Detecting functional rare variants by collapsing and incorporating functional annotation in Genetic Analysis Workshop 17 mini-exome data

Association studies using tag SNPs have been successful in detecting disease-associated common variants. However, common variants, with rare exceptions, explain only at most 5–10% of the heritability resulting from genetic factors, which leads to the common disease/rare variants assumption. Indeed, recent studies using sequencing technologies have demonstrated that common diseases can be due to rare variants that could not be systematically studied earlier. Unfortunately, methods for common variants are not optimal if applied to rare variants. To identify rare variants that affect disease risk, several investigators have designed new approaches based on the idea of collapsing different rare variants inside the same genomic block (e.g., the same gene or pathway) to enrich the signal. Here, we consider three different collapsing methods in the multimarker regression model and compared their performance on the Genetic Analysis Workshop 17 data using the consistency of results across different simulations and the cross-validation prediction error rate. The comparison shows that the proportion collapsing method seems to outperform the other two methods and can find both truly associated rare and common variants. Moreover, we explore one way of incorporating the functional annotations for the variants in the data that collapses nonsynonymous and synonymous variants separately to allow for different penalties on them. The incorporation of functional annotations led to higher sensitivity and specificity levels when the detection results were compared with the answer sheet. The initial analysis was performed without knowledge of the simulating model

Efficient assembly of very short oligonucleotides using T4 DNA Ligase

<p>Abstract</p> <p>Background</p> <p>In principle, a pre-constructed library of all possible short oligonucleotides could be used to construct many distinct gene sequences. In order to assess the feasibility of such an approach, we characterized T4 DNA Ligase activity on short oligonucleotide substrates and defined conditions suitable for assembly of a plurality of oligonucleotides.</p> <p>Findings</p> <p>Ligation by T4 DNA Ligase was found to be dependent on the formation of a double stranded DNA duplex of at least five base pairs surrounding the site of ligation. However, ligations could be performed effectively with overhangs smaller than five base pairs and oligonucleotides as small as octamers, in the presence of a second, complementary oligonucleotide. We demonstrate the feasibility of simultaneous oligonucleotide phosphorylation and ligation and, as a proof of principle for DNA synthesis through the assembly of short oligonucleotides, we performed a hierarchical ligation procedure whereby octamers were combined to construct a target 128-bp segment of the beta-actin gene.</p> <p>Conclusions</p> <p>Oligonucleotides as short as 8 nucleotides can be efficiently assembled using T4 DNA Ligase. Thus, the construction of synthetic genes, without the need for custom oligonucleotide synthesis, appears feasible.</p

Delayed gastric emptying and reduced postprandial small bowel water content of equicaloric whole meal bread versus rice meals in healthy subjects: novel MRI insights

BACKGROUND/OBJECTIVES: Postprandial bloating is a common symptom in patients with functional gastrointestinal (GI) diseases. Whole meal bread (WMB) often aggravates such symptoms though the mechanisms are unclear. We used magnetic resonance imaging (MRI) to monitor the intragastric fate of a WMB meal (11% bran) compared to a rice pudding (RP) meal. SUBJECTS/METHODS: 12 healthy volunteers completed this randomised crossover study. They fasted overnight and after an initial MRI scan consumed a glass of orange juice with a 2267 kJ WMB or an equicaloric RP meal. Subjects underwent serial MRI scans every 45 min up to 270 min to assess gastric volumes and small bowel water content and completed a GI symptom questionnaire. RESULTS: The MRI intragastric appearance of the two meals was markedly different. The WMB meal formed a homogeneous dark bolus with brighter liquid signal surrounding it. The RP meal separated into an upper, liquid layer and a lower particulate layer allowing more rapid emptying of the liquid compared to solid phase (sieving). The WMB meal had longer gastric half emptying times (132±8 min) compared to the RP meal (104±7 min), P<0.008. The WMB meal was associated with markedly reduced MRI-visible small bowel free mobile water content compared to the RP meal, P<0.0001. CONCLUSIONS: WMB bread forms a homogeneous bolus in the stomach which inhibits gastric sieving and hence empties slower than the equicaloric rice meal. These properties may explain why wheat causes postprandial bloating and could be exploited to design foods which prolong satiation

Rare variant collapsing in conjunction with mean log p-value and gradient boosting approaches applied to Genetic Analysis Workshop 17 data

