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SummaryType 1 diabetes-associated single nucleotide polymorphisms are mainly located in non-coding regions of the human genome. Single nucleotide polymorphisms located in long non-coding RNAs may result in the disruption of their secondary structure, affecting their function.Here, we functionally characterized the virus-induced type 1 diabetes-associated lncRNAARGI(Antiviral Response Gene Inducer).ARGIupregulation in pancreatic β cells leads to the transcriptional activation of antiviral and pro-inflammatory genes. Upon a viral insult,ARGIis upregulated in the nuclei of pancreatic β cells and binds to CTCF to interact with the regulatory regions ofIFNβand interferon-stimulated genes, promoting their transcriptional activation in an allele-specific manner.The presence of the risk allele for type 1 diabetes inARGIinduces an hyperactivation of type I IFN response in β cells, an expression signature that is present in the pancreas of diabetic patients. These data shed light on the molecular mechanisms by which type 1 diabetes-related single nucleotide polymorphisms in long non-coding RNAs influence pathogenesis at the pancreatic β cell level.

The role of microbiota in colorectal cancer has been studied since alterations in its composition were observed. In addition, there are more and more pieces of evidence that microbiota could be implicated in colorectal cancer progression. Thus, the components of the microbiota could be biomarkers for the diagnosis and prognosis of colorectal cancer. In addition, it is important to address how the microbiota interacts with the host and how the host shapes the microbiota, in order to understand the biological pathways and mechanisms involved in their relationship and the consequences of their interactions in colorectal cancer. Thereby, it could be possible to find feasible measures and treatments to prevent or better diagnose colorectal cancer. In this review, we will try to summarize the role of the microbiota in colorectal cancer and its interactions with the host and the host genetics, coming to some conclusions that could be useful to find the gaps in our knowledge and propose future steps in this field.

BACKGROUND The results obtained to date concerning food groups, diet quality and colorectal cancer (CRC) risk vary according to criteria used and the study populations. AIM To study the relationships between food groups, diet quality and CRC risk, in an adult population of the Basque Country (North of Spain). METHODS This observational study included 308 patients diagnosed with CRC and 308 ageand sex-matched subjects as controls. During recruitment, dietary, anthropometric, lifestyle, socioeconomic, demographic and health status information was collected. Adherence to the dietary recommendations was evaluated utilizing the Healthy Eating Index for the Spanish Diet and the MedDietScore. Conditional logistic regressions were used to evaluate the associations of food group intakes, diet quality scores, categorized in tertiles, with CRC risk. RESULTS The adjusted models for potential confounding factors showed a direct association between milk and dairy products consumption, in particular high-fat cheeses [odds ratio (OR) third tertile vs first tertile = 1.87, 95% confidence intervals (CI): 1.11-3.16], and CRC risk. While the consumption of fiber-containing foods, especially whole grains (OR third tertile vs first tertile = 0.62, 95%CI: 0.39-0.98), and fatty fish (OR third tertile vs first tertile = 0.53, 95%CI: 0.27-0.99) was associated with a lower risk for CRC. Moreover, higher MD adherence was associated with a reduced CRC risk in adjusted models (OR third tertile vs first tertile = 0.40, 95%CI: 0.20-0.80). CONCLUSION Direct associations were found for high-fat cheese, whereas an inverse relation was reported for fiber-containing foods and fatty fish, as well as adherence to a Mediterranean dietary pattern. Supported by the Department of Health and Consumer Affairs, Basque Government, No. 2011111153; Saiotek, Basque Government, No. S-PE12UN058; Pre-doctoral grant from the Basque Government, No. PRE_2015_2_0084; and United States Department of Agriculture—Agricultural Research Service, No. 58-1950-4-003.

Background Risk prediction is an essential part of preventative medicine and in recent years genomic information has become an interesting factor in risk models. Polygenic risk scores (PRS) combine the effect of many genetic variations into a single score which has been shown to have predictive value for many diseases. This study aimed to investigate the association between PRS for endometriosis and the clinical presentation of the disease. Methods Women with endometriosis (N = 172) were identified at the Department of Gynecology. All participants answered questionnaires regarding sociodemographic factors, lifestyle habits and medical history, registered bowel symptoms on the Visual Analog Scale for Irritable Bowel Syndrome and passed blood samples. DNA was extracted and samples were genotyped, and a PRS was calculated based on previous genome-wide association studies of endometriosis. Inflammatory proteins and TSH receptor antibodies (TRAb) in serum were analyzed. Results Inverse associations were identified between PRS and spread of endometriosis, involvement of the gastrointestinal tract and hormone treatment. However, significance was lost when calculated as p for trend and the specificity and sensitivity were low. There were no correlations between PRS and TRAb or inflammatory proteins. Conclusion The findings indicate that specific PRS should be developed to predict clinical presentations in patient with endometriosis.

