Time of injury in the light of acute and hazardous usual alcohol consumption - A study among emergency department patients.

Purpose: To investigate how prior-to-injury and usual alcohol consumption relate to time of injury. Patients and methods: The associations between injury time of day and day of week and prior-to-injury (labeled as "acute") alcohol intake and hazardous usual alcohol consumption (considered from the point of view of both heavy episodic drinking [HED] and risky volumes of consumption) are assessed using interview data from a randomized sample of 486 injured patients treated in a Swiss emergency department (ED; Lausanne University Hospital). Results: Acute consumption was associated with both injury time of day and day of week, HED with day of week only, and risky volume with none. Conclusions: Acute consumption and HED, but not risky volume of consumption, show specific time distributions for injuries. These findings highlight the potential importance of considering the time dimension of an injury when providing emergency care and have additional implications for interventions aimed at influencing the alcohol consumption of injured patients presenting to the ED.

Non-invasive diagnostic biomarkers for estimating the onset time of permanent cerebral ischemia.

The treatments for ischemic stroke can only be administered in a narrow time-window. However, the ischemia onset time is unknown in ~30% of stroke patients (wake-up strokes). The objective of this study was to determine whether MR spectra of ischemic brains might allow the precise estimation of cerebral ischemia onset time. We modeled ischemic stroke in male ICR-CD1 mice using a permanent middle cerebral artery filament occlusion model with laser Doppler control of the regional cerebral blood flow. Mice were then subjected to repeated MRS measurements of ipsilateral striatum at 14.1 T. A striking initial increase in γ-aminobutyric acid (GABA) and no increase in glutamine were observed. A steady decline was observed for taurine (Tau), N-acetyl-aspartate (NAA) and similarly for the sum of NAA+Tau+glutamate that mimicked an exponential function. The estimation of the time of onset of permanent ischemia within 6 hours in a blinded experiment with mice showed an accuracy of 33±10 minutes. A plot of GABA, Tau, and neuronal marker concentrations against the ratio of acetate/NAA allowed precise separation of mice whose ischemia onset lay within arbitrarily chosen time-windows. We conclude that (1)H-MRS has the potential to detect the clinically relevant time of onset of ischemic stroke.

Matrix-assisted laser desorption ionization time-of-flight mass spectrometry, a revolution in clinical microbial identification.

Until recently, matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) techniques for the identification of microorganisms remained confined to research laboratories. In the last 2 years, the availability of relatively simple to use MALDI-TOF MS devices, which can be utilized in clinical microbiology laboratories, has changed the laboratory workflows for the identification of pathogens. Recently, the first prospective studies regarding the performance in routine bacterial identification showed that MALDI-TOF MS is a fast, reliable and cost-effective technique that has the potential to replace and/or complement conventional phenotypic identification for most bacterial strains isolated in clinical microbiology laboratories. For routine bacterial isolates, correct identification by MALDI-TOF MS at the species level was obtained in 84.1-93.6% of instances. In one of these studies, a protein extraction step clearly improved the overall valid identification yield, from 70.3% to 93.2%. This review focuses on the current state of use of MALDI-TOF MS for the identification of routine bacterial isolates and on the main difficulties that may lead to erroneous or doubtful identifications.

Quantification of glucuronidated and sulfated steroids in human urine by ultra-high pressure liquid chromatography quadrupole time-of-flight mass spectrometry.

The urinary steroid profile is constituted by anabolic androgenic steroids, including testosterone and its relatives, that are extensively metabolized into phase II sulfated or glucuronidated steroids. The use of liquid chromatography coupled to mass spectrometry (LC-MS) is an issue for the direct analysis of conjugated steroids, which can be used as urinary markers of exogenous steroid administration in doping analysis, without hydrolysis of the conjugated moiety. In this study, a sensitive and selective ultra high-pressure liquid chromatography coupled to quadrupole time-of-flight mass spectrometer (UHPLC-QTOF-MS) method was developed to quantify major urinary metabolites simultaneously after testosterone intake. The sample preparation of the urine (1 mL) was performed by solid-phase extraction on Oasis HLB sorbent using a 96-well plate format. The conjugated steroids were analyzed by UHPLC-QTOF-MS(E) with a single-gradient elution of 36 min (including re-equilibration time) in the negative electrospray ionization mode. MS(E) analysis involved parallel alternating acquisitions of both low- and high-collision energy functions. The method was validated and applied to samples collected from a clinical study performed with a group of healthy human volunteers who had taken testosterone, which were compared with samples from a placebo group. Quantitative results were also compared to GC-MS and LC-MS/MS measurements, and the correlations between data were found appropriate. The acquisition of full mass spectra over the entire mass range with QTOF mass analyzers gives promise of the opportunity to extend the steroid profile to a higher number of conjugated steroids.

