Darned in 2013: inclusion of model organisms and linking with Wikipedia

DARNED (DAtabase of RNA EDiting, available at http://darned.ucc.ie) is a centralized repository of reference genome coordinates corresponding to RNA nucleotides having altered templated identities in the process of RNA editing. The data in DARNED are derived from published datasets of RNA editing events. RNA editing instances have been identified with various methods, such as bioinformatics screenings, deep sequencing and/or biochemical techniques. Here we report our current progress in the development and expansion of the DARNED. In addition to novel database features the DARNED update describes inclusion of Drosophila melanogaster and Mus musculus RNA editing events and the launch of a community-based annotation in the RNA WikiProject

Evidence of efficient stop codon readthrough in four mammalian genes

Stop codon readthrough is used extensively by viruses to expand their gene expression. Until recent discoveries in Drosophila, only a very limited number of readthrough cases in chromosomal genes had been reported. Analysis of conserved protein coding signatures that extend beyond annotated stop codons identified potential stop codon readthrough of four mammalian genes. Here we use a modified targeted bioinformatic approach to identify a further three mammalian readthrough candidates. All seven genes were tested experimentally using reporter constructs transfected into HEK-293T cells. Four displayed efficient stop codon readthrough, and these have UGA immediately followed by CUAG. Comparative genomic analysis revealed that in the four readthrough candidates containing UGA-CUAG, this motif is conserved not only in mammals but throughout vertebrates with the first six of the seven nucleotides being universally conserved. The importance of the CUAG motif was confirmed using a systematic mutagenesis approach. One gene, OPRL1, encoding an opiate receptor, displayed extremely efficient levels of readthrough (∼31%) in HEK-293T cells. Signals both 5′ and 3′ of the OPRL1 stop codon contribute to this high level of readthrough. The sequence UGA-CUA alone can support 1.5% readthrough, underlying its importance.National Institutes of Health (U.S.) (NIH-1-R01-HG004037-07)National Institutes of Health (U.S.) (NSF-DBI-0644282)National Institutes of Health (U.S.) (NIH-U41-HG007234

GWIPS-viz: development of a ribo-seq genome browser

We describe the development of GWIPS-viz (http://gwips.ucc.ie), an online genome browser for viewing ribosome profiling data. Ribosome profiling (ribo-seq) is a recently developed technique that provides genome-wide information on protein synthesis (GWIPS) in vivo. It is based on the deep sequencing of ribosome-protected messenger RNA (mRNA) fragments, which allows the ribosome density along all mRNA transcripts present in the cell to be quantified. Since its inception, ribo-seq has been carried out in a number of eukaryotic and prokaryotic organisms. Owing to the increasing interest in ribo-seq, there is a pertinent demand for a dedicated ribo-seq genome browser. GWIPS-viz is based on The University of California Santa Cruz (UCSC) Genome Browser. Ribo-seq tracks, coupled with mRNA-seq tracks, are currently available for several genomes: human, mouse, zebrafish, nematode, yeast, bacteria (Escherichia coli K12, Bacillus subtilis), human cytomegalovirus and bacteriophage lambda. Our objective is to continue incorporating published ribo-seq data sets so that the wider community can readily view ribosome profiling information from multiple studies without the need to carry out computational processing

Epidemiological and Molecular Characterization of an Invasive Group A Streptococcus32.2 Outbreak

Anemm32.2 invasive group A streptococcus (iGAS) outbreak occurred in Liverpool from January 2010 to September 2012. This genotype had not previously been identified in Liverpool, but was responsible for 32% (14/44) of all iGAS cases reported during this time period. We performed a case-case comparison ofemm32.2 iGAS cases with non-emm32.2 control iGAS cases identified in the Liverpool population over the same time period to assess patient risk factors foremm32.2 iGAS infection. Theemm32.2 iGAS cases were confined to the adult population. We show that homelessness, intravenous drug use, and alcohol abuse predisposed patients toemm32.2 iGAS disease; however, no obvious epidemiological linkage between the patients withemm32.2 iGAS could be identified. Comparative whole-genome sequencing analysis ofemm32.2 iGAS and non-emm32.2 control isolates was also performed to identify pathogen factors which might have driven the outbreak. We identified 19 genes, five of which had previously been implicated in virulence, which were present in all of theemm32.2 iGAS isolates but not present in any of the non-emm32.2 control isolates. We report that a novelemm32.2 genotype emerged in Liverpool in 2010 and identified a specific subset of genes, which could have allowed this novelemm32.2 genotype to persist in a disadvantaged population in the region over a 3-year period.</p

