Background

The human intestinal microbiota is a crucial factor in the pathogenesis of various diseases, such as metabolic syndrome or inflammatory bowel disease (IBD). Yet, knowledge about the role of environmental factors such as smoking (which is known to influence theses aforementioned disease states) on the complex microbial composition is sparse. We aimed to investigate the role of smoking cessation on intestinal microbial composition in 10 healthy smoking subjects undergoing controlled smoking cessation.

Methods

During the observational period of 9 weeks repetitive stool samples were collected. Based on abundance of 16S rRNA genes bacterial composition was analysed and compared to 10 control subjects (5 continuing smokers and 5 non-smokers) by means of Terminal Restriction Fragment Length Polymorphism analysis and high-throughput sequencing.

Results

Profound shifts in the microbial composition after smoking cessation were observed with an increase of Firmicutes and Actinobacteria and a lower proportion of Bacteroidetes and Proteobacteria on the phylum level. In addition, after smoking cessation there was an increase in microbial diversity.

Conclusions

These results indicate that smoking is an environmental factor modulating the composition of human gut microbiota. The observed changes after smoking cessation revealed to be similar to the previously reported differences in obese compared to lean humans and mice respectively, suggesting a potential pathogenetic link between weight gain and smoking cessation. In addition they give rise to a potential association of smoking status and the course of IBD.

Several studies examined the fine-scale structure of human genetic variation in Europe. However, the European sets analyzed represent mainly northern, western, central, and southern Europe. Here, we report an analysis of approximately 166,000 single nucleotide polymorphisms in populations from eastern (northeastern) Europe: four Russian populations from European Russia, and three populations from the northernmost Finno-Ugric ethnicities (Veps and two contrast groups of Komi people). These were compared with several reference European samples, including Finns, Estonians, Latvians, Poles, Czechs, Germans, and Italians. The results obtained demonstrated genetic heterogeneity of populations living in the region studied. Russians from the central part of European Russia (Tver, Murom, and Kursk) exhibited similarities with populations from central–eastern Europe, and were distant from Russian sample from the northern Russia (Mezen district, Archangelsk region). Komi samples, especially Izhemski Komi, were significantly different from all other populations studied. These can be considered as a second pole of genetic diversity in northern Europe (in addition to the pole, occupied by Finns), as they had a distinct ancestry component. Russians from Mezen and the Finnic-speaking Veps were positioned between the two poles, but differed from each other in the proportions of Komi and Finnic ancestries. In general, our data provides a more complete genetic map of Europe accounting for the diversity in its most eastern (northeastern) populations.

Instability in the composition of gut bacterial communities (dysbiosis) has been linked to common human intestinal disorders, such as Crohn’s disease and colorectal cancer. Here, we show that dysbiosis caused by Nod2 deficiency gives rise to a reversible, communicable risk of colitis and colitis-associated carcinogenesis in mice. Loss of either Nod2 or RIP2 resulted in a proinflammatory microenvironment that enhanced epithelial dysplasia following chemically induced injury. The condition could be improved by treatment with antibiotics or an anti–interleukin-6 receptor–neutralizing antibody. Genotype-dependent disease risk was communicable via maternally transmitted microbiota in both Nod2-deficient and WT hosts. Furthermore, reciprocal microbiota transplantation reduced disease risk in Nod2-deficient mice and led to long-term changes in intestinal microbial communities. Conversely, disease risk was enhanced in WT hosts that were recolonized with dysbiotic fecal microbiota from Nod2-deficient mice. Thus, we demonstrated that licensing of dysbiotic microbiota is a critical component of disease risk. Our results demonstrate that NOD2 has an unexpected role in shaping a protective assembly of gut bacterial communities and suggest that manipulation of dysbiosis is a potential therapeutic approach in the treatment of human intestinal disorders.

In most adult patients, hepatitis B is a self-limiting disease leading to life-long protective immunity, which is the consequence of a robust adaptive immune response occurring weeks after HBV infection. Intriguingly, HBV-specific T cells can be detected shortly after infection but the mechanisms underlying this early immune priming and its consequences for subsequent control of viral replication are poorly understood. Using primary human and murine hepatocytes and mouse models of transgenic and adenoviral HBV expression, we show that HBV-expressing hepatocytes produce endoplasmic reticulum (ER)-associated endogenous antigenic lipids including lysophospholipids that are generated by HBV-induced secretory phospholipases and lead to activation of natural killer T (NKT) cells. The absence of NKT cells, CD1d or a defect in ER-associated transfer of lipids onto CD1d results in diminished HBV-specific T and B cell responses and delayed viral control. NKT cells may therefore contribute to control of HBV infection through sensing of HBV-induced modified self-lipids.

