in all linear d-c-motors with brushes. An optimal design is: the piston cross-sectional area of 10.5 cm2 cylinder longitudinal length of 10 cm. The total efficiency could be up to 25 percent by improving the gasket to reduce the frictional force.
Measurement of Wall Motion and Wall Shear in a Compliant Arterial Cast
Authors: Deters O. J. , Bargeron C. B. , Mark F. F. , Friedman M. H.
Published: June 2009

Initial measurements of the time-varying wall shear rate at two sites in a compliant cast of a human aortic bifurcation are presented. The shear rates were derived from flow velocities measured by laser Doppler velocimetry (LDV) near the moving walls of the cast. To derive these shear rate values, the distance from the velocimeter sampling volume to the cast wall must be known. The time variation of this distance was obtained from LDV measurements of the velocity of the wall itself.
Oscillometric Determination of Diastolic, Mean and Systolic Blood Pressure—A Numerical Model
Authors: Forster F. K. , Turney D.
Published: June 2009

A theoretical model of oscillometric blood pressure measurement is presented. Particular emphasis is paid to the collapse behavior of the artery, and an exponential volume-pressure curve is used. The results of this study suggest that mean blood pressure can be accurately predicted from the peak of the oscillometric curve if corrections related to the cuff pressure waveform are applied. It is also shown, however, that systolic and diastolic pressure may not in general be accurately determined from fixed amplitude ratios based on the oscillometric peak due to the sensitivity of the method to variations in blood pressure waveform, pulse pressure, and arterial compliance. No simple procedures are found to correct for these effects.
Asymptotic and Dimensionless Analysis of the Response of Living Tissue to Surgical Pulsed CO2 Lasers
Authors: Armon E. , Laufer G.
Published: June 2009

The mathematical model of the interaction between the radiation of pulsed CO2 lasers and tissue was revisited. Asymptotic calculations were employed to determine upper and lower bounds for the evaporated volume and crater depth. Dimensionless time variables for conduction, beam attenuation by tissue vapors and for damage were introduced. Optimal exposure parameters were identified through a dimensionless analysis.
Finite Deformation of Soft Tissue: Analysis of a Mixture Model in Uni-Axial Compression
Authors: Holmes M. H.
Published: June 2009

The dynamic finite deformational behavior of a biphasic model for soft hydrated tissue is examined. In the case of uni-axial confined compression the displacement and stress fields are derived for steady-state permeation, creep, and stress-relaxation. It is shown how to use the results of this analysis to obtain the constitutive relations, as well as the associated material parameters, from the corresponding experiments. It is also shown that the solutions from the theory go much farther, giving a detailed account of the deformation and iteraction of the fluid and solid phases in the tissue.
Escherichia coli
O157:H7 Strain Origin, Lineage, and Shiga Toxin 2 Expression Affect Colonization of Cattle
Authors: Lowe Ross M. S. , Baines Danica , Selinger L. Brent , Thomas James E. , McAllister Tim A. , Sharma Ranjana
Published: June 2009

Enterohemorrhagic
Escherichia coli
O157:H7 has evolved into an important human pathogen with cattle as the main reservoir. The recent discovery of
E. coli
O157:H7-induced pathologies in challenged cattle has suggested that previously discounted bacterial virulence factors may contribute to the colonization of cattle. The objective of the present study was to examine the impact of lineage type, cytotoxin activity, and cytotoxin expression on the amount of
E. coli
O157:H7 colonization of cattle tissue and cells in vitro. Using selected bovine- and human-origin strains, we determined that lineage type predicted the amount of
E. coli
O157:H7 strain colonization: lineage I > intermediate lineages > lineage II. All
E. coli
O157:H7 strain colonization was dose dependent, with threshold colonization at 10
3
to 10
5
CFU and maximum colonization at 10
7
CFU. We also determined that an as-yet-unknown factor of strain origin was the most dominant predictor of the amount of strain colonization in vitro. The amount of
E. coli
O157:H7 colonization was also influenced by strain cytotoxin activity and the inclusion of cytotoxins from lineage I or intermediate lineage strains increased colonization of a lineage II strain. There was a higher level of expression of the Shiga toxin 1 gene (
stx
1
) in human-origin strains than in bovine-origin strains. In addition, lineage I strains expressed higher levels of the Shiga toxin 2 gene (
stx
2
). The present study supports a role for strain origin, lineage type, cytotoxin activity, and
stx
2
expression in modulating the amount of
E. coli
O157:H7 colonization of cattle.
Detection and Enumeration ofClostridium difficileSpores in Retail Beef and Pork
Authors: Weese J. Scott , Avery Brent P. , Rousseau J. , Reid-Smith Richard J.
Published: June 2009

