bluuebunny/crossref_metadata_2025 huggingface dataset:
Evolutionary diversification of satellite DNA sequences fromLeymus(Poaceae: Triticeae)
Authors: Anamthawat-Jónsson K. , Wenke T. , Thórsson Æ. Th. , Sveinsson S. , Zakrzewski F. , Schmidt T.
Published: April 2009

The genus Leymus (lymegrass) comprises about 30 polyploid, perennial, temperate grass species in the tribe Triticeae (family Poaceae). Previous studies indicated a large diversity in the Leymus genome, and therefore, the aim of this study was to isolate new repetitive DNA sequences that can be used for differentiating Leymus species and elucidating their genomic relationships. A C0t-1 DNA plasmid library was generated from genomic DNA of American tetraploid species Leymus triticoides . A family of highly repetitive satellite DNA sequences, designated Lt1, was obtained from this library. The Lt1 family consisted of 380 bp SacI repeating units arranging in tandem arrays. A 120 bp MspI subfamily was discovered within this family, indicating that cytosine methylation may have played an important role in the evolution of satellite sequences. The Lt1 satellite was localized in the subtelomeric heterochromatic blocks of L. triticoides chromosomes, which are present on all chromosomes and often on both arms. The Lt1 sequences are abundant in L. triticoides but absent in its closely related species Leymus racemosus . Significant homology was found between the Lt1 family and numerous repetitive sequences from Poaceae species, indicating that the Lt1 is an ancient family of tandemly repeated sequences in grasses.
Physical localization and genetic mapping of the fertility restoration geneRfoin canola (Brassica napusL.)
Authors: Feng Jiuhuan , Primomo Valerio , Li Zenglu , Zhang Yongping , Jan Chao-Chien , Tulsieram Lomas , Xu Steven S.
Published: April 2009

The Ogu cytoplasm for male sterility and its fertility restorer gene Rfo in canola ( Brassica napus L.) were originally introgressed from radish ( Raphanus sativus L.) and have been widely used for canola hybrid production and breeding. The objective of this study was to determine the physical location of the Rfo locus in the canola genome using fluorescence in situ hybridization and genetic mapping. For physical localization of the Rfo gene, two bacterial artificial chromosome (BAC) clones, G62 and B420, which were closely linked to the Rfo gene, were used as probes to hybridize with the somatic metaphase chromosomes of a canola hybrid variety, PHI-46 (46H02), containing the Rfo fragment. The results showed that both clones were physically located at the end of one large metacentric chromosome. By simultaneous use of two BAC clones and 45S rDNA repeated sequences as the probes, we demonstrated that the large metacentric chromosome probed with the two BAC clones did not carry 45S rDNA repeated sequences. The chromosome was 3.65 ± 0.74 µm in average length (20 cells) and ranked second in size among the chromosomes without 45S rDNAs. The centromere index of the chromosome (20 cells) was calculated as 43.74 ± 4.19. A comparison with previously reported putative karyotypes of B. napus (AACC) and its diploid ancestors Brassica rapa L. (AA) and Brassica oleracea L. (CC) suggests that the chromosome carrying the Rfo fragment might belong to one of three large metacentric chromosomes of the C genome. Genetic mapping has confirmed the localization of the Rfo fragment to the distal region of linkage group N19, which corresponds to the C genome in B. napus. This study has provided the evidence of the location of the Rfo gene on canola chromosomes and established a basic framework for further physical mapping and manipulation of the gene.
A randomized controlled trial of the effects of flaxseed lignan complex on metabolic syndrome composite score and bone mineral in older adults
Authors: Cornish Stephen M. , Chilibeck Philip D. , Paus-Jennsen Lisa , Biem H. Jay , Khozani Talaei , Senanayake Vijitha , Vatanparast Hassanali , Little Jonathan P. , Whiting Susan J. , Pahwa Punam
Published: April 2009

A randomized double-blind placebo controlled study design was used to assess the effects of flaxseed lignan complex supplementation during exercise training on a metabolic syndrome composite score and osteoporosis risk in older adults. A total of 100 subjects (≥50 years) were randomized to receive flaxseed lignan (543 mg·day–1 in a 4050 mg complex) or placebo while completing a 6 month walking program (30–60 min·day–1, 5–6 days·week–1). Fasting serum glucose, triacylglycerol (TAG), high-density lipoprotein (HDL) cholesterol, low-density lipoprotein cholesterol, total cholesterol, interleukin-6, and tumor necrosis factor-α were measured every 2 months, while body composition, bone mineral density, and resting blood pressure were assessed at baseline and at 6 months. A composite Z score of 6 risk factors for metabolic syndrome (fasting glucose, HDL cholesterol, TAG, abdominal adiposity, blood pressure, and inflammatory cytokines) was calculated at baseline and at 6 months. Men taking placebo increased metabolic syndrome composite Z score (p < 0.05), but there were no changes in the other groups. A significant group × sex × time interaction was noted for TAG (p = 0.017) and diastolic blood pressure (p = 0.046), with men taking flaxseed lignan decreasing diastolic blood pressure relative to men taking placebo, and men taking placebo increasing TAG relative to men taking flax lignan. There were no differences between groups for change in bone measures, body composition, lipoproteins, or cytokines. Males taking the flaxseed lignan complex reduced metabolic syndrome score relative to men taking placebo, but a similar trend was not seen in females. Flaxseed lignan had no effect on bone mineral density or content, body composition, lipoproteins, glucose, or inflammation.
Canada’s physical activity guide recommendations are a low benchmark for Manitoba adults
Authors: Ready A. Elizabeth , Butcher Janice E. , Dear James B. , Fieldhouse Paul , Harlos Sande , Katz Alan , Moffatt Michael , Rodrigue Maureen , Schmalenberg Jan , Gardiner Phillip F.
Published: April 2009

