Genetic screening of Fabry patients with EcoTILLING and HRM technology

<p>Abstract</p> <p>Background</p> <p>Anderson-Fabry disease (FD) is caused by a deficit of the α-galactosidase A enzyme which leads to the accumulation of complex sphingolipids, especially globotriaosylceramide (Gb3), in all the cells of the body, causing the onset of a multi-systemic disease with poor prognosis in adulthood. In this article, we describe two alternative methods for screening the <it>GLA </it>gene which codes for the α-galactosidase A enzyme in subjects with probable FD in order to test analysis strategies which include or rely on initial pre-screening.</p> <p>Findings</p> <p>We analyzed 740 samples using EcoTILLING, comparing two mismatch-specific<ul/>endonucleases, CEL I and ENDO-1, while conducting a parallel screening of the same samples using HRM (High Resolution Melting). Afterwards, all samples were subjected to direct sequencing. Overall, we identified 12 different genetic variations: -10C>T, -12G>A, -30G>A, IVS2-76_80del5, D165H, C172Y, IVS4+16A>G, IVS4 +68 A>G, c.718_719delAA, D313Y, IVS6-22C>T, G395A. This was consistent with the high genetic heterogeneity found in FD patients and carriers. All of the mutations were detected by HRM, whereas 17% of the mutations were not found by EcoTILLING. The results obtained by EcoTILLING comparing the CEL I and ENDO-1 endonucleases were perfectly overlapping.</p> <p>Conclusion</p> <p>On the basis of its simplicity, flexibility, repeatability, and sensitivity, we believe that<ul/>HRM analysis of the <it>GLA </it>gene is a reliable presequencing screening tool. This method can be applied to any genomic feature to identify known and unknown genetic alterations, and it is ideal for conducting screening and population studies.</p

"Delirium Day": A nationwide point prevalence study of delirium in older hospitalized patients using an easy standardized diagnostic tool

Background: To date, delirium prevalence in adult acute hospital populations has been estimated generally from pooled findings of single-center studies and/or among specific patient populations. Furthermore, the number of participants in these studies has not exceeded a few hundred. To overcome these limitations, we have determined, in a multicenter study, the prevalence of delirium over a single day among a large population of patients admitted to acute and rehabilitation hospital wards in Italy. Methods: This is a point prevalence study (called "Delirium Day") including 1867 older patients (aged 65 years or more) across 108 acute and 12 rehabilitation wards in Italian hospitals. Delirium was assessed on the same day in all patients using the 4AT, a validated and briefly administered tool which does not require training. We also collected data regarding motoric subtypes of delirium, functional and nutritional status, dementia, comorbidity, medications, feeding tubes, peripheral venous and urinary catheters, and physical restraints. Results: The mean sample age was 82.0 \ub1 7.5 years (58 % female). Overall, 429 patients (22.9 %) had delirium. Hypoactive was the commonest subtype (132/344 patients, 38.5 %), followed by mixed, hyperactive, and nonmotoric delirium. The prevalence was highest in Neurology (28.5 %) and Geriatrics (24.7 %), lowest in Rehabilitation (14.0 %), and intermediate in Orthopedic (20.6 %) and Internal Medicine wards (21.4 %). In a multivariable logistic regression, age (odds ratio [OR] 1.03, 95 % confidence interval [CI] 1.01-1.05), Activities of Daily Living dependence (OR 1.19, 95 % CI 1.12-1.27), dementia (OR 3.25, 95 % CI 2.41-4.38), malnutrition (OR 2.01, 95 % CI 1.29-3.14), and use of antipsychotics (OR 2.03, 95 % CI 1.45-2.82), feeding tubes (OR 2.51, 95 % CI 1.11-5.66), peripheral venous catheters (OR 1.41, 95 % CI 1.06-1.87), urinary catheters (OR 1.73, 95 % CI 1.30-2.29), and physical restraints (OR 1.84, 95 % CI 1.40-2.40) were associated with delirium. Admission to Neurology wards was also associated with delirium (OR 2.00, 95 % CI 1.29-3.14), while admission to other settings was not. Conclusions: Delirium occurred in more than one out of five patients in acute and rehabilitation hospital wards. Prevalence was highest in Neurology and lowest in Rehabilitation divisions. The "Delirium Day" project might become a useful method to assess delirium across hospital settings and a benchmarking platform for future surveys