Background: Irritable bowel syndrome (IBS) is a common gastrointestinal condition which entails a high burden in the quality of life (QoL) of patients. Nutritional interventions have been proposed to alleviate symptoms, since still no effective treatments exist for IBS. Objectives: Our aim is to analyse the feasibility of the use of starch- and sucrose-reduced diet (SSRD). Design: In this study, we used a SSRD accompanied by nutritional and culinary recommendations to measure the effects in IBS patients with diarrhoea. Methods: In all, 34 participants completed a 4-week nutritional intervention based on SSRD. Symptoms, QoL and dietary habits were assessed by several questionnaires that were completed at the beginning, daily, after 2 weeks, at the end, and after 2 months. Results: 85.29% of the participants reached the primary endpoint [reduction of 50 points or more in IBS-symptom severity scale (SSS)], and 58.82% the secondary endpoint (reduction of 50% or more in IBS-SSS). The relief of symptoms and improvement of the QoL were significant after 2 weeks of intervention, at the end and after 2 months. Dietary habits were consistent with the diet and high adherence was achieved. Conclusions: SSRD and individualized nutritional and culinary guidance improved symptoms and QoL of IBS patients with diarrhoea, with a high adherence.

BACKGROUND Functional dyspepsia (FD) is a common gastrointestinal condition of poorly understood pathophysiology. While symptoms' overlap with other conditions may indicate common pathogenetic mechanisms, genetic predisposition is suspected but has not been adequately investigated. METHODS Using healthcare, questionnaire, and genetic data from three large population-based biobanks (UK Biobank, EGCUT, and MGI), we surveyed FD comorbidities, heritability, and genetic correlations across a wide spectrum of conditions and traits in 10,078 cases and 351,282 non-FD controls of European ancestry. KEY RESULTS In UK Biobank, 281 diagnoses were detected at increased prevalence in FD, based on healthcare records. Among these, gastrointestinal conditions (OR = 4.0, p 0.344), mostly overlapping with those also enriched in FD patients. Suggestive (p

Summary Gut dysmotility is associated with constipation, diarrhea, and functional gastrointestinal disorders like irritable bowel syndrome (IBS), although its molecular underpinnings are poorly characterized. We studied stool frequency (defined by the number of bowel movements per day, based on questionnaire data) as a proxy for gut motility in a GWAS meta-analysis including 167,875 individuals from UK Biobank and four smaller population-based cohorts. We identify 14 loci associated with stool frequency (p ≤ 5.0 × 10−8). Gene set and pathway analyses detected enrichment for genes involved in neurotransmitter/neuropeptide signaling and preferentially expressed in enteric motor neurons controlling peristalsis. PheWAS identified pleiotropic associations with dysmotility syndromes and the response to their pharmacological treatment. The genetic architecture of stool frequency correlates with that of IBS, and UK Biobank participants from the top 1% of stool frequency polygenic score distribution were associated with 5× higher risk of IBS with diarrhea. These findings pave the way for the identification of actionable pathological mechanisms in IBS and the dysmotility syndromes., Graphical abstract, Highlights Genetics of gut motility via data on stool frequency in 167,875 individuals GWAS identifies 14 loci associated with stool frequency Candidate genes enriched in neurotransmitter signaling and enteric motor neurons Polygenic scores predict increased risk of irritable bowel syndrome, Bonfiglio et al. investigate the genetics of human gut motility using genotype and questionnaire data on stool frequency in 167,875 individuals from the UK Biobank and four other cohorts. They identify 14 loci associated with stool frequency as well as candidate genes enriched in neurotransmitter signaling and preferentially expressed in enteric neurons controlling peristalsis. The genetic architecture of stool frequency correlates with that of irritable bowel syndrome (IBS), and stool frequency polygenic scores were predictive of IBS risk.

ObjectiveGut dysmotility is associated with constipation, diarrhea, and functional gastrointestinal disorders (FGID) like irritable bowel syndrome (IBS). Its molecular underpinnings, and their anomalies in FGID disorders are poorly characterized, hence we sought to gain mechanistic insight through a large-scale genetic investigation.DesignWe used stool frequency (STL-FRQ) as a (surrogate) quantitative trait to study the genetics of gut motility, exploiting questionnaire and genotype data from UK Biobank and four smaller population-based cohorts (LifeLines-Deep, Genes for Good, Flemish Gut Flora Project and PopCol), in a GWAS meta-analysis spanning 8,817,117 high-quality SNP markers and 167,875 individuals of European descent.ResultsWe identify 13 genome-wide significant loci (P≤5.0×10−8) harboring prioritized genes that are: i) involved in sensory perception and neurotransmitter/neuropeptide signaling; ii) enriched for their expression in enteric motor neurons associated with the control of peristalsis (P=7.0×10−8) iii) previously linked to other traits and conditions, including GI motility and dysmotility syndromes, and the response to their pharmacological treatment. The genetic architecture of STL-FRQ most strongly correlates with that of IBS (rg=0.42; P=1.1×10−3). In UK Biobank, the risk of IBS with diarrhea was 4x higher in individuals from the top 1% of the distribution of polygenic scores (PGS) computed based on STL-FRQ GWAS summary statistics (ORs=4.14; P=1.2×10−97).ConclusionWe identify loci harboring genes with a plausible role in GI motility, possibly acting via neurotransmission and similar pathways in specialized enteric neurons. The demonstrated relevance of these findings to IBS warrants further study for the identification of actionable pathomechanisms in the dysmotility syndromes.

