Rapid detection of SMARCB1 sequence variation using high resolution melting

Background : Rhabdoid tumors are rare cancers of early childhood arising in the kidney, central nervous system and other organs. The majority are caused by somatic inactivating mutations or deletions affecting the tumor suppressor locus SMARCB1 [OMIM 601607]. Germ-line SMARCB1 inactivation has been reported in association with rhabdoid tumor, epitheloid sarcoma and familial schwannomatosis, underscoring the importance of accurate mutation screening to ascertain recurrence and transmission risks. We describe a rapid and sensitive diagnostic screening method, using high resolution melting (HRM), for detecting sequence variations in SMARCB1. Methods : Amplicons, encompassing the nine coding exons of SMARCB1, flanking splice site sequences and the 5' and 3' UTR, were screened by both HRM and direct DNA sequencing to establish the reliability of HRM as a primary mutation screening tool. Reaction conditions were optimized with commercially available HRM mixes. Results : The false negative rate for detecting sequence variants by HRM in our sample series was zero. Nine amplicons out of a total of 140 (6.4%) showed variant melt profiles that were subsequently shown to be false positive. Overall nine distinct pathogenic SMARCB1 mutations were identified in a total of 19 possible rhabdoid tumors. Two tumors had two distinct mutations and two harbored SMARCB1 deletion. Other mutations were nonsense or frame-shifts. The detection sensitivity of the HRM screening method was influenced by both sequence context and specific nucleotide change and varied from 1: 4 to 1:1000 (variant to wild-type DNA). A novel method involving digital HRM, followed by re-sequencing, was used to confirm mutations in tumor specimens containing associated normal tissue. Conclusions : This is the first report describing SMARCB1 mutation screening using HRM. HRM is a rapid, sensitive and inexpensive screening technology that is likely to be widely adopted in diagnostic laboratories to facilitate whole gene mutation screening.

Reduced preprandial dipping accounts for rapid elevation of blood pressure and renal sympathetic nerve activity in rabbits fed a high-fat diet

Consumption of a high-fat diet (HFD) by rabbits results in increased blood pressure (BP), heart rate (HR), and renal sympathetic nerve activity (RSNA) within 1 wk. Here, we determined how early this activation occurred and whether it was related to changes in cardiovascular and neural 24-h rhythms. Rabbits were meal-fed a HFD for 3 wks, then a normal-fat diet (NFD) for 1 wk. BP, HR, and RSNA were measured daily in the home cage via implanted telemeters. Baseline BP, HR, and RSNA over 24 h were 71 ± 1 mm Hg, 205 ± 4 beats/min and 7 ± 1 normalized units (nu). The 24-h pattern was entrained to the feeding cycle and values increased from preprandial minimum to postprandial maximum by 4 ± 1 mm Hg, 51 ± 6 beats/min, and 1.6 ± .6 nu each day. Feeding of a HFD markedly diminished the preprandial dip after 2 d (79–125% of control; p < 0.05) and this reduction lasted for 3 wks of HFD. Twenty-four-hour BP, HR, and RSNA concurrently increased by 2%, 18%, and 22%, respectively. Loss of preprandial dipping accounted for all of the BP increase and 50% of the RSNA increase over 3 wks and the 24-h rhythm became entrained to the light-dark cycle. Resumption of a NFD did not alter the BP preprandial dip. Thus, elevated BP induced by a HFD and mediated by increased sympathetic nerve activity results from a reduction in preprandial dipping, from the first day. Increased calories, glucose, insulin, and leptin may account for early changes, whereas long-term loss of dipping may be related to increased sensitivity of sympathetic pathways.

