The rpoS gene is predominantly inactivated during laboratory storage and undergoes source-sink evolution in Escherichia coli species.

The rpoS gene codes for an alternative RNA polymerase sigma factor, which acts as a general regulator of the stress response. Inactivating alleles of rpoS in collections of natural Escherichia coli isolates have been observed at very variable frequencies, from less than 1% to more than 70% of strains. rpoS is easily inactivated in nutrient-deprived environments such as stab storage, which makes it difficult to determine the true frequency of rpoS inactivation in nature. We studied the evolutionary history of rpoS and compared it to the phylogenetic history of bacteria in two collections of 82 human commensal and extraintestinal E. coli strains. These strains were representative of the phylogenetic diversity of the species and differed only by their storage conditions. In both collections, the phylogenetic histories of rpoS and of the strains were congruent, indicating that horizontal gene transfer had not occurred at the rpoS locus, and rpoS was under strong purifying selection, with a ratio of the nonsynonymous mutation rate (Ka) to the synonymous substitution rate (Ks) substantially smaller than 1. Stab storage was associated with a high frequency of inactivating alleles, whereas almost no amino acid sequence variation was observed in RpoS in the collection studied directly after isolation of the strains from the host. Furthermore, the accumulation of variations in rpoS was typical of source-sink dynamics. In conclusion, rpoS is rarely inactivated in natural E. coli isolates within their mammalian hosts, probably because such strains rapidly become evolutionary dead ends. Our data should encourage bacteriologists to freeze isolates immediately and to avoid the use of stab storage.

Rapid birth-and-death evolution of the xenobiotic metabolizing NAT gene family in vertebrates with evidence of adaptive selection.

BACKGROUND: The arylamine N-acetyltransferases (NATs) are a unique family of enzymes widely distributed in nature that play a crucial role in the detoxification of aromatic amine xenobiotics. Considering the temporal changes in the levels and toxicity of environmentally available chemicals, the metabolic function of NATs is likely to be under adaptive evolution to broaden or change substrate specificity over time, making NATs a promising subject for evolutionary analyses. In this study, we trace the molecular evolutionary history of the NAT gene family during the last ~450 million years of vertebrate evolution and define the likely role of gene duplication, gene conversion and positive selection in the evolutionary dynamics of this family. RESULTS: A phylogenetic analysis of 77 NAT sequences from 38 vertebrate species retrieved from public genomic databases shows that NATs are phylogenetically unstable genes, characterized by frequent gene duplications and losses even among closely related species, and that concerted evolution only played a minor role in the patterns of sequence divergence. Local signals of positive selection are detected in several lineages, probably reflecting response to changes in xenobiotic exposure. We then put a special emphasis on the study of the last ~85 million years of primate NAT evolution by determining the NAT homologous sequences in 13 additional primate species. Our phylogenetic analysis supports the view that the three human NAT genes emerged from a first duplication event in the common ancestor of Simiiformes, yielding NAT1 and an ancestral NAT gene which in turn, duplicated in the common ancestor of Catarrhini, giving rise to NAT2 and the NATP pseudogene. Our analysis suggests a main role of purifying selection in NAT1 protein evolution, whereas NAT2 was predicted to mostly evolve under positive selection to change its amino acid sequence over time. These findings are consistent with a differential role of the two human isoenzymes and support the involvement of NAT1 in endogenous metabolic pathways. CONCLUSIONS: This study provides unequivocal evidence that the NAT gene family has evolved under a dynamic process of birth-and-death evolution in vertebrates, consistent with previous observations made in fungi.

Evaluation of the Bayesian method to derive migration patterns from changes in surname distributions over time.

Known migration in The Netherlands between the periods 1950-1969 and 2007, for 4.5 million individuals, was used to estimate the origin of migration by means of a Bayesian method on the basis of surname distributions in these two periods. Results of the method depend on the geographic specificity of the surnames and tend to be positioned between population density and actual probability of migration origin. An optimum in the correlation between estimated and actual percentages of origin of migration, and their differentiation as expressed by the correlation between the estimated and actual entropy across 40 distinguished areas, was found after a few iterations. The optimal correlation was 0.806 (Spearman), which shows that the Bayesian method provides a reasonable proxy of the rank order of a migrant's origin.