Canada’s Physical Activity Guide to Healthy Active Living (CPAG) is the national reference for messaging on physical activity for health benefits, yet few studies have examined population activity levels in relation to its recommendations. As part of the province-wide in motion initiative, we obtained a baseline measurement of the physical activity levels of adult Manitobans. Physical activity levels were benchmarked against CPAG recommendations and were compared with criteria used in previous surveys. A stratified random sample of adults from the 9 Regional Health Authorities outside of Winnipeg, and from the 12 Community Areas within the Winnipeg Health Region, was surveyed by telephone. Respondents (n = 6536) reported all light, moderate, and vigorous physical activity of 10 min or more in the previous week. Intensity levels were corrected to reflect standard MET equivalents, using the Ainsworth Compendium. A total of 69.5% of respondents met the minimum CPAG requirements; however, only 29.1% of those did so with vigorous activity. Relative to energy expenditure, 18.3% were classified as inactive (<1.50 kcal·kg–1·day–1 (KKD)), 16.4% as moderately active (1.50 to 2.99 KKD), and 65.3% as active (≥3.00 KKD). When assessed against the CPAG recommendations, which promote integration of physical activity into one’s daily routine, a higher proportion of Manitobans met recommended physical activity levels than that reported in previous surveys, which focused on leisure activity. Given the corresponding increase in levels of obesity and chronic disease, and equivocal nutrient intake data, we recommend that the CPAG recommendations be reviewed, especially with respect to the inclusion of routine baseline activities of daily living.
Evaluation of true maximal oxygen uptake based on a novel set of standardized criteria
Authors: Midgley Adrian W. , Carroll Sean , Marchant David , McNaughton Lars R. , Siegler Jason
Published: April 2009

In this study, criteria are used to identify whether a subject has elicited maximal oxygen uptake. We evaluated the validity of traditional maximal oxygen uptake criteria and propose a novel set of criteria. Twenty athletes completed a maximal oxygen uptake test, consisting of an incremental phase and a subsequent supramaximal phase to exhaustion (verification phase). Traditional and novel maximal oxygen uptake criteria were evaluated. Novel criteria were: oxygen uptake plateau defined as the difference between modelled and actual maximal oxygen uptake >50% of the regression slope of the individual oxygen uptake–workrate relationship; as in the first criterion, but for maximal verification oxygen uptake; and a difference of ≤4 beats·min–1 between maximal heart rate values in the 2 phases. Satisfying the traditional oxygen uptake plateau criterion was largely an artefact of the between-subject variation in the oxygen uptake–workrate relationship. Secondary criteria, supposedly an indicator of maximal effort, were often satisfied long before volitional exhaustion, even at intensities as low as 61% maximal oxygen uptake. No significant mean differences were observed between the incremental and verification phases for oxygen uptake (t = 0.4; p = 0.7) or heart rate (t = 0.8; p = 0.5). The novel oxygen uptake plateau criterion, maximal oxygen uptake verification criterion, and maximal heart rate verification criterion were satisfied by 17, 18, and 18 subjects, respectively. The small individual absolute differences in oxygen uptake between incremental and verification phases observed in most subjects provided additional confidence that maximal oxygen uptake was elicited. Current maximal oxygen uptake criteria were not valid and novel criteria should be further explored.
Oxidative stress in the brain of nicotine-induced toxicity: protective role ofAndrographis paniculataNees and vitamin E
Authors: Das Subhasis , Gautam N. , Dey Sankar Kumar , Maiti Tarasankar , Roy Somenath
Published: April 2009

Mitochondria are the crossroads of several crucial cellular activities; they produce considerable quantities of superoxide radical and hydrogen peroxide, which can damage important macromolecules. Nicotine affects a variety of cellular processes, from induction of gene expression to modulation of enzymatic activities. The aim of this study was to elucidate the protective effects of andrographolide (ANDRO) aqueous extract (AE-Ap) of Andrographis paniculata, and vitamin E on nicotine-induced brain mitochondria. In this investigation, nicotine (1 mg·kg body mass–1·day–1) was treated, for the period of 7 days, simultaneously with 2 A. paniculata products, ANDRO and AE-Ap (250 mg·kg body mass–1·day–1); and vitamin E (50 mg·kg body mass–1·day–1) was supplemented in different group of male Wistar rats. The activities of mitochondrial electron transport chain (Mito–ETC) complexes (I, II, III), nitric oxide production, superoxide anion, catalase, glutathione reductase, glutathione peroxidase, glutathione-S-transferase, and concentrations of reduced glutathione and oxidized glutathione were measured in discrete regions of brain (the cerebral hemisphere, cerebellum, diencephalons, and brain stem). The study revealed that nicotine inhibits the Mito–ETC complexes and produces nitric oxide, which suppressed the mitochondrial oxidative stress scavenger system in different brain regions. In these circumstances, lipid peroxidation and protein oxidation were noted in different discrete regions of brain mitochondria. ANDRO, AE-Ap, and vitamin E showed the protective potentiality against nicotine toxicity. The analysis of such alterations is important in determining the basis of normal dysfunction in the brain associated with nicotine toxicity, which could be ameliorated by A. paniculata and vitamin E, and may help to develop therapeutic means against nicotine-induced disorders.
Nutritional influences on maternal autonomic function during pregnancy
Authors: Caulfield Laura E. , Zavaleta Nelly , Chen Ping , Merialdi Mario , DiPietro Janet A.
Published: April 2009

