Genotype-guided diagnostic reassessment after exome sequencing in neuromuscular disorders: experiences with a two-step approach.

BACKGROUND AND PURPOSE: Next-generation sequencing has greatly improved the diagnostic success rates for genetic neuromuscular disorders (NMDs). Nevertheless, most patients still remain undiagnosed, and there is a need to maximize the diagnostic yield. METHODS: A retrospective study was conducted on 72 patients with NMDs who underwent exome sequencing (ES), partly followed by genotype-guided diagnostic reassessment and secondary investigations. The diagnostic yields that would have been achieved by appropriately chosen narrow and comprehensive gene panels were also analysed. RESULTS: The initial diagnostic yield of ES was 30.6% (n = 22/72 patients). In an additional 15.3% of patients (n = 11/72) ES results were of unknown clinical significance. After genotype-guided diagnostic reassessment and complementary investigations, the yield was increased to 37.5% (n = 27/72). Compared to ES, targeted gene panels (<25 kilobases) reached a diagnostic yield of 22.2% (n = 16/72), whereas comprehensive gene panels achieved 34.7% (n = 25/72). CONCLUSION: Exome sequencing allows the detection of pathogenic variants missed by (narrowly) targeted gene panel approaches. Diagnostic reassessment after genetic testing further enhances the diagnostic outcomes for NMDs.

The vaginal metabolome and microbiota of cervical HPV-positive and HPV-negative women: a cross-sectional analysis.

OBJECTIVE: Characterise the vaginal metabolome of cervical HPV-infected and uninfected women. DESIGN: Cross-sectional. SETTING: The Center for Health Behavior Research at the University of Maryland School of Public Health. SAMPLE: Thirty-nine participants, 13 categorised as HPV-negative and 26 as HPV-positive (any genotype; HPV+ ), 14 of whom were positive with at least one high-risk HPV strain (hrHPV). METHOD: Self-collected mid-vaginal swabs were profiled for bacterial composition by 16S rRNA gene amplicon sequencing, metabolites by both gas and liquid chromatography mass spectrometry, and 37 types of HPV DNA. MAIN OUTCOME MEASURES: Metabolite abundances. RESULTS: Vaginal microbiota clustered into Community State Type (CST) I (Lactobacillus crispatus-dominated), CST III (Lactobacillus iners-dominated), and CST IV (low-Lactobacillus, 'molecular-BV'). HPV+ women had higher biogenic amine and phospholipid concentrations compared with HPV- women after adjustment for CST and cigarette smoking. Metabolomic profiles of HPV+ and HPV- women differed in strata of CST. In CST III, there were higher concentrations of biogenic amines and glycogen-related metabolites in HPV+ women than in HPV- women. In CST IV, there were lower concentrations of glutathione, glycogen, and phospholipid-related metabolites in HPV+ participants than in HPV- participants. Across all CSTs, women with hrHPV strains had lower concentrations of amino acids, lipids, and peptides compared with women who had only low-risk HPV (lrHPV). CONCLUSIONS: The vaginal metabolome of HPV+ women differed from HPV- women in terms of several metabolites, including biogenic amines, glutathione, and lipid-related metabolites. If the temporal relation between increased levels of reduced glutathione and oxidised glutathione and HPV incidence/persistence is confirmed in future studies, anti-oxidant therapies may be considered as a non-surgical HPV control intervention. TWEETABLE ABSTRACT: Metabolomics study: Vaginal microenvironment of HPV+ women may be informative for non-surgical interventions.

Altered Relationship between Working Memory and Brain Microstructure after Mild Traumatic Brain Injury.

