48-spot single-molecule FRET setup with periodic acceptor excitation

Single-molecule Förster resonance energy transfer (smFRET) allows measuring distances between donor and acceptor fluorophores on the 3–10 nm range. Solution-based smFRET allows measurement of binding-unbinding events or conformational changes of dye-labeled biomolecules without ensemble averaging and free from surface perturbations. When employing dual (or multi) laser excitation, smFRET allows resolving the number of fluorescent labels on each molecule, greatly enhancing the ability to study heterogeneous samples. A major drawback to solution-based smFRET is the low throughput, which renders repetitive measurements expensive and hinders the ability to study kinetic phenomena in real-time. Here we demonstrate a high-throughput smFRET system that multiplexes acquisition by using 48 excitation spots and two 48-pixel single-photon avalanche diode array detectors. The system employs two excitation lasers allowing separation of species with one or two active fluorophores. The performance of the system is demonstrated on a set of doubly labeled double-stranded DNA oligonucleotides with different distances between donor and acceptor dyes along the DNA duplex. We show that the acquisition time for accurate subpopulation identification is reduced from several minutes to seconds, opening the way to high-throughput screening applications and real-time kinetics studies of enzymatic reactions such as DNA transcription by bacterial RNA polymerase

2011 SOSORT guidelines: Orthopaedic and Rehabilitation treatment of idiopathic scoliosis during growth

<p>Abstract</p> <p>Background</p> <p>The International Scientific Society on Scoliosis Orthopaedic and Rehabilitation Treatment (SOSORT), that produced its first Guidelines in 2005, felt the need to revise them and increase their scientific quality. The aim is to offer to all professionals and their patients an evidence-based updated review of the actual evidence on conservative treatment of idiopathic scoliosis (CTIS).</p> <p>Methods</p> <p>All types of professionals (specialty physicians, and allied health professionals) engaged in CTIS have been involved together with a methodologist and a patient representative. A review of all the relevant literature and of the existing Guidelines have been performed. Documents, recommendations, and practical approach flow charts have been developed according to a Delphi procedure. A methodological and practical review has been made, and a final Consensus Session was held during the 2011 Barcelona SOSORT Meeting.</p> <p>Results</p> <p>The contents of the document are: methodology; generalities on idiopathic scoliosis; approach to CTIS in different patients, with practical flow-charts; literature review and recommendations on assessment, bracing, physiotherapy, Physiotherapeutic Specific Exercises (PSE) and other CTIS. Sixty-five recommendations have been given, divided in the following topics: Bracing (20 recommendations), PSE to prevent scoliosis progression during growth (8), PSE during brace treatment and surgical therapy (5), Other conservative treatments (3), Respiratory function and exercises (3), Sports activities (6), Assessment (20). No recommendations reached a Strength of Evidence level I; 2 were level II; 7 level III; and 20 level IV; through the Consensus procedure 26 reached level V and 10 level VI. The Strength of Recommendations was Grade A for 13, B for 49 and C for 3; none had grade D.</p> <p>Conclusion</p> <p>These Guidelines have been a big effort of SOSORT to paint the actual situation of CTIS, starting from the evidence, and filling all the gray areas using a scientific method. According to results, it is possible to understand the lack of research in general on CTIS. SOSORT invites researchers to join, and clinicians to develop good research strategies to allow in the future to support or refute these recommendations according to new and stronger evidence.</p

Ocorrência, flutuação populacional e danos de percevejo castanho em cultivares de pastagens.

O percevejo castanho é um inseto polífago que ataca inúmeras plantas hospedeiras, e é considerado uma importante praga de gramíneas forrageiras, principalmente devido ao difícil controle. Este trabalho objetivou avaliar a ocorrência e a flutuação populacional de percevejos castanhos ao longo do tempo em área altamente infestada, a preferência do percevejo castanho por diferentes cultivares de pastagens, bem como os danos sofridos por estas em decorrência do seu ataque. As cultivares avaliadas foram Brachiaria brizantha cvs. Marandu, BRS Piatã e BRS Paiaguás, Panicum maximum cvs. Massai, BRS Zuri e BRS Tamani e Andropogon gayanus cv. Baeti, e a pastagem degradada ao redor da área experimental serviu de testemunha. O delineamento utilizado foi em blocos casualizados, com 7 tratamentos (cultivares) e 8 repetições. Em parcelas estabelecidas em área infestada, foram realizadas amostragens mensais da população de percevejos castanhos (ninfas e adultos) e avaliações de dano, por meio de atribuição de notas visuais às plantas e estimativas de perdas na produção, por meio dos parâmetros acúmulo total de MS (kg/ha de MS) e taxa de acúmulo diária de forragem (kg/ha/dia de MS). Os resultados indicaram que não houve diferença na infestação por percevejos castanhos entre as cultivares avaliadas. A distribuição de percevejos ao longo do ano foi uniforme, com sobreposição de gerações. O capim andropogon sofreu menos dano de percevejo castanho em solo arenoso de baixa/média fertilidade em comparação com as cultivares Massai, BRS Tamani, Marandu, BRS Zuri, BRS Paiaguás e BRS Piatã.bitstream/item/211563/1/Ocorrencia-flutuacao-populacional-e-danos-de-percevejo.pd

