Richness of human gut microbiome correlates with metabolic markers.

We are facing a global metabolic health crisis provoked by an obesity epidemic. Here we report the human gut microbial composition in a population sample of 123 non-obese and 169 obese Danish individuals. We find two groups of individuals that differ by the number of gut microbial genes and thus gut bacterial richness. They contain known and previously unknown bacterial species at different proportions; individuals with a low bacterial richness (23% of the population) are characterized by more marked overall adiposity, insulin resistance and dyslipidaemia and a more pronounced inflammatory phenotype when compared with high bacterial richness individuals. The obese individuals among the lower bacterial richness group also gain more weight over time. Only a few bacterial species are sufficient to distinguish between individuals with high and low bacterial richness, and even between lean and obese participants. Our classifications based on variation in the gut microbiome identify subsets of individuals in the general white adult population who may be at increased risk of progressing to adiposity-associated co-morbidities.

Enterotypes of the human gut microbiome.

Our knowledge of species and functional composition of the human gut microbiome is rapidly increasing, but it is still based on very few cohorts and little is known about variation across the world. By combining 22 newly sequenced faecal metagenomes of individuals from four countries with previously published data sets, here we identify three robust clusters (referred to as enterotypes hereafter) that are not nation or continent specific. We also confirmed the enterotypes in two published, larger cohorts, indicating that intestinal microbiota variation is generally stratified, not continuous. This indicates further the existence of a limited number of well-balanced host-microbial symbiotic states that might respond differently to diet and drug intake. The enterotypes are mostly driven by species composition, but abundant molecular functions are not necessarily provided by abundant species, highlighting the importance of a functional analysis to understand microbial communities. Although individual host properties such as body mass index, age, or gender cannot explain the observed enterotypes, data-driven marker genes or functional modules can be identified for each of these host properties. For example, twelve genes significantly correlate with age and three functional modules with the body mass index, hinting at a diagnostic potential of microbial markers.

A human gut microbial gene catalogue established by metagenomic sequencing.

To understand the impact of gut microbes on human health and well-being it is crucial to assess their genetic potential. Here we describe the Illumina-based metagenomic sequencing, assembly and characterization of 3.3 million non-redundant microbial genes, derived from 576.7 gigabases of sequence, from faecal samples of 124 European individuals. The gene set, approximately 150 times larger than the human gene complement, contains an overwhelming majority of the prevalent (more frequent) microbial genes of the cohort and probably includes a large proportion of the prevalent human intestinal microbial genes. The genes are largely shared among individuals of the cohort. Over 99% of the genes are bacterial, indicating that the entire cohort harbours between 1,000 and 1,150 prevalent bacterial species and each individual at least 160 such species, which are also largely shared. We define and describe the minimal gut metagenome and the minimal gut bacterial genome in terms of functions present in all individuals and most bacteria, respectively.

Ancient human genome sequence of an extinct Palaeo-Eskimo.

We report here the genome sequence of an ancient human. Obtained from approximately 4,000-year-old permafrost-preserved hair, the genome represents a male individual from the first known culture to settle in Greenland. Sequenced to an average depth of 20x, we recover 79% of the diploid genome, an amount close to the practical limit of current sequencing technologies. We identify 353,151 high-confidence single-nucleotide polymorphisms (SNPs), of which 6.8% have not been reported previously. We estimate raw read contamination to be no higher than 0.8%. We use functional SNP assessment to assign possible phenotypic characteristics of the individual that belonged to a culture whose location has yielded only trace human remains. We compare the high-confidence SNPs to those of contemporary populations to find the populations most closely related to the individual. This provides evidence for a migration from Siberia into the New World some 5,500 years ago, independent of that giving rise to the modern Native Americans and Inuit.

Sugarcane genome sequencing by methylation filtration provides tools for genomic research in the genus Saccharum.

