Instability of 8E5 calibration standard revealed by digital PCR risks inaccurate quantification of HIV DNA in clinical samples by qPCR

Establishing a cure for HIV is hindered by the persistence of latently infected cells which constitute the viral reservoir. Real-time qPCR, used for quantification of this reservoir by measuring HIV DNA, requires external calibration; a common choice of calibrator is the 8E5 cell line, which is assumed to be stable and to contain one HIV provirus per cell. In contrast, digital PCR requires no external calibration and potentially provides ‘absolute’ quantification. We compared the performance of qPCR and dPCR in quantifying HIV DNA in 18 patient samples. HIV DNA was detected in 18 by qPCR and in 15 by dPCR, the difference being due to the smaller sample volume analysed by dPCR. There was good quantitative correlation (R2 = 0.86) between the techniques but on average dPCR values were only 60% of qPCR values. Surprisingly, investigation revealed that this discrepancy was due to loss of HIV DNA from the 8E5 cell calibrant. 8E5 extracts from two other sources were also shown to have significantly less than one HIV DNA copy per cell and progressive loss of HIV from 8E5 cells during culture was demonstrated. We therefore suggest that the copy number of HIV in 8E5 extracts be established by dPCR prior to use as calibrator

Differing Perception of DNA Evidence and Intelligence Capabilities in Criminal Investigations

The ability to predict physical characteristics from DNA presents significant opportunities for forensic science. Giving scientists an ability to make predictions about the donor of genetic material at a crime scene can then give investigators new intelligence leads for cold cases where DNA evidence has not identified any person of interest. However, the interpretation of this new form of intelligence requires careful analysis. The responses to an online survey, conducted in 2018-19, were used to examine how actors in the criminal justice system assess and interpret different types of DNA evidence and intelligence. The groups of focus for the survey were investigators, legal practitioners and the general public (as potential jurors). Several statistically significant effects were identified based on occupation and whether an individual had prior exposure to new DNA technology. Monitoring how those involved in interpreting reports from different types of DNA evidence and intelligence interpret them helps to ensure that decisions are made based on a sound understanding of their capabilities and limitations and may inform broader training and awareness strategies

Low unspliced cell-associated HIV RNA in early treated adolescents living with HIV on long suppressive ART

Introduction Initiation of antiretroviral treatment (ART) in patients early after HIV-infection and long-term suppression leads to low or undetectable levels of HIV RNA and cell-associated (CA) HIV DNA and RNA. Both CA-DNA and CA-RNA, overestimate the size of the HIV reservoir but CA-RNA as well as p24/cell-free viral RNA can be indicators of residual viral replication. This study describes HIV RNA amounts and levels of cytokines/soluble markers in 40 well-suppressed adolescents who initiated ART early in life and investigated which viral markers may be informative as endpoints in cure clinical trials within this population. Methods Forty adolescents perinatally infected with HIV on suppressive ART for >5 years were enrolled in the CARMA study. HIV DNA and total or unspliced CA-RNA in PBMCs were analyzed by qPCR/RT-qPCR and dPCR/RT-dPCR. Cell-free HIV was determined using an ultrasensitive viral load (US-VL) assay. Plasma markers and p24 were analyzed by digital ELISA and correlations between total and unspliced HIV RNA and clinical markers, including age at ART, Western Blot score, levels of cytokines/inflammation markers or HIV CA-DNA, were tested. Results CA-RNA was detected in two thirds of the participants and was comparable in RT-qPCR and RT-dPCR. Adolescents with undetectable CA-RNA showed significantly lower HIV DNA compared to individuals with detectable CA-RNA. Undetectable unspliced CA-RNA was positively associated with age at ART initiation and Western Blot score. We found that a higher concentration of TNF-alpha was predictive of higher CA-DNA and CA-RNA. Other clinical characteristics like US-VL, time to suppression, or percent CD4+ T-lymphocytes were not predictive of the CA-RNA in this cross-sectional study. Conclusions Low CA-DNA after long-term suppressive ART is associated with lower CA-RNA, in concordance with other reports. Patients with low CA-RNA levels in combination with low CA-DNA and low Western Blot scores should be further investigated to characterize candidates for treatment interruption trials. Unspliced CA-RNA warrants further investigation as a marker that can be prioritized in paediatric clinical trials where the sample volume can be a significant limitation