Objectives: Using a novel candidate SNP approach, we aimed to identify a possible genetic basis for the higher glioma incidence in Whites relative to East Asians and African-Americans. Methods:  We hypothesized that genetic regions containing SNPs with extreme differences in allele frequencies across ethnicities are most likely to harbor susceptibility variants. We used International HapMap Project data to identify 3,961 candidate SNPs with the largest allele frequency differences in Whites compared to East Asians and Africans and tested these SNPs for association with glioma risk in a set of White cases and controls. Top SNPs identified in the discovery dataset were tested for association with glioma in five independent replication datasets. Results: No SNP achieved statistical significance in either the discovery or replication datasets after accounting for multiple testing or conducting meta-analysis. However, the most strongly associated SNP, rs879471, was found to be in linkage disequilibrium with a previously identified risk SNP, rs6010620, in RTEL1. We estimate rs6010620 to account for a glioma incidence rate ratio of 1.34 for Whites relative to East Asians. Conclusion: We explored genetic susceptibility to glioma using a novel candidate SNP method which may be applicable to other diseases with appropriate epidemiologic patterns.

Psoriatic arthritis (PsA) is a chronic inflammatory musculoskeletal disease affecting up to 30% of psoriasis vulgaris (PsV) cases and approximately 0.25% to 1% of the general population. To identify common susceptibility loci, we performed a meta-analysis of three imputed genome-wide association studies (GWAS) on psoriasis, stratified for PsA. A total of 1,160,703 SNPs were analyzed in the discovery set consisting of 535 PsA cases and 3,432 controls from Germany, the United States and Canada. We followed up two SNPs in 1,931 PsA cases and 6,785 controls comprising six independent replication panels from Germany, Estonia, the United States and Canada. In the combined analysis, a genome-wide significant association was detected at 2p16 near the REL locus encoding c-Rel (rs13017599, P=1.18×10−8, OR=1.27, 95% CI=1.18–1.35). The rs13017599 polymorphism is known to associate with rheumatoid arthritis (RA), and another SNP near REL (rs702873) was recently implicated in PsV susceptibility. However, conditional analysis indicated that rs13017599, rather than rs702873, accounts for the PsA association at REL. We hypothesize that c-Rel, as a member of the Rel/NF-κB family, is associated with PsA in the context of disease pathways that involve other identified PsA and PsV susceptibility genes including TNIP1, TNFAIP3 and NFκBIA.

Background

Genome-wide association studies of two main forms of inflammatory bowel diseases (IBD), Crohn’s disease (CD) and ulcerative colitis (UC), have identified 99 susceptibility loci, but these explain only ∼23% of the genetic risk. Part of the ‘hidden heritability’ could be in transmissible genetic effects in which mRNA expression in the offspring depends on the parental origin of the allele (genomic imprinting), since children whose mothers have CD are more often affected than children with affected fathers. We analyzed parent-of-origin (POO) effects in Dutch and Indian cohorts of IBD patients.

Methods

We selected 28 genetic loci associated with both CD and UC, and tested them for POO effects in 181 Dutch IBD case-parent trios. Three susceptibility variants in NOD2 were tested in 111 CD trios and a significant finding was re-evaluated in 598 German trios. The UC-associated gene, BTNL2, reportedly imprinted, was tested in 70 Dutch UC trios. Finally, we used 62 independent Indian UC trios to test POO effects of five established Indian UC risk loci.

Results

We identified POO effects for NOD2 (L1007fs; OR = 21.0, P-value = 0.013) for CD; these results could not be replicated in an independent cohort (OR = 0.97, P-value = 0.95). A POO effect in IBD was observed for IL12B (OR = 3.2, P-value = 0.019) and PRDM1 (OR = 5.6, P-value = 0.04). In the Indian trios the IL10 locus showed a POO effect (OR = 0.2, P-value = 0.03).

Conclusions

Little is known about the effect of genomic imprinting in complex diseases such as IBD. We present limited evidence for POO effects for the tested IBD loci. POO effects explain part of the hidden heritability for complex genetic diseases but need to be investigated further.