The aim of the study is to identify additional celiac disease associated loci in the major histocompatibility complex (MHC) independent from classical HLA risk alleles (HLA-DR3-DQ2) and to characterize their potential functional impact in celiac disease pathogenesis at the intestinal level.We performed a high-resolution single-nucleotide polymorphism (SNP) genotyping of the MHC region, comparing HLA-DR3 homozygous celiac patients and non-celiac controls carrying a single copy of the B8-DR3-DQ2 conserved extended haplotype. Expression level of potential novel risk genes was determined by RT-PCR in intestinal biopsies and in intestinal and immune cells isolated from control and celiac individuals. Small interfering RNA-driven silencing of selected genes was performed in the intestinal cell line T84.MHC genotyping revealed 2 associated SNPs, one located in TRIM27 gene and another in the non-coding gene HCG14. After stratification analysis, only HCG14 showed significant association independent from HLA-DR-DQ loci. Expression of HCG14 was slightly downregulated in epithelial cells isolated from duodenal biopsies of celiac patients, and eQTL analysis revealed that polymorphisms in HCG14 region were associated with decreased NOD1 expression in duodenal intestinal cells.We have successfully employed a conserved extended haplotype-matching strategy and identified a novel additional celiac disease risk variant in the lncRNA HCG14. This lncRNA seems to regulate the expression of NOD1 in an allele-specific manner. Further functional studies are needed to clarify the role of HCG14 in the regulation of gene expression and to determine the molecular mechanisms by which the risk variant in HCG14 contributes to celiac disease pathogenesis.

Long noncoding RNAs (lncRNAs) have emerged as critical regulators of inflammation. To further understand the interaction between inflammatory signaling pathways and lncRNAs, we characterized the function of cardiac and apoptosis-related lncRNA (Carlr), an lncRNA expressed in both mouse and human cells of diverse tissues. Carlr expression is increased following NF-κB signaling in macrophages, with concomitant translocation to, and enrichment of, the transcript in the cytoplasm. Knockdown of Carlr results in impaired expression of NF-κB pathway genes and influences the interaction between macrophages and intestinal cells in an inflammatory environment. In human celiac disease patient samples, increased levels of the Carlr transcript were detected in the cytoplasm, alongside elevated expression of NF-κB pathway genes. These findings suggest that increased Carlr expression and/or cytoplasmic localization is required for efficient NF-κB signaling and is associated with the inflamed tissue state observed in human celiac disease.

[Background] Respiratory failure is a key feature of severe Covid-19 and a critical driver of mortality, but for reasons poorly defined affects less than 10% of SARS-CoV-2 infected patients., [Methods] We included 1,980 patients with Covid-19 respiratory failure at seven centers in the Italian and Spanish epicenters of the SARS-CoV-2 pandemic in Europe (Milan, Monza, Madrid, San Sebastian and Barcelona) for a genome-wide association analysis. After quality control and exclusion of population outliers, 835 patients and 1,255 population-derived controls from Italy, and 775 patients and 950 controls from Spain were included in the final analysis. In total we analyzed 8,582,968 single-nucleotide polymorphisms (SNPs) and conducted a metaanalysis of both case-control panels., [Results] We detected cross-replicating associations with rs11385942 at chromosome 3p21.31 and rs657152 at 9q34, which were genome-wide significant (P, [Conclusions] We herein report the first robust genetic susceptibility loci for the development of respiratory failure in Covid-19. Identified variants may help guide targeted exploration of severe Covid19 pathophysiology., The IKMB's core facilities received infrastructure support by the Deutsche Forschungsgemeinschaft (DFG) Cluster of Excellence "Precision Medicine in Chronic Inflammation" (PMI, EXC2167). The project also received support through a philanthropic donation by Stein Erik Hagen and Canica AS. L.V. was funded by the Fondazione IRCCS Ca’ Granda «COVID-19 Biobank» research grant. This work was also supported by the Ministero dell’Istruzione, dell’Università e della Ricerca – MIUR project "Dipartimenti di Eccellenza 2018 – 2022" (n° D15D18000410001) to the Department of Medical Sciences, University of Torino. The IKMB authors received financial support from the UKSH Foundation "Gutes Tun!" (special thanks to Alexander Eck, Jenspeter Horst and Jens Scholz) and the German Federal Ministry of Education and Research (BMBF; grant ID 01KI20197). HLA-Typing was performed and supported by the Stefan-MorschStiftung. M.A.H was supported by the Spanish Ministry of Science and Innovation ‘JdC fellowship IJC2018-035131-I.