In addition to methods that can identify common variants associated with susceptibility to common diseases, there has been increasing interest in approaches that can identify rare genetic variants. We use the simulated data provided to the participants of Genetic Analysis Workshop 17 (GAW17) to identify both rare and common single-nucleotide polymorphisms and pathways associated with disease status. We apply a rare variant collapsing approach and the usual association tests for common variants to identify candidates for further analysis using pathway-based and tree-based ensemble approaches. We use the mean log p-value approach to identify a top set of pathways and compare it to those used in simulation of GAW17 dataset. We conclude that the mean log p-value approach is able to identify those pathways in the top list and also related pathways. We also use the stochastic gradient boosting approach for the selected subset of single-nucleotide polymorphisms. When compared the result of this tree-based method with the list of single-nucleotide polymorphisms used in dataset simulation, in addition to correct SNPs we observe number of false positives

Community prevalence of SARS-CoV-2 in England from April to November, 2020: results from the ONS Coronavirus Infection Survey

Background: Decisions about the continued need for control measures to contain the spread of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) rely on accurate and up-to-date information about the number of people testing positive for SARS-CoV-2 and risk factors for testing positive. Existing surveillance systems are generally not based on population samples and are not longitudinal in design. Methods: Samples were collected from individuals aged 2 years and older living in private households in England that were randomly selected from address lists and previous Office for National Statistics surveys in repeated cross-sectional household surveys with additional serial sampling and longitudinal follow-up. Participants completed a questionnaire and did nose and throat self-swabs. The percentage of individuals testing positive for SARS-CoV-2 RNA was estimated over time by use of dynamic multilevel regression and poststratification, to account for potential residual non-representativeness. Potential changes in risk factors for testing positive over time were also assessed. The study is registered with the ISRCTN Registry, ISRCTN21086382. Findings: Between April 26 and Nov 1, 2020, results were available from 1 191 170 samples from 280 327 individuals; 5231 samples were positive overall, from 3923 individuals. The percentage of people testing positive for SARS-CoV-2 changed substantially over time, with an initial decrease between April 26 and June 28, 2020, from 0·40% (95% credible interval 0·29–0·54) to 0·06% (0·04–0·07), followed by low levels during July and August, 2020, before substantial increases at the end of August, 2020, with percentages testing positive above 1% from the end of October, 2020. Having a patient-facing role and working outside your home were important risk factors for testing positive for SARS-CoV-2 at the end of the first wave (April 26 to June 28, 2020), but not in the second wave (from the end of August to Nov 1, 2020). Age (young adults, particularly those aged 17–24 years) was an important initial driver of increased positivity rates in the second wave. For example, the estimated percentage of individuals testing positive was more than six times higher in those aged 17–24 years than in those aged 70 years or older at the end of September, 2020. A substantial proportion of infections were in individuals not reporting symptoms around their positive test (45–68%, dependent on calendar time. Interpretation: Important risk factors for testing positive for SARS-CoV-2 varied substantially between the part of the first wave that was captured by the study (April to June, 2020) and the first part of the second wave of increased positivity rates (end of August to Nov 1, 2020), and a substantial proportion of infections were in individuals not reporting symptoms, indicating that continued monitoring for SARS-CoV-2 in the community will be important for managing the COVID-19 pandemic moving forwards. Funding: Department of Health and Social Care

Ct threshold values, a proxy for viral load in community SARS-CoV-2 cases, demonstrate wide variation across populations and over time

Background: Information on SARS-CoV-2 in representative community surveillance is limited, particularly cycle threshold (Ct) values (a proxy for viral load). Methods: We included all positive nose and throat swabs 26 April 2020 to 13 March 2021 from the UK’s national COVID-19 Infection Survey, tested by RT-PCR for the N, S, and ORF1ab genes. We investigated predictors of median Ct value using quantile regression. Results: Of 3,312,159 nose and throat swabs, 27,902 (0.83%) were RT-PCR-positive, 10,317 (37%), 11,012 (40%), and 6550 (23%) for 3, 2, or 1 of the N, S, and ORF1ab genes, respectively, with median Ct = 29.2 (~215 copies/ml; IQR Ct = 21.9–32.8, 14–56,400 copies/ml). Independent predictors of lower Cts (i.e. higher viral load) included self-reported symptoms and more genes detected, with at most small effects of sex, ethnicity, and age. Single-gene positives almost invariably had Ct > 30, but Cts varied widely in triple-gene positives, including without symptoms. Population-level Cts changed over time, with declining Ct preceding increasing SARS-CoV-2 positivity. Of 6189 participants with IgG S-antibody tests post-first RT-PCR-positive, 4808 (78%) were ever antibody-positive; Cts were significantly higher in those remaining antibody negative. Conclusions: Marked variation in community SARS-CoV-2 Ct values suggests that they could be a useful epidemiological early-warning indicator. Funding: Department of Health and Social Care, National Institutes of Health Research, Huo Family Foundation, Medical Research Council UK; Wellcome Trust