BACKGROUND AND PURPOSE: Working memory impairment is one of the most troubling and persistent symptoms after mild traumatic brain injury (MTBI). Here we investigate how working memory deficits relate to detectable WM microstructural injuries to discover robust biomarkers that allow early identification of patients with MTBI at the highest risk of working memory impairment. MATERIALS AND METHODS: Multi-shell diffusion MR imaging was performed on a 3T scanner with 5 b-values. Diffusion metrics of fractional anisotropy, diffusivity and kurtosis (mean, radial, axial), and WM tract integrity were calculated. Auditory-verbal working memory was assessed using the Wechsler Adult Intelligence Scale, 4th ed, subtests: 1) Digit Span including Forward, Backward, and Sequencing; and 2) Letter-Number Sequencing. We studied 19 patients with MTBI within 4 weeks of injury and 20 healthy controls. Tract-Based Spatial Statistics and ROI analyses were performed to reveal possible correlations between diffusion metrics and working memory performance, with age and sex as covariates. RESULTS: ROI analysis found a significant positive correlation between axial kurtosis and Digit Span Backward in MTBI (Pearson r = 0.69, corrected P = .04), mainly present in the right superior longitudinal fasciculus, which was not observed in healthy controls. Patients with MTBI also appeared to lose the normal associations typically seen in fractional anisotropy and axonal water fraction with Letter-Number Sequencing. Tract-Based Spatial Statistics results also support our findings. CONCLUSIONS: Differences between patients with MTBI and healthy controls with regard to the relationship between microstructure measures and working memory performance may relate to known axonal perturbations occurring after injury.

Cigarette smoking is associated with an altered vaginal tract metabolomic profile.

Cigarette smoking has been associated with both the diagnosis of bacterial vaginosis (BV) and a vaginal microbiota lacking protective Lactobacillus spp. As the mechanism linking smoking with vaginal microbiota and BV is unclear, we sought to compare the vaginal metabolomes of smokers and non-smokers (17 smokers/19 non-smokers). Metabolomic profiles were determined by gas and liquid chromatography mass spectrometry in a cross-sectional study. Analysis of the 16S rRNA gene populations revealed samples clustered into three community state types (CSTs) ---- CST-I (L. crispatus-dominated), CST-III (L. iners-dominated) or CST-IV (low-Lactobacillus). We identified 607 metabolites, including 12 that differed significantly (q-value < 0.05) between smokers and non-smokers. Nicotine, and the breakdown metabolites cotinine and hydroxycotinine were substantially higher in smokers, as expected. Among women categorized to CST-IV, biogenic amines, including agmatine, cadaverine, putrescine, tryptamine and tyramine were substantially higher in smokers, while dipeptides were lower in smokers. These biogenic amines are known to affect the virulence of infective pathogens and contribute to vaginal malodor. Our data suggest that cigarette smoking is associated with differences in important vaginal metabolites, and women who smoke, and particularly women who are also depauperate for Lactobacillus spp., may have increased susceptibilities to urogenital infections and increased malodor.

Note: Laser-cut molybdenum grids for a retarding field energy analyzer.

A retarding field energy analyzer (RFEA) with grids created by laser-cutting a honeycomb mesh in a 50 µm thick molybdenum foil is presented. The flat grids span an area of 1 cm2 and have high transmission (20 µm wide walls between 150 µm wide meshes). The molybdenum grids were tested in a 3-grid RFEA configuration with an analyzer depth of 0.87 mm.

Hereditary spastic paraplegia caused by compound heterozygous mutations outside the motor domain of the KIF1A gene.

BACKGROUND AND PURPOSE: Hereditary spastic paraplegia is a clinically and genetically heterogeneous group of rare, inherited disorders causing an upper motor neuron syndrome with (complex) or without (pure) additional neurological symptoms. Mutations in the KIF1A gene have already been associated with recessive and dominant forms of hereditary spastic paraplegia (SPG30) in a few cases. METHODS: All family members included in the study were examined neurologically. Whole-exome sequencing was used in affected individuals to identify the responsible candidate gene. Conventional Sanger sequencing was conducted to validate familial segregation. RESULTS: A family of Macedonian origin with two affected siblings, one with slowly progressive and the other one with a more complex and rapidly progressing hereditary spastic paraplegia is reported. In both affected individuals, two novel pathogenic mutations outside the motor domain of the KIF1A gene were found (NM_001244008.1:c.2909G>A, p.Arg970His and c.1214dup, p.Asn405Lysfs*40) that segregate with the disease within the family establishing the diagnosis of autosomal recessive SPG30. CONCLUSIONS: This report provides the first evidence that mutations outside the motor domain of the gene can cause (recessive) SPG30 and extends the genotype-phenotype association for KIF1A-related diseases.