The objective of this study was to identify influences on longitudinal changes in autonomic function during pregnancy. We studied 195 low-income Peruvian women who were subjects in a randomized controlled trial of zinc supplementation and fetal neurobehavioral development. Maternal cardiorespiratory status and electrodermal activity were assessed at 20, 24, 28, 32, 36, and 38 weeks’ gestation over a 50-min session. At baseline (10–16 weeks), and at 28 and 36 weeks’ gestation, mass and height, arm and calf circumferences, and skinfold thicknesses were assessed, and blood was taken to measure hemoglobin, hematocrit, and plasma zinc concentration. Measures of heart period (HP), respiratory period (RP), respiratory sinus arrhythmia (RSA), skin conductance level (SCL), and mean arterial pressure (MAP) were analyzed using longitudinal models. Having a hemoglobin concentration <10.5 g·dL–1 at 28 weeks was associated with shorter HP and RP and lower RSA, and change in hemoglobin from baseline to 36 weeks was associated with decreased SCL. Further, 3 measures were associated with variation in fat or fat-free mass in the calf; RSA was negatively associated with changes in calf muscle area over pregnancy, while changes in calf fat area were positively associated with MAP and SCL. Variability in autonomic function during the second half of pregnancy is associated with hematologic status and changes in lower limb body composition across pregnancy.
Non-high-density-lipoprotein cholesterol and cardiovascular risk factors among adolescents with and without impaired fasting glucose
Authors: Liu Jian , Joshi Divya , Sempos Christopher T.
Published: April 2009

To evaluate how non-high-density-lipoprotein (non-HDL) is associated with impaired fasting glucose (IFG) and clustered metabolic risk (MR) factors among adolescents, we pooled 2764 adolescents, aged 12–19 years, from the National Health and Nutrition Examination Survey from 3 time periods (1999–2000, 2001–2002, and 2003–2004) who were free of diabetes and had fasted overnight for this analysis. IFG was defined as 100 ≤ glucose ≤ 125 mg·dL–1. Age- and sex-specific cut-offs were used for 4 MR factors: higher levels of triglycerides, waist circumference, blood pressure, and lower levels of HDL. Clustered MR was defined as having any 2 of the 4 factors. Overall, approximately 11% of adolescents had IFG. The mean level of non-HDL cholesterol was much higher in those with IFG than in those without IFG, with adjustment for certain confounding variables (121.4 vs. 110.1 mg·dL–1; p < 0.05). This difference could still be observed in adolescents with one or more clustered MR factors. However, there were no statistical differences in low-density-lipoprotein (LDL) level. Compared with those who were without IFG and not at high levels of non-HDL — after adjustment for age, sex, race, current smoking, and body mass index — the odds of having clustered MR factors were 1.08 (95% CI, 0.65–1.82) for those with IFG and low non-HDL cholesterol, 3.55 (2.29–5.48) for those without IFG but with high non-HDL cholesterol, and 10.10 (3.67–27.80) for those with both IFG and high non-HDL cholesterol. Moreover, those with IFG and at increased risk of obesity were more likely to have higher levels of non-HDL cholesterol (odds ratio (95% CI): 4.41 (2.28–8.50)), compared with those without IFG and not at increased risk of obesity. In summary, prediabetic adolescents with IFG and high levels of non-HDL cholesterol are more likely to have clustered MR factors. Thus, the levels of non-HDL cholesterol may be an important indicator in monitoring cardiovascular disease risk among adolescents with IFG.
Age and physical activity status effects on appetite and mood state in older humans
Authors: Apolzan John W. , Flynn Michael G. , McFarlin Brian K. , Campbell Wayne W.
Published: April 2009

This study examined the influences of age and chronic physical activity status on appetite and mood state. Groups of younger inactive, younger active, older inactive, and older active men and women completed questionnaires each waking hour, rating appetite and mood state for 1 day. Maximal oxygen consumption was 20% lower in older than in younger (p < 0.001) subjects, and 32% lower in inactive than in active (p < 0.001) subjects. Mean hunger (older, 4 ± 1; younger, 5 ± 1 arbitrary units (AU); p < 0.01) and desire to eat (older, 3 ± 1; younger, 4 ± 1 AU; p < 0.01) were lower in older than in younger subjects. Nadir arousal was higher for the active subjects (active, 3 ± 1; inactive, 2 ± 1 AU; p < 0.05). Nadir arousal, nadir pleasantness, and mean pleasantness were higher for the older subjects (p < 0.05). Physical activity status does not influence appetite or the age-associated declines in hunger or desire to eat. The increased nadir arousal of the physically active and older groups is consistent with these subjects experiencing less extreme sleepiness.
Physiological responses and energy cost during a simulation of a Muay Thai boxing match
Authors: Crisafulli Antonio , Vitelli Stefano , Cappai Ivo , Milia Raffaele , Tocco Filippo , Melis Franco , Concu Alberto
Published: April 2009

