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PubMed2006

Changes in the community structure and activity of betaproteobacterial ammonia-oxidizing sediment bacteria along a freshwater-marine gradient.

To determine whether the distribution of estuarine ammonia-oxidizing bacteria (AOB) was influenced by salinity, the community structure of betaproteobacterial ammonia oxidizers (AOB) was characterized along a salinity gradient in sediments of the Ythan estuary, on the east coast of Scotland, UK, by denaturant gradient gel electrophoresis (DGGE), cloning and sequencing of 16S rRNA gene fragments. Ammonia-oxidizing bacteria communities at sampling sites with strongest marine influence were dominated by Nitrosospira cluster 1-like sequences and those with strongest freshwater influence were dominated by Nitrosomonas oligotropha-like sequences. Nitrosomonas sp. Nm143 was the prevailing sequence type in communities at intermediate brackish sites. Diversity indices of AOB communities were similar at marine- and freshwater-influenced sites and did not indicate lower species diversity at intermediate brackish sites. The presence of sequences highly similar to the halophilic Nitrosomonas marina and the freshwater strain Nitrosomonas oligotropha at identical sampling sites indicates that AOB communities in the estuary are adapted to a range of salinities, while individual strains may be active at different salinities. Ammonia-oxidizing bacteria communities that were dominated by Nitrosospira cluster 1 sequence types, for which no cultured representative exists, were subjected to stable isotope probing (SIP) with 13C-HCO3-, to label the nucleic acids of active autotrophic nitrifiers. Analysis of 13C-associated 16S rRNA gene fragments, following CsCl density centrifugation, by cloning and DGGE indicated sequences highly similar to the AOB Nitrosomonas sp. Nm143 and Nitrosomonas cryotolerans and to the nitrite oxidizer Nitrospira marina. No sequence with similarity to the Nitrosospira cluster 1 clade was recovered during SIP analysis. The potential role of Nitrosospira cluster 1 in autotrophic ammonia oxidation therefore remains uncertain.

Ammonia↗
PubMed2005

Isolation of lipid- and polysaccharide-degrading micro-organisms from subtropical forest soil, and analysis of lipolytic strain Bacillus sp. CR-179.

AIMS: To isolate the micro-organisms from three soil samples obtained from a subtropical forest of Puerto Iguazu (Argentina), to analyse them for detection of the biotechnologically interesting enzymatic activities lipase, esterase, cellulase, xylanase and pectinase, and to identify the most active strain. METHODS AND RESULTS: A total of 724 strains were isolated using different culture media and temperatures, and 449 of them showed at least one of the hydrolytic activities pursued. Lipolytic activity of the lipid-degrading strains was further determined using MUF-butyrate and MUF-oleate as substrates. The alkalophilic strain CR-179, one of the most active for all the enzymatic activities assayed, was characterized and preliminarily identified by morphological, physiological and 16S rDNA tests, as a Bacillus sp. closely related to Bacillus subtilis. CONCLUSIONS: Highly hydrolytic strains were isolated from all soil samples, suggesting the existence of a microbial community well-adapted to nutrient recycling. Strain CR-179, one of the most active, has been preliminarily identified as a Bacillus sp. SIGNIFICANCE AND IMPACT OF THE STUDY: A collection of hydrolytic strains with high biotechnological potential was obtained. Presence of sequences codifying for a lipolytic system related to the B. subtilis group lipases was revealed by PCR for the best lipolytic strain.

Bacillus↗
PubMed1998

Semiautomatic and robotic methods for determining serum haptoglobin levels.

The assay of Makimura and Suzuki for serum haptoglobin has been adapted to use one tube with both semiautomated and robotic methods. The adaptation permits the determination of haptoglobin in several hundred serum samples per day. Both methods were strongly correlated with the original two-tube method and had within-assay variability of 4.4% for the semiautomated and 4.8% for the robotic method. Day-to-day variability averaged 6.0% for the semiautomated and 5.5% for the robotic method. The y-intercepts for both assays were higher than that of the original two-tube method, and thus, reference values are higher than those reported previously.

