Vertical transmission of bacteria from mother to infant at birth is postulated to initiate a life-long host-microbe symbiosis, playing an important role in early infant development. However, only the tracking of strictly defined unique microbial strains can clarify where the intestinal bacteria come from, how long the initial colonisers persist and whether colonisation by other strains from the environment can replace existing ones. Using rare single nucleotide variants in fecal metagenomes of infants and their family members, we show strong evidence of selective and persistent transmission of maternal strain populations to the vaginally born infant, and their occasional replacement by strains from the environment, including those from family members, in later childhood. Only strains from the classes Actinobacteria and Bacteroidia, which are essential components of the infant microbiome, are transmitted from the mother and persist for at least one year. In contrast, maternal strains of Clostridia, a dominant class in the mother's gut microbiome, are not observed in the infant. Caesarean-born infants show a striking lack of maternal transmission at birth. After the first year, strain influx from the family environment occurs, and continues even in adulthood. Fathers appear to be more frequently donors of novel strains to other family members than receivers. Thus, the infant gut is seeded by selected maternal bacteria, which expand to form a stable community, with a rare but stable continuing strain influx over time.

Despite increasing concerns over inappropriate use of antibiotics in medicine and food production, population-level resistance transfer into the human gut microbiota has not been demonstrated beyond individual case studies. To determine the "antibiotic resistance potential" for entire microbial communities, we employ metagenomic data and quantify the totality of known resistance genes in each community (its resistome) for 68 classes and subclasses of antibiotics. In 252 fecal metagenomes from three countries, we show that the most abundant resistance determinants are those for antibiotics also used in animals, and for antibiotics that have been available longer. Resistance genes are also more abundant in samples from Spain, Italy and France than from Denmark, the US, or Japan. Where comparable country-level data on antibiotic use in both humans and animals are available, differences in these statistics match the observed resistance potential differences. The results are robust over time as the antibiotic resistance determinants of individuals persist in the human gut flora for at least a year.

The human gut microbiome is a complex ecosystem composed mainly of uncultured bacteria. It plays an essential role in the catabolism of dietary fibers, the part of plant material in our diet that is not metabolized in the upper digestive tract, because the human genome does not encode adequate carbohydrate active enzymes (CAZymes). We describe a multi-step functionally based approach to guide the in-depth pyrosequencing of specific regions of the human gut metagenome encoding the CAZymes involved in dietary fiber breakdown. High-throughput functional screens were first applied to a library covering 5.4 × 10(9) bp of metagenomic DNA, allowing the isolation of 310 clones showing beta-glucanase, hemicellulase, galactanase, amylase, or pectinase activities. Based on the results of refined secondary screens, sequencing efforts were reduced to 0.84 Mb of nonredundant metagenomic DNA, corresponding to 26 clones that were particularly efficient for the degradation of raw plant polysaccharides. Seventy-three CAZymes from 35 different families were discovered. This corresponds to a fivefold target-gene enrichment compared to random sequencing of the human gut metagenome. Thirty-three of these CAZy encoding genes are highly homologous to prevalent genes found in the gut microbiome of at least 20 individuals for whose metagenomic data are available. Moreover, 18 multigenic clusters encoding complementary enzyme activities for plant cell wall degradation were also identified. Gene taxonomic assignment is consistent with horizontal gene transfer events in dominant gut species and provides new insights into the human gut functional trophic chain.

The increasing number of sequenced genomes makes it possible to infer the evolutionary history of functional modules, i.e., groups of proteins that contribute jointly to the same cellular function in a given species. Here we identify and analyze those prokaryotic functional modules, whose composition remains largely unchanged during evolution, and study their properties. Such "cohesive" modules have a large number of internal functional connections, encode genes that tend to be in close proximity in prokaryotic genomes, and correspond to physical complexes or complex functional systems like the flagellar apparatus. Cohesive modules are enriched in processes such as energy and amino acid metabolism, cell motility, and intracellular trafficking, or secretion. By grouping genes into modules we achieve a more precise estimate of their age and find that the young modules are often horizontally transferred between species and are enriched in functions involved in interactions with the environment, implying that they play an important role in the adaptation of species to new environments.