The family name as socio-cultural feature and genetic metaphor: from concepts to methods.

A recent workshop entitled "The Family Name as Socio-Cultural Feature and Genetic Metaphor: From Concepts to Methods" was held in Paris in December 2010, sponsored by the French National Centre for Scientific Research (CNRS) and by the journal Human Biology. This workshop was intended to foster a debate on questions related to the family names and to compare different multidisciplinary approaches involving geneticists, historians, geographers, sociologists and social anthropologists. This collective paper presents a collection of selected communications.

Role of intraspecies recombination in the spread of pathogenicity islands within the Escherichia coli species.

Horizontal gene transfer is a key step in the evolution of bacterial pathogens. Besides phages and plasmids, pathogenicity islands (PAIs) are subjected to horizontal transfer. The transfer mechanisms of PAIs within a certain bacterial species or between different species are still not well understood. This study is focused on the High-Pathogenicity Island (HPI), which is a PAI widely spread among extraintestinal pathogenic Escherichia coli and serves as a model for horizontal transfer of PAIs in general. We applied a phylogenetic approach using multilocus sequence typing on HPI-positive and -negative natural E. coli isolates representative of the species diversity to infer the mechanism of horizontal HPI transfer within the E. coli species. In each strain, the partial nucleotide sequences of 6 HPI-encoded genes and 6 housekeeping genes of the genomic backbone, as well as DNA fragments immediately upstream and downstream of the HPI were compared. This revealed that the HPI is not solely vertically transmitted, but that recombination of large DNA fragments beyond the HPI plays a major role in the spread of the HPI within E. coli species. In support of the results of the phylogenetic analyses, we experimentally demonstrated that HPI can be transferred between different E. coli strains by F-plasmid mediated mobilization. Sequencing of the chromosomal DNA regions immediately upstream and downstream of the HPI in the recipient strain indicated that the HPI was transferred and integrated together with HPI-flanking DNA regions of the donor strain. The results of this study demonstrate for the first time that conjugative transfer and homologous DNA recombination play a major role in horizontal transfer of a pathogenicity island within the species E. coli.

On the use of phylogeny-based tests to detect association between quantitative traits and haplotypes.

With the increasing availability of genetic data, several SNPs in a candidate gene can be combined into haplotypes to test for association with a quantitative trait. When the number of SNPs increases, the number of haplotypes can become very large and there is a need to group them together. The use of the phylogenetic relationships between haplotypes provides a natural and efficient way of grouping. Moreover, it allows us to identify disease or quantitative trait-related loci. In this article, we describe ALTree-q, a phylogeny-based approach to test for association between quantitative traits and haplotypes and to identify putative quantitative trait nucleotides (QTN). This study focuses on ALTree-q association test which is based on one-way analyses of variance (ANOVA) performed at the different levels of the tree. The statistical properties (type-one error and power rates) were estimated through simulations under different genetic models and were compared to another phylogeny-based test, TreeScan, (Templeton, 2005) and to a haplotypic omnibus test consisting in a one-way ANOVA between all haplotypes. For dominant and additive models ALTree-q is usually the most powerful test whereas TreeScan performs better under a recessive model. However, power depends strongly on the recurrence rate of the QTN, on the QTN allele frequency, and on the linkage disequilibrium between the QTN and other markers. An application of the method on Thrombin Activatable Fibronolysis Inhibitor Antigen levels in European and African samples confirms a possible association with polymorphisms of the CPB2 gene and identifies several QTNs.

The treeness of the tree of historical trees of life.

This paper compares and categorizes historical ideas about trees showing relationships among biological entities. The hierarchical structure of a tree is used to test the global consistency of similarities among these ideas; in other words we assess the "treeness" of the tree of historical trees. The collected data are figures and ideas about trees showing relationships among biological entities published or drawn by naturalists from 1555 to 2012. They are coded into a matrix of 235 historical trees and 141 descriptive attributes. From the most parsimonious "tree" of historical trees, treeness is measured by consistency index, retention index and homoplasy excess ratio. This tree is used to create sets or categories of trees, or to study the circulation of ideas. From an unrooted network of historical trees, treeness is measured by the delta-score. This unrooted network is used to measure and visualize treeness. The two approaches show a rather good treeness of the data, with respectively a retention idex of 0.83 and homoplasy excess ratio of 0.74, on one hand, and a delta-score of 0.26 on the other hand. It is interpreted as due to vertical transmission, i.e. an inheritance of shared ideas about biological trees among authors. This tree of trees is then used to test categories previously made. For instance, cladists and gradists are « paraphyletic ¼. The branches of this tree of trees suggest new categories of tree-thinkers that could have been overlooked by historians or systematists.