Recent studies have identifiedClostridium difficilein food animals and retail meat, and concern has been raised about the potential for food to act as a source ofC. difficileinfection in humans. Previous studies of retail meat have relied on enrichment culture alone, thereby preventing any assessment of the level of contamination in meat. This study evaluated the prevalence ofC. difficilecontamination of retail ground beef and ground pork in Canada. Ground beef and ground pork were purchased from retail outlets in four Canadian provinces. Quantitative and enrichment culture was performed.Clostridium difficilewas isolated from 28/230 (12%) samples overall: 14/115 (12%) ground beef samples and 14/115 (12%) ground pork samples (P= 1.0). For ground beef, 10/14 samples (71%) were positive by enrichment culture only. Of the 4 ground beef samples that were positive by direct culture, 20 spores/g were present in 2 while 120 and 240 spores/g were present in 1 each. For ground pork, 10/14 (71%) samples were positive by enrichment culture only. Of the 4 ground pork samples that were positive by direct culture, 20 spores/g were present in 3 while 60 spores/g were present in 1. Ribotype 078 predominated, consistent with some previous studies ofC. difficilein food animals. Ribotype 027/North American pulsotype 1 was also identified in both retail beef and pork. This study has identified relatively common contamination of retail ground beef and pork withC. difficilespores; however, the levels of contamination were very low.
Prevalence and Genotypes of Human Noroviruses in Tropical Urban Surface Waters and Clinical Samples in Singapore
Authors: Aw Tiong Gim , Gin Karina Yew-Hoong , Ean Oon Lynette Lin , Chen Eileen Xueqin , Woo Chee Hoe
Published: June 2009

The prevalence and genotypes of norovirus genogroup I (GI) and GII in tropical urban catchment waters and an estuarine bay were studied. A comparative analysis was performed with environmental isolates of noroviruses and concurrently identified clinical isolates in Singapore during gastroenteritis outbreaks between August 2006 to January 2007. Noroviruses in environmental water samples were concentrated by using ultrafiltration techniques and then analyzed by reverse transcription-seminested PCR assay targeting the partial capsid region of noroviruses and DNA sequencing. Among the 60 water samples collected, noroviruses were detected in 43 (71.7%) of these samples. Of these 43 norovirus-positive samples, the coexistence of both GI and GII strains was identified in 23 (53.5%) water samples. The phylogenetic analysis revealed multiple genotypes of noroviruses GI and GII in environmental water samples. GI and GII strains were clustered into seven and nine (including two unclassified) genotypes, respectively. The major norovirus genotypes in environmental water samples were GI/2 and GI/4 and GII/4. Genotyping of the 21 norovirus-positive clinical samples showed that all of the strains belonged to the GII/4 cluster. The environmental and clinical norovirus GII/4 isolates showed high levels of nucleotide sequence identity to each other and to the novel GII/4 variant associated with global epidemics of gastroenteritis during 2006. This study suggests the emergence and circulation of multiple novel norovirus GI and GII genotypes in water environments. Further comprehensive surveillance of water environments for noroviruses and routine clinical reporting is warranted.
Characterization of the Central Metabolic Pathways in
Thermoanaerobacter
sp. Strain X514 via Isotopomer-Assisted Metabolite Analysis
Authors: Feng Xueyang , Mouttaki Housna , Lin Lu , Huang Rick , Wu Bing , Hemme Christopher L. , He Zhili , Zhang Baichen , Hicks Leslie M. , Xu Jian , Zhou Jizhong , Tang Yinjie J.
Published: June 2009