Brace technology thematic series: the progressive action short brace (PASB)

<p>Abstract</p> <p>Background</p> <p>The Progressive Action Short Brace (PASB) is a custom-made thoraco-lumbar-sacral orthosis (TLSO), devised in 1976 by Dr. Lorenzo Aulisa (Institute of Orthopedics at the Catholic University of the Sacred Heart, Rome, Italy). The PASB was designed to overcome the limits imposed by the trunk anatomy. Indeed, the particular geometry of the brace is able to generate internal forces that modify the elastic reaction of the spine. The PASB is indicated for the conservative treatment of lumbar and thoraco-lumbar scoliosis. The aim of this article is to explain the biomechanic principles of the PASB and the rationale underlying its design. Recently published studies reporting the results of PASB-based treatment of adolescent scoliotic patients are also discussed.</p> <p>Description and principles</p> <p>On the coronal plane, the upper margin of the PASB, at the side of the curve concavity, prevents the homolateral bending of the scoliotic curve. The opposite upper margin ends just beneath the apical vertebra. The principle underlying such configuration is that the deflection of the inferior tract of a curved elastic structure, fixed at the bottom end, causes straightening of its upper tract. Therefore, whenever the patient bends towards the convexity of the scoliotic curve, the spine is deflected. On the sagittal plane, the inferior margins of the PASB reach the pelvitrochanteric region, in order to stabilize the brace on the pelvis. The transverse section of the brace above the pelvic grip consists of asymmetrical ellipses. This allows the spine to rotate towards the concave side only, leading to the continuous generation of derotating moments. On the sagittal plane, the brace is contoured so as to reduce the lumbar lordosis. The PASB, by allowing only those movements counteracting the progression of the curve, is able to produce corrective forces that are not dissipated. Therefore, the brace is based on the principle that a constrained spine dynamics can achieve the correction of a curve by inverting the abnormal load distribution during skeletal growth.</p> <p>Results</p> <p>Since its introduction in 1976, several studies have been published supporting the validity of the biomechanical principles to which the brace is inspired. In this article, we present the outcome of a case series comprising 110 patients with lumbar and thoraco-lumbar curves treated with PASB brace. Antero-posterior radiographs were used to estimate the curve magnitude (C_M) and the torsion of the apical vertebra (T_A) at 5 time points: beginning of treatment (t₁), one year after the beginning of treatment (t₂), intermediate time between t₁and t₄(t₃), end of weaning (t₄), 2-year minimum follow-up from t₄(t₅). The average C_Mvalue was 29.3°Cobb at t₁and 13.0°Cobb at t₅. T_Awas 15.8° Perdroille at t₁and 5.0° Perdriolle at t₅. These results support the efficacy of the PASB in the management of scoliotic patients with lumbar and thoraco-lumbar curves.</p> <p>Conclusion</p> <p>The results obtained in patients treated with the PASB confirm the validity of our original biomechanical approach. The efficacy of the PASB derives not only from its unique biomechanical features but also from the simplicity of its design, construction and management.</p

Treatment of thoraco-lumbar curves in adolescent females affected by idiopathic scoliosis with a progressive action short brace (PASB): assessment of results according to the SRS committee on bracing and nonoperative management standardization criteria