Rapid determination of protein contents in microencapsulated fish oil supplements by ATR-FTIR spectroscopy and partial least square regression (PLSR) analysis

Following the recent success in quantitative analysis of essential fatty acid compositions in a commercial microencapsulated fish oil (?EFO) supplement, we extended the application of portable attenuated total reflection Fourier transform infrared (ATR-FTIR) spectroscopic technique and partial least square regression (PLSR) analysis for rapid determination of total protein contents-the other major component in most commercial ?EFO powders. In contrast to the traditional chromatographic methodology used in a routine amino acid analysis (AAA), the ATR-FTIR spectra of the ?EFO powder can be acquired directly from its original powder form with no requirement of any sample preparation, making the technique exceptionally fast, noninvasive, and environmentally friendly as well as being cost effective and hence eminently suitable for routine use by industry. By optimizing the spectral region of interest and number of latent factors through the developed PLSR strategy, a good linear calibration model was produced as indicated by an excellent value of coefficient of determination R2 = 0.9975, using standard ?EFO powders with total protein contents in the range of 140-450 mg/g. The prediction of the protein contents acquired from an independent validation set through the optimized PLSR model was highly accurate as evidenced through (1) a good linear fitting (R2 = 0.9759) in the plot of predicted versus reference values, which were obtained from a standard AAA method, (2) lowest root mean square error of prediction (11.64 mg/g), and (3) high residual predictive deviation (6.83) ranked in very good level of predictive quality indicating high robustness and good predictive performance of the achieved PLSR calibration model. The study therefore demonstrated the potential application of the portable ATR-FTIR technique when used together with PLSR analysis for rapid online monitoring of the two major components (i.e., oil and protein contents) in finished ?EFO powders in the actual manufacturing setting.

Corner Inlet and Nooramunga Habitat Mapping Project – Part 2 : High resolution mapping and change detection analysis using 2011 Rapid Eye Satellite Image compared to 2009 ALOS Satellite Image

Building on a habitat mapping project completed in 2011, Deakin University was commissioned by Parks Victoria (PV) to apply the same methodology and ground-truth data to a second, more recent and higher resolution satellite image to create habitat maps for areas within the Corner Inlet and Nooramunga Marine and Coastal Park and Ramsar area. A ground-truth data set using in situ video and still photographs was used to develop and assess predictive models of benthic marine habitat distributions incorporating data from both RapidEye satellite imagery (corrected for atmospheric and water column effects by CSIRO) and LiDAR (Light Detection and Ranging) bathymetry. This report describes the results of the mapping effort as well as the methodology used to produce these habitat maps.

Overall accuracies of habitat classifications were good, with error rates similar to or better than the earlier classification (>73 % and kappa values > 0.58 for both study localities). The RapidEye classification failed to accurately detect Pyura and reef habitat classes at the Corner Inlet locality, possibly due to differences in spectral frequencies. For comparison, these categories were combined into a ‘non-seagrass’ category, similar to the one used at the Nooramunga locality in the original classification. Habitats predicted with highest accuracies differed from the earlier classification and were Posidonia in Corner Inlet (89%), and bare sediment (no-visible seagrass class) in Nooramunga (90%). In the Corner Inlet locality reef and Pyura habitat categories were not distinguishable in the repeated classification and so were combined with bare sediments. The majority of remaining classification errors were due to the misclassification of Zosteraceae as bare sediment and vice versa. Dominant habitats were the same as those from the 2011 classification with some differences in extent. For the Corner Inlet study locality the no-visible seagrass category remained the most extensive (9059 ha), followed by Posidonia (5,513 ha) and Zosteraceae (5,504 ha). In Nooramunga no-visible seagrass (6,294 ha), Zosteraceae (3,122 ha) and wet saltmarsh (1,562 ha) habitat classes were most dominant.

Change detection analyses between the 2009 and 2011 imagery were undertaken as part of this project, following the analyses presented in Monk et al. (2011) and incorporating error estimates from both classifications. These analyses indicated some shifts in classification between Posidonia and Zosteraceae as well as a general reduction in the area of Zosteraceae. Issues with classification of mixed beds were apparent, particularly in the main Posidonia bed at Nooramunga where a mosaic of Zosteraceae and Posidonia was seen that was not evident in the ALOS classification. Results of a reanalysis of the 1998-2009 change detection illustrating effects of binning of mixed beds is also provided as an appendix.

This work has been successful in providing baseline maps at an improved level of detail using a repeatable method meaning that any future changes in intertidal and shallow water marine habitats may be assessed in a consistent way with quantitative error assessments. In wider use, these maps should also allow improved conservation planning, advance fisheries and catchment management, and progress infrastructure planning to limit impacts on the Inlet environment.