Evaluating NAT2PRED for inferring the individual acetylation status from unphased genotype data.

BACKGROUND: Genetically determined differences in N-acetylation capacity have proved to be important determinants of both the effectiveness of therapeutic response and the development of adverse drug reactions and toxicity during drug treatment. NAT2PRED is a web-server that allows a fast determination of NAT2 acetylation phenotype from genotype data without taking the extra step of reconstructing haplotypes for each individual (publicly available at http://nat2pred.rit.albany.edu). However, the classification accuracy of NAT2PRED needs to be assessed before its application can be advocated at a large scale. METHODS: The ability of NAT2PRED to classify individuals according to their acetylation status (slow, intermediate and rapid acetylators) was evaluated in a worldwide dataset composed of 56 population samples (8,489 individuals) from four continental regions. RESULTS: NAT2PRED correctly identified slow acetylators with a sensitivity above 99% for all populations outside sub-Saharan Africa. Nevertheless, NAT2PRED showed a poor ability to distinguish between intermediate and rapid acetylators, with a classification error rate reaching up to 10% in the non-African samples. CONCLUSION: NAT2PRED is an excellent tool to infer the individual acetylation status from NAT2 genotype data when the main interest is to distinguish slow acetylators from the others. This should facilitate the determination of the individual acetylation status in routine clinical practice and lead to better monitoring of risks associated with cancer and adverse drug reactions.

aes, the gene encoding the esterase B in Escherichia coli, is a powerful phylogenetic marker of the species.

BACKGROUND: Previous studies have established a correlation between electrophoretic polymorphism of esterase B, and virulence and phylogeny of Escherichia coli. Strains belonging to the phylogenetic group B2 are more frequently implicated in extraintestinal infections and include esterase B2 variants, whereas phylogenetic groups A, B1 and D contain less virulent strains and include esterase B1 variants. We investigated esterase B as a marker of phylogeny and/or virulence, in a thorough analysis of the esterase B-encoding gene. RESULTS: We identified the gene encoding esterase B as the acetyl-esterase gene (aes) using gene disruption. The analysis of aes nucleotide sequences in a panel of 78 reference strains, including the E. coli reference (ECOR) strains, demonstrated that the gene is under purifying selection. The phylogenetic tree reconstructed from aes sequences showed a strong correlation with the species phylogenetic history, based on multi-locus sequence typing using six housekeeping genes. The unambiguous distinction between variants B1 and B2 by electrophoresis was consistent with Aes amino-acid sequence analysis and protein modelling, which showed that substituted amino acids in the two esterase B variants occurred mostly at different sites on the protein surface. Studies in an experimental mouse model of septicaemia using mutant strains did not reveal a direct link between aes and extraintestinal virulence. Moreover, we did not find any genes in the chromosomal region of aes to be associated with virulence. CONCLUSION: Our findings suggest that aes does not play a direct role in the virulence of E. coli extraintestinal infection. However, this gene acts as a powerful marker of phylogeny, illustrating the extensive divergence of B2 phylogenetic group strains from the rest of the species.

Selecting predictive markers for pharmacogenetic traits: tagging vs. data-mining approaches.

OBJECTIVE: The tagging approach appears as a promising tool to test the association of genetic variants with complex traits such as disease susceptibility or drug response. However, since tag markers are selected only on the basis of inter-marker LD properties, regardless of any phenotypes, it remains unclear to what extent they can be useful to predict variable drug responses, once typed in clinical material. We undertook a study to provide further insights into the usefulness of the tagging approach for selecting phenotype-associated markers relevant to drug response. METHODS: Several tagging methods were applied to the genotyping data of two drug-metabolizing enzymes, NAT2 and CYP2D6, and the ability of the selected tagging markers to predict the individual metabolizer status was empirically evaluated. We also assessed the impact of LD levels, tagging thresholds and allele frequencies on tagging efficiency. RESULTS: We found that the functional variation was adequately represented by the selected tagging markers, these latter providing a classification accuracy for the individual metabolizer status close to the maximal 100% value observed with the entire set of polymorphisms. CONCLUSION: The tagging approach is an interesting approach to select candidate gene markers predictive of drug response in pharmacogenomic studies.