SPAD-based asynchronous-readout array detectors for image-scanning microscopy

Fluorescence microscopy and derived techniques are continuously looking for photodetectors able to guarantee increased sensitivity, high spatial and temporal resolution, and ease of integration into modern microscopy architectures. Recent advances in single-photon avalanche diodes (SPADs) fabricated with industry-standard microelectronic processes allow the development of new detection systems tailored to address the requirements of advanced imaging techniques (such as image-scanning microscopy). To this aim, we present the complete design and characterization of two bidimensional SPAD arrays composed of 25 fully independent and asynchronously operated pixels, both having fill factor of about 50% and specifically designed for being integrated into existing laser scanning microscopes. We used two different microelectronics technologies to fabricate our detectors: the first technology exhibiting very low noise (roughly 200 dark counts per second at room temperature) and the second one showing enhanced detection efficiency (more than 60% at a wavelength of 500 nm). Starting from the silicon-level device structures and moving towards the in-pixel and readout electronics description, we present performance assessments and comparisons between the two detectors. Images of a biological sample acquired after their integration into our custom image-scanning microscope finally demonstrate their exquisite on-field performance in terms of spatial resolution and contrast enhancement. We envisage that this work can trigger the development of a new class of SPAD-based detector arrays able to substitute the typical single-element sensor used in fluorescence laser scanning microscopy

SPAD-based asynchronous-readout array detectors for image-scanning microscopy

Fluorescence microscopy and derived techniques are continuously looking for photodetectors able to guarantee increased sensitivity, high spatial and temporal resolution, and ease of integration into modern microscopy architectures. Recent advances in single-photon avalanche diodes (SPADs) fabricated with industry-standard microelectronic processes allow the development of new detection systems tailored to address the requirements of advanced imaging techniques (such as image-scanning microscopy). To this aim, we present the complete design and characterization of two bidimensional SPAD arrays composed of 25 fully independent and asynchronously operated pixels, both having fill factor of about 50% and specifically designed for being integrated into existing laser scanning microscopes. We used two different microelectronics technologies to fabricate our detectors: the first technology exhibiting very low noise (roughly 200 dark counts per second at room temperature) and the second one showing enhanced detection efficiency (more than 60% at a wavelength of 500 nm). Starting from the silicon-level device structures and moving towards the in-pixel and readout electronics description, we present performance assessments and comparisons between the two detectors. Images of a biological sample acquired after their integration into our custom image-scanning microscope finally demonstrate their exquisite on-field performance in terms of spatial resolution and contrast enhancement. We envisage that this work can trigger the development of a new class of SPAD-based detector arrays able to substitute the typical single-element sensor used in fluorescence laser scanning microscopy

Nanostructured TiO2 Grown by Low-Temperature Reactive Sputtering for Planar Perovskite Solar Cells

Low-temperature nanostructured electron-transporting layers (ETLs) for perovskite solar cells are grown by reactive sputtering at 160 degrees C with thickness in the range 22-76 nm and further stabilization in air at 180 degrees C to improve the lattice structure and to consequently reduce charge recombination during solar cell operation. In addition, the post-deposition treatment aims at leveling differences among samples to ensure material reproducibility. Nanostructured TiO2 has a further added value in promoting the structural coupling with the perovskite layer and establishing conformal interfaces in favor of the charge extraction from the active material. Nanostructuring of the ETLs also allows the shaping of the band gap width and position with a beneficial impact on the electrical parameters, as tested in standard architecture containing methylammonium lead iodide perovskites. A balance among parameters is achieved using a 40-nm-thick TiO2 ETL with a maximum efficiency of similar to 5% reached without surface treatments or additional layers. The proposed growth methodology would be compatible with the use of flexible substrates after appropriated ETL structural adaptation. It can be likewise applied in perovskite/silicon-heterojunction tandem solar cells to fulfill the industrial demand for clean, solvent-free, reproducible, reliable, and high-throughput processes

Observational case-control study of the prevalence of chronic cerebrospinal venous insufficiency in multiple sclerosis: results from the CoSMo study.

Background: Chronic cerebrospinal venous insufficiency (CCSVI) has been proposed as a possible cause of multiple sclerosis (MS). Objectives: The CoSMo study evaluated the association between CCSVI and MS. Methods: The primary end-point of this multicentric, case-control study was to compare the prevalence of CCSVI between patients with MS, patients with other neurodegenerative diseases (ONDs) and healthy controls (HCs). Color-coded duplex sonography was performed by a sonologist and the images were sent to one of three central sonologists for a second reading. Agreement between local and central sonologists or, in case of disagreement, the predominant judgment among the three central readers, was required for a diagnosis of CCSVI. All readings, data collection and analysis were blinded. Results: The study involved 35 MS centers across Italy and included 1874 subjects aged 18-55. 1767 (94%) were evaluable: 1165 MS patients, 226 patients with ONDs and 376 HCs. CCSVI prevalence was 3.26%, 3.10% and 2.13% for the MS, OND and HC groups, respectively. No significant difference in CCSVI prevalence was found amongst the three cohorts (MS versus HC, OR = 1.55, 95%CI = 0.72-3.36, ρ= 0.30; OND versus HC, OR = 1.47, 95%CI = 0.53-4.11, ρ= 0.46; MS versus OND, OR = 1.05, 95%CI = 0.47-2.39, ρ = 0.99). High negative and low positive agreement was found between the local and centralized readers. Conclusions: CCSVI is not associated with MS. © 2013 The Author(s)