Emergency department rapid response systems: the case for a standardized approach to deteriorating patients

Objectives: 

The effect of a rapid heating rate, mechanical vibration and surfactant chemistry on the structure-property relationships of epoxy/clay nanocomposites

The role of processing conditions and intercalant chemistry in montmorillonite clays on the dispersion, morphology and mechanical properties of two epoxy/clay nanocomposite systems was investigated in this paper. This work highlights the importance of employing complementary techniques (X-ray diffraction, small angle X-ray scattering, optical microscopy and transmission electron microscopy) to correlate nanomorphology to macroscale properties. Materials were prepared using an out of autoclave manufacturing process equipped to generate rapid heating rates and mechanical vibration. The results suggested that the quaternary ammonium surfactant on C30B clay reacted with the epoxy during cure, while the primary ammonium surfactant (I.30E) catalysed the polymerisation reaction. These effects led to important differences in nanocomposite clay morphologies. The use of mechanical vibration at 4 Hz prior to matrix gelation was found to facilitate clay dispersion and to reduce the area fraction of I.30E clay agglomerates in addition to increasing flexural strength by over 40%.

Developing a simple, rapid method for identifying and monitoring jellyfish aggregations from the air

Within the marine environment, aerial surveys have historically centred on apex predators, such as pinnipeds, cetaceans and sea birds. However, it is becoming increasingly apparent that the utility of this technique may also extend to subsurface species such as pre-spawning fish stocks and aggregations of jellyfish that occur close to the surface. In light of this, we tested the utility of aerial surveys to provide baseline data for 3 poorly understood scyphozoan jellyfish found throughout British and Irish waters: Rhizostoma octopus, Cyanea capillata and Chrysaora hysoscella. Our principal objectives were to develop a simple sampling protocol to identify and quantify surface aggregations, assess their consistency in space and time, and consider the overall applicability of this technique to the study of gelatinous zooplankton. This approach provided a general understanding of range and relative abundance for each target species, with greatest suitability to the study of R. octopus. For this species it was possible to identify and monitor extensive, temporally consistent and previously undocumented aggregations throughout the Irish Sea, an area spanning thousands of square kilometres. This finding has pronounced implications for ecologists and fisheries managers alike and, moreover, draws attention to the broad utility of aerial surveys for the study of gelatinous aggregations beyond the range of conventional ship-based techniques.

The timing of Rapid-Response Team activations: a multicentre international study

Background:
Most studies of Rapid-Response Teams (RRTs) assess their effect on outcomes of all hospitalised patients. Little information exists on RRT activation patterns or why RRT calls are needed. Triage error may necessitate RRT review of ward patients shortly after hospital admission. RRT diurnal activation rates may reflect the likely frequency of caregiver visits.

Objectives:
To study the timing of RRT calls in relation to time of day and day of week, and their frequency and outcomes in relation to days after hospital admission.

Methods:
We prospectively studied RRT calls over 1 month in seven hospitals during 2009, collecting data on patient age, sex, admitting unit, admission source, limitations of medical therapy (LOMTs), and admission and discharge dates. We assessed the timing of RRT calls in relation to hospital admission and circadian variation; and differences in characteristics and outcomes of calls occurring early (Days 0 and 1) versus late (after Day 7) after hospital admission.

Results:
There were 652 RRT calls for 518 patients. Calls were more likely on Mondays (P=0.018) and during work hours (P<0.0001) but less likely on weekends (P=0.003) or overnight (P<0.001). There were 177 early calls (27.1%) and 198 late calls (30.4%). Early calls involved younger patients (median ages, 67.5 years [early calls] v 73 years [late calls]; P= 0.01), fewer LOMTs (P=0.029), and lower in hospital mortality (12.8% [early calls] v 32.3% [late calls]; P<0.0001). The mortality difference remained in patients without LOMTs (5.6% [early calls] v 19.6% [late calls]; P=0.003).

Conclusions:
About one-quarter of RRT calls occurred shortly after hospital admission, and were more common when caregivers were around. Early calls may partially reflect suboptimal triage, though the associated mortality appeared low. Late calls may reflect suboptimal end-of-life care planning, and the associated mortality was high. There is a need to further assess the epidemiology of RRT calls at different phases of the hospital stay.