Molecular evolution studies are usually based on the analysis of individual genes and thus reflect only small-range variations in genomic sequences. A complementary approach is to study the evolutionary history of rearrangements in entire genomes based on the analysis of gene orders. The progress in whole genome sequencing provides an unprecedented level of detailed sequence data to infer genome rearrangements through comparative approaches. The comparative analysis of recently sequenced rodent genomes with the human genome revealed evidence for a larger number of rearrangements than previously thought and led to the reconstruction of the putative genomic architecture of the murid rodent ancestor, while the architecture of the ancestral mammalian genome and the rate of rearrangements in the human lineage remained unknown. Sequencing the chicken genome provides an opportunity to reconstruct the architecture of the ancestral mammalian genome by using chicken as an outgroup. Our analysis reveals a very low rate of rearrangements and, in particular, interchromosomal rearrangements in chicken, in the early mammalian ancestor, or in both. The suggested number of interchromosomal rearrangements between the mammalian ancestor and chicken, during an estimated 500 million years of evolution, only slightly exceeds the number of interchromosomal rearrangements that happened in the mouse lineage, over the course of about 87 million years.

Orthologous genes that maintain a single-copy status in a broad range of species may indicate a selection against gene duplication. If this is the case, then duplicates of such genes that do survive may have escaped the dosage control by rapid and sizable changes in their function. To test this hypothesis and to develop a strategy for the identification of novel gene functions, we have analyzed 22 primate-specific intrachromosomal duplications of genes with a single-copy ortholog in all other completely sequenced metazoans. When comparing this set to genes not exposed to the single-copy status constraint, we observed a higher tendency of the former to modify their gene structure, often through complex genomic rearrangements. The analysis of the most dramatic of these duplications, affecting approximately 10% of human Chromosome 2, enabled a detailed reconstruction of the events leading to the appearance of a novel gene family. The eight members of this family originated from the highly conserved nucleoporin RanBP2 by several genetic rearrangements such as segmental duplications, inversions, translocations, exon loss, and domain accretion. We have experimentally verified that at least one of the newly formed proteins has a cellular localization different from RanBP2's, and we show that positive selection did act on specific domains during evolution.

We screened all intergenic regions in the human genome to identify pseudogenes with a combination of homology searches and a functionality test using the ratio of silent to replacement nucleotide substitutions (KA/KS). We identified 19,724 regions of which 95% +/- 3% are estimated to evolve neutrally and thus are likely to encode pseudogenes. Half of these have no detectable truncation in their pseudocoding regions and therefore are not identifiable by methods that require the presence of truncations to prove nonfunctionality. A comparative analysis with the mouse genome showed that 70% of these pseudogenes have a retrotranspositional origin (processed), and the rest arose by segmental duplication (nonprocessed). Although the spread of both types of pseudogenes correlates with chromosome size, nonprocessed pseudogenes appear to be enriched in regions with high gene density. It is likely that the human pseudogenes identified here represent only a small fraction of the total, which probably exceeds the number of genes.

A systematic computational analysis of protein sequences containing known nuclear domains led to the identification of 28 novel domain families. This represents a 26% increase in the starting set of 107 known nuclear domain families used for the analysis. Most of the novel domains are present in all major eukaryotic lineages, but 3 are species specific. For about 500 of the 1200 proteins that contain these new domains, nuclear localization could be inferred, and for 700, additional features could be predicted. For example, we identified a new domain, likely to have a role downstream of the unfolded protein response; a nematode-specific signalling domain; and a widespread domain, likely to be a noncatalytic homolog of ubiquitin-conjugating enzymes.