The global distribution and diversity of protein vaccine candidate antigens in the highly virulent Streptococcus pnuemoniae serotype 1

Serotype 1 is one of the most common causes of pneumococcal disease worldwide. Pneumococcal protein vaccines are currently being developed as an alternate intervention strategy to pneumococcal conjugate vaccines. Pre-requisites for an efficacious pneumococcal protein vaccine are universal presence and minimal variation of the target antigen in the pneumococcal population, and the capability to induce a robust human immune response. We used in silico analysis to assess the prevalence of seven protein vaccine candidates (CbpA, PcpA, PhtD, PspA, SP0148, SP1912, SP2108) among 445 serotype 1 pneumococci from 26 different countries, across four continents. CbpA (76%), PspA (68%), PhtD (28%), PcpA (11%) were not universally encoded in the study population, and would not provide full coverage against serotype 1. PcpA was widely present in the European (82%), but not in the African (2%) population. A multi-valent vaccine incorporating CbpA, PcpA, PhtD and PspA was predicted to provide coverage against 86% of the global population. SP0148, SP1912 and SP2108 were universally encoded and we further assessed their predicted amino acid, antigenic and structural variation. Multiple allelic variants of these proteins were identified, different allelic variants dominated in different continents; the observed variation was predicted to impact the antigenicity and structure of two SP0148 variants, one SP1912 variant and four SP2108 variants, however these variants were each only present in a small fraction of the global population (<2%). The vast majority of the observed variation was predicted to have no impact on the efficaciousness of a protein vaccine incorporating a single variant of SP0148, SP1912 and/or SP2108 from S. pneumoniae TIGR4. Our findings emphasise the importance of taking geographic differences into account when designing global vaccine interventions and support the continued development of SP0148, SP1912 and SP2108 as protein vaccine candidates against this important pneumococcal serotype

Population genetic structure, antibiotic resistance, capsule switching and evolution of invasive pneumococci before conjugate vaccination in Malawi

INTRODUCTION: Pneumococcal infections cause a high death toll in Sub Saharan Africa (SSA) but the recently rolled out pneumococcal conjugate vaccines (PCV) will reduce the disease burden. To better understand the population impact of these vaccines, comprehensive analysis of large collections of pneumococcal isolates sampled prior to vaccination is required. Here we present a population genomic study of the invasive pneumococcal isolates sampled before the implementation of PCV13 in Malawi. MATERIALS AND METHODS: We retrospectively sampled and whole genome sequenced 585 invasive isolates from 2004 to 2010. We determine the pneumococcal population genetic structure and assessed serotype prevalence, antibiotic resistance rates, and the occurrence of serotype switching. RESULTS: Population structure analysis revealed 22 genetically distinct sequence clusters (SCs), which consisted of closely related isolates. Serotype 1 (ST217), a vaccine-associated serotype in clade SC2, showed highest prevalence (19.3%), and was associated with the highest MDR rate (81.9%) followed by serotype 12F, a non-vaccine serotype in clade SC10 with an MDR rate of 57.9%. Prevalence of serotypes was stable prior to vaccination although there was an increase in the PMEN19 clone, serotype 5 ST289, in clade SC1 in 2010 suggesting a potential undetected local outbreak. Coalescent analysis revealed recent emergence of the SCs and there was evidence of natural capsule switching in the absence of vaccine induced selection pressure. Furthermore, majority of the highly prevalent capsule-switched isolates were associated with acquisition of vaccine-targeted capsules. CONCLUSIONS: This study provides descriptions of capsule-switched serotypes and serotypes with potential to cause serotype replacement post-vaccination such as 12F. Continued surveillance is critical to monitor these serotypes and antibiotic resistance in order to design better infection prevention and control measures such as inclusion of emerging replacement serotypes in future conjugate vaccines

Understanding pneumococcal serotype 1 biology through population genomic analysis