The generalised anxiety stigma scale (GASS): psychometric properties in a community sample

<p>Abstract</p> <p>Background</p> <p>Although there is substantial concern about negative attitudes to mental illness, little is known about the stigma associated with Generalised Anxiety Disorder (GAD) or its measurement. The aim of this study was to develop a multi-item measure of Generalised Anxiety Disorder stigma (the GASS).</p> <p>Methods</p> <p>Stigma items were developed from a thematic analysis of web-based text about the stigma associated with GAD. Six hundred and seventeen members of the public completed a survey comprising the resulting 20 stigma items and measures designed to evaluate construct validity. Follow-up data were collected for a subset of the participants (n = 212).</p> <p>Results</p> <p>The factor structure comprised two components: Personal Stigma (views about Generalised Anxiety Disorder); and Perceived Stigma (views about the beliefs of most others in the community). There was evidence of good construct validity and reliability for each of the Generalised Anxiety Stigma Scale (GASS) subscales.</p> <p>Conclusions</p> <p>The GASS is a promising brief measure of the stigma associated with Generalised Anxiety Disorder.</p

A population study comparing screening performance of prototypes for depression and anxiety with standard scales

<p>Abstract</p> <p>Background</p> <p>Screening instruments for mental disorders need to be short, engaging, and valid. Current screening instruments are usually questionnaire-based and may be opaque to the user. A prototype approach where individuals identify with a description of an individual with typical symptoms of depression, anxiety, social phobia or panic may be a shorter, faster and more acceptable method for screening. The aim of the study was to evaluate the accuracy of four new prototype screeners for predicting depression and anxiety disorders and to compare their performance with existing scales.</p> <p>Methods</p> <p>Short and ultra-short prototypes were developed for Major Depressive Disorder (MDD), Generalised Anxiety Disorder (GAD), Panic Disorder (PD) and Social Phobia (SP). Prototypes were compared to typical short and ultra-short self-report screening scales, such as the Centre for Epidemiology Scale, CES-D and the GAD-7, and their short forms. The Mini International Neuropsychiatric Interview (MINI) version 6 <abbrgrp><abbr bid="B1">1</abbr></abbrgrp> was used as the gold standard for obtaining clinical criteria through a telephone interview. From a population sample, 225 individuals who endorsed a prototype and 101 who did not were administered the MINI. Receiver operating characteristic (ROC) curves were plotted for the short and ultra short prototypes and for the short and ultra short screening scales.</p> <p>Results</p> <p>The study found that the rates of endorsement of the prototypes were commensurate with prevalence estimates. The short-form and ultra short scales outperformed the short and ultra short prototypes for every disorder except GAD, where the GAD prototype outperformed the GAD 7.</p> <p>Conclusions</p> <p>The findings suggest that people may be able to self-identify generalised anxiety more accurately than depression based on a description of a prototypical case. However, levels of identification were lower than expected. Considerable benefits from this method of screening may ensue if our prototypes can be improved for Major Depressive Disorder, Social Phobia and Panic Disorder.</p