Journal Article↗
PubMed1998

Robust mean and covariance structure analysis.

Covariance structure analysis is used to evaluate hypothesized influences among unmeasured latent and observed variables. As implemented, it is not robust to outliers and bad data. Several robust methods in model fitting and testing are proposed. These include direct estimation of M-estimators of structured parameters and a two-stage procedure based on robust M- and S-estimators of population covariances. The large sample properties of these estimators are obtained. The equivalence between a direct M-estimator and a two-stage estimator based on an M-estimator of population covariance is established when sampling from an elliptical distribution. Two test statistics are presented in judging the adequacy of a hypothesized model; both are asymptotically distribution free if using distribution free weight matrices. So these test statistics possess both finite sample and large sample robustness. The two-stage procedures can be easily adapted into standard software packages by modifying existing asymptotically distribution free procedures. To demonstrate the two-stage procedure, S-estimator and M-estimators under different weight functions are calculated for some real data sets.

Analysis of Variance↗
PubMed2006

The mean matters: effects of statistically defined nonspeech spectral distributions on speech categorization.

Adjacent speech, and even nonspeech, contexts influence phonetic categorization. Four experiments investigated how preceding sequences of sine-wave tones influence phonetic categorization. This experimental paradigm provides a means of investigating the statistical regularities of acoustic events that influence online speech categorization and, reciprocally, reveals regularities of the sound environment tracked by auditory processing. The tones comprising the sequences were drawn from distributions sampling different acoustic frequencies. Results indicate that whereas the mean of the distributions predicts contrastive shifts in speech categorization, variability of the distributions has little effect. Moreover, speech categorization is influenced by the global mean of the tone sequence, without significant influence of local statistical regularities within the tone sequence. Further arguing that the effect is strongly related to the average spectrum of the sequence, notched noise spectral complements of the tone sequences produce a complementary effect on speech categorization. Lastly, these effects are modulated by the number of tones in the acoustic history and the overall duration of the sequence, but not by the density with which the distribution defining the sequence is sampled. Results are discussed in light of stimulus-specific adaptation to statistical regularity in the acoustic input and a speculative link to talker normalization is postulated.

Acoustic Stimulation↗
PubMed2006

Detection of norovirus capsid protein in authentic standards and in stool extracts by matrix-assisted laser desorption ionization and nanospray mass spectrometry.

Mass spectrometry (MS) represents a rapid technique for the identification of microbial monocultures, and its adaptation to the detection of pathogens in real-world samples is a public health and homeland security priority. Norovirus, a leading cause of gastroenteritis in the world, is difficult to monitor because it cannot be cultured outside the human body. The detection of norovirus capsid protein was explored using three common MS-based methods: scanning of intact proteins, peptide mass fingerprinting, and peptide sequencing. Detection of intact target protein was limited by poor selectivity and sensitivity. Detection of up to 16 target peptides by peptide mass fingerprinting allowed for the reproducible and confident (P < 0.05) detection of the 56-kDa norovirus capsid protein in the range of 0.1 x 10(-12) to 50 x 10(-12) mol in authentic standards of recombinant norovirus virus-like particles (VLPs). To explore assay performance in complex matrixes, a non-gel-based, rapid method (2 to 3 h) for virus extraction from human stool was evaluated (72% +/- 12% recovery), and additional analyses were performed on norovirus-free stool extracts fortified with VLPs. Whereas peptide mass fingerprinting was rendered impractical by sample interferences, peptide sequencing using nanospray tandem MS facilitated unambiguous identification of > or =250 fmol of capsid protein in stool extracts. This is the first report on MS-based detection of norovirus, accomplished by using structurally identical, noninfective VLPs at clinically relevant concentrations. It represents an important milestone in the development of assays for surveillance of this category B bioterrorism agent.

Amino Acid Sequence↗
PubMed2003

Detection of antiendothelial cell antibodies by an enzyme-linked immunosorbent assay using antigens from cell lysate: minimal interference with antinuclear antibodies and rheumatoid factors.