Thermoanaerobacter
sp. strain X514 has great potential in biotechnology due to its capacity to ferment a range of C
5
and C
6
sugars to ethanol and other metabolites under thermophilic conditions. This study investigated the central metabolism of strain X514 via
13
C-labeled tracer experiments using either glucose or pyruvate as both carbon and energy sources. X514 grew on minimal medium and thus contains complete biosynthesis pathways for all macromolecule building blocks. Based on genome annotation and isotopic analysis of amino acids, three observations can be obtained about the central metabolic pathways in X514. First, the oxidative pentose phosphate pathway in X514 is not functional, and the tricarboxylic acid cycle is incomplete under fermentative growth conditions. Second, X514 contains (
Re
)-type citrate synthase activity, although no gene homologous to the recently characterized (
Re
)-type citrate synthase of
Clostridium kluyveri
was found. Third, the isoleucine in X514 is derived from acetyl coenzyme A and pyruvate via the citramalate pathway rather than being synthesized from threonine via threonine ammonia-lyase. The functionality of the citramalate synthase gene (
cimA
[Teth514_1204]) has been confirmed by enzymatic activity assays, while the presence of intracellular citramalate has been detected by mass spectrometry. This study demonstrates the merits of combining
13
C-assisted metabolite analysis, enzyme assays, and metabolite detection not only to examine genome sequence annotations but also to discover novel enzyme activities.
Effect of Nitrate and Acetylene on
nirS
,
cnorB
, and
nosZ
Expression and Denitrification Activity in
Pseudomonas mandelii
Authors: Saleh-Lakha Saleema , Shannon Kelly E. , Henderson Sherri L. , Zebarth Bernie J. , Burton David L. , Goyer Claudia , Trevors Jack T.
Published: June 2009

Nitrate acts as an electron acceptor in the denitrification process. The effect of nitrate in the range of 0 to 1,000 mg/liter on
Pseudomonas mandelii nirS
,
cnorB
, and
nosZ
gene expression was studied, using quantitative reverse transcription-quantitative PCR. Denitrification activity was measured by using the acetylene blockage method and gas chromatography. The effect of acetylene on gene expression was assessed by comparing denitrification gene expression in
P. mandelii
culture grown in the presence or absence of acetylene. The higher the amount of NO
3
−
present, the greater the induction and the longer the denitrification genes remained expressed.
nirS
gene expression reached a maximum at 2, 4, 4, and 6 h in cultures grown in the presence of 0, 10, 100, and 1,000 mg of KNO
3
/liter, respectively, while induction of
nirS
gene ranged from 12- to 225-fold compared to time zero.
cnorB
gene expression also followed a similar trend.
nosZ
gene expression did not respond to NO
3
−
treatment under the conditions tested. Acetylene decreased
nosZ
gene expression but did not affect
nirS
or
cnorB
gene expression. These results showed that
nirS
and
cnorB
responded to nitrate concentrations; however, significant denitrification activity was only observed in culture with 1,000 mg of KNO
3
/liter, indicating that there was no relationship between gene expression and denitrification activity under the conditions tested.
Pig Manure Contamination Marker Selection Based on the Influence of Biological Treatment on the Dominant Fecal Microbial Groups
Authors: Marti Romain , Dabert Patrick , Pourcher Anne-Marie
Published: June 2009