Search for gravitational waves from binary inspirals in S3 and S4 LIGO data

We report on a search for gravitational waves from the coalescence of compact binaries during the third and fourth LIGO science runs. The search focused on gravitational waves generated during the inspiral phase of the binary evolution. In our analysis, we considered three categories of compact binary systems, ordered by mass: (i) primordial black hole binaries with masses in the range 0.35 M(sun) < m1, m2 < 1.0 M(sun), (ii) binary neutron stars with masses in the range 1.0 M(sun) < m1, m2 < 3.0 M(sun), and (iii) binary black holes with masses in the range 3.0 M(sun)< m1, m2 < m_(max) with the additional constraint m1+ m2 < m_(max), where m_(max) was set to 40.0 M(sun) and 80.0 M(sun) in the third and fourth science runs, respectively. Although the detectors could probe to distances as far as tens of Mpc, no gravitational-wave signals were identified in the 1364 hours of data we analyzed. Assuming a binary population with a Gaussian distribution around 0.75-0.75 M(sun), 1.4-1.4 M(sun), and 5.0-5.0 M(sun), we derived 90%-confidence upper limit rates of 4.9 yr^(-1) L10^(-1) for primordial black hole binaries, 1.2 yr^(-1) L10^(-1) for binary neutron stars, and 0.5 yr^(-1) L10^(-1) for stellar mass binary black holes, where L10 is 10^(10) times the blue light luminosity of the Sun.Comment: 12 pages, 11 figure

All-sky search for periodic gravitational waves in LIGO S4 data

We report on an all-sky search with the LIGO detectors for periodic gravitational waves in the frequency range 50-1000 Hz and with the frequency's time derivative in the range -1.0E-8 Hz/s to zero. Data from the fourth LIGO science run (S4) have been used in this search. Three different semi-coherent methods of transforming and summing strain power from Short Fourier Transforms (SFTs) of the calibrated data have been used. The first, known as "StackSlide", averages normalized power from each SFT. A "weighted Hough" scheme is also developed and used, and which also allows for a multi-interferometer search. The third method, known as "PowerFlux", is a variant of the StackSlide method in which the power is weighted before summing. In both the weighted Hough and PowerFlux methods, the weights are chosen according to the noise and detector antenna-pattern to maximize the signal-to-noise ratio. The respective advantages and disadvantages of these methods are discussed. Observing no evidence of periodic gravitational radiation, we report upper limits; we interpret these as limits on this radiation from isolated rotating neutron stars. The best population-based upper limit with 95% confidence on the gravitational-wave strain amplitude, found for simulated sources distributed isotropically across the sky and with isotropically distributed spin-axes, is 4.28E-24 (near 140 Hz). Strict upper limits are also obtained for small patches on the sky for best-case and worst-case inclinations of the spin axes.Comment: 39 pages, 41 figures An error was found in the computation of the C parameter defined in equation 44 which led to its overestimate by 2^(1/4). The correct values for the multi-interferometer, H1 and L1 analyses are 9.2, 9.7, and 9.3, respectively. Figure 32 has been updated accordingly. None of the upper limits presented in the paper were affecte

Search for Gravitational Waves Associated with 39 Gamma-Ray Bursts Using Data from the Second, Third, and Fourth LIGO Runs

We present the results of a search for short-duration gravitational-wave bursts associated with 39 gamma-ray bursts (GRBs) detected by gamma-ray satellite experiments during LIGO's S2, S3, and S4 science runs. The search involves calculating the crosscorrelation between two interferometer data streams surrounding the GRB trigger time. We search for associated gravitational radiation from single GRBs, and also apply statistical tests to search for a gravitational-wave signature associated with the whole sample. For the sample examined, we find no evidence for the association of gravitational radiation with GRBs, either on a single-GRB basis or on a statistical basis. Simulating gravitational-wave bursts with sine-gaussian waveforms, we set upper limits on the root-sum-square of the gravitational-wave strain amplitude of such waveforms at the times of the GRB triggers. We also demonstrate how a sample of several GRBs can be used collectively to set constraints on population models. The small number of GRBs and the significant change in sensitivity of the detectors over the three runs, however, limits the usefulness of a population study for the S2, S3, and S4 runs. Finally, we discuss prospects for the search sensitivity for the ongoing S5 run, and beyond for the next generation of detectors.Comment: 24 pages, 10 figures, 14 tables; minor changes to text and Fig. 2; accepted by Phys. Rev.