The objective of the present work was to set up a routine test adapted to screening for antiendothelial cell antibodies (AECAs) in serum samples with minimal interference from antinuclear antibodies (ANAs) or rheumatoid factors (RFs). We compared the titers of AECAs titrated following two enzyme-linked immunosorbent assays (ELISAs): (i) an ELISA with ethanol-fixed EA.hy926 monolayers as the antigenic substrate and (ii) an ELISA with nucleus-depleted lysates prepared from EA.hy926 cells and normalized for protein (1.0 to 1.7 mg/ml) and DNA (< or =0.1 microg/ml) contents as a surrogate substrate (postnuclear supernatant ELISA [PNS-ELISA]). The AECA titers in 51 serum samples, including 28 samples containing ANAs, were compared. A significantly positive correlation (r = 0.77; P < 0.001) between the two series was shown only for the ANA-negative serum samples. Conversely, ANAs or RFs in samples were shown not to interfere in tests for AECAs by the PNS-ELISA. AECAs recognize their antigenic targets in postnuclear supernatants, which is representative of the endothelial antigenic content, with improvement of the reliability of the assay, a prerequisite to application of the assay for their evaluation in clinical practice.

Antibodies, Antinuclear↗
PubMed2000

Nutrient tasting and signaling mechanisms in the gut V. Mechanisms of immunologic sensation of intestinal contents.

Immune perception of intestinal contents reflects a functional dualism with systemic hyporesponsiveness to dietary antigens and resident microflora (oral tolerance) and active immune responses to mucosal pathogens. This facilitates optimal absorption of dietary nutrients while conserving immunologic resources for episodic pathogenic challenge. Discrimination between dangerous and harmless antigens within the enteric lumen requires continual sampling of the microenvironment by multiple potential pathways, innate and adaptive recognition mechanisms, bidirectional lymphoepithelial signaling, and rigorous control of effector responses. Errors in these processes disrupt mucosal homeostasis and are associated with food hypersensitivity and mucosal inflammation. Mechanisms of mucosal immune perception and handling of dietary proteins and other antigens have several practical and theoretical implications including vaccine design, therapy of systemic autoimmunity, and alteration of enteric flora with probiotics.

Animals↗
PubMed2003

Stroke and Aphasia Quality of Life Scale-39 (SAQOL-39): evaluation of acceptability, reliability, and validity.

BACKGROUND AND PURPOSE: Health-related quality of life (HRQL) is a key outcome in stroke clinical trials. Stroke-specific HRQL scales (eg, SS-QOL, SIS) have generally been developed with samples of stroke survivors that exclude people with aphasia. We adapted the SS-QOL for use with people with aphasia to produce the Stroke and Aphasia Quality of Life Scale (SAQOL). We report results from the psychometric evaluation of the initial 53-item SAQOL and the item-reduced SAQOL-39. METHODS: We studied 95 people with long-term aphasia to evaluate the acceptability, reliability, and validity of the SAQOL and SAQOL-39 using standard psychometric methods. RESULTS: A total of 83 of 95 (87%) were able to complete the SAQOL by self-report; their results are reported here. Results supported the reliability and validity of the overall score on the 53-item SAQOL, but there was little support for hypothesized subdomains. Using factor analysis, we derived a shorter version (SAQOL-39) that identified 4 subdomains (physical, psychosocial, communication, and energy). The SAQOL-39 demonstrated good acceptability, internal consistency (Cronbach's alpha=0.74 to 0.94), test-retest reliability (intraclass correlation coefficient=0.89 to 0.98), and construct validity (corrected domain-total correlations, r=0.38 to 0.58; convergent, r=0.55 to 0.67; discriminant, r=0.02 to 0.27 validity). CONCLUSIONS: The SAQOL-39 is an acceptable, reliable, and valid measure of HRQL in people with long-term aphasia. Further testing is needed to evaluate the responsiveness of the SAQOL-39 and to investigate its usefulness in evaluative research and routine clinical practice.

Adult↗
PubMedDate not supplied

Specific somatoform disorder in the general population.

