Burkholderia pseudomallei:another emerging pathogen in cystic fibrosis

BACKGROUND: Burkholderia pseudomallei is an important cause of acute fulminant pneumonia and septicaemia in tropical regions of northern Australia and south east Asia. Subacute and chronic forms of the disease also occur. There have been three recent reports of adults with cystic fibrosis (CF) who presumably acquired B pseudomallei infection during extended vacations or residence in either Thailand or northern Australia.

METHODS: The clinical course, molecular characteristics, serology and response to treatment are described in four adult CF patients infected with B pseudomallei. Polymerase chain reaction (PCR) based methods were used to confirm B pseudomallei and exclude B cepacia complex. Genotyping was performed using randomly amplified polymorphic DNA (RAPD) PCR and pulsed field gel electrophoresis (PFGE).

RESULTS: Four patients are described with a mean duration of infection of 32 months. All but one patient lived in tropical Queensland. Two patients (with the longest duration of infection) deteriorated clinically and one subsequently died of respiratory failure. Both responded to intravenous treatment specifically targeting B pseudomallei. Another patient suffered two severe episodes of acute bronchopneumonia following acquisition of B pseudomallei. Eradication of the organism was not possible in any of the cases. PFGE of a sample isolate from each patient revealed the strains to be unique and RAPD analysis showed retention of the same strain within an individual over time.

CONCLUSIONS: These findings support a potential pathogenic role for B pseudomallei in CF lung disease, producing both chronic infection and possibly acute bronchopneumonia. Identical isolates are retained over time and are unique, consistent with likely environmental acquisition and not person to person spread. B pseudomallei is emerging as a significant pathogen for patients with CF residing and holidaying in the tropics.

Efficacy and safety of peri-prostatic local anaesthetic injection in tram-rectal biopsy of the prostrate:A prospective randomised study

Objective: To assess the efficacy and safety of periprostatic lignocaine injection in trans-rectal ultrasound (TRUS) -guided biopsy of the prostate gland.

Methods: Ninety- six men (mean age 65 years, range 47-74) undergoing TRUS biopsy were randomised into the local anaesthetic (LA) or placebo group. Six to twelve biopsy cores were taken, the majority being 10 cores. Patients were asked to fill in the expected pain score on a visual analogue scale (VAS) prior to the procedure. They also completed the actual pain experienced on VAS after the biopsy. The incidence of complications was documented.

Results: The age, mean prostate specific antigen (PSA) were comparable in both groups. The expected pain score was also comparable (5.2 +/- 1.6 in LA, 5.0 +/- 1.4 in Placebo). In the LA group, the mean actual pain score was 3.0 +/- 1.8 and in the placebo group it was 6.5 +/- 2.2 (P = 0.0001). When patients were asked whether they would undergo the procedure again in the same way, 100% of the LA group and only 64% of the placebo group responded 'yes'(P=0.002 using Fisher's test). The complication rates were not significantly different between the two groups.

Conclusion: Peri-prostatic injection of local anaesthetic is safe and reduces discomfort significantly, and should be routinely offered to patients.

Toward Characterization Of The Type IIP Supernova Progenitor Population: A Statistical Sample Of Light Curves From Pan-STARRS1

In recent years, wide-field sky surveys providing deep multi-band imaging have presented a new path for indirectly characterizing the progenitor populations of core-collapse supernovae (SN): systematic light curve studies. We assemble a set of 76 grizy-band Type IIP SN light curves from Pan-STARRS1, obtained over a constant survey program of 4 years and classified using both spectroscopy and machine learning-based photometric techniques. We develop and apply a new Bayesian model for the full multi-band evolution of each light curve in the sample. We find no evidence of a sub-population of fast-declining explosions (historically referred to as "Type IIL" SNe). However, we identify a highly significant relation between the plateau phase decay rate and peak luminosity among our SNe IIP. These results argue in favor of a single parameter, likely determined by initial stellar mass, predominantly controlling the explosions of red supergiants. This relation could also be applied for supernova cosmology, offering a standardizable candle good to an intrinsic scatter of 0.2 mag. We compare each light curve to physical models from hydrodynamic simulations to estimate progenitor initial masses and other properties of the Pan-STARRS1 Type IIP SN sample. We show that correction of systematic discrepancies between modeled and observed SN IIP light curve properties and an expanded grid of progenitor properties, are needed to enable robust progenitor inferences from multi-band light curve samples of this kind. This work will serve as a pathfinder for photometric studies of core-collapse SNe to be conducted through future wide field transient searches.