We investigate the co-occurrence of domain families in eukaryotic proteins to predict protein cellular localization. Approximately half (300) of SMART domains form a "small-world network", linked by no more than seven degrees of separation. Projection of the domains onto two-dimensional space reveals three clusters that correspond to cellular compartments containing secreted, cytoplasmic, and nuclear proteins. The projection method takes into account the existence of "bridging" domains, that is, instances where two domains might not occur with each other but frequently co-occur with a third domain; in such circumstances the domains are neighbors in the projection. While the majority of domains are specific to a compartment ("locale"), and hence may be used to localize any protein that contains such a domain, a small subset of domains either are present in multiple locales or occur in transmembrane proteins. Comparison with previously annotated proteins shows that SMART domain data used with this approach can predict, with 92% accuracy, the localizations of 23% of eukaryotic proteins. The coverage and accuracy will increase with improvements in domain database coverage. This method is complementary to approaches that use amino-acid composition or identify sorting sequences; these methods may be combined to further enhance prediction accuracy.

In the course of evolution, genomes are shaped by processes like gene loss, gene duplication, horizontal gene transfer, and gene genesis (the de novo origin of genes). Here we reconstruct the gene content of ancestral Archaea and Proteobacteria and quantify the processes connecting them to their present day representatives based on the distribution of genes in completely sequenced genomes. We estimate that the ancestor of the Proteobacteria contained around 2500 genes, and the ancestor of the Archaea around 2050 genes. Although it is necessary to invoke horizontal gene transfer to explain the content of present day genomes, gene loss, gene genesis, and simple vertical inheritance are quantitatively the most dominant processes in shaping the genome. Together they result in a turnover of gene content such that even the lineage leading from the ancestor of the Proteobacteria to the relatively large genome of Escherichia coli has lost at least 950 genes. Gene loss, unlike the other processes, correlates fairly well with time. This clock-like behavior suggests that gene loss is under negative selection, while the processes that add genes are under positive selection.

Sequence database searching methods such as BLAST, are invaluable for predicting molecular function on the basis of sequence similarities among single regions of proteins. Searches of whole databases however, are not optimized to detect multiple homologous regions within a single polypeptide. Here we have used the prospero algorithm to perform self-comparisons of all predicted Drosophila melanogaster gene products. Predicted repeats, and their homologs from all species, were analyzed further to detect hitherto unappreciated evolutionary relationships. Results included the identification of novel tandem repeats in the human X-linked retinitis pigmentosa type-2 gene product, repeated segments in cystinosin, associated with a defect in cystine transport, and 'nested' homologous domains in dysferlin, whose gene is mutated in limb girdle muscular dystrophy. Novel signaling domain families were found that may regulate the microtubule-based cytoskeleton and ubiquitin-mediated proteolysis, respectively. Two families of glycosyl hydrolases were shown to contain internal repetitions that hint at their evolution via a piecemeal, modular approach. In addition, three examples of fruit fly genes were detected with tandem exons that appear to have arisen via internal duplication. These findings demonstrate how completely sequenced genomes can be exploited to further understand the relationships between molecular structure, function, and evolution.

Various new methods have been proposed to predict functional interactions between proteins based on the genomic context of their genes. The types of genomic context that they use are Type I: the fusion of genes; Type II: the conservation of gene-order or co-occurrence of genes in potential operons; and Type III: the co-occurrence of genes across genomes (phylogenetic profiles). Here we compare these types for their coverage, their correlations with various types of functional interaction, and their overlap with homology-based function assignment. We apply the methods to Mycoplasma genitalium, the standard benchmarking genome in computational and experimental genomics. Quantitatively, conservation of gene order is the technique with the highest coverage, applying to 37% of the genes. By combining gene order conservation with gene fusion (6%), the co-occurrence of genes in operons in absence of gene order conservation (8%), and the co-occurrence of genes across genomes (11%), significant context information can be obtained for 50% of the genes (the categories overlap). Qualitatively, we observe that the functional interactions between genes are stronger as the requirements for physical neighborhood on the genome are more stringent, while the fraction of potential false positives decreases. Moreover, only in cases in which gene order is conserved in a substantial fraction of the genomes, in this case six out of twenty-five, does a single type of functional interaction (physical interaction) clearly dominate (>80%). In other cases, complementary function information from homology searches, which is available for most of the genes with significant genomic context, is essential to predict the type of interaction. Using a combination of genomic context and homology searches, new functional features can be predicted for 10% of M. genitalium genes.