The objective of this study was to identify a microbial marker for pig manure contamination. We quantified the persistence of four dominant bacterial groups from the pig intestinal tract throughout manure handling at 10 livestock operations (including aerobic digestion) by using molecular typing. The partial 16S rRNA genes of
Bacteroides
-
Prevotella
,
Eubacterium
-
Clostridiaceae
,
Bacillus
-
Streptococcus
-
Lactobacillus
(BSL), and
Bifidobacterium
group isolates were amplified and analyzed by capillary electrophoresis single-strand conformation polymorphism. The most dominant bacterial populations were identified by cloning and sequencing their 16S rRNA genes. The results showed that
Bifidobacterium
spp. and, to a lesser extent, members of the BSL group, were less affected by the aerobic treatment than either
Eubacterium
-
Clostridiaceae
or
Bacteroides
-
Prevotella
. Two
Bifidobacterium
species found in raw manure were still present in manure during land application, suggesting that they can survive outside the pig intestinal tract and also survive aerobic treatment. The 16S-23S rRNA internal transcribed spacer of one species,
Bifidobacterium thermacidophilum
subsp.
porcinum
, was sequenced, and a specific pair of primers was designed for its detection in the environment. With this nested PCR assay, this potential marker was not detected in samples from 30 bovine, 30 poultry, and 28 human fecal samples or in 15 urban wastewater effluents. As it was detected in runoff waters after spreading of pig manure, we propose this marker as a suitable microbial indicator of pig manure contamination.
Identification of anaoxSystem That Requires Cytochromecin the Highly Arsenic-Resistant BacteriumOchrobactrum triticiSCII24
Authors: Branco Rita , Francisco Romeu , Chung Ana Paula , Morais Paula Vasconcelos
Published: June 2009

Microbial biotransformations have a major impact on environments contaminated with toxic elements, including arsenic, resulting in an increasing interest in strategies responsible for how bacteria cope with arsenic. In the present work, we investigated the metabolism of this metalloid in the bacteriumOchrobactrum triticiSCII24. This heterotrophic organism contains two differentarsoperons and is able to oxidize arsenite to arsenate. The presence of arsenite oxidase genes in this organism was evaluated, and sequence analysis revealed structural genes for an As(III) oxidase (aoxAB), ac-type cytochrome (cytC), and molybdopterin biosynthesis (moeA). Two other genes coding for a two-component signal transduction pair (aoxRS) were also identified upstream from the previous gene cluster. The involvement ofaoxgenes in As(III) oxidation was confirmed by functionally expressing them intoO. tritici5bvl1, a non-As(III) oxidizer. Experiments showed that the As(III) oxidation process inO. triticirequires not only the enzyme arsenite oxidase but also the cytochromecencoded in the operon. The fundamental role of this cytochromec, reduced in the presence of arsenite in strain SCII24 but not in anO. triticiΔaoxBmutant, is surprising, since to date this feature has not been found in other organisms. In this strain the presence of anaoxsystem does not seem to confer an additional arsenite resistance capability; however, it might act as part of an As(III)-detoxifying strategy. Such mechanisms may have played a crucial role in the development of early stages of life on Earth and may one day be exploited as part of a potential bioremediation strategy in toxic environments.
Glycerol-3-Phosphate Acyltransferases Gat1p and Gat2p Are Microsomal Phosphoproteins with Differential Contributions to Polarized Cell Growth
Authors: Bratschi Martin W. , Burrowes David P. , Kulaga Adam , Cheung Jing F. , Alvarez Ana L. , Kearley Jennifer , Zaremberg Vanina
Published: June 2009

Glycerol-3-phosphate acyltransferase (GPAT) catalyzes the initial step in the synthesis of all glycerolipids. It is the committed and rate-limiting step and is redundant in
Saccharomyces cerevisiae
, mammals, and plants. GPAT controls the formation of lipid intermediates that serve not only as precursors of more-complex lipids but also as intracellular signaling molecules.
Saccharomyces cerevisiae
possesses two GPATs, encoded by the
GAT1
and
GAT2
genes. Metabolic analysis of yeast lacking either
GAT1
or
GAT2
indicated partitioning of the two main branches of phospholipid synthesis at the initial and rate-limiting GPAT step. We are particularly interested in identifying molecular determinants mediating lipid metabolic pathway partitioning; therefore, as a starting point, we have performed a detailed study of Gat1p and Gat2p cellular localization. We have compared Gat1p and Gat2p localization by fluorescence microscopy and subcellular fractionation using equilibrium density gradients. Our results indicate Gat1p and Gat2p overlap mostly in their localization and are in fact microsomal GPATs, localized to both perinuclear and cortical endoplasmic reticula in actively proliferating cells. A more detailed analysis suggests a differential enrichment of Gat1p and Gat2p in distinct ER fractions. Furthermore, overexpression of these enzymes in the absence of endogenous GPATs induces proliferation of distinct ER arrays, differentially affecting cortical ER morphology and polarized cell growth. In addition, our studies also uncovered a dynamic posttranslational regulation of Gat1p and Gat2p and a compensation mechanism through phosphorylation that responds to a cellular GPAT imbalance.
Cooperative Regulation of
ADE3
Transcription by Gcn4p and Bas1p in
Saccharomyces cerevisiae
Authors: Joo Yoo Jin , Kim Jung-Ae , Baek Joung Hee , Seong Ki Moon , Han Kyung-Duk , Song Jae Mahn , Choi Jin Young , Kim Joon
Published: June 2009

