Removing noise from pyrosequenced amplicons

Background In many environmental genomics applications a homologous region of DNA from a diverse sample is first amplified by PCR and then sequenced. The next generation sequencing technology, 454 pyrosequencing, has allowed much larger read numbers from PCR amplicons than ever before. This has revolutionised the study of microbial diversity as it is now possible to sequence a substantial fraction of the 16S rRNA genes in a community. However, there is a growing realisation that because of the large read numbers and the lack of consensus sequences it is vital to distinguish noise from true sequence diversity in this data. Otherwise this leads to inflated estimates of the number of types or operational taxonomic units (OTUs) present. Three sources of error are important: sequencing error, PCR single base substitutions and PCR chimeras. We present AmpliconNoise, a development of the PyroNoise algorithm that is capable of separately removing 454 sequencing errors and PCR single base errors. We also introduce a novel chimera removal program, Perseus, that exploits the sequence abundances associated with pyrosequencing data. We use data sets where samples of known diversity have been amplified and sequenced to quantify the effect of each of the sources of error on OTU inflation and to validate these algorithms

Genome-Wide Association Study in BRCA1 Mutation Carriers Identifies Novel Loci Associated with Breast and Ovarian Cancer Risk

BRCA1-associated breast and ovarian cancer risks can be modified by common genetic variants. To identify further cancer risk-modifying loci, we performed a multi-stage GWAS of 11,705 BRCA1 carriers (of whom 5,920 were diagnosed with breast and 1,839 were diagnosed with ovarian cancer), with a further replication in an additional sample of 2,646 BRCA1 carriers. We identified a novel breast cancer risk modifier locus at 1q32 for BRCA1 carriers (rs2290854, P = 2.7×10-8, HR = 1.14, 95% CI: 1.09-1.20). In addition, we identified two novel ovarian cancer risk modifier loci: 17q21.31 (rs17631303, P = 1.4×10-8, HR = 1.27, 95% CI: 1.17-1.38) and 4q32.3 (rs4691139, P = 3.4×10-8, HR = 1.20, 95% CI: 1.17-1.38). The 4q32.3 locus was not associated with ovarian cancer risk in the general population or BRCA2 carriers, suggesting a BRCA1-specific associat

The effect of a first kidney transplant on a subsequent transplant outcome: An experimental and clinical study

The effect of a first kidney transplant on a subsequent transplant outcome: An experimental and clinical study.BackgroundSecond kidney transplantations have a roughly similar clinical outcome to first transplantations. Nevertheless, the effect of the presence of the first, nonfunctional transplant at the time of the second transplantation may also influence its outcome and has not yet been specifically studied.MethodsWe analyzed the effect of the presence of a first graft on the outcome of a second graft in a rodent allograft model and in a cohort of 240 human second kidney allograft recipients.ResultsIn rodents, 100 days subsequent to the rejection of the first graft, we observed an increase in blood but not spleen CD4+CD25+ T cells, whereas no differences were observed in transcriptional patterns. Adoptive transfer of day 100 splenocytes did not prolong graft survival. Moreover, the presence of a first rejected graft does not prolong the survival of a second graft performed at a later date. In the human context, a higher incidence of patients with anti-HLA immunization and a higher % of PRA were observed in retransplant recipients with primary allograft nephrectomy. Despite a relatively low statistical power, our data do not suggest significant differences in graft outcome between recipients of second transplants with primary allograft nephrectomy and those without.ConclusionCollectively, the data from both an experimental model and a large cohort of human recipients of a second graft do not suggest a beneficial effect of the presence of a first rejected graft at the time of a second transplantation

Gas Seepage along the Edge of the Aquitaine Shelf (France): Origin and Local Fluxes

During the scientific expedition GAZCOGNE2 at the Bay of Biscay nine gas seeps were sampled for the first time and their flux was measured using an in situ pressure-preservation sampler (PEGAZ, ©IFREMER). Overall, three sites were investigated to determine the nature and the origin of the gases bubbling at the seafloor and forming acoustic plumes into the water column, as this was the question raised fromthe first geologic study of the area.This has guided our study and accordingly corresponds to themain purpose of the present article.Thus, themolecular and isotopic (D and 13C) analyses revealed that the gas seeps were primarily composed of methane. Both methane and ethane are of microbial origin, and the former has been generated by microbial reduction of carbon dioxide. Heavier hydrocarbons accounted for less than 0.06% mol of the total amount. Despite the microbial origin of methane, the samples exhibit subtle differences with respect to the 13CCH4 values, which varied between −72.7 and −66.1‰. It has been suggested that such a discrepancy was predominantly governed by the occurrence of anaerobic methane oxidation. The PEGAZ sampler also enabled us to estimate the local gas fluxes fromthe sampled streams. The resulting values are extremely heterogeneous between seeps, ranging from 35 to 368mLn⋅min−1. Assuming a steady discharge, the mean calculated methane emission for the nine seeps is of 38 kmol⋅yr−1. Considering the extent of the seep area, this very local estimate suggests that the Aquitaine Shelf is a very appropriate place to study methane discharge and its fate on continental shelves

