925 resultados para MICROARRAY, CLUSTER ANALYSIS, Q-PCR, RT-PCR, RESISTANCE TRAINING, AEROBIC TRAINING


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Physical exercise programmes are routinely prescribed in clinical practice to treat impairments, improve activity and participation in daily life because of their known physiological, health and psychological benefits (RCP, 2009). Progressive resistance exercise is a type of exercise prescribed specifically to improve skeletal muscle strength (Latham et al., 2004). The effectiveness of progressive resistance exercise varies considerably between studies and populations. This thesis focuses on how training parameters influence the delivery of progressive resistance exercise. In order to appropriately evaluate the influence of training parameters, this thesis argues the need to record training performance and the total work completed by participants as prescribed by training protocols. In the first study, participants were taken through a series of protocols differentiated by the intensity and volume of training. Training intensity was defined as a proportion of the mean peak torque achieved during maximal voluntary contractions and was set at 80% and 40% respectively of the MVC mean peak torque. Training volume was defined as the total external work achieved over the training period. Measures of training performance were developed to accurately report the intensity, repetitions and work completed during the training period. A second study evaluated training performance of the training protocols over repeated sessions. These protocols were then applied to 3 stroke survivors. Study 1 found sedentary participants could achieve a differentiated training intensity. Participants completing the high and low intensity protocols trained at 80% and 40% respectively of the MVC mean peak torque. The total work achieved in the high intensity low repetition protocol was lower than the total work achieved in the low intensity high repetition protocol. With repeated practice, study 2 found participants were able to improve in their ability to perform manoeuvres as shown by a reduction in the variation of the mean training intensity achieving total work as specified by the protocol to a lower margin of error. When these protocols were applied to 3 stroke survivors, they were able to achieve the specified training intensity but they were not able to achieve the total work as expected for the protocol. This is likely to be due to an inability in achieving a consistent force throughout the contraction. These results demonstrate evaluation of training characteristics and support the need to record and report training performance characteristics during progressive resistance exercise, including the total work achieved, in order to elucidate the influence of training parameters on progressive resistance exercise. The lack of accurate training performance may partly explain the inconsistencies between studies on optimal training parameters for progressive resistance exercise.

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Introduction: It is widely accepted that obesity is associated with endothelial dysfunction. In a recent paper, we have also found circuit resistance training may reduce visceral fat in obese aged women. Accordingly, the current study was conducted to ascertain the effects of circuit resistance training on markers of endothelial dysfunction in this population group. Methods: In the present interventional study, a total of 48 obese aged women were recruited from the community. Twenty-four of them were randomly assigned to perform a 12-week resistance circuit training programme, 3-days per week. This training was circularly performed in 6 stations: arm curl, leg extension, seated row, leg curl, triceps extension and leg press. The Jamar handgrip electronic dynamometer was used to assess maximal handgrip strength of the dominant hand. Lastly, serum samples were analysed using an immunoassay (ELISA) for endothelin-1, intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1). Results: When compared to baseline, resistance training significantly reduced serum levels of endothelin-1 (2.28 ± 0.7 vs. 1.98 ± 1.1 pg/ml; p = 0.019; d = 0.67) and ICAM-1 (290 ± 69 vs. 255 ± 76 ng/ml; p = 0.004; d = 0.92) in the experimental group. No significant changes in any of the tested outcomes were found in the control group. Conclusion: A short-term circuit resistance program improved endothelial dysfunction in aged obese women. Further studies on this topic are still required to consolidate this approach in clinical application.

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The primary aim of this study was to compare rating of perceived exertion (RPE) values measuring repetitions in reserve (RIR) at particular intensities of 1 repetition maximum (RM) in experienced (ES) and novice squatters (NS). Furthermore, this investigation compared average velocity between ES and NS at the same intensities. Twenty-nine individuals (24.0 ± 3.4 years) performed a 1RM squat followed by a single repetition with loads corresponding to 60, 75, and 90% of 1RM and an 8-repetition set at 70% 1RM. Average velocity was recorded at 60, 75, and 90% 1RM and on the first and last repetitions of the 8-repetition set. Subjects reported an RPE value that corresponded to an RIR value (RPE-10 = 0-RIR, RPE-9 = 1-RIR, and so forth). Subjects were assigned to one of the 2 groups: (a) ES (n = 15, training age: 5.2 ± 3.5 years) and (b) NS (n = 14, training age: 0.4 ± 0.6 years). The mean of the average velocities for ES was slower (p ≤ 0.05) than NS at 100% and 90% 1RM. However, there were no differences (p > 0.05) between groups at 60, 75%, or for the first and eighth repetitions at 70% 1RM. In addition, ES recorded greater RPE at 1RM than NS (p = 0.023). In ES, there was a strong inverse relationship between average velocity and RPE at all percentages (r = −0.88, p < 0.001), and a strong inverse correlation in NS between average velocity and RPE at all intensities (r = −0.77, p = 0.001). Our findings demonstrate an inverse relationship between average velocity and RPE/RIR. Experienced squatter group exhibited slower average velocity and higher RPE at 1RM than NS, signaling greater efficiency at high intensities. The RIR-based RPE scale is a practical method to regulate daily training load and provide feedback during a 1RM test.