Inflammatory bowel disease (IBD) is a chronic inflammatory condition of the gastrointestinal tract that develops due to the interaction between genetic and environmental factors. More than 160 loci have been associated with IBD, but the functional implication of many of the associated genes remains unclear. N6-Methyladenosine (m6A) is the most abundant internal modification in mRNA. m6A methylation regulates many aspects of mRNA metabolism, playing important roles in the development of several pathologies. Interestingly, SNPs located near or within m6A motifs have been proposed as possible contributors to disease pathogenesis. We hypothesized that certain IBD-associated SNPs could regulate the function of genes involved in IBD development via m6A-dependent mechanisms. We used online available GWAS, m6A and transcriptome data to find differentially expressed genes that harbored m6A-SNPs associated with IBD. Our analysis resulted in five candidate genes corresponding to two of the major IBD subtypes: UBE2L3 and SLC22A4 for Crohn&rsquo, s Disease and TCF19, C6orf47 and SNAPC4 for Ulcerative Colitis. Further analysis using in silico predictions and co-expression analyses in combination with in vitro functional studies showed that our candidate genes seem to be regulated by m6A-dependent mechanisms. These findings provide the first indication of the implication of RNA methylation events in IBD pathogenesis.

Celiac disease (CeD) is a complex immune-mediated inflammatory condition triggered by the ingestion of gluten in genetically predisposed individuals. Literature suggests that alterations in gut microbiota composition and function precede the onset of CeD. Considering that microbiota is partly determined by host genetics, we speculated that the genetic makeup of CeD patients could elicit disease development through alterations in the intestinal microbiota. To evaluate potential causal relationships between gut microbiota and CeD, we performed a two-sample Mendelian randomization analysis (2SMR). Exposure data were obtained from the raw results of a previous genome-wide association study (GWAS) of gut microbiota and outcome data from summary statistics of CeD GWAS and Immunochip studies. We identified a number of putative associations between gut microbiota single nucleotide polymorphisms (SNPs) associated with CeD. Regarding bacterial composition, most of the associated SNPs were related to Firmicutes phylum, whose relative abundance has been previously reported to be altered in CeD patients. In terms of functional units, we linked a number of SNPs to several bacterial metabolic pathways that seemed to be related to CeD. Overall, this study represented the first 2SMR approach to elucidate the relationship between microbiome and CeD. This research was funded by the Basque Department of Health, grant numbers GVSAN2018/111086 and GVSAN2019/111085 to J.R.B. and N.F.-J., respectively.

Celiac disease (CD) patients present a loss of intestinal barrier function due to structural alterations in the tight junction (TJ) network, the most apical unions between epithelial cells. The association of TJ-related gene variants points to an implication of this network in disease susceptibility. This work aims to characterize the functional implication of TJ-related, disease-associated loci in CD pathogenesis. We performed an association study of 8 TJ-related gene variants in a cohort of 270 CD and 91 non-CD controls. The expression level of transcripts located in the associated SNP region was analyzed by RT-PCR in several human tissues and in duodenal biopsies of celiac patients and non-CD controls. (si)RNA-driven silencing combined with gliadin in the Caco2 intestinal cell line was used to analyze the implication of transcripts from the associated region in the regulation of TJ genes. We replicated the association of rs6962966*A variant [p = 0.0029; OR = 1.88 (95%1.24-2.87)], located in an intron of TJ-related MAGI2 coding gene and upstream of RP4-587D13.2 transcript, bioinformatically classified as a long non-coding RNA (lncRNA). The expression of both genes is correlated and constitutively downregulated in CD intestine. Silencing of lncRNA decreases the levels of MAGI2 protein. At the same time, silencing of MAGI2 affects the expression of several TJ-related genes. The associated region is functionally altered in disease, probably affecting CD-related TJ genes. This work was partially funded by the Basque Department of Education grant IT1281-19 and ISCIII Research Project PI16/00258, cofunded by the European Union ERDF, A way to make Europe to JB. AC-R is supported by an Ikerbasque Fellowship and funded by a research project grant 2017111082 from the Basque Goverment. IS was funded by a research project grant 2015111068 from the Basque Department of Health. AJ-M and AO-G are predoctoral fellows funded by FPI grants from the Basque Department of Education, Universities and Research and IR-G and MS are predoctoral fellows funded by the University of Basque Country.

Neonatal diabetes mellitus (NDM) is a rare form of diabetes diagnosed within the first 6 months of life. Genetic studies have allowed the identification of several genes linked to the development of NDM; however, genetic causes for ∼20% of the cases remain to be clarified. Most cases of NDM involve isolated diabetes, but sometimes NDM appears in association with other pathological conditions, including autoimmune diseases. Recent reports have linked activating mutations in STAT3 with early-onset autoimmune disorders that include diabetes of autoimmune origin, but the functional impact of STAT3-activating mutations have not been characterized at the pancreatic β-cell level. By using whole-exome sequencing, we identified a novel missense mutation in the binding domain of the STAT3 protein in a patient with NDM. The functional analyses showed that the mutation results in an aberrant activation of STAT3, leading to deleterious downstream effects in pancreatic β-cells. The identified mutation leads to hyperinhibition of the transcription factor Isl-1 and, consequently, to a decrease in insulin expression. These findings represent the first functional indication of a direct link between an NDM-linked activating mutation in STAT3 and pancreatic β-cell dysfunction.