Efficient T cell activation requires an optimal dwell-time of interaction between the TCR and the pMHC complex.

Cytotoxic T cell (CTL) activation by antigen requires the specific detection of peptide-major histocompatibility class I (pMHC) molecules on the target-cell surface by the T cell receptor (TCR). We examined the effect of mutations in the antigen-binding site of a Kb-restricted TCR on T cell activation, antigen binding and dissociation from antigen.These parameters were also examined for variants derived from a Kd-restricted peptide that was recognized by a CTL clone. Using these two independent systems, we show that T cell activation can be impaired by mutations that either decrease or increase the binding half-life of the TCR-pMHC interaction. Our data indicate that efficient T cell activation occurs within an optimal dwell-time range of TCR-pMHC interaction. This restricted dwell-time range is consistent with the exclusion of either extremely low or high affinity T cells from the expanded population during immune responses.

Impact of Matrix-Assisted Laser Desorption Ionization Time-of-Flight Mass Spectrometry on the Clinical Management of Patients With Gram-negative Bacteremia: A Prospective Observational Study.

Background. Early identification of pathogens from blood cultures using matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) mass spectrometry may optimize the choice of empirical antibiotic therapy in the setting of bloodstream infections. We aimed to assess the impact of this new technology on the use of antibiotic treatment in patients with gram-negative bacteremia. Methods. We conducted a prospective observational study from January to December 2010 to evaluate the sequential and separate impacts of Gram stain reporting and MALDI-TOF bacterial identification performed on blood culture pellets in patients with gram-negative bacteremia. The primary outcome was the impact of MALDI-TOF on empirical antibiotic choice. Results. Among 202 episodes of gram-negative bacteremia, Gram stain reporting had an impact in 42 cases (20.8%). MALDI-TOF identification led to a modification of empirical therapy in 71 of all 202 cases (35.1%), and in 16 of 27 cases (59.3%) of monomicrobial bacteremia caused by AmpC-producing Enterobacteriaceae. The most frequently observed impact was an early appropriate broadening of the antibiotic spectrum in 31 of 71 cases (43.7%). In total, 143 of 165 episodes (86.7%) of monomicrobial bacteremia were correctly identified at genus level by MALDI-TOF. Conclusions. In a low prevalence area for extended spectrum betalactamases (ESBL) and multiresistant gram-negative bacteria, MALDI-TOF performed on blood culture pellets had an impact on the clinical management of 35.1% of all gram-negative bacteremia cases, demonstrating a greater impact than Gram stain reporting. Thus, MALDI-TOF could become a vital second step beside Gram stain in guiding the empirical treatment of patients with bloodstream infection.

Is it the grain size or the characteristic pore size that controls the induced polarization relaxation time of clean sands and sandstones?

There is a wide range of evidence to suggest that permeability can be constrained through of induced polarization measurements. For clean sands and sandstones, current mechanistic models of induced polarization predict a relationship between the low-frequency time constant inferred from induced polarization measurements and the grain diameter. A number of observations do, however, disagree with this and indicate that the observed relaxation behavior is rather governed by the so-called dynamic pore radius L. To test this hypothesis, we have developed a set of new scaling relationships, which allow the relaxation time to be computed from the pore size and the permeability to be computed from both the Cole-Cole time constant and the formation factor. Moreover, these new scaling relationships can be also used to predict the dependence of the Cole-Cole time constant as a function of the water saturation under unsaturated conditions. Comparative tests of the proposed new relationships with regard to various published experimental results for saturated clean sands and sandstones as well as for partially saturated clean sandstones, do indeed confirm that the dynamic pore radius L is a much more reliable indicator of the observed relaxation behavior than grain-size-based models.

Fast analysis of doping agents in urine by ultra-high-pressure liquid chromatography-quadrupole time-of-flight mass spectrometry I. Screening analysis.