The authors assessed the validity of the recently proposed diagnosis for specific somatoform disorder in the general population. German versions of the DSM-IV adapted Composite International Diagnostic Interview were administered to a representative sample of 4075 individuals. Multivariate analyses were used to compare impairment, life satisfaction, and use of health care. A total of 803 of 4075 subjects (19.7%) with undifferentiated somatization disorder were identified, which included 51 subjects (1.3%) who met criteria for specific somatoform disorder. Subjects with specific somatoform disorder were more impaired, had lower life satisfaction, and had higher use of health care than subjects with undifferentiated somatization disorder only. The proposed diagnosis of specific somatoform disorder demonstrated a high validity independent of comorbid depressive and anxiety disorders.

Activities of Daily Living↗
PubMed1987

A cytochemical stain for glutathione in rat hepatocytes cultured on plastic.

Thiol groups of glutathione react with the organomercurial azo dye mercury orange at a faster rate than with -SH groups of proteins. This property makes possible visualization of glutathione in cells without appreciable interference from other -SH groups. To render this method useful for cytochemical localization of glutathione in plastic cultured cells, it was necessary to adapt this reaction to the specific characteristics of the biological samples to be assayed. First, the choice of a solvent that would allow a convenient solubility of the dye and at the same time be compatible with the plastic culture plate was crucial. Second, to avoid diffusion of glutathione out of the cell the procedure for staining cells was also important. Satisfactory results were obtained after 30-40 sec reaction with 50 microM mercury orange in acetone/water 9:1, v/v, at room temperature. Glutathione-mercury orange complexes exhibited orange fluorescence on excitation with blue light. No diffusion of glutathione out of the cells was observed, and the hepatocytes stained with the dye showed orange fluorescence which paralleled their glutathione content.

Animals↗
PubMedDate not supplied

Correlative measurement of four biological contaminants on cotton lint, and their implications for occupational health.

Four biological contaminants of cotton fibers (gram-negative bacterial cells, endotoxin, fungal cells, and (1-3)-beta-D-glucan) were measured in 13 cotton lint samples from international origins, using traditional microbiological spread plating and adaptation of the Limulus amoebocyte lysate (LAL) assay. Correlations were evaluated using Spearman's rank correlation analyses. Contamination levels ranged from 713 +/- 212 to 216,830 +/- 30,413 CFU/g gram-negative bacteria; 281 +/- 29 to 9,250 +/- 820 CFU/g fungal cells; 8.30 +/- 0.89 to 137.89 +/- 21.55 ng/g endotoxin; and 15.96 +/- 5.18 to 2,964.42 +/- 313.90 LAL-reactive units/g glucan. Positive correlations existed between all contaminants; however, they were significant only between fungal cells and glucan (p < 0.05) and between endotoxin and glucan (p < 0.01). The highly significant positive correlation between endotoxin and glucan has implications for the health risk posed by the cotton-production environment, as simultaneous inhalation of these agents may cause or exacerbate lung inflammation.

Cotton Fiber↗
PubMed2002

Prediction of coronary heart disease preventive behaviors in women: a test of the health belief model.

Coronary heart disease (CHD) risk increases with age, and this increase is more dramatic in women than men. The incidence of CHD is lower in premenopausal women compared with men. After menopause, the risk of mortality from CHD increases in women. The purpose of this study was to test predictors of CHD preventive behaviors using an adapted form of the Health Belief Model (HBM) in a sample of 178 women. The predictor variables were perceptions of susceptibility to CHD, perceptions of seriousness of CHD, general health motivation, social support, and knowledge of risk factors of CHD. Regression results revealed that susceptibility to CHD, seriousness of CHD, knowledge of risk factors of CHD, and general health motivation together explained 76% of the variance of CHD behaviors. Implications for future research and for improving education about consistent adoption of CHD preventive behaviors are discussed.

Aged↗
PubMed2005

Strategies for measurement of biotransformation enzyme gene expression.