<p>Abstract</p> <p>Background</p> <p>The effectiveness of conservative treatment of scoliosis is controversial. Some studies suggest that brace is effective in stopping curve progression, whilst others did not report such an effect.</p> <p>The purpose of the present study was to effectiveness of Progressive Action Short Brace (PASB) in the correction of thoraco-lumbar curves, in agreement with the Scoliosis Research Society (SRS) Committee on Bracing and Nonoperative Management Standardisation Criteria.</p> <p>Methods</p> <p>Fifty adolescent females (mean age 11.8 ± 0.5 years) with thoraco-lumbar curve and a pre-treatment Risser score ranging from 0 to 2 have been enrolled. The minimum duration of follow-up was 24 months (mean: 55.4 ± 44.5 months). Antero-posterior radiographs were used to estimate the curve magnitude (C_M) and the torsion of the apical vertebra (T_A) at 5 time points: beginning of treatment (t₁), one year after the beginning of treatment (t₂), intermediate time between t₁and t₄(t₃), end of weaning (t₄), 2-year minimum follow-up from t₄(t₅). Three situations were distinguished: curve correction, curve stabilisation and curve progression.</p> <p>The Kruskal Wallis and Spearman Rank Correlation tests have been used as statistical tests.</p> <p>Results</p> <p>C_Mmean value was 29,30 ± 5,16 SD at t₁and 14,67 ± 7,65 SD at t₅. T_Awas 12.70 ± 6,14 SD at t₁and 8,95 ± 5,82 at t₅. The variation between measures of Cobb and Perdriolle degrees at t_1,2,3,4,5and between C_Mt₅-t₁and T_At₅-t₁were significantly different.</p> <p>Curve correction was accomplished in 94% of patients, whereas a curve stabilisation was obtained in 6% of patients.</p> <p>Conclusion</p> <p>The PASB, due to its peculiar biomechanical action on vertebral modelling, is highly effective in correcting thoraco-lumbar curves.</p

Antibody Vh Repertoire Differences between Resolving and Chronically Evolving Hepatitis C Virus Infections

Despite the production of neutralizing antibodies to hepatitis C virus (HCV), many patients fail to clear the virus and instead develop chronic infection and long-term complications. To understand how HCV infection perturbs the antibody repertoire and to identify molecular features of antibody genes associated with either viral clearance or chronic infection, we sequenced the V(D)J region of naïve and memory B cells of 6 persons who spontaneously resolved an HCV infection (SR), 9 patients with a newly diagnosed chronically evolving infection (CE), and 7 healthy donors. In both naïve and memory B cells, the frequency of use of particular antibody gene subfamilies and segments varied among the three clinical groups, especially between SR and CE. Compared to CE, SR antibody genes used fewer VH, D and JH gene segments in naïve B cells and fewer VH segments in memory B cells. SR and CE groups significantly differed in the frequency of use of 7 gene segments in naïve B cell clones and 3 gene segments in memory clones. The nucleotide mutation rates were similar among groups, but the pattern of replacement and silent mutations in memory B cell clones indicated greater antigen selection in SR than CE. Greater clonal evolution of SR than CE memory B cells was revealed by analysis of phylogenetic trees and CDR3 lengths. Pauciclonality of the peripheral memory B cell population is a distinguishing feature of persons who spontaneously resolved an HCV infection. This finding, previously considered characteristic only of patients with HCV-associated lymphoproliferative disorders, suggests that the B cell clones potentially involved in clearance of the virus may also be those susceptible to abnormal expansion

Determination of quality of life in adolescents with idiopathic scoliosis subjected to conservative treatment.

<p>Abstract</p> <p>Introduction and objectives</p> <p>Physical deformities caused by adolescent idiopathic scoliosis (AIS) coupled with conservative treatment of AIS with orthesis unavoidably impacts on patients' quality of life (QoL). The present study aimed at evaluating the QoL in patients affected by AIS treated with brace. The study also sought to determine the ability of different QoL questionnaires to monitor QoL over the course of treatment.</p> <p>Materials and methods</p> <p>Data were collected in 108 consecutive patients (96 females, 16 males) affected by AIS admitted to the outpatient orthopaedic clinic of the Catholic University of the Sacred Heart in Rome (Italy). Patients were subjected to full-time (i.e., 22 hrs per day) conservative treatment with the progressive action short brace (PASB), the Lyon brace or a combination of PASB + Lyon brace. Three instruments were used for QoL determination: the Scoliosis Research Society 22 (SRS-22), Bad Sobernheim Stress Questionnaire (BSSQ) and the Brace Questionnaire (BrQ).</p> <p>Results</p> <p>A significant correlation was detected among the 3 scores (p < 0.001). The BrQ possesses a higher capacity to detect changes in QoL in relation to the patient gender, type of brace, curve severity at baseline and at the completion of treatment, and curve type. Overall, boys displayed a higher QoL than girls. In all 3 questionnaires, higher QoL scores were determined in patients treated with the PASB compared with those using the Lyon brace. QoL scores were significantly correlated with the curve severity. Higher QoL scores were obtained by participants with thoraco-lumbar curves as compared with those with other curves.</p> <p>Conclusions</p> <p>The 3 questionnaires are effective in capturing changes in QoL in AIS patients subjected to conservative treatment. However, the BrQ possesses a higher discriminatory capacity compared with the other questionnaires tested. PASB-based treatment is associated with better QoL than the Lyon bracing.</p

Performance of current guidelines for diagnosis of macrophage activation syndrome complicating systemic juvenile idiopathic arthritis