Many economically important crops have large and complex genomes that hamper their sequencing by standard methods such as whole genome shotgun (WGS). Large tracts of methylated repeats occur in plant genomes that are interspersed by hypomethylated gene-rich regions. Gene-enrichment strategies based on methylation profiles offer an alternative to sequencing repetitive genomes. Here, we have applied methyl filtration with McrBC endonuclease digestion to enrich for euchromatic regions in the sugarcane genome. To verify the efficiency of methylation filtration and the assembly quality of sequences submitted to gene-enrichment strategy, we have compared assemblies using methyl-filtered (MF) and unfiltered (UF) libraries. The use of methy filtration allowed a better assembly by filtering out 35% of the sugarcane genome and by producing 1.5× more scaffolds and 1.7× more assembled Mb in length compared with unfiltered dataset. The coverage of sorghum coding sequences (CDS) by MF scaffolds was at least 36% higher than by the use of UF scaffolds. Using MF technology, we increased by 134× the coverage of gene regions of the monoploid sugarcane genome. The MF reads assembled into scaffolds that covered all genes of the sugarcane bacterial artificial chromosomes (BACs), 97.2% of sugarcane expressed sequence tags (ESTs), 92.7% of sugarcane RNA-seq reads and 98.4% of sorghum protein sequences. Analysis of MF scaffolds from encoded enzymes of the sucrose/starch pathway discovered 291 single-nucleotide polymorphisms (SNPs) in the wild sugarcane species, S. spontaneum and S. officinarum. A large number of microRNA genes was also identified in the MF scaffolds. The information achieved by the MF dataset provides a valuable tool for genomic research in the genus Saccharum and for improvement of sugarcane as a biofuel crop.

A comparative analysis of the intestinal metagenomes present in guinea pigs (Cavia porcellus) and humans (Homo sapiens).

BACKGROUND: Guinea pig (Cavia porcellus) is an important model for human intestinal research. We have characterized the faecal microbiota of 60 guinea pigs using Illumina shotgun metagenomics, and used this data to compile a gene catalogue of its prevalent microbiota. Subsequently, we compared the guinea pig microbiome to existing human gut metagenome data from the MetaHIT project. RESULTS: We found that the bacterial richness obtained for human samples was lower than for guinea pig samples. The intestinal microbiotas of both species were dominated by the two phyla Bacteroidetes and Firmicutes, but at genus level, the majority of identified genera (320 of 376) were differently abundant in the two hosts. For example, the guinea pig contained considerably more of the mucin-degrading Akkermansia, as well as of the methanogenic archaea Methanobrevibacter than found in humans. Most microbiome functional categories were less abundant in guinea pigs than in humans. Exceptions included functional categories possibly reflecting dehydration/rehydration stress in the guinea pig intestine. Finally, we showed that microbiological databases have serious anthropocentric biases, which impacts model organism research. CONCLUSIONS: The results lay the foundation for future gastrointestinal research applying guinea pigs as models for humans.

Twenty-eight divergent polysaccharide loci specifying within- and amongst-strain capsule diversity in three strains of Bacteroides fragilis.

Comparison of the complete genome sequence of Bacteroides fragilis 638R, originally isolated in the USA, was made with two previously sequenced strains isolated in the UK (NCTC 9343) and Japan (YCH46). The presence of 10 loci containing genes associated with polysaccharide (PS) biosynthesis, each including a putative Wzx flippase and Wzy polymerase, was confirmed in all three strains, despite a lack of cross-reactivity between NCTC 9343 and 638R surface PS-specific antibodies by immunolabelling and microscopy. Genomic comparisons revealed an exceptional level of PS biosynthesis locus diversity. Of the 10 divergent PS-associated loci apparent in each strain, none is similar between NCTC 9343 and 638R. YCH46 shares one locus with NCTC 9343, confirmed by mAb labelling, and a second different locus with 638R, making a total of 28 divergent PS biosynthesis loci amongst the three strains. The lack of expression of the phase-variable large capsule (LC) in strain 638R, observed in NCTC 9343, is likely to be due to a point mutation that generates a stop codon within a putative initiating glycosyltransferase, necessary for the expression of the LC in NCTC 9343. Other major sequence differences were observed to arise from different numbers and variety of inserted extra-chromosomal elements, in particular prophages. Extensive horizontal gene transfer has occurred within these strains, despite the presence of a significant number of divergent DNA restriction and modification systems that act to prevent acquisition of foreign DNA. The level of amongst-strain diversity in PS biosynthesis loci is unprecedented.