To identify candidate genes in celiac disease (CD), we reanalyzed the whole Immunochip CD cohort using a different approach that clusters individuals based on immunoancestry prior to disease association analysis, rather than by geographical origin. We detected 636 new associated SNPs (P

BACKGROUND & AIMS: Genetic factors are believed to affect risk for irritable bowel syndrome (IBS), but there have been no sufficiently powered and adequately sized studies. To identify DNA variants associated with IBS risk, we performed a genome-wide association study (GWAS) of the large UK Biobank population-based cohort, which includes genotype and health data from 500,000 participants. METHODS: We studied 7,287,191 high-quality single nucleotide polymorphisms in individuals who self-reported a doctor's diagnosis of IBS (cases; n = 9576) compared to the remainder of the cohort (controls; n = 336,499) (mean age of study subjects, 40-69 years). Genome-wide significant findings were further investigated in 2045 patients with IBS from tertiary centers and 7955 population controls from Europe and the United States, and a small general population sample from Sweden (n = 249). Functional annotation of GWAS results was carried out by integrating data from multiple biorepositories to obtain biological insights from the observed associations. RESULTS: We identified a genome-wide significant association on chromosome 9q31.2 (single nucleotide polymorphism rs10512344; P = 3.57 × 10-8) in a region previously linked to age at menarche, and 13 additional loci of suggestive significance (P < 5.0×10-6). Sex-stratified analyses revealed that the variants at 9q31.2 affect risk of IBS in women only (P = 4.29 × 10-10 in UK Biobank) and also associate with constipation-predominant IBS in women (P = .015 in the tertiary cohort) and harder stools in women (P = .0012 in the population-based sample). Functional annotation of the 9q31.2 locus identified 8 candidate genes, including the elongator complex protein 1 gene (ELP1 or IKBKAP), which is mutated in patients with familial dysautonomia. CONCLUSIONS: In a sufficiently powered GWAS of IBS, we associated variants at the locus 9q31.2 with risk of IBS in women. This observation may provide additional rationale for investigating the role of sex hormones and autonomic dysfunction in IBS. ispartof: GASTROENTEROLOGY vol:155 issue:1 pages:168-179 ispartof: location:United States status: published

The major histocompatibility complex (MHC)-containing genes are among the most polymorphic in vertebrates. MHC genes code for proteins that are critical in the immune system response. In this study, the polymorphism of the second exon of the MHC class II DRB gene was characterized in the Eastern woodchuck (Marmota monax). Woodchucks chronically infected with the woodchuck hepatitis virus (WHV) represent the best available animal model for the study of chronic hepatitis B infection in humans. In the genotyped animals we found fifteen alleles, which were expressed in two independent loci and that were named DRB1A and DRB1B in this work. The 15 alleles investigated showed an elevated divergence. A significant excess of non-synonymous substitutions was detected, which could indicate that a historical positive selection is acting in the woodchuck DRB1 genes. This hypothesis was confirmed in our study by the high variability in or near the antigen binding sites (ABS) and by the results obtained in sequence variability analyses. This analysis identified the presence of a microsatellite sequence that is located at the start of the second intron, which could further allow the development of a fast and cheap semiautomatic sequencing method.

In this work, we attempt to identify the location and expression of bovine ERVs and the nature of nearby genes. As many as 1610 bovine genes contain a bovine ERV (BoERV) inserted into their introns. Most of these BoERVs present an antisense orientation, which could be a consequence of the detrimental effects of the sense insertion. Based on the overrepresentation of Gene Ontology terms, we determined that some genes located in the vicinity of BoERVs are related to viral response and chromatin assembly. In addition, we identified some genes that belonged to IFNs that are inserted in or between BoERVs, pointing out a possible role of BoERVs in some gene duplication. Based on EST database mining, significant expression was detected for BoERV genes in the thyroid and in embryos only. Transcription factor binding sites for Rxra, a steroid and thyroid hormone receptor, were detected in three of the most expressed BoERVs in the thyroid glands.