Muay Thai is a martial art that requires complex skills and tactical excellence for success. However, the energy demand during a Muay Thai competition has never been studied. This study was devised to obtain an understanding of the physiological capacities underlying Muay Thai performance. To that end, the aerobic energy expenditure and the recruitment of anaerobic metabolism were assessed in 10 male athletes during a simulation match of Muay Thai. Subjects were studied while wearing a portable gas analyzer, which was able to provide data on oxygen uptake, carbon dioxide production, and heart rate (HR). The excess of CO2 production (CO2 excess) was also measured to obtain an index of anaerobic glycolysis. During the match, group energy expenditure was, on average (mean ± standard error of the mean), 10.75 ± 1.58 kcal·min–1, corresponding to 9.39 ± 1.38 metabolic equivalents. Oxygen uptake and HRs were always above the level of the anaerobic threshold assessed in a preliminary incremental test. CO2 excess showed an abrupt increase in the first round, and reached a value of 636 ± 66.5 mL·min–1. This parameter then gradually decreased throughout the simulation match. These data suggest that Muay Thai is a physically demanding activity with great involvement of both the aerobic metabolism and anaerobic glycolysis. In particular, it appears that, after an initial burst of anaerobic glycolysis, there was a progressive increase in the aerobic energy supply. Thus, training protocols should include exercises that train both aerobic and anaerobic energetic pathways.
Physical activity levels of older community-dwelling adults are influenced by summer weather variables
Authors: Brandon Caitlin A. , Gill Dawn P. , Speechley Mark , Gilliland Jason , Jones Gareth R.
Published: April 2009

Adequate daily physical activity (PA) is important for maintaining functional capacity and independence in older adults. However, most older adults in Canada do not engage in enough PA to sustain fitness and functional independence. Environmental influences, such as warmer daytime temperatures, may influence PA participation; however, few studies have examined the effect of summertime temperatures on PA levels in older adults. This investigation measured the influence of summertime weather variables on PA in 48 community-dwelling older adults who were randomly recruited from a local seniors’ community centre. Each participant wore an accelerometer for a single 7-consecutive-day period (between 30 May and 9 August 2006) during waking hours, and completed a PA logbook to remark on major daily PA events. Local weather variables were collected from a national weather service and compared with PA counts per minute. Regression analysis revealed a curvilinear relationship between log-transformed PA and mean daily temperature (r2 = 0.025; p < 0.05). Linear mixed effects models that accounted for repeated measures nested within individuals were performed for monthly periods, meteorological variables, sex, age, and estimated maximal oxygen consumption, with PA as the dependent variable. Age and Air Quality Index remained significant variables within the model. Higher fitness levels had no effect on allowing individuals to perform more vigorous PA in warmer temperatures.
Many adult Canadians are not meeting current calcium recommendations from food and supplement intake
Authors: Vatanparast Hassanali , Dolega-Cieszkowski Jadwiga H. , Whiting Susan J.
Published: April 2009

The objective of this study was to determine trends in calcium intake from foods of Canadian adults from 1970–1972 to 2004. We compiled the calcium intake of adults (aged ≥19 years) from foods from Nutrition Canada (1970–1972; n = 7036); 9 provincial nutrition surveys (1990–1999; n = 16 915); and the 2004 Canadian Community Health Survey 2.2 (n = 20 197). Where possible, we used published confidence intervals to test for significant differences in calcium intake. In 2004, the mean calcium intake of Canadians was below Dietary Reference Intake recommendations for most adults, with the greatest difference in older adults (≥51 years), in part because the recommended calcium intake for this group is higher (1200 mg) than that for younger adults (1000 mg). The calcium intake of males in every age category was greater than that of females. Calcium intake increased from 1970 to 2004, yet, despite the introduction of calcium-fortified beverages to the market in the late 1990s, increases in calcium intake between 1970 and 2004 were modest. Calcium intakes in provinces were mostly similar in the 1990s and in 2004, except for women in Newfoundland and Labrador, who consumed less, especially in the 1990s, and for young men in 2004 in Prince Edward Island, who consumed more. When supplemental calcium intake was added, mean intakes remained below recommended levels, except for males 19–30 years, but the prevalence of adequacy increased in all age groups, notably for women over 50 years. The calcium intake of Canadian adults remains in need of improvement, despite fortification and supplement use.
Stimulation of muscle anabolism by resistance exercise and ingestion of leucine plus protein
Authors: Tipton Kevin D. , Elliott Tabatha A. , Ferrando Arny A. , Aarsland Asle A. , Wolfe Robert R.
Published: April 2009

Leucine is known to stimulate muscle protein synthesis and anabolism. However, evidence for the efficacy of additional leucine to enhance the response of muscle anabolism to resistance exercise and protein ingestion is unclear. Thus, we investigated the response of net muscle protein balance to ingestion of additional leucine with protein in association with resistance exercise. Two groups of untrained subjects performed an intense bout of leg resistance exercise following ingestion of 1 of 2 drinks: flavored water (PL); or 16.6 g of whey protein + 3.4 g of leucine (W+L). Arteriovenous amino acid balance across the leg was measured to assess the anabolic response of muscle in each group. Arterial amino acid concentrations increased in response to ingestion of W+L. Amino acid concentrations peaked between 60 and 120 min after ingestion, and then declined to baseline values. Valine concentration decreased to levels significantly lower than baseline. Net balance of leucine, threonine, and phenylalanine did not change following PL ingestion, but increased and remained elevated above baseline for 90–120 min following W+L ingestion. Leucine (138 ± 37 and –23 ± 23 mg), phenylalanine (58 ± 28 and –38 ± 14 mg), and threonine (138 ± 37 and –23 ± 23 mg) uptake was greater for W+L than for PL over the 5.5 h following drink ingestion. Our results indicate that the whey protein plus leucine in healthy young volunteers results in an anabolic response in muscle that is not greater than the previously reported response to whey protein alone.
Pro-apoptotic effects of parathyroid hormone in intestinal cells
Authors: Calvo Natalia , German Olga , Russo de Boland Ana , Gentili Claudia
Published: April 2009