Publisher Copyright: Copyright © 2014 by the American College of Rheumatology.Results The study sample included 362 patients with systemic JIA and MAS, 404 patients with active systemic JIA without MAS, and 345 patients with systemic infection. The best capacity to differentiate MAS from systemic JIA without MAS was found when the preliminary MAS guidelines were applied. The 3/5-adapted HLH-2004 guidelines performed better than the 4/5-adapted guidelines in distinguishing MAS from active systemic JIA without MAS. The 3/5-adapted HLH-2004 guidelines and the preliminary MAS guidelines with the addition of ferritin levels ≥500 ng/ml discriminated best between MAS and systemic infections. Conclusion The preliminary MAS guidelines showed the strongest ability to identify MAS in systemic JIA. The addition of hyperferritinemia enhanced their capacity to differentiate MAS from systemic infections. The HLH-2004 guidelines are likely not appropriate for identification of MAS in children with systemic JIA. Objective To compare the capacity of the 2004 diagnostic guidelines for hemophagocytic lymphohistiocytosis (HLH-2004) with the capacity of the preliminary diagnostic guidelines for systemic juvenile idiopathic arthritis (JIA)-associated macrophage activation syndrome (MAS) to discriminate MAS complicating systemic JIA from 2 potentially confusable conditions, represented by active systemic JIA without MAS and systemic infection. Methods International pediatric rheumatologists and hemato-oncologists were asked to retrospectively collect clinical information from patients with systemic JIA-associated MAS and confusable conditions. The ability of the guidelines to differentiate MAS from the control diseases was evaluated by calculating the sensitivity and specificity of each set of guidelines and the kappa statistics for concordance with the physician's diagnosis. Owing to the fact that not all patients were assessed for hemophagocytosis on bone marrow aspirates and given the lack of data on natural killer cell activity and soluble CD25 levels, the HLH-2004 guidelines were adapted to enable the diagnosis of MAS when 3 of 5 of the remaining items (3/5-adapted) or 4 of 5 of the remaining items (4/5-adapted) were present.publishersversionPeer reviewe

The TESS-Keck Survey. XV. Precise Properties of 108 TESS Planets and Their Host Stars

We present the stellar and planetary properties for 85 TESS Objects of Interest (TOIs) hosting 108 planet candidates which comprise the TESS-Keck Survey (TKS) sample. We combine photometry, high-resolution spectroscopy, and Gaia parallaxes to measure precise and accurate stellar properties. We then use these parameters as inputs to a lightcurve processing pipeline to recover planetary signals and homogeneously fit their transit properties. Among these transit fits, we detect significant transit-timing variations among at least three multi-planet systems (TOI-1136, TOI-1246, TOI-1339) and at least one single-planet system (TOI-1279). We also reduce the uncertainties on planet-to-star radius ratios $R_p/R_\star$ across our sample, from a median fractional uncertainty of 8.8$\%$ among the original TOI Catalog values to 3.0$\%$ among our updated results. With this improvement, we are able to recover the Radius Gap among small TKS planets and find that the topology of the Radius Gap among our sample is broadly consistent with that measured among Kepler planets. The stellar and planetary properties presented here will facilitate follow-up investigations of both individual TOIs and broader trends in planet properties, system dynamics, and the evolution of planetary systems.Comment: Accepted at The Astronomical Journal; 21 pages, 9 figure

The TESS-Keck Survey. XXII. A Sub-Neptune Orbiting TOI-1437

Exoplanet discoveries have revealed a dramatic diversity of planet sizes across a vast array of orbital architectures. Sub-Neptunes are of particular interest; due to their absence in our own solar system, we rely on demographics of exoplanets to better understand their bulk composition and formation scenarios. Here, we present the discovery and characterization of TOI-1437 b, a sub-Neptune with a 18.84 day orbit around a near-solar analog (M⋆ = 1.10 ± 0.10 M☉, R⋆=1.17 ± 0.12 R☉). The planet was detected using photometric data from the Transiting Exoplanet Survey Satellite (TESS) mission and radial velocity (RV) follow-up observations were carried out as a part of the TESS-Keck Survey using both the HIRES instrument at Keck Observatory and the Levy Spectrograph on the Automated Planet Finder telescope. A combined analysis of these data reveal a planet radius of Rp = 2.24 ± 0.23 R⊕ and a mass measurement of Mp = 9.6 ± 3.9 M⊕). TOI-1437 b is one of few (∼50) known transiting sub-Neptunes orbiting a solar-mass star that has a RV mass measurement. As the formation pathway of these worlds remains an unanswered question, the precise mass characterization of TOI-1437 b may provide further insight into this class of planet