8 páginas, 1 tabla, 5 figuras, Benthic Prorocentrum species can produce toxins that adversely affect animals and human health. They are known to co-occur with other bloom-forming, potentially toxic, benthic dinoflagellates of the genera Ostreopsis, Coolia, and Gambierdiscus. In this study, we report on the presence of P. elegans M.Faust and P. levis M.A.Faust, Kibler, Vandersea, P.A. Tester & Litaker from the southeastern Bay of Biscay. Sampling was carried out in the Summer-Autumn 2010–2012 along the Atlantic coast of the Iberian Peninsula, but these two species were only found in the northeastern part of the Peninsula. Strains were isolated from macroalgae collected from rocky-shore areas bordering accessible beaches. Morphological traits of isolated strains were analyzed by LM and SEM, whereas molecular analyses were performed using the LSU and internal transcribed spacer (ITS)1-5.8S-ITS2 regions of the rDNA. A bioassay with Artemia fransciscana and liquid chromatography–high-resolution mass spectrometry analyses were used to check the toxicity of the species, whose results were negative. The strains mostly corresponded to their species original morphological characterization, which is supported by the phylogenetic analyses in the case of P. levis, whereas for P. elegans, this is the first known molecular characterization. This is also the second known report of P. elegans, Financial support for this research was provided by the Department for Environment of Bizkaiko Foru Aldundia, the Bilbao-Bizkaia Water Consortium, and the Basque Government (project IT-417-07). A grant from the University of the Basque Country to H. David and a specialization fellowship for PhD researchers awarded by the University of the Basque Country to A. Laza-Martínez also supported this study. And also Project PIE 201270E032.

Neonatal diabetes mellitus (NDM) is a rare form of diabetes diagnosed within the first 6 months of life. Genetic studies have allowed the identification of several genes linked to the development of NDM; however, genetic causes for ∼20% of the cases remain to be clarified. Most cases of NDM involve isolated diabetes, but sometimes NDM appears in association with other pathological conditions, including autoimmune diseases. Recent reports have linked activating mutations in

Potential relationships between amino acid motifs of various alleles of the ovine major histocompatibility complex DR (Ovar-DR) molecule and occurrence of clinical OPA caused by JSRV were investigated in a case–control study. Latxa sheep (n = 132) screened for presence/absence of pulmonary OPA lesions were typed for their Ovar-DRB1 2nd exon alleles by PCR and sequence-based typing (PCR-SBT). The polymorphic amino acid residues derived from the obtained 34 DRB1 protein variants were subjected to a logistic regression-based association study. The amino acids at several positions showed significant associations with the presence/absence of pulmonary OPA lesions; some of the residues were located within the peptide binding cleft of the DRB molecule, including pockets P1, P4, P7 and P9.

Summary Host-commensal relationships in the skin are a complex system governed by variables related to the host, the bacteria and the environment. A disruption of this system may lead to new steady states, which, in turn, may lead to disease. We have studied one such disruption by characterizing the skin micro- biota in healthy and immunodepressed (ID) mice. A detailed anatomopathological study failed to reveal any difference between the skin of healthy and ID mice. We sequenced the 16S rDNA V1-V2 gene region to saturation in 10 healthy and 10 ID 8 week- old mice, and found than all of the healthy and two of the ID mice had bacterial communities that were similar in composition to that of human skin, although, presumably because of the uniform raising conditions, less interindividual variation was found in mice. However, eight ID mice showed microbiota dominated by Staphylococcus epidermidis. Quantita- tive PCR amplification of 16S rDNA gene and of the Staphylococcus-specific TstaG region confirmed the previous results and indicated that the quantitative levels of Staphylococcus were similar in both groups while the total number of 16S copies was greater in the healthy mice. Thus, it is possible that, under long-term immunodeficiency, which removes the acquired but not the native immune system, S. epi- dermidis may inhibit the growth of other bacteria but does not cause a pathogenic state.

Ovine pulmonary adenocarcinoma (OPA) and Maedi-Visna (Maedi) are two chronic respiratory diseases of retroviral origin which occur worldwide. It is known that different host genetic factors influence the outcome of viral infections. To determine if variation in the Mhc-DRB1 gene was associated with progression to these ovine diseases, sheep lungs with and without OPA and Maedi lesions were collected. A sequence-based method was applied and 40 different alleles were detected in the sample analysed. In the allele-by-allele association analysis, allele DRB1*0325 had a significant association with susceptibility to Maedi (P = 0.045). For OPA, DRB1*0143 and DRB1*0323 were significantly associated with susceptibility (P = 0.024 and P = 0.029), and allele DRB1*0702 was significantly associated with resistance (P = 0.012). Based on these results, the Mhc-DRB1 alleles were classified by effect in three categories-susceptible (S), resistant (R) and neutral (N)-and animals were reassigned the genotypes as S/S, S/R, S/N, R/R, R/N and N/N. In a second analysis, penalised logistic regression models including a flock effect were run. In Maedi, significant association was detected for the N/S heterozygote (P = 0.0007), but not for the S/S homozygote, probably as a result of the low number of S/S animals. In OPA, association was detected for both the S/S and R/R homozygotes (P = 0.005 and P = 0.047). This allele grouping method may be applied in association studies with highly variable genes. This is the first study demonstrating significant associations between sheep Mhc-DRB1 alleles and susceptibility to OPA and Maedi. Therefore, both diseases are suitable candidates for more comprehensive genetic studies.