Background

Many hypothesis-driven genetic studies require the ability to comprehensively and efficiently target specific regions of the genome to detect sequence variations. Often, sample availability is limited requiring the use of whole genome amplification (WGA). We evaluated a high-throughput microdroplet-based PCR approach in combination with next generation sequencing (NGS) to target 384 discrete exons from 373 genes involved in cancer. In our evaluation, we compared the performance of six non-amplified gDNA samples from two HapMap family trios. Three of these samples were also preamplified by WGA and evaluated. We tested sample pooling or multiplexing strategies at different stages of the tested targeted NGS (T-NGS) workflow.

Results

The results demonstrated comparable sequence performance between non-amplified and preamplified samples and between different indexing strategies [sequence specificity of 66.0% ± 3.4%, uniformity (coverage at 0.2× of the mean) of 85.6% ± 0.6%]. The average genotype concordance maintained across all the samples was 99.5% ± 0.4%, regardless of sample type or pooling strategy. We did not detect any errors in the Mendelian patterns of inheritance of genotypes between the parents and offspring within each trio. We also demonstrated the ability to detect minor allele frequencies within the pooled samples that conform to predicted models.

Conclusion

Our described PCR-based sample multiplex approach and the ability to use WGA material for NGS may enable researchers to perform deep resequencing studies and explore variants at very low frequencies and cost.

The bivalve Arctica islandica is extremely long lived (>400 years) and can tolerate long periods of hypoxia and anoxia. European populations differ in maximum life spans (MLSP) from 40 years in the Baltic to >400 years around Iceland. Characteristic behavior of A. islandica involves phases of metabolic rate depression (MRD) during which the animals burry into the sediment for several days. During these phases the shell water oxygen concentrations reaches hypoxic to anoxic levels, which possibly support the long life span of some populations. We investigated gene regulation in A. islandica from a long-lived (MLSP 150 years) German Bight population and the short-lived Baltic Sea population, experimentally exposed to different oxygen levels. A new A. islandica transcriptome enabled the identification of genes important during hypoxia/anoxia events and, more generally, gene mining for putative stress response and (anti-) aging genes. Expression changes of a) antioxidant defense: Catalase, Glutathione peroxidase, manganese and copper-zinc Superoxide dismutase; b) oxygen sensing and general stress response: Hypoxia inducible factor alpha, Prolyl hydroxylase and Heat-shock protein 70; and c) anaerobic capacity: Malate dehydrogenase and Octopine dehydrogenase, related transcripts were investigated. Exposed to low oxygen, German Bight individuals suppressed transcription of all investigated genes, whereas Baltic Sea bivalves enhanced gene transcription under anoxic incubation (0 kPa) and, further, decreased these transcription levels again during 6 h of re-oxygenation. Hypoxic and anoxic exposure and subsequent re-oxygenation in Baltic Sea animals did not lead to increased protein oxidation or induction of apoptosis, emphasizing considerable hypoxia/re-oxygenation tolerance in this species. The data suggest that the energy saving effect of MRD may not be an attribute of Baltic Sea A. islandica chronically exposed to high environmental variability (oxygenation, temperature, salinity). Contrary, higher physiological flexibility and stress hardening may predispose these animals to perform a pronounced stress response at the expense of life span.

Scientists working with single-nucleotide variants (SNVs), inferred by next-generation sequencing software, often need further information regarding true variants, artifacts and sequence coverage gaps. In clinical diagnostics, e.g. SNVs must usually be validated by visual inspection or several independent SNV-callers. We here demonstrate that 0.5–60% of relevant SNVs might not be detected due to coverage gaps, or might be misidentified. Even low error rates can overwhelm the true biological signal, especially in clinical diagnostics, in research comparing healthy with affected cells, in archaeogenetic dating or in forensics. For these reasons, we have developed a package called pibase, which is applicable to diploid and haploid genome, exome or targeted enrichment data. pibase extracts details on nucleotides from alignment files at user-specified coordinates and identifies reproducible genotypes, if present. In test cases pibase identifies genotypes at 99.98% specificity, 10-fold better than other tools. pibase also provides pair-wise comparisons between healthy and affected cells using nucleotide signals (10-fold more accurately than a genotype-based approach, as we show in our case study of monozygotic twins). This comparison tool also solves the problem of detecting allelic imbalance within heterozygous SNVs in copy number variation loci, or in heterogeneous tumor sequences.

Background & Aims

A limited number of genetic risk factors have been reported in primary sclerosing cholangitis (PSC). To discover further genetic susceptibility factors for PSC, we followed up on a second tier of single nucleotide polymorphisms (SNPs) from a genome-wide association study (GWAS).