Rapid quantification of microRNAs in plasma using a fast real time PCR system

The ability to rapidly detect circulating small RNAs, in particular microRNAs (miRNAs), would further increase their already established potential as biomarkers in a range of conditions. One rate-limiting factor is the time taken to perform quantitative real time PCR amplification. We therefore evaluated the ability of a novel thermal cycler to perform this step in less than 10 minutes. Quantitative PCR was performed on an xxpress® thermal cycler (BJS Biotechnologies, Perivale, UK), which employs a resistive heating system and forced air cooling to achieve thermal ramp rates of 10 °C/s, and a conventional peltier-controlled LightCycler 480 system (Roche, Basel, Switzerland) ramping at 4.8 °C/s. The threshold cycle (Ct) for detection of 18S rDNA from a standard genomic DNA sample was significantly more variable across the block (F-test, p=2.4x10-25) for the xxpress (20.01±0.47SD) than the LightCycler (19.87±0.04SD). RNA was extracted from human plasma, reverse transcribed and a panel of miRNAs amplified and detected using SYBR green (Kapa Biosystems, Wilmington, Ma, USA). The sensitivity of both systems was broadly comparable and both detected a panel of miRNAs reliably and indicated similar relative abundances. The xxpress thermal cycler facilitates rapid qPCR detection of small RNAs and brings point-of care diagnostics based upon circulating miRNAs a step closer to reality.

MarcoPolo-R near earth asteroid sample return mission

MarcoPolo-R is a sample return mission to a primitive Near-Earth Asteroid (NEA) proposed in collaboration with NASA. It will rendezvous with a primitive NEA, scientifically characterize it at multiple scales,and return a unique sample to Earth unaltered by the atmospheric entry process or terrestrial weathering. MarcoPolo-R will return bulk samples (up to 2 kg) from an organic-rich binary asteroid to Earth for laboratory analyses, allowing us to: explore the origin of planetary materials and initial stages of habitable planet formation; identify and characterize the organics and volatiles in a primitive asteroid; understand the unique geomorphology, dynamics and evolution of a binaryNEA. This project is based on the previous Marco Polo mission study,which was selected for the Assessment Phase of the first round of Cosmic Vision. Its scientific rationale was highly ranked by ESA committees andit was not selected only because the estimated cost was higher than theallotted amount for an M class mission. The cost of Marco Polo-R will be reduced to within the ESA medium mission budget by collaboration withAPL (John Hopkins University) and JPL in the NASA program for coordination with ESA's Cosmic Vision Call. The baseline target is a binary asteroid (175706) 1996 FG3, which offers a very efficient operational and technical mission profile. A binary target also providesenhanced science return. The choice of this target will allow newinvestigations to be performed more easily than at a single object, andalso enables investigations of the fascinating geology and geophysics ofasteroids that are impossible at a single object. Several launch windows have been identified in the time-span 2020-2024. A number of otherpossible primitive single targets of high scientific interest have beenidentified covering a wide range of possible launch dates. The baselinemission scenario of Marco Polo-R to 1996 FG3 is as follows: a singleprimary spacecraft provided by ESA, carrying the Earth Re-entry Capsule, sample acquisition and transfer system provided by NASA, will be launched by a Soyuz-Fregat rocket from Kourou into GTO and using two space segment stages. Two similar missions with two launch windows, in 2021 and 2022 and for both sample return in 2029 (with mission durationof 7 and 8 years), have been defined. Earlier or later launches, in 2020 or 2024, also offer good opportunities. All manoeuvres are carried out by a chemical propulsion system. MarcoPolo-R takes advantage of three industrial studies completed as part of the previous Marco Polo mission (see ESA/SRE (2009)3, Marco Polo Yellow Book) and of the expertise of the consortium led by Dr. A.F. Cheng (PI of the NASA NEAR Shoemaker mission) of the JHU-APL, including JPL, NASA ARC, NASA LaRC, and MIT.

Measurement method and apparatus for analysing the metal or metal ion conten of a sample

2. K. Gow, P.K.J Robertson, P.M. Pollard, and K. Christidis

Measuring and modelling air mass flow rate in the injection stretch blow moulding process

The injection stretch blow moulding process involves the inflation and stretching of a hot preform into a mould to form bottles. A critical process variable and an essential input for process simulations is the rate of pressure increase within the preform during forming, which is regulated by an air flow restrictor valve. The paper describes a set of experiments for measuring the air flow rate within an industrial ISBM machine and the subsequent modelling of it with the FEA package ABAQUS. Two rigid containers were inserted into a Sidel SBO1 blow moulding machine and subjected to different supply pressures and air flow restrictor settings. The pressure and air temperature were recorded for each experiment enabling the mass flow rate of air to be determined along with an important machine characteristic known as the ‘dead volume’. The experimental setup was simulated within the commercial FEA package ABAQUS/Explicit using a combination of structural, fluid and fluid link elements that idealize the air flowing through an orifice behaving as an ideal gas under isothermal conditions. Results between experiment and simulation are compared and show a good correlation.