Endogenous retroviruses (ERVs) are remnants of retroviral infections that are present in a large number of vertebrate genomes. Based on the proposal that the rat could act as a reservoir of retroviruses, rat ERVs were analysed in silico using a whole-genome approach. To enrich the detected ERV groups, we applied an upgraded approach based on the hidden Markov model. We found 2637 elements that were classified into the following groups: 9 groups of Class I; 15 of Class II, 7 of them previously described; 1 of Class III; and 3 groups whose classification was unclear but were distantly related to Class I. Sixteen ERV groups seemed to be specific to rat. The high number of rat-specific groups might be related to the contact of rats with retroviruses and their role as a reservoir. In addition, the env gene of the more extended groups seemed to be undetectable.

The variability of the sheep major histocompatibility complex (MHC) class II DRB1 locus has been analyzed in this work. Exon 2 of Ovar-DRB1 was amplified by polymerase chain reaction (PCR) with the primers designed by Amills et al. for goats. In a total of 187 sheep of Latxa breed, we identified by PCR-single-strand conformation polymorphism (SSCP) 19 alleles, eight of them previously unpublished. Moreover, the observed heterozygosity reached 91-95%. A new allelic type named DRB*14 was defined, which brings to light once more the discussion as to the ancestor of the domestic sheep. Although all the defined alleles can be discriminated by PCR-SSCP, sequence-based typing is proposed as the technique of choice for sheep DRB1 gene typing.

Garcia-Etxebarria, Koldo et al., While most patients affected by the influenza A(H1N1) pandemic experienced mild symptoms, a small fraction required hospitalization, often without concomitant factors that could explain such a severe course. We hypothesize that host genetic factors could contribute to aggravate the disease. To test this hypothesis, we compared the allele frequencies of 547,296 genome-wide single nucleotide polymorphisms (SNPs) between 49 severe and 107 mild confirmed influenza A cases, as well as against a general population sample of 549 individuals. When comparing severe vs. mild influenza A cases, only one SNP was close to the conventional p = 5×10−8. This SNP, rs28454025, sits in an intron of the GSK233 gene, which is involved in a neural development, but seems not to have any connections with immunological or inflammatory functions. Indirectly, a previous association reported with CD55 was replicated. Although sample sizes are low, we show that the statistical power in our design was sufficient to detect highly-penetrant, quasi-Mendelian genetic factors. Hence, and assuming that rs28454025 is likely to be a false positive, no major genetic factor was detected that could explain poor influenza A course., Fundign for this work came from Instituto de Salud Carlos III (Madrid, Spain; www.isciii.es) grant GR09/0032.

[Backgroun] Metagenomics is the genomic study of uncultured environmental samples, which has been greatly facilitated by the advent of shotgun-sequencing technologies. One of the main focuses of metagenomics is the discovery of previously uncultured microorganisms, which makes the assignment of sequences to a particular taxon a challenge and a crucial step. Recently, several methods have been developed to perform this task, based on different methodologies such as sequence composition or sequence similarity. The sequence composition methods have the ability to completely assign the whole dataset. However, their use in metagenomics and the study of their performance with real data is limited. In this work, we assess the consistency of three different methods (BLAST + Lowest Common Ancestor, Phymm, and Naïve Bayesian Classifier) in assigning real and simulated sequence reads., [Results] Both in real and in simulated data, BLAST + Lowest Common Ancestor (BLAST + LCA), Phymm, and Naïve Bayesian Classifier consistently assign a larger number of reads in higher taxonomic levels than in lower levels. However, discrepancies increase at lower taxonomic levels. In simulated data, consistent assignments between all three methods showed greater precision than assignments based on Phymm or Bayesian Classifier alone, since the BLAST + LCA algorithm performed best. In addition, assignment consistency in real data increased with sequence read length, in agreement with previously published simulation results., [Conclusions] The use and combination of different approaches is advisable to assign metagenomic reads. Although the sensitivity could be reduced, the reliability can be increased by using the reads consistently assigned to the same taxa by, at least, two methods, and by training the programs using all available information., This work was financed by the MICINN (Spanish Ministry of Science and Innovation) grant SAF2010-16240. MGG was supported by a predoctoral fellowship from MICINN.

The genus Pseudo-nitzschia contains potentially toxic species of problematic taxonomy, making it one of the most intensively studied diatom genera. The study of 35 clonal strains isolated from the Bilbao estuary, an area that experiences recurrent blooms of Pseudo-nitzschia, revealed the presence of two new species, P. abrensis and P. plurisecta, differing from their congeners in both morphology and gene sequence. The morphological features were analyzed by LM and EM, whereas molecular analyses were based on the internal transcribed spacer (ITS) and large subunit (LSU) regions of the rDNA. P. plurisecta appears closely related to P. cuspidata/P. pseudodelicatissima in the phylogenetic tree, whereas P. abrensis forms a moderately supported clade with P. heimii/P. subpacifica and P. caciantha/P. circumpora. Comparison of the secondary structure of ITS2 regions reveals marked differences in the most highly conserved regions among related taxa. Morphologically, the new species differ from their closest congeners in the arrangement of the poroid sectors and the density of valve striae and fibulae. The two species share similar pigment composition, and belong to the group of Pseudo-nitzschia species containing only chlorophyll c2 and c3 .