Methods

We analyzed 45 SNPs in 1221 PSC cases and 3508 controls. The association results from the replication analysis and the original GWAS (715 PSC cases and 2962 controls) were combined in a meta-analysis comprising 1936 PSC cases and 6470 controls. We performed an analysis of bile microbial community composition in 39 PSC patients by 16S rRNA sequencing.

Results

Seventeen SNPs representing 12 distinct genetic loci achieved nominal significance (Preplication<0.05) in the replication. The most robust novel association was detected at chromosome 1p36 (rs3748816; Pcombined=2.1×10−8) where the MMEL1 and TNFRSF14 genes represent potential disease genes. Eight additional novel loci showed suggestive evidence of association (Prepl<0.05). FUT2 at chromosome 19q13 (rs602662; Pcomb=1.9×10−6, rs281377; Pcomb = 2.1×10−6 and rs601338; Pcomb=2.7×10−6) is notable due to its implication in altered susceptibility to infectious agents. We found that FUT2 secretor status and genotype defined by rs601338 significantly influences biliary microbial community composition in PSC patients.

Conclusions

We identify multiple new PSC risk loci by extended analysis of a PSC GWAS. FUT2 genotype needs to be taken into account when assessing the influence from microbiota on biliary pathology in PSC.

Background

Compared to classical genotyping, targeted next-generation sequencing (tNGS) can be custom-designed to interrogate entire genomic regions of interest, in order to detect novel as well as known variants. To bring down the per-sample cost, one approach is to pool barcoded NGS libraries before sample enrichment. Still, we lack a complete understanding of how this multiplexed tNGS approach and the varying performance of the ever-evolving analytical tools can affect the quality of variant discovery. Therefore, we evaluated the impact of different software tools and analytical approaches on the discovery of single nucleotide polymorphisms (SNPs) in multiplexed tNGS data. To generate our own test model, we combined a sequence capture method with NGS in three experimental stages of increasing complexity (E. coli genes, multiplexed E. coli, and multiplexed HapMap BRCA1/2 regions).

Results

We successfully enriched barcoded NGS libraries instead of genomic DNA, achieving reproducible coverage profiles (Pearson correlation coefficients of up to 0.99) across multiplexed samples, with <10% strand bias. However, the SNP calling quality was substantially affected by the choice of tools and mapping strategy. With the aim of reducing computational requirements, we compared conventional whole-genome mapping and SNP-calling with a new faster approach: target-region mapping with subsequent ‘read-backmapping’ to the whole genome to reduce the false detection rate. Consequently, we developed a combined mapping pipeline, which includes standard tools (BWA, SAMtools, etc.), and tested it on public HiSeq2000 exome data from the 1000 Genomes Project. Our pipeline saved 12 hours of run time per Hiseq2000 exome sample and detected ~5% more SNPs than the conventional whole genome approach. This suggests that more potential novel SNPs may be discovered using both approaches than with just the conventional approach.

Conclusions

We recommend applying our general ‘two-step’ mapping approach for more efficient SNP discovery in tNGS. Our study has also shown the benefit of computing inter-sample SNP-concordances and inspecting read alignments in order to attain more confident results.

Background

The purpose of this study was to examine the differing perspectives and perceptual gaps relating to ulcerative colitis (UC) symptoms and their management between patients and healthcare professionals (HCPs).

Methods

Structured, cross-sectional, Web-based questionnaires designed to assess a variety of disease indices were completed by adult patients with UC and HCPs involved in the care of patients with UC from Canada, France, Germany, Ireland, Spain, and the United Kingdom.

Results

Surveys were completed by 775 patients, 475 physicians, and 50 nurses. Patient self-reported classification of disease severity revealed generally greater severity (mild, 32%; moderate, 53%) compared with physician and nurse estimates of UC severity among their caseloads (mild, 52% and 49%; moderate, 34% and 37%, respectively). Patients reported that an average of 5.5 (standard deviation, 11.0) flares (self-defined) occurred over the past year, compared with 3.4 and 3.8 flares per year estimated by physicians and nurses. Perceived flare triggers differed between patients (stress ranked first) and HCPs (natural disease course ranked first). Fifty-five percent of patients stated that UC symptoms over the past year had affected their quality of life, while physicians and nurses estimated that 35% to 37% of patients would have a reduced quality of life over the same period. Patients ranked urgency and pain as the most bothersome symptoms, while physicians and nurses ranked urgency and stool frequency highest. About half of patients (47%) defined remission as experiencing no symptoms; by comparison, 62% to 63% of HCPs defined remission as requiring the complete absence of symptoms. HCPs (doctors/nurses in general practice and/or hospital) were regarded by patients as their main source of UC information by 72%; however, 59% reported not arranging regular visits to see their HCPs.