Apoptosis, a form of programmed cell death, is a process fundamental to normal growth and development, immune response, tissue remodeling after injury or insult, and homeostasis of the intestinal epithelium. Recently, it has become apparent that apoptosis is a crucial process in skeletal development and homeostasis, and that signaling by the parathyroid hormone (PTH) receptor can either promote or suppress apoptosis depending on the cellular context. In this study, we evaluated the role of PTH in intestinal apoptosis using human colonic Caco-2 cells. To that end, Caco-2 cells were exposed to PTH (10−8 mol/L) for 1–5 days. Evaluation of cell survival by use of resazurin staining, 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTS) staining, and crystal violet staining revealed that PTH treatment diminishes the number of viable cells. Assessment of cells after PTH treatment by use of propidium iodide showed that the hormone increased the number of red-stained (dead) cells (178%, 5 days). Moreover, we found that the hormone induced disruption of actin filaments with changes to cellular shape, alteration of cell-to-cell junctions, externalization of membrane phosphatidylserine, chromatin condensation, and DNA fragmentation of the nucleus, which are morphological features consistent with apoptosis. In addition, PTH treatment revealed a cytosolic staining pattern of 14-3-3. However, the significance of such differential localization for 14-3-3 function remains unknown. Taken together, the present study suggests that PTH promotes apoptosis in Caco-2 intestinal cells.
Interconversion of a pair of active-site residues inEscherichia colicystathionine γ-synthase,E. colicystathionine β-lyase, andSaccharomyces cerevisiaecystathionine γ-lyase and development of tools for the investigation of their mechanisms and reaction specificity
Authors: Farsi Ali , Lodha Pratik H. , Skanes Jennifer E. , Los Heidi , Kalidindi Navya , Aitken Susan M.
Published: April 2009

Cystathionine γ-synthase (CGS) and cystathionine β-lyase (CBL), which comprise the transsulfuration pathway of bacteria and plants, and cystathionine γ-lyase (CGL), the second enzyme of the fungal and animal reverse transsulfuration pathway, share ∼30% sequence identity and are almost indistinguishable in overall structure. One difference between the active site of Escherichia coli CBL and those of E. coli CGS and Saccharomyces cerevisiae CGL is the replacement of a pair of aromatic residues, F55 and Y338, of the former by acidic residues in CGS (D45 and E325) and CGL (E48 and E333). A series of interconverting, site-directed mutants of these 2 residues was constructed in CBL (F55D, Y338E, F55D/Y338E), CGS (D45F, E325Y and D45F/E325Y) and CGL (E48A,D and E333A,D,Y) to probe the role of these residues as determinants of reaction specificity. Mutation of either position results in a reduction in catalytic efficiency, as exemplified by the 160-fold reduction in the kcat/Kml-Cysof eCGS-D45F and the 2850- and 30-fold reductions in the kcat/Kml-Cthof the eCBL-Y338E and the yCGL-E333A,Y mutants, respectively. However, the in vivo reaction specificity of the mutants was not altered, compared with the corresponding wild-type enzymes. The ΔmetB and ΔmetC strains, the optimized CBL and CGL assay conditions, and the efficient expression and affinity purification systems described provide the necessary tools to enable the continued exploration of the determinants of reaction specificity in the enzymes of the transsulfuration pathways.
A glycosylated antitumor ether lipid kills cells via paraptosis-like cell death
Authors: Samadder Pranati , Bittman Robert , Byun Hoe-Sup , Arthur Gilbert
Published: April 2009

Glycosylated antitumor ether lipids (GAELs) have superior anticancer properties relative to the alkyllysophospholipid class, but there have been no studies of the mechanisms of these compounds. The prototype GAEL, 1-O-hexadecyl-2-O-methyl-3-O-(2′-amino-2′-deoxy-β-d-glucopyranosyl)-sn-glycerol (Gln), effectively killed mouse embryonic fibroblasts (MEFs) lacking key molecules involved in caspase-dependent apoptosis, and cell death was not prevented by caspase inhibitors. Gln did not cause a loss of mitochondrial membrane potential, even in rounded-up dying cells. Gln stimulated the appearance and accumulation of LC3-II, a protein marker for autophagy, in a variety of cells, including wild-type MEFs, but not in MEFs lacking ATG5, a key protein required for autophagy. Gln induced LC3 puncta formation in Chinese hamster ovary cells stably expressing a LC3–green fluorescent protein fusion protein. Thus, Gln appears to induce autophagy. Autophagy was mTOR-independent and was not inhibited by 3-methyladenine or wortmannin. Although Gln is toxic, cellular ability to undergo autophagy was not essential for its toxicity. Furthermore, the GAEL analog 2-deoxy-C-Glc induced LC3 puncta formation but did not kill the cells. Gln, but not 2-deoxy-C-Glc, caused the accumulation of cytoplasmic acidic vacuoles in the cells. Our data suggest that GAELs may activate autophagy; however, GAELs do not kill cells by apoptosis or autophagy but rather by a paraptosis-like cell death mechanism.
Characterization of a group 1 late embryogenesis abundant protein in encysted embryos of the brine shrimpArtemia franciscana
Authors: Sharon Michelle A. , Kozarova Anna , Clegg James S. , Vacratsis Panayiotis O. , Warner Alden H.
Published: April 2009