Complete genome sequence of the sugarcane nitrogen-fixing endophyte Gluconacetobacter diazotrophicus Pal5.

BACKGROUND: Gluconacetobacter diazotrophicus Pal5 is an endophytic diazotrophic bacterium that lives in association with sugarcane plants. It has important biotechnological features such as nitrogen fixation, plant growth promotion, sugar metabolism pathways, secretion of organic acids, synthesis of auxin and the occurrence of bacteriocins. RESULTS: Gluconacetobacter diazotrophicus Pal5 is the third diazotrophic endophytic bacterium to be completely sequenced. Its genome is composed of a 3.9 Mb chromosome and 2 plasmids of 16.6 and 38.8 kb, respectively. We annotated 3,938 coding sequences which reveal several characteristics related to the endophytic lifestyle such as nitrogen fixation, plant growth promotion, sugar metabolism, transport systems, synthesis of auxin and the occurrence of bacteriocins. Genomic analysis identified a core component of 894 genes shared with phylogenetically related bacteria. Gene clusters for gum-like polysaccharide biosynthesis, tad pilus, quorum sensing, for modulation of plant growth by indole acetic acid and mechanisms involved in tolerance to acidic conditions were identified and may be related to the sugarcane endophytic and plant-growth promoting traits of G. diazotrophicus. An accessory component of at least 851 genes distributed in genome islands was identified, and was most likely acquired by horizontal gene transfer. This portion of the genome has likely contributed to adaptation to the plant habitat. CONCLUSION: The genome data offer an important resource of information that can be used to manipulate plant/bacterium interactions with the aim of improving sugarcane crop production and other biotechnological applications.

Protein expression profile of Gluconacetobacter diazotrophicus PAL5, a sugarcane endophytic plant growth-promoting bacterium.

This is the first broad proteomic description of Gluconacetobacter diazotrophicus, an endophytic bacterium, responsible for the major fraction of the atmospheric nitrogen fixed in sugarcane in tropical regions. Proteomic coverage of G. diazotrophicus PAL5 was obtained by two independent approaches: 2-DE followed by MALDI-TOF or TOF-TOF MS and 1-DE followed by chromatography in a C18 column online coupled to an ESI-Q-TOF or ESI-IT mass spectrometer. The 583 identified proteins were sorted into functional categories and used to describe potential metabolic pathways for nucleotides, amino acids, carbohydrates, lipids, cofactors and energy production, according to the Enzyme Commission of Enzyme Nomenclature (EC) and Kyoto Encyclopedia of genes and genomes (KEGG) databases. The identification of such proteins and their possible insertion in conserved biochemical routes will allow comparisons between G. diazotrophicus and other bacterial species. Furthermore, the 88 proteins classified as conserved unknown or unknown constitute a potential target for functional genomic studies, aiming at the understanding of protein function and regulation of gene expression. The knowledge of metabolic fundamentals and coordination of these actions are crucial for the rational, safe and sustainable interference on crops. The entire dataset, including peptide sequence information, is available as Supporting Information and is the major contribution of this work.

Whole transcriptome expression of trigeminal ganglia compared to dorsal root ganglia in Rattus Norvegicus.

The trigeminal ganglia (TG) subserving the head and the dorsal root ganglia (DRG) subserving the rest of the body are homologous handling sensory neurons. Differences exist, as a number of signaling substances cause headache but no pain in the rest of the body. To date, very few genes involved in this difference have been identified. We aim to reveal basal gene expression levels in TG and DRG and detect genes that are differentially expressed (DE) between TG and DRG. RNA-Sequencing from six naïve rats describes the whole transcriptome expression profiles of TG and DRG. Differential expression analysis was followed by pathway analysis to identify DE processes between TG and DRG. In total, 64 genes had higher and 55 genes had lower expressed levels in TG than DRG. Higher expressed genes, including S1pr5 and Gjc2, have been related to phospholipase activity. The lower expressed genes, including several Hox genes and Slc5a7, have been related to tyrosine and phenylalanine metabolism. Tissue-specific expression was identified for Gabra6 and Gabrd in TG, and for several Hox genes in DRG. Furthermore, genes that were known to be associated with headache/migraine were mostly moderately to highly expressed in one or both tissues. We present a comprehensive overview of the expression profiles of whole tissue comparison of TG and DRG. Further, we showed DE genes/pathways between TG and DRG, including several known migraine-associated genes. This study provides a basis for further pain-related studies using TG and DRG in rats.