The analysis of gene expression is an integral part of any research characterizing gene function. A wide variety of techniques have been developed for this purpose, each with their own advantages and limitations. This chapter seeks to provide an overview of some of the most recent as well as conventional methods to quantitate gene expression. These approaches include Northern blot analysis, ribonuclease protection assay (RPA), reverse transcription polymerase chain reaction, expressed sequence tag (EST) sequencing, differential display, cDNA arrays, and the serial analysis of gene expression (SAGE). Current applications of the information derived from gene expression studies require assays to be adaptable for the quantitative analysis of a large number of samples and end points within a short period coupled with cost effectiveness. A comparison of some of these features of each analytical approach as well as their advantages and disadvantages has also been provided.

Animals↗
PubMed2001

Applicability of the CERAD neuropsychological battery to Brazilian elderly.

There is a limited choice of psychometric tests for Portuguese speaking people which have been evaluated in well defined groups. A Portuguese version of CERAD neuropsychological battery was applied to a control group of healthy elderly (CG) (mean age 75.1 years/ education 7.9 years), 31 Alzheimer disease (AD) patients classified by clinical dementia rating (CDR) as CDR1 (71.4/ 9.0) and 12 AD patients CDR 2 (74.1/ 9.3). Cut-off points were: verbal fluency-11; modified Boston naming-12; Mini-mental State Examination (MMSE) -26; word list memory-13; constructional praxis-9; word recall-3, word recognition-7; praxis recall-4. There was a significant difference between CG and AD-CDR1 (p<0.0001) for all tests. There was a less significant difference for constructional praxis and no difference for Boston naming. Comparison between AD-CDR1 and AD-CDR2 showed difference only for MMSE, verbal fluency, and Boston naming. The performance of CG was similar to that of a US control sample with comparable education level. These results indicate that this adaptation may be useful for the diagnosis of mild dementia but further studies are needed to define cut-offs for illiterates/low education people.

Aged↗
PubMed2003

Spreading of epithelial cells on machined and sandblasted titanium surfaces: an in vitro study.

BACKGROUND: The purpose of this investigation was to determine the influence of the surface structure of dental implants on epithelial cell spreading and growth in vitro. Cell morphology on machined and sandblasted titanium surfaces was investigated. METHODS: A total of 10 machined and 10 sandblasted discs and 10 glass coverslips were used for the present study. Samples were analyzed using scanning electron microscopy (SEM) and the cell spreading area was determined using a video image analysis system. RESULTS: After 24 hours incubation, keratinocytes grown on sandblasted titanium samples displayed numerous, long, and branched or dendritic filopodia closely adapted to the surface roughness. Filopodia varied from 3 to 12 microm in length and 0.1 to 0.3 microm in width. Cells cultured on a machined surface did not present such cytoplasmic extensions and displayed a round morphology. Keratinocytes seeded on glass coverslips were flat and edged by filopodia (maximum length 7 to 8 microm) on the spreading site of the cluster. Though cell morphology is comparable with that observed on sandblasted specimens, cytoplasmic extensions suggestive of strong adhesion and spreading attitude were less pronounced. CONCLUSION: These results indicate that sandblasted surfaces are the optimal substrata for epithelial cell adhesion and spreading.

Carcinoma, Squamous Cell↗
PubMed2002

Effect of high-oil corn or added corn oil on ruminal biohydrogenation of fatty acids and conjugated linoleic acid formation in beef steers fed finishing diets.