La polypose associée à MUTYH : synthèse et actualisation des recommandations françaises établies en 2012 sous l’égide de l’Institut national du cancer (INCa)

International audienceMUTYH-associated polyposis (MUTYH-associated polyposis, MAP) is an autosomal recessive inheritance disorder related to bi-allelic constitutional pathogenic variants of the MUTYH gene which was first described in 2002. In 2011, a group of French experts composed of clinicians and biologists, performed a summary of the available data on this condition and drew up recommendations concerning the indications and the modalities of molecular analysis of the MUTYH gene in index cases and their relatives, as well as the management of affected individuals. In view of recent developments, some recommendations have become obsolete, in particular with regard to the molecular analysis strategy since MUTYH gene has been recently included in a consensus panel of 14 genes predisposing to colorectal cancer. This led us to revise all the points of the previous expertise. We report here the revised version of this work which successively considers the phenotype and the tumor risks associated with this genotype, the differential diagnoses, the indication criteria and the strategy of the molecular analysis and the recommendations for the management of affected individuals. We also discuss the phenotype and the tumor risks associated with mono-allelic pathogenic variants of MUTYH gene

Ibuprofen and ketoprofen potentiate UVA-induced cell death by a photosensitization process

International audienceNonsteroidal 2-arylproprionic acids are widely used, over-the-counter, anti-inflammatory drugs. Photosensitivity is a commonly overlooked adverse effect of these drugs. Based on the combined use of cell viability assays and molecular modeling, we prove and rationalize the photochemical pathways triggering photosensitization for two drugs, ibuprofen and ketoprofen. As its parent compound benzophenone, ketoprofen produces singlet oxygen, upon triplet manifold population. However, ibuprofen and ketoprofen photodissociate and hence may generate two highly reactive radicals. The formation of metastable aggregates between the two drugs and B-DNA is also directly probed by molecular dynamics. Our approach characterizes the coupled influence of the drug's intrinsic photochemistry and the interaction pattern with DNA. The photosensitization activity of nonsteroidal 2-arylproprionic acids, being added to gels and creams for topical use, should be crucially analyzed and rationalized to enact the proper preventive measures

Intra-database validation of case-identifying algorithms using reconstituted electronic health records from healthcare claims data

BACKGROUND: Diagnosis performances of case-identifying algorithms developed in healthcare database are usually assessed by comparing identified cases with an external data source. When this is not feasible, intra-database validation can present an appropriate alternative. OBJECTIVES: To illustrate through two practical examples how to perform intra-database validations of case-identifying algorithms using reconstituted Electronic Health Records (rEHRs). METHODS: Patients with 1) multiple sclerosis (MS) relapses and 2) metastatic castration-resistant prostate cancer (mCRPC) were identified in the French nationwide healthcare database (SNDS) using two case-identifying algorithms. A validation study was then conducted to estimate diagnostic performances of these algorithms through the calculation of their positive predictive value (PPV) and negative predictive value (NPV). To that end, anonymized rEHRs were generated based on the overall information captured in the SNDS over time (e.g. procedure, hospital stays, drug dispensing, medical visits) for a random selection of patients identified as cases or non-cases according to the predefined algorithms. For each disease, an independent validation committee reviewed the rEHRs of 100 cases and 100 non-cases in order to adjudicate on the status of the selected patients (true case/ true non-case), blinded with respect to the result of the corresponding algorithm. RESULTS: Algorithm for relapses identification in MS showed a 95% PPV and 100% NPV. Algorithm for mCRPC identification showed a 97% PPV and 99% NPV. CONCLUSION: The use of rEHRs to conduct an intra-database validation appears to be a valuable tool to estimate the performances of a case-identifying algorithm and assess its validity, in the absence of alternative