Conclusions

This large survey identified important differences between patients' and HCPs' perceptions of the impact of UC symptoms on patients' lives. Notably, HCPs may underestimate the effect of specific UC symptoms on patients and may fail to recognize issues that are important to patients.

The gene tyrosine hydroxylase 1 (TH01) has been suggested as a candidate for human longevity. A previous study has shown an association between longevity and specific alleles of the TH01 short tandem repeat (STR) polymorphism in an Italian population. This STR locus is also widely used in forensic genetics. If the TH01–longevity association could be confirmed in independent samples, this finding would have important ramifications for the use of this polymorphism in a forensic context. In the present study, we sought to replicate the previous association result by investigating 471 long-lived individuals (96–110 years) and 462 younger controls (19–75 years) from Germany. In the analyzed samples, the association between TH01 and longevity was not replicated. However, the obtained TH01 allele frequencies were consistent with published data. We observed considerable differences in the allele distribution between Germans and Italians, in particular with regard to allele 9.3, which displayed a previously undetected decreasing West–East and North–South cline across Europe. The discrepant TH01–longevity association results in Germans and Italians could therefore be due to population-specific effects. This finding highlights the need to take into consideration population genetic data when dealing with association studies.

While gliomas are the most common primary brain tumors, their etiology is largely unknown. To identify novel risk loci for glioma, we conducted genome-wide association (GWA) analysis of two case–control series from France and Germany (2269 cases and 2500 controls). Pooling these data with previously reported UK and US GWA studies provided data on 4147 glioma cases and 7435 controls genotyped for 424 460 common tagging single-nucleotide polymorphisms. Using these data, we demonstrate two statistically independent associations between glioma and rs11979158 and rs2252586, at 7p11.2 which encompasses the EGFR gene (population-corrected statistics, Pc = 7.72 × 10−8 and 2.09 × 10−8, respectively). Both associations were independent of tumor subtype, and were independent of EGFR amplification, p16INK4a deletion and IDH1 mutation status in tumors; compatible with driver effects of the variants on glioma development. These findings show that variation in 7p11.2 is a determinant of inherited glioma risk.

The fatty-acid-binding protein-2 (FABP2) gene has been proposed as a candidate gene for diabetes because the encoded protein is involved in fatty acid absorption and therefore may affect insulin sensitivity and glucose metabolism. The rare haplotype (B) of its promoter was shown to be associated with a lower risk for type 2 diabetes. The aim of this study was to investigate whether a polymorphism in the FABP2 promoter does affect the metabolic response to either an medium-chain triacylglycerol (MCT) or an long-chain triacylglycerol (LCT) diet, which were suggested to differ in transport mechanisms, in affinity to FABP2, in activating transcription factors binding to the FABP2 promoter and in their effects on insulin sensitivity. We studied 82 healthy male subjects varying in the FABP2 promoter (42 homozygous for common haplotype (A), 40 homozygous for the rare haplotype (B)) in an interventional study with either an MCT or LCT diet over 2 weeks to examine gene–nutrient interaction. The saturation grade of MCT was adjusted to that of the LCT fat. We determined glucose, insulin, triacylglycerols (TGs), chylomicron triacylglycerols and cholesterol before and after a standardised mixed meal before and after the intervention. HDL cholesterol increased in all groups, which was most pronounced in subjects homozygous for the common promoter haplotype A who received MCT diet (P = 0.001), but not significant in homozygous rare haplotype B subjects who received MCT fat. Subjects homozygous for FABP2 haplotype A showed a significant decrease in fasting and postprandial glucose (P = 0.01, 0.04, respectively) and a decrease in insulin resistance (HOMA-IR, P = 0.04) during LCT diet. After correction for multiple testing, those effects did not remain significant. Fasting and postprandial triacylglycerols, LDL cholesterol, chylomicron TGs and cholesterol were not affected by genotype or diet. MCT diet increased HDL cholesterol dependent on the FABP2 promoter haplotype. The effects of the promoter haplotype B could be mediated by PPARγ, which is upregulated by medium-chain fatty acids.