Three Angus steers (410 kg) cannulated in the proximal duodenum were used in a replicated 3 x 3 Latin square to evaluate the effects of dietary lipid level and oil source on ruminal biohydrogenation and conjugated linoleic acid (CLA) outflow. Dietary treatments included: 1) typical corn (TC; 79.2% typical corn), 2) high-oil corn (HOC; 79.2% high-oil corn), and 3) the TC diet with corn oil added to supply an amount of lipid equal to the HOC diet (OIL; 76.9% TC + 2.4% corn oil). Duodenal samples were collected for 4 d following 10-d diet adaptation periods. Data were analyzed with animal, square, period, and treatment in the model and planned, nonorthogonal contrasts were used to test the effects of dietary lipid content (TC vs HOC and OIL) and oil source (HOC vs OIL) on ruminal biohydrogenation. Intake and duodenal flow of total long-chain fatty acids were increased (P < 0.05) by over 63% for diets containing more lipid regardless of oil source. Apparent ruminal dry matter and long chain fatty acid digestibilities were not altered (P > 0.05) by dietary lipid level or oil source. Ruminal biohydrogenation of total and individual 18-carbon unsaturated fatty acids was greater (P < 0.05) for diets with higher lipid content. Biohydrogenation of oleic acid was greater (P < 0.05) for HOC than OIL, but biohydrogenation of linoleic acid was lower (P < 0.05) for HOC than OIL. Duodenal flows of palmitic, stearic, oleic, linoleic, and arachidic acids were more than 30% greater (P < 0.05) for diets containing more lipid. Flow of all trans-octadecenoic acids was greater (P < 0.05) for diets containing more lipid. Corn oil addition increased (P < 0.05) the flow of trans-10 octadecenoic acid and the trans-10, cis-12 isomer of CLA by threefold compared to feeding high-oil corn. Feeding high-oil corn or adding corn oil to typical corn rations increased intake, biohydrogenation, and duodenal flow of unsaturated long-chain fatty acids. Compared with high-oil corn diets, addition of corn oil increased duodenal flow of trans-10, trans-12 and cis-12 isomers of octadecenoic acid and the trans-10, cis-12 isomer of CLA. The amount of cis-9, trans-11 isomer of conjugated linoleic acid flowing to the duodenum was less than 260 mg/d, a value over 20 times lower than flow of trans-11 vaccenic acid indicating the importance of tissue desaturation for enhanced conjugated linoleic acid content of beef.

Animals↗
PubMed2003

Effects of forage and sunflower oil levels on ruminal biohydrogenation of fatty acids and conjugated linoleic acid formation in beef steers fed finishing diets.

Six Hereford steers (295 kg) cannulated in the proximal duodenum were used to evaluate the effects of forage and sunflower oil level on ruminal biohydrogenation (BH) and conjugated linoleic acid (CLA) outflow. Steers were fed one of six treatment diets in a 3 x 2 factorial arrangement of treatments (grass hay level: 12, 24, or 36% of DM; and sunflower oil level: 2 or 4% of DM) in a 6 x 6 Latin square design. The remainder of the diet was made up of steam rolled corn and protein/mineral supplement. Duodenal samples were collected for 4 d following 10-d diet adaptation periods. Data were analyzed with animal, period, forage level, sunflower oil level, and two-way interaction between forage and sunflower oil level in the model. Dry matter intake showed a quadratic response (P < 0.04), with an increase in DMI as forage level increased from 12 to 24% followed by a decrease in DMI when 36% forage was fed. Flow of fatty acids at the duodenum was higher (P < 0.03) for 4 vs. 2% sunflower oil diets, and similar among forage levels. Apparent ruminal digestibility of NDF increased in a linear manner (P < 0.04) as dietary forage level increased. Ruminal BH of dietary unsaturated 18-C fatty acids, oleic acid, and linoleic acid increased linearly (P < 0.05) as dietary forage level increased. Linoleic acid BH tended (P < 0.07) to be greater for 4 than 2% sunflower oil level. Duodenal flow of pentadecyclic, stearic, linolenic, and arachidic acids increased linearly (P < 0.05) as dietary forage level increased from 12 to 36%. Duodenal flow of linoleic acid decreased in a linear manner (P < 0.03) with increasing dietary forage level. Flow of trans-10 octadecenoate decreased linearly (P < 0.03) as dietary forage level increased, whereas trans-11 vaccenic acid flow to the duodenum increased (P < 0.01) linearly with increased dietary forage. Dietary forage or sunflower oil levels did not alter the outflow of cis-9, trans-11 CLA. Flows of cis-11, trans-13, and cis-9, cis-11 CLA increased linearly (P < 0.05) with increased dietary forage. Flows of cis-11, cis-13, and trans-11, trans-13 CLA decreased linearly (P < 0.05) with increased dietary forage. Increasing dietary forage levels from 12 to 36% in beef cattle finishing diets increased BH of unsaturated 18-C fatty acid and outflow of trans-11 vaccenic acid to duodenum without altering cis-9, trans-11 CLA outflow.

Animal Feed↗