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Background: La fibromialgia è una sindrome cronica caratterizzata da dolore muscolo-scheletrico associato ad altri sintomi quali stanchezza, depressione, ansia e disturbi del sonno. Questa patologia colpisce tra il 2% ed il 4% della popolazione mondiale, con un’incidenza maggiore nel sesso femminile. Chi ne è affetto riferisce un significativo peggioramento delle relazioni sociali e della qualità di vita. Il resistance training, di cui sono noti i molteplici benefici sulla salute fisica e mentale dell’individuo, è stato di recente proposto come strategia terapeutica in questa tipologia di pazienti. Obiettivo: Ricercare in letteratura prove dell’efficacia di un approccio terapeutico basato sul resistance training per ridurre il dolore e migliorare la qualità di vita nei pazienti affetti da fibromialgia. Materiali e metodi: Sono state indagate le banche dati PubMed, PEDro e Cochrane Library utilizzando quattro diverse stringhe di ricerca, ottenute combinando le parole chiave con gli operatori booleani. Per la selezione degli studi è stato impiegato il diagramma di flusso PRISMA. Sono stati inclusi solo gli RCT che proponevano un programma di resistance training non combinato con altre metodiche riabilitative. Risultati: Dei 327 articoli inizialmente individuati sono stati inclusi 3 studi che mettevano a confronto il resistance training con altre metodiche riabilitative. Tutti e tre gli studi hanno dimostrato una riduzione statisticamente significativa dell’intensità del dolore e un miglioramento, seppur non sempre significativo, della qualità di vita. Conclusioni: Gli studi selezionati hanno evidenziato un effetto benefico del resistance training, se effettuato con regolarità e costanza, nel ridurre l’intensità del dolore e migliorare la qualità di vita nei pazienti affetti da fibromialgia. Tuttavia, visto il numero ancora limitato di studi sull’argomento, è auspicabile lo svolgimento di ulteriori trials per poter acquisire maggiori prove di efficacia.

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The taxonomy of the N(2)-fixing bacteria belonging to the genus Bradyrhizobium is still poorly refined, mainly due to conflicting results obtained by the analysis of the phenotypic and genotypic properties. This paper presents an application of a method aiming at the identification of possible new clusters within a Brazilian collection of 119 Bradryrhizobium strains showing phenotypic characteristics of B. japonicum and B. elkanii. The stability was studied as a function of the number of restriction enzymes used in the RFLP-PCR analysis of three ribosomal regions with three restriction enzymes per region. The method proposed here uses Clustering algorithms with distances calculated by average-linkage clustering. Introducing perturbations using sub-sampling techniques makes the stability analysis. The method showed efficacy in the grouping of the species B. japonicum and B. elkanii. Furthermore, two new clusters were clearly defined, indicating possible new species, and sub-clusters within each detected cluster. (C) 2008 Elsevier B.V. All rights reserved.

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The taxonomy of the N(2)-fixing bacteria belonging to the genus Bradyrhizobium is still poorly refined, mainly due to conflicting results obtained by the analysis of the phenotypic and genotypic properties. This paper presents an application of a method aiming at the identification of possible new clusters within a Brazilian collection of 119 Bradryrhizobium strains showing phenotypic characteristics of B. japonicum and B. elkanii. The stability was studied as a function of the number of restriction enzymes used in the RFLP-PCR analysis of three ribosomal regions with three restriction enzymes per region. The method proposed here uses Clustering algorithms with distances calculated by average-linkage clustering. Introducing perturbations using sub-sampling techniques makes the stability analysis. The method showed efficacy in the grouping of the species B. japonicum and B. elkanii. Furthermore, two new clusters were clearly defined, indicating possible new species, and sub-clusters within each detected cluster. (C) 2008 Elsevier B.V. All rights reserved.