Humans and Chimpanzees Display Opposite Patterns of Diversity in Arylamine N-Acetyltransferase Genes.

Among the many genes involved in the metabolism of therapeutic drugs, human arylamine N-acetyltransferases (NATs) genes have been extensively studied, due to their medical importance both in pharmacogenetics and disease epidemiology. One member of this small gene family, NAT2, is established as the locus of the classic human acetylation polymorphism in drug metabolism. Current hypotheses hold that selective processes favoring haplotypes conferring lower NAT2 activity have been operating in modern humans' recent history as an adaptation to local chemical and dietary environments. To shed new light on such hypotheses, we investigated the genetic diversity of the three members of the NAT gene family in seven hominid species, including modern humans, Neanderthals and Denisovans. Little polymorphism sharing was found among hominids, yet all species displayed high NAT diversity, but distributed in an opposite fashion in chimpanzees and bonobos (Pan genus) compared to modern humans, with higher diversity in Pan species at NAT1 and lower at NAT2, while the reverse is observed in humans. This pattern was also reflected in the results returned by selective neutrality tests, which suggest, in agreement with the predicted functional impact of mutations detected in non-human primates, stronger directional selection, presumably purifying selection, at NAT1 in modern humans, and at NAT2 in chimpanzees. Overall, the results point to the evolution of divergent functions of these highly homologous genes in the different primate species, possibly related to their specific chemical/dietary environment (exposome) and we hypothesize that this is likely linked to the emergence of controlled fire use in the human lineage.

Worldwide distribution of NAT2 diversity: implications for NAT2 evolutionary history.

BACKGROUND: The N-acetyltransferase 2 (NAT2) gene plays a crucial role in the metabolism of many drugs and xenobiotics. As it represents a likely target of population-specific selection pressures, we fully sequenced the NAT2 coding region in 97 Mandenka individuals from Senegal, and compared these sequences to extant data on other African populations. The Mandenka data were further included in a worldwide dataset composed of 41 published population samples (6,727 individuals) from four continental regions that were adequately genotyped for all common NAT2 variants so as to provide further insights into the worldwide haplotype diversity and population structure at NAT2. RESULTS: The sequencing analysis of the NAT2 gene in the Mandenka sample revealed twelve polymorphic sites in the coding exon (two of which are newly identified mutations, C345T and C638T), defining 16 haplotypes. High diversity and no molecular signal of departure from neutrality were observed in this West African sample. On the basis of the worldwide genotyping survey dataset, we found a strong genetic structure differentiating East Asians from both Europeans and sub-Saharan Africans. This pattern could result from region- or population-specific selective pressures acting at this locus, as further suggested in the HapMap data by extremely high values of FST for a few SNPs positions in the NAT2 coding exon (T341C, C481T and A803G) in comparison to the empirical distribution of FST values accross the whole 400-kb region of the NAT gene family. CONCLUSION: Patterns of sequence variation at NAT2 are consistent with selective neutrality in all sub-Saharan African populations investigated, whereas the high level of population differentiation between Europeans and East Asians inferred from SNPs could suggest population-specific selective pressures acting at this locus, probably caused by differences in diet or exposure to other environmental signals.

Clustering of haplotypes based on phylogeny: how good a strategy for association testing?