Adaptive super-twisting observer for estimation of random road excitation profile in automotive suspension systems.

The estimation of road excitation profile is important for evaluation of vehicle stability and vehicle suspension performance for autonomous vehicle control systems. In this work, the nonlinear dynamics of the active automotive system that is excited by the unknown road excitation profile are considered for modeling. To address the issue of estimation of road profile, we develop an adaptive supertwisting observer for state and unknown road profile estimation. Under Lipschitz conditions for the nonlinear functions, the convergence of the estimation error is proven. Simulation results with Ford Fiesta MK2 demonstrate the effectiveness of the proposed observer for state and unknown input estimation for nonlinear active suspension system.

The benefits of skull stripping in the normalization of clinical fMRI data.

Establishing a reliable correspondence between lesioned brains and a template is challenging using current normalization techniques. The optimum procedure has not been conclusively established, and a critical dichotomy is whether to use input data sets which contain skull signal, or whether skull signal should be removed. Here we provide a first investigation into whether clinical fMRI benefits from skull stripping, based on data from a presurgical language localization task. Brain activation changes related to deskulled/not-deskulled input data are determined in the context of very recently developed (New Segment, Unified Segmentation) and standard normalization approaches. Analysis of structural and functional data demonstrates that skull stripping improves language localization in MNI space - particularly when used in combination with the New Segment normalization technique.

Low temperature silicon nitride by hot wire chemical vapour deposition for the use in impermeable thin film encapsulation on flexible substrates.

High quality non porous silicon nitride layers were deposited by hot wire chemical vapour deposition at substrate temperatures lower than 110 degrees C. The layer properties were investigated using FTIR, reflection/transmission measurements and 1:6 buffered HF etching rate. A Si-H peak position of 2180 cm(-1) in the Fourier transform infrared absorption spectrum indicates a N/Si ratio around 1.2. Together with a refractive index of 1.97 at a wavelength of 632 nm and an extinction coefficient of 0.002 at 400 nm, this suggests that a transparent high density silicon nitride material has been made below 110 degrees C, which is compatible with polymer films and is expected to have a high impermeability. To confirm the compatibility with polymer films a silicon nitride layer was deposited on poly(glycidyl methacrylate) made by initiated chemical vapour deposition, resulting in a highly transparent double layer.

Clinical fMRI: evidence for a 7T benefit over 3T.

Despite there being an increasing number of installations of ultra high field MR systems (>3T) in clinical environments, no functional patient investigations have yet examined possible benefits for functional diagnostics. Here we performed presurgical localization of the primary motor hand area on 3T and 7T Siemens scanners with identical investigational procedures and comparable system specific sequence optimizations. Results from 17 patients showed significantly higher functional sensitivity of the 7T system measured via percent signal change, mean t-values, number of suprathreshold voxels and contrast to noise ratio. On the other hand, 7T data suffered from a significant increase of artifacts (ghosting, head motion). We conclude that ultra high field systems provide a clinically relevant increase of functional sensitivity for patient investigations.

FMRI correlates of apraxia in Parkinson's disease patients OFF medication.

Impairment of hand dexterity in Parkinson's disease (PD) is usually attributed to bradykinesia. Recently, behavioral studies illustrated that decreased dexterity might also be due to limb-kinetic apraxia (LkA), as demonstrated by impaired performance in a coin rotation task. Here, we provide a first investigation on whether functional magnetic resonance imaging (fMRI) may reveal specific brain activation patterns for PD patients with impaired performance in a coin rotation task. We compared coin rotation as an apraxia task to simple finger tapping as a bradykinesia task in ten PD patients OFF medication and matched healthy controls. In addition to a tendency for general overactivation, PD patients showed a perirolandic dissociation with precentral overactivation and postcentral underactivation. This finding significantly separated PD patients from healthy controls.