Late embryogenesis abundant (LEA) proteins are hydrophilic molecules that are believed to function in desiccation and low-temperature tolerance in some plants and plant propagules, certain prokaryotes, and several animal species. The brine shrimp Artemia franciscana can produce encysted embryos (cysts) that enter diapause and are resistant to severe desiccation. This ability is based on biochemical adaptations, one of which appears to be the accumulation of the LEA protein that is the focus of this study. The studies described herein characterize a 21 kDa protein in encysted Artemia embryos as a group 1 LEA protein. The amino acid sequence of this protein and its gene have been determined and entered into the NCBI database (no. EF656614). The LEA protein consists of 182 amino acids and it is extremely hydrophilic, with glycine (23%), glutamine (17%), and glutamic acid (12.6%) being the most abundant amino acids. This protein also consists of 8 tandem repeats of a 20 amino acid sequence, which is characteristic of group 1 LEA proteins from non-animal species. The LEA protein and its gene are expressed only in encysted embryos and not in larvae or adults. Evidence is presented to show that the LEA protein functions in the prevention of drying-induced protein aggregation, which supports its functional role in desiccation tolerance. This report describes, for the first time, the purification and characterization of a group 1 LEA protein from an animal species.
Structure of the antigenic repeating pentasaccharide unit of the LPS O-polysaccharide of Cronobacter sakazakii implicated in the Tennessee outbreak
Authors: MacLean Leann L. , Pagotto Franco , Farber Jeffrey M. , Perry Malcolm B.
Published: April 2009

Strains of the Gram-negative bacterium Cronobacter (Enterobacter) sakazakii have been identified as emerging opportunistic pathogens that can cause enterocolitis, bacteraemia, meningitis, and brain abscess, and have been particularly associated with meningitis in neonates where infant-milk formulae has been epidemiologically linked to the disease. A study of the lipopolysaccharides produced by clinical isolates using chemical, 2D 1H and 13C NMR, and MS methods revealed that the O-polysaccharide produced by C. sakazakii (3290), a clinical strain from the Tennessee outbreak, was a branched polymer of repeating pentasaccharide units composed of 2-acetamido-2-deoxy-d-galactose, 3-(N-acetyl-l-alanylamido)-3-deoxy-d-quinovose, d-glucuronic acid, and d-glucose present in the molar ratio 1:1:1:2 and had the structure: [Formula: see text]The O-PS structure provides a unique specific structurally defined marker for the clinical tracking of this pathogen.
Influence of vitamins and osmolites on growth and bacteriocin production byLactobacillus salivariusCRL 1328 in a chemically defined medium
Authors: Vera Pingitore Esteban , Hebert Elvira María , Sesma Fernando , Nader-Macías María Elena
Published: April 2009

The aim of this study was to analyze the influence of vitamins, glycerol, and salts on the growth and bacteriocin production by Lactobacillus salivarius CRL 1328, a human vagina isolate, by using a chemically defined medium to determine the optimal conditions for salivaricin production. The single omission of d-biotin, thiamine, p-aminobenzoic acid, folic acid, or cyanocobalamin did not affect the bacterial growth, whereas the removal of nicotinic acid, riboflavin, and pyridoxal produced a decrease of about 30% in the growth rate. Maximum salivaricin activity was observed after the addition of 5 or 10 g/L of NaCl. On the basis of the nutritional requirements and the levels of salivaricin production, a new optimized and simplified defined medium (SDM–NaCl) for L. salivarius CRL 1328 bacteriocin production was formulated. The kinetics of salivaricin production in SDM–NaCl and in the complex media LAPTg revealed that bacteriocin production was growth linked. A combination of tricine – sodium dodecyl sulfate polyacrylamide gel electrophoresis (Tricine–SDS–PAGE), Lumitein protein gel staining, and a bioassay for antibacterial activity indicated that the molecular mass of salivaricin CRL 1328 is about 4.5 kDa. The partially purified bacteriocin, obtained from SDM–NaCl after concentration, allowed for the design of a relatively simple method for the recovery of a biologically active protein.
Identification and potential enzyme capacity of flavobacteria isolated from the rhizosphere of barley (Hordeum vulgare L.)
Authors: Johansen Jens Efsen , Nielsen Preben , Binnerup Svend Jørgen
Published: April 2009

The diversity of 99 flavobacterium-like isolates from a barley rhizosphere is described. They were identified on 1/10 strength tryptic soy agar by their yellowish colour and a flexirubin reaction after exposure to 10% KOH.16S rDNA partial sequencing identified the majority (70%) of isolates as Flavobacterium . Twelve percent of the isolates belonged to other genera in the phylum Bacteroidetes. Finally 17% of the isolates did not belong to the phylum Bacteroidetes. Most of the Flavobacterium isolates were affiliated to various aquatic, validly named species and likely represent a bulk of undescribed soil Flavobacterium species found in especially high numbers in the rhizosphere of young plant roots. Most Flavobacterium isolates showed gliding motility on CY agar and VY/2 agar, whereas none of the other isolates shared this feature. A high percentage of Flavobacterium strains produced enzymes involved in polysaccharide and protein digestion as well as extracellular phosphatases, compared with strains related to other genera in the phylum Bacteroidetes.
Repetitive element (REP)-polymerase chain reaction (PCR) analysis of Escherichia coli isolates from recreational waters of southeastern Lake Huron
Authors: Kon Tanya , Weir Susan C. , Howell E. Todd , Lee Hung , Trevors Jack T.
Published: April 2009