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Sulfate-reducing prokaryotes (SRP) are ubiquitous and quantitatively important members in many ecosystems, especially in marine sediments. However their abundance and diversity in subsurface marine sediments is poorly understood. In this study, the abundance and diversity of the functional genes for the enzymes adenosine 5'-phosphosulfate reductase (aprA) and dissimilatory sulfite reductase (dsrA) of SRP in marine sediments of the Peru continental margin and the Black Sea were analyzed, including samples from the deep biosphere (ODP site 1227). For aprA quantification a Q-PCR assay was designed and evaluated. Depth profiles of the aprA and dsrA copy numbers were almost equal for all sites. Gene copy numbers decreased concomitantly with depth from around 10(8)/g sediment close to the sediment surface to less than 10(5)/g sediment at 5 mbsf. The 16S rRNA gene copy numbers of total bacteria were much higher than those of the functional genes at all sediment depths and used to calculate the proportion of SRP to the total Bacteria. The aprA and dsrA copy numbers comprised in average 0.5-1% of the 16S rRNA gene copy numbers of total bacteria in the sediments up to a depth of ca. 40 mbsf. In the zone without detectable sulfate in the pore water from about 40-121 mbsf (Peru margin ODP site 1227), only dsrA (but not aprA) was detected with copy numbers of less than 10(4)/g sediment, comprising ca. 14% of the 16S rRNA gene copy numbers of total bacteria. In this zone, sulfate might be provided for SRP by anaerobic sulfide oxidation. Clone libraries of aprA showed that all isolated sequences originate from SRP showing a close relationship to aprA of characterized species or form a new cluster with only distant relation to aprA of isolated SRP. For dsrA a high diversity was detected, even up to 121 m sediment depth in the deep biosphere.

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Salmonella spp. are the etiologic agents of salmonellosis, a worldwide spread zoonoses causing foodborne outbreaks and clinical diseases. By serological identification, Salmonella enterica subsp. enterica serotype 1,4,[5],12:i:- accounted for 8.8% of human and 1.6% of nonhuman Salmonella strains isolated in São Paulo State, during 1991-2000. A total of 28.6% of them amplified a fragment corresponding to H:1,2 (flagellar phase two) through PCR analysis and were further assigned as S. Typhimurium. Antimicrobial resistance was detected in 36.3% of the 369 PCR-negative strains tested, including the multiresistance to ampicillin, chloramphenicol, sulfonamides, tetracycline, and streptomycin.

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We have constructed cDNA microarrays for soybean (Glycine max L. Merrill), containing approximately 4,100 Unigene ESTs derived from axenic roots, to evaluate their application and utility for functional genomics of organ differentiation in legumes. We assessed microarray technology by conducting studies to evaluate the accuracy of microarray data and have found them to be both reliable and reproducible in repeat hybridisations. Several ESTs showed high levels (>50 fold) of differential expression in either root or shoot tissue of soybean. A small number of physiologically interesting, and differentially expressed sequences found by microarray analysis were verified by both quantitative real-time RT-PCR and Northern blot analysis. There was a linear correlation (r(2) = 0.99, over 5 orders of magnitude) between microarray and quantitative real-time RT-PCR data. Microarray analysis of soybean has enormous potential not only for the discovery of new genes involved in tissue differentiation and function, but also to study the expression of previously characterised genes, gene networks and gene interactions in wild-type, mutant or transgenic; plants.

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Xanthomonas axonopodis pv. passiflorae causes bacterial spot in passion fruit. It attacks the purple and yellow passion fruit as well as the sweet passion fruit. The diversity of 87 isolates of pv. passiflorae collected from across 22 fruit orchards in Brazil was evaluated using molecular profiles and statistical procedures, including an unweighted pair-group method with arithmetical averages-based dendrogram, analysis of molecular variance (AMOVA), and an assigning test that provides information on genetic structure at the population level. Isolates from another eight pathovars were included in the molecular analyses and all were shown to have a distinct repetitive sequence-based polymerase chain reaction profile. Amplified fragment length polymorphism technique revealed considerable diversity among isolates of pv. passiflorae, and AMOVA showed that most of the variance (49.4%) was due to differences between localities. Cluster analysis revealed that most genotypic clusters were homogeneous and that variance was associated primarily with geographic origin. The disease adversely affects fruit production and may kill infected plants. A method for rapid diagnosis of the pathogen, even before the disease symptoms become evident, has value for producers. Here, a set of primers (Xapas) was designed by exploiting a single-nucleotide polymorphism between the sequences of the intergenic 16S-23S rRNA spacer region of the pathovars. Xapas was shown to effectively detect all pv. passiflorae isolates and is recommended for disease diagnosis in passion fruit orchards.