Haplotypes are now widely used in association studies between markers and disease susceptibility locus. However, when a large number of markers are considered, the number of possible haplotypes increases leading to two problems: an increased number of degrees of freedom that may result in a lack of power and the existence of rare haplotypes that may be difficult to take into account in the statistical analysis. In a recent paper, Durrant et al proposed a method, CLADHC, to group haplotypes based on distance matrices and showed that this could considerably increase the power of the association test as compared to either single-locus analysis or haplotype analysis without prior grouping. Although the authors considered different one-disease-locus susceptibility models in their simulations, they did not study the impact of the linkage disequilibrium (LD) pattern and of the susceptibility allele frequency on their conclusions. Here, we show, using haplotype data from five regions of the genome of different lengths and with different LD patterns, that, when a single disease susceptibility locus is simulated, the prior grouping of haplotypes based on the algorithm of Durrant et al does not increase the power of association testing except in very particular situations of LD patterns and allele frequencies.

Inferring haplotypes at the NAT2 locus: the computational approach.

BACKGROUND: Numerous studies have attempted to relate genetic polymorphisms within the N-acetyltransferase 2 gene (NAT2) to interindividual differences in response to drugs or in disease susceptibility. However, genotyping of individuals single-nucleotide polymorphisms (SNPs) alone may not always provide enough information to reach these goals. It is important to link SNPs in terms of haplotypes which carry more information about the genotype-phenotype relationship. Special analytical techniques have been designed to unequivocally determine the allocation of mutations to either DNA strand. However, molecular haplotyping methods are labour-intensive and expensive and do not appear to be good candidates for routine clinical applications. A cheap and relatively straightforward alternative is the use of computational algorithms. The objective of this study was to assess the performance of the computational approach in NAT2 haplotype reconstruction from phase-unknown genotype data, for population samples of various ethnic origin. RESULTS: We empirically evaluated the effectiveness of four haplotyping algorithms in predicting haplotype phases at NAT2, by comparing the results with those directly obtained through molecular haplotyping. All computational methods provided remarkably accurate and reliable estimates for NAT2 haplotype frequencies and individual haplotype phases. The Bayesian algorithm implemented in the PHASE program performed the best. CONCLUSION: This investigation provides a solid basis for the confident and rational use of computational methods which appear to be a good alternative to infer haplotype phases in the particular case of the NAT2 gene, where there is near complete linkage disequilibrium between polymorphic markers.

On the use of haplotype phylogeny to detect disease susceptibility loci.

BACKGROUND: The cladistic approach proposed by Templeton has been presented as promising for the study of the genetic factors involved in common diseases. This approach allows the joint study of multiple markers within a gene by considering haplotypes and grouping them in nested clades. The idea is to search for clades with an excess of cases as compared to the whole sample and to identify the mutations defining these clades as potential candidate disease susceptibility sites. However, the performance of this approach for the study of the genetic factors involved in complex diseases has never been studied. RESULTS: In this paper, we propose a new method to perform such a cladistic analysis and we estimate its power through simulations. We show that under models where the susceptibility to the disease is caused by a single genetic variant, the cladistic test is neither really more powerful to detect an association nor really more efficient to localize the susceptibility site than an individual SNP testing. However, when two interacting sites are responsible for the disease, the cladistic analysis greatly improves the probability to find the two susceptibility sites. The impact of the linkage disequilibrium and of the tree characteristics on the efficiency of the cladistic analysis are also discussed. An application on a real data set concerning the CARD15 gene and Crohn disease shows that the method can successfully identify the three variant sites that are involved in the disease susceptibility. CONCLUSION: The use of phylogenies to group haplotypes is especially interesting to pinpoint the sites that are likely to be involved in disease susceptibility among the different markers identified within a gene.

Scaling of free-ranging primate energetics with body mass predicts low energy expenditure in humans.

Studies of how a mammal's daily energy expenditure scales with its body mass suggest that humans, whether Westerners, agro-pastoralists, or hunter-gatherers, all have much lower energy expenditures for their body mass than other mammals. However, non-human primates also differ from other mammals in several life history traits suggestive of low energy use. Judging by field metabolic rates of free-ranging strepsirhine and haplorhine primates with different lifestyle and body mass, estimated using doubly labeled water, primates have lower energy expenditure than other similar-sized eutherian mammals. Daily energy expenditure in humans fell along the regression line of non-human primates. The results suggest that thrifty energy use could be an ancient strategy of primates. Although physical activity is a major component of energy balance, our results suggest a need to revise the basis for establishing norms of energy expenditure in modern humans.

Analysis of the French National Registry of unrelated bone marrow donors, using surnames as a tool for improving geographical localisation of HLA haplotypes.