Host-commensal relationships in the skin are a complex system governed by variables related to the host, the bacteria and the environment. A disruption of this system may lead to new steady states, which, in turn, may lead to disease. We have studied one such disruption by characterizing the skin microbiota in healthy and immunodepressed (ID) mice. A detailed anatomopathological study failed to reveal any difference between the skin of healthy and ID mice. We sequenced the 16S rDNA V1-V2 gene region to saturation in 10 healthy and 10 ID 8 week-old mice, and found than all of the healthy and two of the ID mice had bacterial communities that were similar in composition to that of human skin, although, presumably because of the uniform raising conditions, less interindividual variation was found in mice. However, eight ID mice showed microbiota dominated by Staphylococcus epidermidis. Quantitative PCR amplification of 16S rDNA gene and of the Staphylococcus-specific TstaG region confirmed the previous results and indicated that the quantitative levels of Staphylococcus were similar in both groups while the total number of 16S copies was greater in the healthy mice. Thus, it is possible that, under long-term immunodeficiency, which removes the acquired but not the native immune system, S.epidermidis may inhibit the growth of other bacteria but does not cause a pathogenic state.

The identification of the conventionally accepted species of Clavulina (Cantharellales, Basidiomycota) in Europe (Clavulina amethystina, Clavulina cinerea, Clavulina cristata, and Clavulina rugosa) is often difficult and many specimens are not straightforwardly assignable to any of those four species, which is why some authors have questioned their identity. In order to assess the status of those species, a morphological examination was combined with the molecular analysis of the ITS region. The same six major clades were obtained in the Bayesian and parsimony phylogenetic analyses, and all six clades were well-supported at least by one of the analyses. Morphological characters, such as the overall branching pattern, the presence and intensity of grey colour, the cristation of the apices, and basidiospore size and shape were to various extents correlated with the phylogenetic signal obtained from the ITS region. The congruence between the molecular analyses and morphology, rather than geographical origin, suggests the existence of several species that can be delimited using a combined phylogenetic and morphological species recognition. The analyses revealed that C. cristata and C. rugosa are well-delimited species. In contrast, more than one taxa could be subsumed under the names C. amethystina and C. cinerea, the taxonomical complexity of which is discussed. The ITS region is proved to be adequate to separate phylogenetic species of Clavulina.

Major histocompatibility complex class II locus DRB variation was investigated by single-strand conformation polymorphism analysis and sequence analysis in the two subspecies of Pyrenean chamois (Rupicapra pyrenaica) endemic to the Iberian Peninsula. Low levels of genetic variation were detected in both subspecies, with seven different alleles in R. p. pyrenaica and only three in the R. p. parva. After applying the rarefaction method to cope with the differences in sample size, the low allele number of parva was highlighted. The low allelic repertoire of the R. p. parva subspecies is most likely the result of bottlenecks caused by hunting pressure and recent parasitic infections by sarcoptic mange. A phylogenetic analysis of both Pyrenean chamois and DRB alleles from 10 different caprinid species revealed that the chamois alleles form two monophyletic groups. In comparison with other Caprinae DRB sequences, the Rupicapra alleles displayed a species-specific clustering that reflects a large temporal divergence of the chamois from other caprinids, as well as a possible difference in the selective environment for these species.

In the last decade, an extensive effort has been made to characterize the human microbiota, due to its clinical and economic interests. However, a metagenomic approach to the skin microbiota is hampered by the high proportion of host DNA that is recovered. In contrast with the burgeoning field of gut metagenomics, skin metagenomics has been hindered by the absence of an efficient method to avoid sequencing the host DNA. We present here a method for recovering microbial DNA from skin samples, based on a combination of molecular techniques. We have applied this method to mouse skin, and have validated it by standard, quantitative PCR and amplicon sequencing of 16S rRNA. The taxonomic diversity recovered was not altered by this new method, as proved by comparing the phylogenetic structure revealed by 16S rRNA sequencing in untreated vs. treated samples. As proof of concept, we also present the first two mouse skin metagenomes, which allowed discovering new taxa (not only prokaryotes but also viruses and eukaryots) not reachable by 16S rRNA sequencing, as well as to characterize the skin microbiome functional landscape. Our method paves the way for the development of skin metagenomics, which will allow a much deeper knowledge of the skin microbiome and its relationship with the host, both in a healthy state and in relation to disease. © 2013 Garcia-Garcerà et al., This work was funded by the Spanish Ministry of Science and Innovation (MICINN)[grant numbers SAF2010-16240 and BFU2009-12895-CO2-01]. MGG was supported by a predoctoral fellowship from the Spanish Ministry of Science and Innovation [Grant number BES-2008-006029].