Design characteristic of randomised controlled trials for geographic atrophy in age-related macular degeneration: selection of outcomes and sample size calculation

PurposeThe selection of suitable outcomes and sample size calculation are critical factors in the design of a randomised controlled trial (RCT). The goal of this study was to identify the range of outcomes and information on sample size calculation in RCTs on geographic atrophy (GA).MethodsWe carried out a systematic review of age-related macular degeneration (AMD) RCTs. We searched MEDLINE, EMBASE, Scopus, Cochrane Library, www.controlled-trials.com, and www.ClinicalTrials.gov. Two independent reviewers screened records. One reviewer collected data and the second reviewer appraised 10% of collected data. We scanned references lists of selected papers to include other relevant RCTs.ResultsLiterature and registry search identified 3816 abstracts of journal articles and 493 records from trial registries. From a total of 177 RCTs on all types of AMD, 23 RCTs on GA were included. Eighty-one clinical outcomes were identified. Visual acuity (VA) was the most frequently used outcome, presented in 18 out of 23 RCTs and followed by the measures of lesion area. For sample size analysis, 8 GA RCTs were included. None of them provided sufficient Information on sample size calculations.ConclusionsThis systematic review illustrates a lack of standardisation in terms of outcome reporting in GA trials and issues regarding sample size calculation. These limitations significantly hamper attempts to compare outcomes across studies and also perform meta-analyses.

Systematic Uncertainties Associated with the Cosmological Analysis of the First PAN-STARRS1 Type Ia Supernova Sample

We probe the systematic uncertainties from the 113 Type Ia supernovae (SN Ia) in the Pan-STARRS1 (PS1) sample along with 197 SN Ia from a combination of low-redshift surveys. The companion paper by Rest et al. describes the photometric measurements and cosmological inferences from the PS1 sample. The largest systematic uncertainty stems from the photometric calibration of the PS1 and low-z samples. We increase the sample of observed Calspec standards from 7 to 10 used to define the PS1 calibration system. The PS1 and SDSS-II calibration systems are compared and discrepancies up to ∼0.02 mag are recovered. We find uncertainties in the proper way to treat intrinsic colors and reddening produce differences in the recovered value of w up to 3%. We estimate masses of host galaxies of PS1 supernovae and detect an insignificant difference in distance residuals of the full sample of 0.037 ± 0.031 mag for host galaxies with high and low masses. Assuming flatness and including systematic uncertainties in our analysis of only SNe measurements, we find w = -1.120^+0.360_-0.206(Stat)^+0.269_-0.291(Sys). With additional constraints from Baryon acoustic oscillation, cosmic microwave background (CMB) (Planck) and H₀ measurements, we find w = -1.166^+0.072_-0.069 and Ω_m = 0.280^+0.013_-0.012 (statistical and systematic errors added in quadrature). The significance of the inconsistency with w = -1 depends on whether we use Planck or Wilkinson Microwave Anisotropy Probe measurements of the CMB: w_{BAO+H0+SN+WMAP} = -1.124^+0.083_-0.065.

Prevalence of the prothrombin G20210A mutation in the Irish populations:use of a novel polymerase chain reaction approach

The prothrombin G20210A polymorphism is associated with a threefold-increased risk of venous thrombosis. There is considerable variation in the reported prevalence of this polymorphism within normal populations, ranging from 0 to 6.5%. The prevalence within the Irish population has not been determined. A restriction fragment length polymorphism (RFLP)-based assay is commonly used for the detection of the prothrombin 20210A allele. This assay does not include a control restriction digest fragment and, consequently, failure of the enzyme activity or lack of addition of enzyme to the sample cannot be distinguished from wild-type prothrombin. We developed a RFLP-based assay, which incorporates an invariant digest site, resulting in the generation of a control digest fragment. Furthermore, we developed a nested polymerase chain reaction (PCR) method for the amplification and digestion of poor-quality or low-concentration DNA. In the Irish population studied, five of 385 (1.29%) were heterozygous and one patient was homozygous for the prothrombin 20210A polymorphism. This is the first reported data on an Irish or Celtic population and suggests that the allele frequency is similar to Anglo-Saxon populations. The nested PCR method successfully amplified and digested 100/100 (100%) of the archived samples; none of these samples could be analyzed by the standard single-round PCR method. In conclusion, nested PCR should be considered in the analysis of archived samples. Single-round PCR is appropriate for recently collected samples; however, an invariant control digest site should be incorporated in RFLP-based assays to validate the integrity of the digestion enzyme and limit the risk of false-negative results.