The first statistical analysis of the French National Registry of volunteer bone marrow donors estimated the probabilities of haplotype frequencies separately for each of the 20 administrative regions of France. Here we propose to use donors' surnames to increase the accuracy of location of the donor's geographical origin. This approach allows us to estimate haplotype frequencies for administrative entities (90 departments) smaller than regions and to correct for bias resulting from recent mobility. We analysed 30,777 donors typed for HLA-A,B and 17,745 donors typed for HLA-A,B,DR,DQ. By using the donors' surnames, we identified common and rare haplotypes (those found in only one department) and estimated the degree of HLA polymorphism at the department level. We also identified departments with a distinctive genetic structure (for example, Paris, Corsica, Pyrenees and Meurthe-et-Moselle). By providing a more accurate geographical distribution of HLA polymorphisms in France, this study will enable us to optimise policies for recruiting bone marrow donors and to improve the fit between the donor file and patients' needs.

Arylamine N-acetyltransferase 2 (NAT2) genetic diversity and traditional subsistence: a worldwide population survey.

Arylamine N-acetyltransferase 2 (NAT2) is involved in human physiological responses to a variety of xenobiotic compounds, including common therapeutic drugs and exogenous chemicals present in the diet and the environment. Many questions remain about the evolutionary mechanisms that have led to the high prevalence of slow acetylators in the human species. Evidence from recent surveys of NAT2 gene variation suggests that NAT2 slow-causing variants might have become targets of positive selection as a consequence of the shift in modes of subsistence and lifestyle in human populations in the last 10,000 years. We aimed to test more extensively the hypothesis that slow acetylation prevalence in humans is related to the subsistence strategy adopted by the past populations. To this end, published frequency data on the most relevant genetic variants of NAT2 were collected from 128 population samples (14,679 individuals) representing different subsistence modes and dietary habits, allowing a thorough analysis at both a worldwide and continent scale. A significantly higher prevalence of the slow acetylation phenotype was observed in populations practicing farming (45.4%) and herding (48.2%) as compared to populations mostly relying on hunting and gathering (22.4%) (P = 0.0007). This was closely mirrored by the frequency of the slow 590A variant that was found to occur at a three-fold higher frequency in food producers (25%) as compared to hunter-gatherers (8%). These findings are consistent with the hypothesis that the Neolithic transition to subsistence economies based on agricultural and pastoral resources modified the selective regime affecting the NAT2 acetylation pathway. Furthermore, the vast amount of data collected enabled us to provide a comprehensive and up-to-date description of NAT2 worldwide genetic diversity, thus building up a useful resource of frequency data for further studies interested in epidemiological or anthropological research questions involving NAT2.

SNP selection at the NAT2 locus for an accurate prediction of the acetylation phenotype.

PURPOSE: Genetic polymorphisms in the N-acetyltransferase 2 gene determine the individual acetylator status, which influences both the toxicity and efficacy profile of acetylated drugs. Determination of an individual's acetylation phenotype prior to initiation of therapy, through DNA-based tests, should permit to improve therapy response and reduce adverse events. However, due to extensive linkage disequilibrium between markers within NAT2, the genotyping of closely spaced markers yields highly redundant data: testing them all is expensive and often unnecessary. The objective of this study is to establish the optimal strategy to define, in the genetic context of a given ethnic group, the most informative set of single-nucleotide polymorphisms that best enables accurate prediction of acetylation phenotype. METHODS: Three classification methods have been investigated (classification trees, artificial neural networks and multifactor dimensionality reduction method) in order to find the optimal set of single-nucleotide polymorphisms enabling the most efficient classification of individuals in rapid and slow acetylators. RESULTS: Our results show that, in almost all population samples, only one or two single-nucleotide polymorphisms would be enough to obtain a good predictive capacity with no or only a modest reduction in power relative to direct assays of all common markers. In contrast, in Black African populations, where lower levels of linkage disequilibrium are observed at NAT2, a larger number of single-nucleotide polymorphisms are required to predict acetylation phenotype. CONCLUSION: The results of this study will be helpful for the design of time- and cost-effective pharmacogenetic tests (adapted to specific populations) that could be used as routine tools in clinical practice.