The general strategy to perform anti-doping analyses of urine samples starts with the screening for a wide range of compounds. This step should be fast, generic and able to detect any sample that may contain a prohibited substance while avoiding false negatives and reducing false positive results. The experiments presented in this work were based on ultra-high-pressure liquid chromatography coupled to hybrid quadrupole time-of-flight mass spectrometry. Thanks to the high sensitivity of the method, urine samples could be diluted 2-fold prior to injection. One hundred and three forbidden substances from various classes (such as stimulants, diuretics, narcotics, anti-estrogens) were analysed on a C(18) reversed-phase column in two gradients of 9min (including two 3min equilibration periods) for positive and negative electrospray ionisation and detected in the MS full scan mode. The automatic identification of analytes was based on retention time and mass accuracy, with an automated tool for peak picking. The method was validated according to the International Standard for Laboratories described in the World Anti-Doping Code and was selective enough to comply with the World Anti-Doping Agency recommendations. In addition, the matrix effect on MS response was measured on all investigated analytes spiked in urine samples. The limits of detection ranged from 1 to 500ng/mL, allowing the identification of all tested compounds in urine. When a sample was reported positive during the screening, a fast additional pre-confirmatory step was performed to reduce the number of confirmatory analyses.

Matrix-assisted laser desorption ionization-time of flight mass spectrometry as an alternative to 16S rRNA gene sequencing for identification of difficult-to-identify bacterial strains.

Conventional methods are sometimes insufficient to identify human bacterial pathogens, and alternative techniques, often molecular, are required. Matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) identified with a valid score 45.9% of 410 clinical isolates from 207 different difficult-to-identify species having required 16S rRNA gene sequencing. MALDI-TOF MS might represent an alternative to 16S rRNA gene sequencing.

Trends over time of virological and immunological characteristics in the Swiss HIV Cohort Study.

BACKGROUND: The long-term outcome of antiretroviral therapy (ART) is not assessed in controlled trials. We aimed to analyse trends in the population effectiveness of ART in the Swiss HIV Cohort Study over the last decade. METHODS: We analysed the odds of stably suppressed viral load (ssVL: three consecutive values <50 HIV-1 RNA copies/mL) and of CD4 cell count exceeding 500 cells/μL for each year between 2000 and 2008 in three scenarios: an open cohort; a closed cohort ignoring the influx of new participants after 2000; and a worst-case closed cohort retaining lost or dead patients as virological failures in subsequent years. We used generalized estimating equations with sex, age, risk, non-White ethnicity and era of starting combination ART (cART) as fixed co-factors. Time-updated co-factors included type of ART regimen, number of new drugs and adherence to therapy. RESULTS: The open cohort included 9802 individuals (median age 38 years; 31% female). From 2000 to 2008, the proportion of participants with ssVL increased from 37 to 64% [adjusted odds ratio (OR) per year 1.16 (95% CI 1.15-1.17)] and the proportion with CD4 count >500 cells/μL increased from 40 to >50% [OR 1.07 (95% CI 1.06-1.07)]. Similar trends were seen in the two closed cohorts. Adjustment did not substantially affect time trends. CONCLUSIONS: There was no relevant dilution effect through new participants entering the open clinical cohort, and the increase in virological/immunological success over time was not an artefact of the study design of open cohorts. This can partly be explained by new treatment options and other improvements in medical care.

Hind limb ischemia in rabbit model: T2-prepared versus time-of-flight MR angiography at 3 T.

PURPOSE: To prospectively compare various parameters of vessels imaged at 3 T by using time-of-flight (TOF) and T2-prepared magnetic resonance (MR) angiography in a rabbit model of hind limb ischemia. MATERIALS AND METHODS: Experiments were approved by the institutional animal care and use committee. Endovascular occlusion of the left superficial femoral artery was induced in 14 New Zealand white rabbits. After 2 weeks, MR angiography and conventional (x-ray) angiography were performed. Vessel sharpness was evaluated visually in the ischemic and nonischemic limbs, and the presence of small collateral vessels was evaluated in the ischemic limbs. Vessel sharpness was also quantified by evaluating the magnitude of signal intensity change at the vessel borders. RESULTS: The sharpness of vessels in the nonischemic limbs was similar between the TOF and the T2-prepared images. In the ischemic limbs, however, T2-prepared imaging, as compared with TOF imaging, generated higher vessel sharpness in arteries with diminished blood flow (mean vessel sharpness: 44% vs 30% for popliteal arteries, 45% vs 28% for saphenous arteries; P < .001 for both comparisons) and enabled better detection of small collateral vessels (93% vs 36% of vessels, P < .001). CONCLUSION: T2-prepared imaging can facilitate high-spatial-resolution MR angiography of small vessels with low blood flow and thus has potential as a tool for noninvasive evaluation of arteriogenic therapies, without use of contrast material. Supplemental material: http://radiology.rsnajnls.org/cgi/content/full/2452062067/DC1.