Sorting out mix-ups. The provenance of tissue sections may be confirmed by PCR using microsatellite markers

Standard identification systems usually ensure that biopsy material is correctly associated with a given patient. Sometimes, as when a tumor is unexpectedly found, the provenance (proof of origin) of a tissue sample may be questioned; the tissue may have been mislabelled or contaminated with tissue from another patient. Techniques used to confirm tissue provenance include comparing either tissue markers of gender or ABO blood groups; however, these methods have weak confirmatory power. Recently, the use of DNA-based polymerase chain reaction (PCR) techniques has been reported. Paired, formalin-fixed, paraffin-embedded, 10 microns tissue sections were selected from 17 patients, 8 of whom had carcinoma, either by dividing a biopsy section, using sequential biopsies, or sequential biopsy and autopsy tissue. The resulting 36 samples were coded before analysis. In two additional cases, 1-mm fragments of tumor from one patient were included in the tissue block of benign tissue from another patient, the tumor fragments were identified on hematoxylin-and-eosin-stained sections, separately scraped off the glass slide, and analyzed. Tissue from two clinical cases, one of suspected mislabelling and one with a suspected carry-over of malignant tissue were also investigated. Short tandem repeat sequences (STR) or microsatellites, are 2-5 base pair repeats that vary in their repeat number between individuals. This variation (polymorphism) can be assessed using a PCR. A panel of markers of 3 STRs; ACPP, INT 2, and CYP 19 (on chromosomes 3, 11, and 15, respectively) were used. DNA was isolated from the samples after xylene deparaffinization and proteinase digestion, and was then amplified in a radioactive PCR using primers selected to give a product size ranging from 136-178 bases. Amplified products were electrophoresed on denaturing polyacrylamide gels, dried, and autoradiographed. DNA segments were successfully extracted from all samples but one, which was fixed in Bouin's fluid. By comparing allele sizes from the panel, all tissue pairs (other than the Bouin's pair) were successfully matched, the 1-mm tumor fragments were correctly assigned, and the two clinical problems were solved. STRs are highly informative and robust markers, well suited to PCR of small portions of tissue sections, and are an effective method to confirm the provenance of benign and malignant biopsy and autopsy material.

RMA with quantile normalization mixes biological signals between different sample groups in microarray data analysis

Quantile normalization (QN) is a technique for microarray data processing and is the default normalization method in the Robust Multi-array Average (RMA) procedure, which was primarily designed for analysing gene expression data from Affymetrix arrays. Given the abundance of Affymetrix microarrays and the popularity of the RMA method, it is crucially important that the normalization procedure is applied appropriately. In this study we carried out simulation experiments and also analysed real microarray data to investigate the suitability of RMA when it is applied to dataset with different groups of biological samples. From our experiments, we showed that RMA with QN does not preserve the biological signal included in each group, but rather it would mix the signals between the groups. We also showed that the Median Polish method in the summarization step of RMA has similar mixing effect. RMA is one of the most widely used methods in microarray data processing and has been applied to a vast volume of data in biomedical research. The problematic behaviour of this method suggests that previous studies employing RMA could have been misadvised or adversely affected. Therefore we think it is crucially important that the research community recognizes the issue and starts to address it. The two core elements of the RMA method, quantile normalization and Median Polish, both have the undesirable effects of mixing biological signals between different sample groups, which can be detrimental to drawing valid biological conclusions and to any subsequent analyses. Based on the evidence presented here and that in the literature, we recommend exercising caution when using RMA as a method of processing microarray gene expression data, particularly in situations where there are likely to be unknown subgroups of samples.

Injection locked oscillator lock detector

A simple circuit that is able to indicate if an injection-locked oscillator is in the locked condition by providing a ‘high’ or ‘low’ output is presented. The detector is compatible with most injection-locked oscillators as all that is required is access to the low-frequency bias circuit, with no direct access needed to the RF/microwave signals. To prove the universal nature of the lock detector it is successfully demonstrated practically for two scenarios: (i) a 1 GHz injection-locked VCO and (ii) a 60 GHz SiGe VCO MMIC.