Contribution of copy number variants to schizophrenia from a genome-wide study of 41,321 subjects.

Copy number variants (CNVs) have been strongly implicated in the genetic etiology of schizophrenia (SCZ). However, genome-wide investigation of the contribution of CNV to risk has been hampered by limited sample sizes. We sought to address this obstacle by applying a centralized analysis pipeline to a SCZ cohort of 21,094 cases and 20,227 controls. A global enrichment of CNV burden was observed in cases (odds ratio (OR) = 1.11, P = 5.7 × 10-15), which persisted after excluding loci implicated in previous studies (OR = 1.07, P = 1.7 × 10-6). CNV burden was enriched for genes associated with synaptic function (OR = 1.68, P = 2.8 × 10-11) and neurobehavioral phenotypes in mouse (OR = 1.18, P = 7.3 × 10-5). Genome-wide significant evidence was obtained for eight loci, including 1q21.1, 2p16.3 (NRXN1), 3q29, 7q11.2, 15q13.3, distal 16p11.2, proximal 16p11.2 and 22q11.2. Suggestive support was found for eight additional candidate susceptibility and protective loci, which consisted predominantly of CNVs mediated by nonallelic homologous recombination.

Transcriptional interactions suggest niche segregation among microorganisms in the human gut.

The human gastrointestinal (GI) tract is the habitat for hundreds of microbial species, of which many cannot be cultivated readily, presumably because of the dependencies between species1. Studies of microbial co-occurrence in the gut have indicated community substructures that may reflect functional and metabolic interactions between cohabiting species2,3. To move beyond species co-occurrence networks, we systematically identified transcriptional interactions between pairs of coexisting gut microbes using metagenomics and microarray-based metatranscriptomics data from 233 stool samples from Europeans. In 102 significantly interacting species pairs, the transcriptional changes led to a reduced expression of orthologous functions between the coexisting species. Specific species-species transcriptional interactions were enriched for functions important for H2 and CO2 homeostasis, butyrate biosynthesis, ATP-binding cassette (ABC) transporters, flagella assembly and bacterial chemotaxis, as well as for the metabolism of carbohydrates, amino acids and cofactors. The analysis gives the first insight into the microbial community-wide transcriptional interactions, and suggests that the regulation of gene expression plays an important role in species adaptation to coexistence and that niche segregation takes place at the transcriptional level.

Linkage and whole genome sequencing identify a locus on 6q25-26 for formal thought disorder and implicate MEF2A regulation.

Formal thought disorder is a major feature of schizophrenia and other psychotic disorders. It is heritable, found in healthy relatives of patients with schizophrenia and other mental disorders but knowledge of specific genetic factors is lacking. The aim of this study was to search for biologically relevant high-risk variants. Formal thought disorder was assessed in participants in the Copenhagen Schizophrenia Linkage Study (N=236), a unique high-risk family study comprised of six large pedigrees. Microsatellite linkage analysis of formal thought disorder was performed and subsequent haplotype analysis of the implicated region using phased microsatellite and SNP genotypes. Whole genome sequencing (N=3) was used in the attempt to identify causative variants in the linkage region. Linkage analysis of formal thought disorder resulted in a single peak at chromosome 6(q26-q27) centred on marker D6S1277, with a maximum LOD score of 4.0. Phasing and fine mapping of the linkage peak identified a 5.5Mb haplotype (chr6:162242322-167753547, hg18) in 31 individuals, all belonging to the same pedigree sharing the haplotype from a common ancestor. The haplotype segregated with increased total thought disorder index score (P=4.9 × 10(-5)) and qualitatively severe forms of thought disturbances. Whole genome sequencing identified a novel nucleotide deletion (chr6:164377205 AG>A, hg18) predicted to disrupt the potential binding of the transcription factor MEF2A. The MEF2A binding site is located between two genes previously reported to associate with schizophrenia, QKI (HGNC:21100) and PDE10A (HGNC:8772). The findings are consistent with MEF2A deregulation conferring risk of formal thought disorder.