We have validated the association of two genes on chromosome 20q13.31–33 with tuberculosis susceptibility. A previous genome-wide linkage study performed by Cooke et al identified the genes melanocortin-3-receptor (MC3R) and cathepsin Z (CTSZ) as possible candidates in tuberculosis susceptibility. MC3R has been implicated in obesity studies and is known to play a role in many biological systems including the regulation of energy homeostasis and fat metabolism. CTSZ has been detected in immune cells, such as macrophages and monocytes, and it is hypothesized that the protein may play a role in the immune response. In our South African population a case–control study confirmed the previously reported association with a single-nucleotide polymorphism (SNP) in CTSZ and found an association in MC3R with a SNP not previously implicated in tuberculosis susceptibility. Six SNPs in MC3R and eight in CTSZ were genotyped and haplotypes were inferred. SNP rs6127698 in the promoter region of MC3R (cases=498; controls=506) and rs34069356 in the 3′UTR of CTSZ (cases=396; controls=298) both showed significant association with tuberculosis susceptibility (P=0.0004 and <0.0001, respectively), indicating that pathways involving these proteins, not previously researched in this disease, could yield novel therapies for tuberculosis.

Primary sclerosing cholangitis (PSC) is a chronic cholestatic liver disease characterized by inflammation and fibrosis of the bile ducts. Both environmental and genetic factors contribute to its pathogenesis. To further clarify its genetic background, we investigated susceptibility loci recently identified for ulcerative colitis (UC) in a large cohort of 1186PSC patients and 1748 controls. Single nucleotide polymorphisms (SNPs) tagging 13 UC susceptibility loci were initially genotyped in 854 PSC patients and 1491 controls from the Benelux (331 cases, 735 controls), Germany (265 cases, 368 controls) and Scandinavia (258 cases, 388 controls). Subsequently, a joint analysis was performed with an independent second Scandinavian cohort (332 cases, 257 controls). SNPs at chromosomes2p16 (p value 4.12×10−4), 4q27 (p value 4.10×10−5) and 9q34 (p value 8.41×10−4) were associated with PSC in the joint analysis after correcting for multiple testing. In PSC patients without inflammatory bowel disease(IBD), SNPs at 4q27and9q34 were nominally associated (p<0.05). We applied additional in silico analyses to identify likely candidate genes at PSC susceptibility loci. To identify non-random, evidence-based links we used GRAIL analysis showing interconnectivity between genes in six out of in total nine PSC-associated regions. Expression quantitative trait analysis from 1469 Dutch and UK individuals demonstrated that five out of nine SNPs had an effect on cis-gene expression. These analyses prioritized IL2, CARD9 and REL as novel candidates.

Conclusion

We have identified three UC susceptibility loci to be associated with PSC, harboring the putative candidate genes REL, IL2 and CARD9. These results add to the scarce knowledge on the genetic background of PSC and imply an important role for both innate and adaptive immunological factors.

Background

Currently, the TNM classification of malignant tumours based on clinicopathological staging remains the standard for colorectal cancer (CRC) prognostication. Recently, we identified the mitochondrial oxidative phosphorylation chain as a consistently overrepresented category in the published gene expression profiling (GEP) studies on CRC prognosis.

Methods

We evaluated associations of putative regulatory single nucleotide polymorphisms (SNPs) in genes from the oxidative phosphorylation chain with survival and disease prognosis in 613 CRC patients from Northern Germany (PopGen cohort).

Results

Two SNPs in the 3′ untranslated region of UQCRB (complex III), rs7836698 and rs10504961, were associated with overall survival (HR = 0.52, 95% CI 0.32–0.85 and HR = 0.64, 95% CI 0.42–0.99, for TT carriers). These associations were restricted to the group of patients with cancer located in the colon (HR = 0.42, 95% CI 0.22–0.82 and HR = 0.46, 95% CI 0.25–0.83). Multivariate analysis indicated that both markers might act as independent prognostic markers. Additionally, the TT carriers were ~2 times more likely to develop tumours in the colon than in the rectum. Two SNPs in COX6B1 (complex IV) were associated with lymph node metastasis in a dominant model (rs6510502, OR = 1.75, 95% CI 1.20–2.57; rs10420252, OR = 1.68, 95% CI 1.11–2.53); rs6510502 was associated also with distant metastasis (OR = 1.67, 95% CI 1.09–2.56 in a dominant model).