The one-carbon response regulon is essential for the biosynthesis of nucleic acids as well as several amino acids. The
ADE3
gene is known to encode a crucial one-carbon regulon enzyme, tetrahydrofolate synthase, which is involved in the biosynthesis of purine and the amino acids methionine and glycine. Therefore, the mechanism through which
ADE3
transcription is regulated appears to be critical for the cross-talk among these metabolic pathways. Even so, the direct involvement of
ADE3
transcription through gene-specific transcription factors has not been shown clearly. In this study, the promoter structure of the
ADE3
gene was investigated in detail, and a genuine Gcn4p responsive element (GCRE) was confirmed among three putative GCRE elements in vivo and in vitro. Through gene deletion studies of Gcn4p and Bas1p, it was established that both factors are involved in the transcriptional regulation of the
ADE3
gene. Direct binding to this GCRE and the occupancy of the
ADE3
promoter by these factors were also confirmed. Taking these results together, we concluded that Gcn4p is responsible for the basal and inducible expression of the
ADE3
gene, while Bas1p is required for its basal expression.
Roles of the Glyoxylate and Methylcitrate Cycles in Sexual Development and Virulence in the Cereal PathogenGibberella zeae
Authors: Lee Seung-Ho , Han You-Kyoung , Yun Sung-Hwan , Lee Yin-Won
Published: June 2009

The glyoxylate and methylcitrate cycles are involved in the metabolism of two- or three-carbon compounds in fungi. To elucidate the role(s) of these pathways inGibberella zeae, which causes head blight in cereal crops, we focused on the functions ofG. zeaeorthologs (GzICL1and GzMCL1) of the genes that encode isocitrate lyase (ICL) and methylisocitrate lyase (MCL), respectively, key enzymes in each cycle. The deletion of GzICL1(ΔGzICL1) caused defects in growth on acetate and in perithecium (sexual fruiting body) formation but not in virulence on barley and wheat, indicating that GzICL1 acts as the ICL of the glyoxylate cycle and is essential for self-fertility inG. zeae. In contrast, the ΔGzMCL1strains failed to grow on propionate but exhibited no major changes in other traits, suggesting that GzMCL1 is required for the methylcitrate cycle inG. zeae. Interestingly, double deletion of both GzICL1and GzMCL1caused significantly reduced virulence on host plants, indicating that both GzICL1and GzMCL1have redundant functions for plant infection inG. zeae. Thus, both GzICL1and GzMCL1may play important roles in determining major mycological and pathological traits ofG. zeaeby participating in different metabolic pathways for the use of fatty acids.
Investigation of Carbon Metabolism in “
Dehalococcoides ethenogenes
” Strain 195 by Use of Isotopomer and Transcriptomic Analyses
Authors: Tang Yinjie J. , Yi Shan , Zhuang Wei-Qin , Zinder Stephen H. , Keasling Jay D. , Alvarez-Cohen Lisa
Published: June 2009