Repetitive element-polymerase chain reaction (REP-PCR) DNA fingerprinting and library-based microbial source tracking (MST) methods were utilized to investigate the potential sources of Escherichia coli pollution in recreational waters of southeastern Lake Huron. In addition to traditional sources such as humans, agriculture, and wildlife, environmentally persistent E. coli isolates were included in the identification library as a separate library unit consisting of the E. coli strains isolated from interstitial water on the beach itself. Our results demonstrated that the dominant source of E. coli pollution of the lake was agriculture, followed by environmentally adapted E. coli strains, wildlife, and then humans. A similar ratio of contributing sources was observed in all samples collected from various locations including the river discharging to the beach in both 2005 and 2006. The high similarity between the compositions of E. coli communities collected simultaneously in the river and in the lake suggests that tributaries were the major overall sources of E. coli to the lake. Our findings also suggest that environmentally adapted strains (EAS) of E. coli should be included as one of the potential sources in future microbial source tracking efforts.
Genetic and phenotypic diversity of geographically different isolates ofGlomus mosseae
Authors: Avio Luciano , Cristani Caterina , Strani Patrizia , Giovannetti Manuela
Published: April 2009

In this work, we combined morphological taxonomy and molecular methods to investigate the intraspecific diversity of Glomus mosseae , whose global distribution has been reviewed by a survey of scientific literature and Web-available records from international germplasm collections (International Culture Collection of Vesicular Arbuscular Mycorrhizal Fungi and International Bank of Glomeromycota). We surveyed 186 publications reporting the occurrence of G. mosseae from at least 474 different sites from 55 countries throughout all continents, producing a geographical map of their distribution. The relationships among G. mosseae isolates originating from Europe (United Kingdom), the United States (Arizona, Florida, and Indiana), Africa (Namibia), and West Asia (Syria) were analyzed. The level of resolution of internal transcribed spacer (ITS) sequences strongly supports the morphological species definition of G. mosseae. An ITS – restriction fragment length polymorphism assay with the enzyme HinfI yielded a unique profile for all G. mosseae isolates, allowing a straightforward identification of this morphospecies. Genetic variability among G. mosseae isolates was revealed by the inter-simple-sequence repeat (ISSR) – polymerase chain reaction: the magnitude of genetic divergence shown by the investigated geographical isolates was higher than 50%, consistent with previous data on vegetative compatibility and functional diversity. The variability of ISSR patterns suggests that intraspecific diversity is much higher than that foreseen by morphology and rDNA regions, and should be further investigated by using other genes, such as those related to functional diversity.
A novel nicotine catabolic plasmid pMH1 in Pseudomonas sp. strain HF-1
Authors: Wang Meizhen , Yang Guiqin , Min Hang , Lv Zhenmei
Published: April 2009

Attempts were made to acquire a plasmid-loss mutant via various methods (spontaneous mutation, SDS, and mitomycin C), among which the method involving mitomycin C (10 µg/mL) has been proven successful. Concomitant with the loss of the plasmid in Pseudomonas sp. strain HF-1, the cured derivative was identified as having a nicotine-negative (Nic–) phenotype, named mutant strain 6-13 (Nic–). After plasmids were transferred from strain HF-1 (named plasmid pMH1) to the mutant strain 6-13, the mutant strain acquired nicotine-degrading ability, called 6-13 transformant (Nic+). There were no differences in growth or nicotine-degrading efficiency between strain HF-1 (wild-type strain) and strain 6-13 transformant. After pMH1 was transferred to Escherichia coli strain Top10 (Nic–), a distant relative of Pseudomonas, it also gained nicotine-degrading ability, showing the highest nicotine degradation efficiency at pH 7.0, the optimal pH for growth of E. coli. The hsp gene, which encodes 6-hydroxy-3-succinoylpyridine hydroxylase, is involved in nicotine degradation in Pseudomonas putida strain S16 and was present in pMH1 but not in pAO1, the well-known nicotine degradation plasmid in Arthrobacter nicotinovorans . It was demonstrated that plasmid pMH1 is a novel nicotine-degrading plasmid.
Investigation of microbial community isolated from indoor artworks and air environment: identification, biodegradative abilities, and DNA typing
Authors: Pangallo Domenico , Chovanová Katarina , Šimonovičová Alexandra , Ferianc Peter
Published: April 2009

This study deals with establishing the characteristics of a microbial community isolated from indoor artworks and the surrounding air environment. It is one of the few studies on microbial degradation of indoor artworks. It shows the potential biodegradative risk that can occur if artworks are not exhibited and conserved in an appropriate environment. The microbial community isolated from the indoor artworks and air environment was examined by cultural and molecular methods. Different plate assays were used to screen the biodegradative activity of the isolated microflora: Remazol Brilliant Blue R, phenol red, and Azure B for the ligninolytic properties; Ostazin brilliant red H-3B for cellulose degradation; CaCO3glucose agar for solubilization activity; and B4 agar for biomineralization. To type the bacterial and fungal isolates, 2 PCR methods, repetitive extragenic palindromes (REP) and random amplified microsatellite polymorphisms (RAMP) were used. The art objects were principally colonized by fungi. The most commonly isolated strains were represented by hyphomycetes of the genera Penicillium , Aspergillus , Cladosporium , and Chaetomium . Members of these genera showed intensive biodegradation activity, both on wood and on stone. Bacteria were predominant in the air, exhibiting complex communities, both in the air and on the artworks. The most frequently isolated genera were Bacillus and Staphylococcus with extensive biodegradation abilities. REP-PCR revealed high variability within strains belonging to the same genus. RAMP is a new PCR-based method, used in this research for the first time to cluster the microfilamentous fungi and to characterize and select especially Penicillium and Aspergillus strains, which were isolated in a large number.
Genetic diversity and temporal variation of the marine Synechococcus community in the subtropical coastal waters of Hong Kong
Authors: Jing Hongmei , Zhang Rui , Pointing Stephen B. , Liu Hongbin , Qian Peiyuan
Published: April 2009