Spectrum of malaria complications in an intensive care unit.

INTRODUCTION: Malaria remains a major health concern in tropical and subtropical countries. A large number of cases of malaria have been reported from the State of Orissa, India. Severe malaria cases are reported throughout the year, but they are more common during the high transmission season. The last decade has witnessed a changing pattern of presentations and complications across the country. Severe falciparum malaria is an important cause of multiple organ failure in Indian intensive care unit (ICU) patients. METHODS: All patients with severe falciparum malaria above the age of 14 years admitted to the ICU were included in this study. The clinical spectrum of severe falciparum malaria in a tertiary care level III ICU was analysed from December 1998 to June 2008. In all, there were 301 patients with severe malaria admitted to the ICU during that period. RESULTS: Most patients (66.9 percent) had a history of fever for less than seven days. The age distribution of the patients was 38.24 +/- 14.24 years. The Sequential Organ Failure Assessment score at admission to the ICU was 10.44 +/- 4.26. The median duration of ICU stay was three days (range 0-15 days) and 42 percent of the patients required ventilator support. Approximately 48 percent and 42 percent of patients required blood component transfusion and renal replacement therapy, respectively. The rate of single organ involvement was relatively low and multi-organ dysfunction was very common. Jaundice with acute renal failure (ARF) was the most common presentation (13.28 percent), followed by cerebral malaria with jaundice and ARF (6.37 percent), and jaundice, ARF and acute respiratory distress syndrome (ARDS) (5.31 percent). The overall mortality rate was 35.4 percent. Multivariate logistic regression analysis was conducted to estimate the association of the complications with mortality. Shock, ARF, seizure and ARDS were associated with higher mortality. CONCLUSION: Severe falciparum malaria is a common cause of multi-organ failure in the ICUs in eastern India. There has been no change in the pattern of presentations over the last ten years in the east Indian state of Orissa. Apart from early diagnosis and treatment, good supportive care is the mainstay for better outcome in these cases.

L-arginine preferentially dilates stenotic segments of coronary arteries thereby increasing coronary flow.

BACKGROUND AND OBJECTIVES: Oxidized LDL cholesterol and cytokines increase arginase and decrease nitric oxide (NO) synthase expression in human endothelial cells, leading to a decrease in NO production. In arteriosclerotic plaques, characterized by increased oxidized LDL and cytokine levels, a sustained local NO reduction might enhance sensitivity of the downstream guanylyl cyclase system towards an acute NO increase. We tested whether application of the NO synthase substrate l-arginine (l-arg, 150 micromol min(-1)) or the NO donor isosorbide dinitrate (ISDN; 0.3 mg) preferentially dilates stenotic coronary artery segments (CS) subsequently increasing poststenotic coronary blood flow (CBF) in patients with coronary artery disease (CAD). DESIGN, SETTING AND SUBJECTS: Changes in coronary diameter and circumferential surface area were assessed by quantitative coronary angiography (QCA) in a nonstenotic upstream segment, the CS, downstream the CS and in a reference vessel (n = 24). CBF was estimated in a subset of 13 patients by QCA and intracoronary Doppler. RESULTS: CS ranged from 62% to 89% (77 +/- 5%). l-arg increased minimal luminal diameter of the stenotic segment from 0.98 +/- 0.06 to 1.14 +/- 0.07 mm (P < 0.05) without affecting other coronary segments. Poststenotic CBF increased by 24 +/- 3%. ISDN dilated all segments again with a predominance of CS (25 +/- 4%) and increased poststenotic CBF by 38 +/- 9%. In a multifactorial anova, a medication with an angiotensin-converting enzyme inhibitor (decreasing inflammation and radical formation) and a ratio of LDL/HDL <3.5 were predictive for an l-arg-induced dilation. CONCLUSION: The increase in poststenotic CBF without affecting nondiseased arteries highlights the therapeutic potential of l-arg in patients with CAD.