Members of the genus “
Dehalococcoides
” are the only known microorganisms that can completely dechlorinate tetrachloroethene and trichloroethene to the innocuous end product, ethene. This study examines the central metabolism in “
Dehalococcoides ethenogenes
” strain 195 via
13
C-labeled tracer experiments. Supported by the genome annotation and the transcript profile, isotopomer analysis of key metabolites clarifies ambiguities in the genome annotation and identifies an unusual biosynthetic pathway in strain 195. First, the
13
C-labeling studies revealed that strain 195 contains complete amino acid biosynthesis pathways, even though current genome annotation suggests that several of these pathways are incomplete. Second, the tricarboxylic acid cycle of strain 195 is confirmed to be branched, and the Wood-Ljungdahl carbon fixation pathway is shown to not be functionally active under our experimental conditions; rather, CO
2
is assimilated via two reactions, conversion of acetyl-coenzyme A (acetyl coenzyme A [acetyl-CoA]) to pyruvate catalyzed by pyruvate synthase (DET0724-0727) and pyruvate conversion to oxaloacetate via pyruvate carboxylase (DET0119-0120). Third, the
13
C-labeling studies also suggested that isoleucine is synthesized from acetyl-CoA and pyruvate via citramalate synthase (CimA, EC 2.3.1.182), rather than from the common pathway via threonine ammonia-lyase (EC 4.3.1.19). Finally, evidence is presented that strain 195 may contain an undocumented citrate synthase (>95%
Re
-type stereospecific), i.e., a novel
Re
-citrate synthase that is apparently different from the one recently reported in
Clostridium kluyveri
.
Identification and Characterization ofgshA, a Gene Encoding the Glutamate-Cysteine Ligase in the Halophilic ArchaeonHaloferax volcanii
Authors: Malki Liron , Yanku Michaela , Borovok Ilya , Cohen Gerald , Mevarech Moshe , Aharonowitz Yair
Published: June 2009

Halophilic archaea were found to contain in their cytoplasm millimolar concentrations of γ-glutamylcysteine (γGC) instead of glutathione. Previous analysis of the genome sequence of the archaeonHalobacteriumsp. strain NRC-1 has indicated the presence of a sequence homologous to sequences known to encode the glutamate-cysteine ligase GshA. We report here the identification of thegshAgene in the extremely halophilic archaeonHaloferax volcaniiand show thatH. volcanii gshAdirects in vivo the synthesis and accumulation of γGC. We also show that theH. volcaniigene when expressed in anEscherichia colistrain lacking functional GshA is able to restore synthesis of glutathione.
Structural Basis for the Differential Binding Affinities of the HsfBD1 and HsfBD2 Domains in the
Haemophilus influenzae
Hsf Adhesin
Authors: Radin Jana N. , Grass Susan A. , Meng Guoyu , Cotter Shane E. , Waksman Gabriel , St. Geme Joseph W.
Published: June 2009

Haemophilus influenzae
is a human-specific gram-negative coccobacillus that causes a variety of human infections ranging from localized respiratory infections to invasive diseases. Hsf is the major nonpilus adhesin in encapsulated strains of
H. influenzae
and belongs to the trimeric autotransporter family of proteins. The Hsf protein contains two highly homologous binding domains, designated HsfBD1 and HsfBD2. In this study we characterized the differential binding properties of HsfBD1 and HsfBD2. In assays using HeLa cells, we found that bacteria expressing either full-length Hsf or HsfBD1 by itself adhered at high levels, while bacteria expressing HsfBD2 by itself adhered at low levels. Immunofluorescence microscopy and a cellular enzyme-linked immunosorbent assay using purified proteins revealed that the binding affinity was significantly higher for HsfBD1 than for HsfBD2. Purified HsfBD1 was able to completely block adherence by bacteria expressing either HsfBD1 or HsfBD2, while purified HsfBD2 was able to block adherence by bacteria expressing HsfBD2 but had minimal activity against bacteria expressing HsfBD1. Conversion of the residue at position 1935 in the HsfBD1 binding pocket from Asp to Glu resulted in HsfBD2-like binding properties, and conversion of the residue at position 569 in the HsfBD2 binding pocket from Glu to Asp resulted in HsfBD1-like binding properties, as assessed by adherence assays with recombinant bacteria and by immunofluorescence microscopy with purified proteins. This work demonstrates the critical role of a single amino acid in the core of the binding pocket in determining the relative affinities of the HsfBD1 and HsfBD2 binding domains.