Background: Pneumococcus kills over one million children annually and over 90 % of these deaths occur in low-income countries especially in Sub-Saharan Africa (SSA) where HIV exacerbates the disease burden. In SSA, serotype 1 pneumococci particularly the endemic ST217 clone, causes majority of the pneumococcal disease burden. To understand the evolution of the virulent ST217 clone, we analysed ST217 whole genomes from isolates sampled from African and Asian countries. Methods: We analysed 226 whole genome sequences from the ST217 lineage sampled from 9 African and 4 Asian countries. We constructed a whole genome alignment and used it for phylogenetic and coalescent analyses. We also screened the genomes to determine presence of antibiotic resistance conferring genes. Results: Population structure analysis grouped the ST217 isolates into five sequence clusters (SCs), which were highly associated with different geographical regions and showed limited intracontinental and intercontinental spread. The SCs showed lower than expected genomic sequence, which suggested strong purifying selection and small population sizes caused by bottlenecks. Recombination rates varied between the SCs but were lower than in other successful clones such as PMEN1. African isolates showed higher prevalence of antibiotic resistance genes than Asian isolates. Interestingly, certain West African isolates harbored a defective chloramphenicol and tetracycline resistance-conferring element (Tn5253) with a deletion in the loci encoding the chloramphenicol resistance gene (cat(pC194)), which caused lower chloramphenicol than tetracycline resistance. Furthermore, certain genes that promote colonisation were absent in the isolates, which may contribute to serotype 1's rarity in carriage and consequently its lower recombination rates. Conclusions: The high phylogeographic diversity of the ST217 clone shows that this clone has been in circulation globally for a long time, which allowed its diversification and adaptation in different geographical regions. Such geographic adaptation reflects local variations in selection pressures in different locales. Further studies will be required to fully understand the biological mechanisms which makes the ST217 clone highly invasive but unable to successfully colonise the human nasopharynx for long durations which results in lower recombination rates.Peer reviewe

African Genomic Medicine Portal: A Web Portal for Biomedical Applications

Genomics data are currently being produced at unprecedented rates, resulting in increased knowledge discovery and submission to public data repositories. Despite these advances, genomic information on African-ancestry populations remains significantly low compared with European- and Asian-ancestry populations. This information is typically segmented across several different biomedical data repositories, which often lack sufficient fine-grained structure and annotation to account for the diversity of African populations, leading to many challenges related to the retrieval, representation and findability of such information. To overcome these challenges, we developed the African Genomic Medicine Portal (AGMP), a database that contains metadata on genomic medicine studies conducted on African-ancestry populations. The metadata is curated from two public databases related to genomic medicine, PharmGKB and DisGeNET. The metadata retrieved from these source databases were limited to genomic variants that were associated with disease aetiology or treatment in the context of African-ancestry populations. Over 2000 variants relevant to populations of African ancestry were retrieved. Subsequently, domain experts curated and annotated additional information associated with the studies that reported the variants, including geographical origin, ethnolinguistic group, level of association significance and other relevant study information, such as study design and sample size, where available. The AGMP functions as a dedicated resource through which to access African-specific information on genomics as applied to health research, through querying variants, genes, diseases and drugs. The portal and its corresponding technical documentation, implementation code and content are publicly available

African Genomic Medicine Portal: A Web Portal for Biomedical Applications

Genomics data are currently being produced at unprecedented rates, resulting in increased knowledge discovery and submission to public data repositories. Despite these advances, genomic information on African-ancestry populations remains significantly low compared with European- and Asian-ancestry populations. This information is typically segmented across several different biomedical data repositories, which often lack sufficient fine-grained structure and annotation to account for the diversity of African populations, leading to many challenges related to the retrieval, representation and findability of such information. To overcome these challenges, we developed the African Genomic Medicine Portal (AGMP), a database that contains metadata on genomic medicine studies conducted on African-ancestry populations. The metadata is curated from two public databases related to genomic medicine, PharmGKB and DisGeNET. The metadata retrieved from these source databases were limited to genomic variants that were associated with disease aetiology or treatment in the context of African-ancestry populations. Over 2000 variants relevant to populations of African ancestry were retrieved. Subsequently, domain experts curated and annotated additional information associated with the studies that reported the variants, including geographical origin, ethnolinguistic group, level of association significance and other relevant study information, such as study design and sample size, where available. The AGMP functions as a dedicated resource through which to access African-specific information on genomics as applied to health research, through querying variants, genes, diseases and drugs. The portal and its corresponding technical documentation, implementation code and content are publicly available