Conclusions

This is the first report suggesting that markers in genes from the mitochondrial oxidative chain might be prognostic factors for CRC. Additional studies replicating the presented findings are needed.

Major depression (MD) is one of the most prevalent psychiatric disorders and a leading cause of loss in work productivity. A combination of genetic and environmental risk factors likely contributes to MD. We present data from a genome-wide association study revealing a neuron-specific neutral amino acid transporter (SLC6A15) as a novel susceptibility gene for MD. Risk allele carrier status in humans and chronic stress in mice were associated with a downregulation of the expression of this gene in the hippocampus, a brain region implicated in the pathophysiology of MD. The same polymorphisms also showed associations with alterations in hippocampal volume and neuronal integrity. Thus, decreased SLC6A15 expression, due to genetic or environmental factors might alter neuronal circuits related to the susceptibility for MD. Our convergent data from human genetics, expression studies, brain imaging and animal models suggest a novel pathophysiological mechanism for MD that may be accessible to drug targeting.

Background

Recent genome-wide association studies (GWAS) have identified several novel loci that reproducibly associate with CAD and/or MI risk. However, known common CAD risk variants explain only 10% of the predicted genetic heritability of the disease, suggesting that important genetic signals remain to be discovered.

Methods and Results

We performed a discovery meta-analysis of 5 GWASs involving 13,949 subjects (7123 cases, 6826 controls) imputed at approximately 5 million SNPs using pilot 1000 Genomes based haplotypes. Promising loci were followed up in an additional 5 studies with 11,032 subjects (5211 cases, 5821 controls). A novel CAD locus on chromosome 6p21.3 in the major histocompatibility complex (MHC) between HCG27 and HLA-C was identified and achieved genome wide significance in the combined analysis (rs3869109; pdiscovery=3.3×10−7, preplication=5.3×10−4 pcombined=1.12×10−9). A sub-analysis combining discovery GWASs showed an attenuation of significance when stringent corrections for European population structure were employed (p=4.1×10-10 versus 3.2×10-7) suggesting the observed signal is partly confounded due to population stratification. This gene dense region plays an important role in inflammation, immunity and self cell recognition. To determine whether the underlying association was driven by MHC class I alleles, we statistically imputed common HLA alleles into the discovery subjects; however, no single common HLA type contributed significantly or fully explained the observed association.

Conclusions

We have identified a novel locus in the MHC associated with CAD. MHC genes regulate inflammation and T cell responses that contribute importantly to the initiation and propagation of atherosclerosis. Further laboratory studies will be required to understand the biological basis of this association and identify the causative allele(s).

Background

Recent genome-wide association studies (GWAS) have identified several novel loci that reproducibly associate with CAD and/or MI risk. However, known common CAD risk variants explain only 10% of the predicted genetic heritability of the disease, suggesting that important genetic signals remain to be discovered.

Methods and Results

We performed a discovery meta-analysis of 5 GWASs involving 13,949 subjects (7123 cases, 6826 controls) imputed at approximately 5 million SNPs using pilot 1000 Genomes based haplotypes. Promising loci were followed up in an additional 5 studies with 11,032 subjects (5211 cases, 5821 controls). A novel CAD locus on chromosome 6p21.3 in the major histocompatibility complex (MHC) between HCG27 and HLA-C was identified and achieved genome wide significance in the combined analysis (rs3869109; pdiscovery=3.3×10−7, preplication=5.3×10−4 pcombined=1.12×10−9). A sub-analysis combining discovery GWASs showed an attenuation of significance when stringent corrections for European population structure were employed (p=4.1×10−10 versus 3.2×10−7) suggesting the observed signal is partly confounded due to population stratification. This gene dense region plays an important role in inflammation, immunity and self cell recognition. To determine whether the underlying association was driven by MHC class I alleles, we statistically imputed common HLA alleles into the discovery subjects; however, no single common HLA type contributed significantly or fully explained the observed association.

Conclusion

We have identified a novel locus in the MHC associated with CAD. MHC genes regulate inflammation and T cell responses that contribute importantly to the initiation and propagation of atherosclerosis. Further laboratory studies will be required to understand the biological basis of this association and identify the causative allele(s).