The phylogenetic diversity of the marine Synechococcus community in the subtropical coastal waters of Hong Kong, China, was examined through intergenic transcribed spacer clone libraries. All the sequences obtained fell within both marine cluster A (MC-A) and B (MC-B), with MC-A phylotypes dominating throughout the year. Distinct phylogenetic lineages specific to Hong Kong waters were detected from both MC-A and MC-B. The highest Synechococcus community diversity occurred in December, but the highest Synechococcus abundance occurred in August. On the other hand, both the abundance and diversity of Synechococcus showed a minimum in February. The remarkable seasonal variations of Synechococcus diversity observed were likely the result of the changes of hydrographic condition modulated by monsoons. Principal component analysis revealed that the in situ abiotic water characteristics, especially salinity and water turbidity, explained much of the variability of the marine Synechococcus population diversity in Hong Kong coastal waters. In addition, the temporal changes of Synechococcus abundance were largely driven by water temperature.
The SR-rich motif in SARS-CoV nucleocapsid protein is important for virus replication
Authors: Tylor Shaun , Andonov Anton , Cutts Todd , Cao Jingxin , Grudesky Elsey , Van Domselaar Gary , Li Xuguang , He Runtao
Published: April 2009

The multimerization/self-interaction of viral proteins is an important step in the process of viral assembly and maturation. Our previous study indicated that the severe acute respiratory syndrome-associated coronavirus (SARS-CoV) nucleocapsid protein forms self-multimers through a serine–arginine (SR)-rich motif (SSRSSSRSRGNSR) by using a mammalian two-hybrid system. To determine the biological relevance of this motif, we constructed a SARS-CoV reverse genetic construct by using a bacterial artificial chromosome (BAC)-based vector controlled by a T7 promoter; and subsequently deleted the SR-rich motif from the N gene. The mutated infectious clone showed reduced level of genome transcription and significantly reduced levels of the infectious virions. These results strongly suggest that the SR-rich motif is critical for effective virus replication.
Regulation of arterial remodeling and angiogenesis by urokinase-type plasminogen activatorThis article is one of a selection of papers from the NATO Advanced Research Workshop on Translational Knowledge for Heart Health (published in part 2 of a 2-part Special Issue).
Authors: Tkachuk Vsevolod A. , Plekhanova Olga S. , Parfyonova Yelena V.
Published: April 2009

A wide variety of disorders are associated with an imbalance in the plasminogen activator system, including inflammatory diseases, atherosclerosis, intimal hyperplasia, the response mechanism to vascular injury, and restenosis. Urokinase-type plasminogen activator (uPA) is a multifunctional protein that in addition to its fibrinolytic and matrix degradation capabilities also affects growth factor bioavailability, cytokine modulation, receptor shedding, cell migration and proliferation, phenotypic modulation, protein expression, and cascade activation of proteases, inhibitors, receptors, and modulators. uPA is the crucial protein for neointimal growth and vascular remodeling. Moreover, it was recently shown to be implicated in the stimulation of angiogenesis, which makes it a promising multipurpose therapeutic target. This review is focused on the mechanisms by which uPA can regulate arterial remodeling, angiogenesis, and cell migration and proliferation after arterial injury and the means by which it modulates gene expression in vascular cells. The role of domain specificity of urokinase in these processes is also discussed.
Involvement of UTP in protection of cardiomyocytes from hypoxic stressThis article is one of a selection of papers from the NATO Advanced Research Workshop on Translational Knowledge for Heart Health (published in part 2 of a 2-part Special Issue).
Authors: Shainberg Asher , Yitzhaki Smadar , Golan Or , Jacobson Kenneth A. , Hochhauser Edith
Published: April 2009

Massive amounts of nucleotides are released during ischemia in the cardiovascular system. Although the effect of the purine nucleotide ATP has been intensively studied in myocardial infarction, the cardioprotective role of the pyrimidine nucleotide UTP is still unclear, especially in the cardiovascular system. The purpose of our study was to elucidate the protective effects of UTP receptor activation and describe the downstream cascade for the cardioprotective effect. Cultured cardiomyocytes and left anterior descending (LAD)-ligated rat hearts were pretreated with UTP and exposed to hypoxia–ischemia. In vitro experiments revealed that UTP reduced cardiomyocyte death induced by hypoxia, an effect that was diminished by suramin. UTP caused several effects that could trigger a cardioprotective response: a transient increase of [Ca2+]i, an effect that was abolished by PPADS or RB2; phosphorylation of the kinases ERK and Akt, which was abolished by U0126 and LY294002, respectively; and reduced mitochondrial calcium elevation after hypoxia. In vivo experiments revealed that UTP maintained ATP levels, improved mitochondrial activity, and reduced infarct size. In conclusion, UTP administrated before ischemia reduced infarct size and improved myocardial function. Reduction of mitochondrial calcium overload can partially explain the protective effect of UTP after hypoxic–ischemic injury.