Effect of Delta variant on viral burden and vaccine effectiveness against new SARS-CoV-2 infections in the UK

The effectiveness of the BNT162b2 and ChAdOx1 vaccines against new severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infections requires continuous re-evaluation, given the increasingly dominant B.1.617.2 (Delta) variant. In this study, we investigated the effectiveness of these vaccines in a large, community-based survey of randomly selected households across the United Kingdom. We found that the effectiveness of BNT162b2 and ChAdOx1 against infections (new polymerase chain reaction (PCR)-positive cases) with symptoms or high viral burden is reduced with the B.1.617.2 variant (absolute difference of 10–13% for BNT162b2 and 16% for ChAdOx1) compared to the B.1.1.7 (Alpha) variant. The effectiveness of two doses remains at least as great as protection afforded by prior natural infection. The dynamics of immunity after second doses differed significantly between BNT162b2 and ChAdOx1, with greater initial effectiveness against new PCR-positive cases but faster declines in protection against high viral burden and symptomatic infection with BNT162b2. There was no evidence that effectiveness varied by dosing interval, but protection was higher in vaccinated individuals after a prior infection and in younger adults. With B.1.617.2, infections occurring after two vaccinations had similar peak viral burden as those in unvaccinated individuals. SARS-CoV-2 vaccination still reduces new infections, but effectiveness and attenuation of peak viral burden are reduced with B.1.617.2

Monitoring populations at increased risk for SARS-CoV-2 infection in the community using population-level demographic and behavioural surveillance

BACKGROUND: The COVID-19 pandemic is rapidly evolving, with emerging variants and fluctuating control policies. Real-time population screening and identification of groups in whom positivity is highest could help monitor spread and inform public health messaging and strategy. METHODS: To develop a real-time screening process, we included results from nose and throat swabs and questionnaires taken 19 July 2020-17 July 2021 in the UK's national COVID-19 Infection Survey. Fortnightly, associations between SARS-CoV-2 positivity and 60 demographic and behavioural characteristics were estimated using logistic regression models adjusted for potential confounders, considering multiple testing, collinearity, and reverse causality. FINDINGS: Of 4,091,537 RT-PCR results from 482,677 individuals, 29,903 (0·73%) were positive. As positivity rose September-November 2020, rates were independently higher in younger ages, and those living in Northern England, major urban conurbations, more deprived areas, and larger households. Rates were also higher in those returning from abroad, and working in healthcare or outside of home. When positivity peaked December 2020-January 2021 (Alpha), high positivity shifted to southern geographical regions. With national vaccine roll-out from December 2020, positivity reduced in vaccinated individuals. Associations attenuated as rates decreased between February-May 2021. Rising positivity rates in June-July 2021 (Delta) were independently higher in younger, male, and unvaccinated groups. Few factors were consistently associated with positivity. 25/45 (56%) confirmed associations would have been detected later using 28-day rather than 14-day periods. INTERPRETATION: Population-level demographic and behavioural surveillance can be a valuable tool in identifying the varying characteristics driving current SARS-CoV-2 positivity, allowing monitoring to inform public health policy. FUNDING: Department of Health and Social Care (UK), Welsh Government, Department of Health (on behalf of the Northern Ireland Government), Scottish Government, National Institute for Health Research

Nietzsche’s Epistemic Perspectivism

Nietzsche offers a positive epistemology, and those who interpret him as a skeptic or a mere pragmatist are mistaken. Instead he supports what he calls per- spectivism. This is a familiar take on Nietzsche, as perspectivism has been analyzed by many previous interpreters. The present paper presents a sketch of the textually best supported and logically most consistent treatment of perspectivism as a first- order epistemic theory. What’s original in the present paper is an argument that Nietzsche also offers a second-order methodological perspectivism aimed at enhancing understanding, an epistemic state distinct from knowledge. Just as Descartes considers and rejects radical skepticism while at the same time adopting methodological skepticism, one could consistently reject perspectivism as a theory of knowledge while accepting it as contributing to our understanding. It is argued that Nietzsche’s perspectivism is in fact two-tiered: knowledge is perspectival because truth itself is, and in addition there is a methodological perspectivism in which distinct ways of knowing are utilized to produce understanding. A review of the manner in which understanding is conceptualized in contemporary epistemology and philosophy of science serves to illuminate how Nietzsche was tackling these ideas