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To evaluate differential gene expression in penile tissue after treatment with the phosphodiesterase 5 (PDE5) inhibitor tadalafil, as of the three clinically available PDE5 inhibitors (sildenafil, tadalafil, and vardenafil) used for the treatment of erectile dysfunction (ED), tadalafil has a long half-life and low incidence of side-effects. In all, 32 adult rats were divided into two groups. The control group received 0.5 mL of drinking water alone, while the tadalafil group was treated with tadalafil at a dose of 0.27 mg/kg. At 4 h after treatment with water or tadalafil the rats were killed and the penile tissue was removed. The total RNA was isolated from the penile tissue from both groups and differentially expressed genes were identified by cDNA microarray analysis. To validate the expression data from the microarray analysis, quantitative real-time polymerase chain reaction (PCR) and immunohistochemistry were used. In all, 153 genes were differentially expressed between the control group and the tadalafil group. We validated the microarray results by quantitative PCR for the insulin-like growth factor binding protein 6 (IGFBP-6) gene and the neuronal calcium sensor 1 (NCS-1) gene, both of which were up-regulated in the tadalafil group, and for the natriuretic peptide receptor 1 (NPR-1) gene that was down-regulated in this group. Immunohistochemistry showed localization of the NCS-1 protein in sinusoid trabeculae of the corpus cavernosum in control and tadalafil-treated rats. There was differential expression in 153 genes after tadalafil treatment. Some of these genes such as IGFBP-6, NPR-1 and NCS-1, might result in new targets in the treatment of ED.

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Existing procedures for the generation of polymorphic DNA markers are not optimal for insect studies in which the organisms are often tiny and background molecular Information is often non-existent. We have used a new high throughput DNA marker generation protocol called randomly amplified DNA fingerprints (RAF) to analyse the genetic variability In three separate strains of the stored grain pest, Rhyzopertha dominica. This protocol is quick, robust and reliable even though it requires minimal sample preparation, minute amounts of DNA and no prior molecular analysis of the organism. Arbitrarily selected oligonucleotide primers routinely produced similar to 50 scoreable polymorphic DNA markers, between individuals of three Independent field isolates of R. dominica. Multivariate cluster analysis using forty-nine arbitrarily selected polymorphisms generated from a single primer reliably separated individuals into three clades corresponding to their geographical origin. The resulting clades were quite distinct, with an average genetic difference of 37.5 +/- 6.0% between clades and of 21.0 +/- 7.1% between individuals within clades. As a prelude to future gene mapping efforts, we have also assessed the performance of RAF under conditions commonly used in gene mapping. In this analysis, fingerprints from pooled DNA samples accurately and reproducibly reflected RAF profiles obtained from Individual DNA samples that had been combined to create the bulked samples.

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Traditional culture-dependent methods to quantify and identify airborne microorganisms are limited by factors such as short-duration sampling times and inability to count nonculturableor non-viable bacteria. Consequently, the quantitative assessment of bioaerosols is often underestimated. Use of the real-time quantitative polymerase chain reaction (Q-PCR) to quantify bacteria in environmental samples presents an alternative method, which should overcome this problem. The aim of this study was to evaluate the performance of a real-time Q-PCR assay as a simple and reliable way to quantify the airborne bacterial load within poultry houses and sewage treatment plants, in comparison with epifluorescencemicroscopy and culture-dependent methods. The estimates of bacterial load that we obtained from real-time PCR and epifluorescence methods, are comparable, however, our analysis of sewage treatment plants indicate these methods give values 270-290 fold greater than those obtained by the ''impaction on nutrient agar'' method. The culture-dependent method of air impaction on nutrient agar was also inadequate in poultry houses, as was the impinger-culture method, which gave a bacterial load estimate 32-fold lower than obtained by Q-PCR. Real-time quantitative PCR thus proves to be a reliable, discerning, and simple method that could be used to estimate airborne bacterial load in a broad variety of other environments expected to carry high numbers of airborne bacteria. [Authors]

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Previous studies have demonstrated that poultry house workers are exposed to very high levels of organic dust and consequently have an increased prevalence of adverse respiratory symptoms. However, the influence of the age of broilers on bioaerosol concentrations has not been investigated. To evaluate the evolution of bioaerosol concentration during the fattening period, bioaerosol parameters (inhalable dust, endotoxin and bacteria) were measured in 12 poultry confinement buildings in Switzerland, at three different stages of the birds' growth; samples of air taken from within the breathing zones of individual poultry house employees as they caught the chickens ready to be transported for slaughter were also analysed. Quantitative polymerase chain reaction (Q-PCR) was used to assess the quantity of total airborne bacteria and total airborne Staphylococcus species. Bioaerosol levels increased significantly during the fattening period of the chickens. During the task of catching mature birds, the mean inhalable dust concentration for a worker was 26 +/- 1.9 mg m(-3) and endotoxin concentration was 6198 +/- 2.3 EU m(-3) air, >6-fold higher than the Swiss occupational recommended value (1000 EU m(-3)). The mean exposure level of bird catchers to total bacteria and Staphylococcus species measured by Q-PCR is also very high, respectively, reaching values of 53 (+/-2.6) x 10(7) cells m(-3) air and 62 (+/-1.9) x 10(6) m(-3) air. It was concluded that in the absence of wearing protective breathing apparatus, chicken catchers in Switzerland risk exposure beyond recommended limits for all measured bioaerosol parameters. Moreover, the use of Q-PCR to estimate total and specific numbers of airborne bacteria is a promising tool for evaluating any modifications intended to improve the safety of current working practices