Background

Antibiotic associated diarrhea and Clostridium difficile infection are frequent complications of broad spectrum antibiotic therapy. Probiotic bacteria are used as therapeutic and preventive agents in these disorders, but the exact functional mechanisms and the mode of action are poorly understood. The effects of clindamycin and the probiotic mixture VSL#3 (containing the 8 bacterial strains Streptococcus thermophilus, Bifidobacterium breve, Bifidobacterium longum, Bifidobacterium infantis, Lactobacillus acidophilus, Lactobacillus plantarum, Lactobacillus paracasei and Lactobacillus delbrueckii subsp. Bulgaricus) consecutively or in combination were investigated and compared to controls without therapy using a standardized human fecal microbiota in a computer-controlled in vitro model of large intestine. Microbial metabolites (short chain fatty acids, lactate, branched chain fatty acids, and ammonia) and the intestinal microbiota were analyzed.

Results

Compared to controls and combination therapy, short chain fatty acids and lactate, but also ammonia and branched chain fatty acids, were increased under probiotic therapy. The metabolic pattern under combined therapy with antibiotics and probiotics had the most beneficial and consistent effect on intestinal metabolic profiles. The intestinal microbiota showed a decrease in several indigenous bacterial groups under antibiotic therapy, there was no significant recovery of these groups when the antibiotic therapy was followed by administration of probiotics. Simultaneous application of anti- and probiotics had a stabilizing effect on the intestinal microbiota with increased bifidobacteria and lactobacilli.

Conclusions

Administration of VSL#3 parallel with the clindamycin therapy had a beneficial and stabilizing effect on the intestinal metabolic homeostasis by decreasing toxic metabolites and protecting the endogenic microbiota from destruction. Probiotics could be a reasonable strategy in prevention of antibiotic associated disturbances of the intestinal homeostasis and disorders.

The marine mussel Mytilus edulis and its closely related sister species are distributed world-wide and play an important role in coastal ecology and economy. The diversification in different species and their hybrids, broad ecological distribution, as well as the filter feeding mode of life has made this genus an attractive model to investigate physiological and molecular adaptations and responses to various biotic and abiotic environmental factors. In the present study we investigated the immune system of Mytilus, which may contribute to the ecological plasticity of this species. We generated a large Mytilus transcriptome database from different tissues of immune challenged and stress treated individuals from the Baltic Sea using 454 pyrosequencing. Phylogenetic comparison of orthologous groups of 23 species demonstrated the basal position of lophotrochozoans within protostomes. The investigation of immune related transcripts revealed a complex repertoire of innate recognition receptors and downstream pathway members including transcripts for 27 toll-like receptors and 524 C1q domain containing transcripts. NOD-like receptors on the other hand were absent. We also found evidence for sophisticated TNF, autophagy and apoptosis systems as well as for cytokines. Gill tissue and hemocytes showed highest expression of putative immune related contigs and are promising tissues for further functional studies. Our results partly contrast with findings of a less complex immune repertoire in ecdysozoan and other lophotrochozoan protostomes. We show that bivalves are interesting candidates to investigate the evolution of the immune system from basal metazoans to deuterostomes and protostomes and provide a basis for future molecular work directed to immune system functioning in Mytilus.

Background

Genetic variation in NOD2 and cigarette smoking are well-established risk factors for the development of Crohn's disease (CD). However, little is known about a potential interaction between these risk factors. We investigated gene-environment interactions between CD-associated NOD2 alleles and cigarette smoking in a large sample of patients with CD.

Methods

Three previously reported CD-associated variants in NOD2 (R702W, G908R, 1007fs) were genotyped in 1636 patients with CD continuously recruited between 1995 and 2010 based on physician referral. Data on history of smoking behaviour was obtained for all participants through a written questionnaire. Using a case-only design, we performed logistic regression analyses to investigate statistical interactions between NOD2 risk alleles and smoking status.

Results

We detected a significant negative interaction between carriership of at least one of the NOD2 risk alleles and history of ever having smoked (OR = 0.71; p = 0.005) as well as smoking at the time of CD diagnosis (OR = 0.68; p = 0.005). Subsequent separate analyses of the three variants revealed a significant negative interaction between the 1007fs variant and history of ever having smoked (OR = 0.64; p = 9 × 10-4) and smoking at the time of CD diagnosis (OR = 0.53; p = 7 × 10-5).

Conclusions

The observed significant negative gene-environment interaction suggests that the risk increase for CD conferred simultaneously by cigarette smoking and the 1007fs NOD2 polymorphism is smaller than expected and may point to a biological interaction. Our findings warrant further investigation in epidemiological and functional studies to elucidate pathophysiology as well as to aid in the development of recommendations for disease prevention.