Detection of metabolite induction in fungal co-cultures on solid media by high-throughput differential ultra-high pressure liquid chromatography-time-of-flight mass spectrometry fingerprinting.

Access to new biological sources is a key element of natural product research. A particularly large number of biologically active molecules have been found to originate from microorganisms. Very recently, the use of fungal co-culture to activate the silent genes involved in metabolite biosynthesis was found to be a successful method for the induction of new compounds. However, the detection and identification of the induced metabolites in the confrontation zone where fungi interact remain very challenging. To tackle this issue, a high-throughput UHPLC-TOF-MS-based metabolomic approach has been developed for the screening of fungal co-cultures in solid media at the petri dish level. The metabolites that were overexpressed because of fungal interactions were highlighted by comparing the LC-MS data obtained from the co-cultures and their corresponding mono-cultures. This comparison was achieved by subjecting automatically generated peak lists to statistical treatments. This strategy has been applied to more than 600 co-culture experiments that mainly involved fungal strains from the Fusarium genera, although experiments were also completed with a selection of several other filamentous fungi. This strategy was found to provide satisfactory repeatability and was used to detect the biomarkers of fungal induction in a large panel of filamentous fungi. This study demonstrates that co-culture results in consistent induction of potentially new metabolites.

Profiling of steroid metabolites after transdermal and oral administration of testosterone by ultra-high pressure liquid chromatography coupled to quadrupole time-of-flight mass spectrometry.

The screening of testosterone (T) misuse for doping control is based on the urinary steroid profile, including T, its precursors and metabolites. Modifications of individual levels and ratio between those metabolites are indicators of T misuse. In the context of screening analysis, the most discriminant criterion known to date is based on the T glucuronide (TG) to epitestosterone glucuronide (EG) ratio (TG/EG). Following the World Anti-Doping Agency (WADA) recommendations, there is suspicion of T misuse when the ratio reaches 4 or beyond. While this marker remains very sensitive and specific, it suffers from large inter-individual variability, with important influence of enzyme polymorphisms. Moreover, use of low dose or topical administration forms makes the screening of endogenous steroids difficult while the detection window no longer suits the doping habit. As reference limits are estimated on the basis of population studies, which encompass inter-individual and inter-ethnic variability, new strategies including individual threshold monitoring and alternative biomarkers were proposed to detect T misuse. The purpose of this study was to evaluate the potential of ultra-high pressure liquid chromatography (UHPLC) coupled with a new generation high resolution quadrupole time-of-flight mass spectrometer (QTOF-MS) to investigate the steroid metabolism after transdermal and oral T administration. An approach was developed to quantify 12 targeted urinary steroids as direct glucuro- and sulfo-conjugated metabolites, allowing the conservation of the phase II metabolism information, reflecting genetic and environmental influences. The UHPLC-QTOF-MS(E) platform was applied to clinical study samples from 19 healthy male volunteers, having different genotypes for the UGT2B17 enzyme responsible for the glucuroconjugation of T. Based on reference population ranges, none of the traditional markers of T misuse could detect doping after topical administration of T, while the detection window was short after oral TU ingestion. The detection ability of the 12 targeted steroids was thus evaluated by using individual thresholds following both transdermal and oral administration. Other relevant biomarkers and minor metabolites were studied for complementary information to the steroid profile, including sulfoconjugated analytes and hydroxy forms of glucuroconjugated metabolites. While sulfoconjugated steroids may provide helpful screening information for individuals with homozygotous UGT2B17 deletion, hydroxy-glucuroconjugated analytes could enhance the detection window of oral T undecanoate (TU) doping.