Search

We investigated the controversial origin of domestic sheep (Ovis aries) using large samples of contemporary and ancient domestic individuals and their closest wild relatives: the Asiatic mouflon (Ovis gmelini), the urial (Ovis vignei) and the argali (Ovis ammon). A phylogeny based on mitochondrial DNA, including 213 new cytochrome-b sequences of wild Ovism confirmed that O. gmelini is the maternal ancestor of sheep and precluded mtDNA contributions from O. vignei (and O. gmelini × O. vignei hybrids) to domestic lineages. We also produced 54 new control region sequences showing shared haplogroups (A, B, C and E) between domestic sheep and wild O. gmelini which localized the domestication center in eastern Anatolia and central Zagros, excluding regions further east where exclusively wild haplogroups were found. This overlaps with the geographic distribution of O. gmelini gmelini, further suggesting that the maternal origin of domestic sheep derives from this subspecies. Additionally, we produced 57 new CR sequences of Neolithic sheep remains from a large area covering Anatolia to Europe, showing the early presence of at least three mitochondrial haplogroups (A, B and D) in Western colonization routes. This confirmed that sheep domestication was a large-scale process that captured diverse maternal lineages (haplogroups).

The extent to which genomic convergence shapes locally adapted phenotypes in different species remains a fundamental question in evolutionary biology. It would help assessing the relative role of historical contingencies versus determinism in evolution. To bring new insights to this debate we set up a framework which aimed to compare the adaptive trajectories of two domesticated mammal species co-distributed in diversified landscapes. We sequenced the genomes of 160 sheep and 161 goats extensively managed along environmental gradients, including temperature, rainfall, seasonality and altitude, to identify genes and biological processes shaping local adaptation. Allele frequencies at adaptive loci were rarely found to vary gradually along environmental gradients, but rather displayed a discontinuous shift at the extremities of environmental clines. Of the more than 430 adaptive genes identified, only 6 were orthologous between sheep and goats and those responded differently to environmental pressures, suggesting different adaptive mechanisms in these two closely related species. Such diversity of adaptive pathways may result from a high number of biological functions involved in adaptation to multiple eco-climatic gradients, and provides more arguments for the role of contingency and stochasticity in adaptation rather than repeatability and determinism.

The way in which living organisms mobilize a combination of long-term adaptive mechanisms and short-term phenotypic plasticity to face environmental variations is still largely unknown. In the context of climate change, understanding the genetic and epigenetic bases for adaptation and plasticity is a major stake for preserving genomic resources and the resilience capacity of livestock populations. We characterized both epigenetic and genetic variations by contrasting 22 sheep and 21 goats from both sides of a climate gradient, focusing on free-ranging populations from Morocco. We produced for each individual Whole-Genome Sequence at 12X coverage and MeDIP-Seq data, to identify regions under selection and those differentially methylated. For both species, the analysis of genetic differences (FST) along the genome between animals from localities with high vs. low temperature annual variations detected candidate genes under selection in relation to environmental perception (5 genes), immunity (4 genes), reproduction (8 genes) and production (11 genes). Moreover, we found for each species one differentially methylated gene, namely AGPTA4 in goat and SLIT3 in sheep, which were both related, among other functions, to milk production and muscle development. In both sheep and goats, the comparison between genomic regions impacted by genetic and epigenetic variations suggests that climatic variations impacted similar biological pathways but different genes.

During domestication processes, changes in selective pressures induce multiple phenotypical, physiological, and behavioral changes in target species. The rise of next-generation sequencing has provided a chance to study the genetics bases of these changes, most of the time based on single nucleotide polymorphisms (SNPs). However, several studies have highlighted the impact of structural variations (SVs) on individual fitness, particularly in domestic species. We aimed at unraveling the role of SVs during the domestication and later improvement of small ruminants by analyzing whole-genome sequences of 40 domestic sheep and 11 of their close wild relatives (Ovis orientalis), and 40 goats and 18 of their close wild relatives (Capra aegagrus). Using a combination of detection tools, we called 45,796 SVs in Ovis and 15,047 SVs in Capra genomes, including insertions, deletions, inversions, copy number variations, and chromosomal translocations. Most of these SVs were previously unreported in small ruminants. 69 and 45 SVs in sheep and goats, respectively, were in genomic regions with neighboring SNPs highly differentiated between wilds and domestics (i.e., putatively related to domestication). Among them, 25 and 20 SVs were close to or overlapping with genes related to physiological and morpho-anatomical traits linked with productivity (e.g., size, meat or milk quality, wool color), reproduction, or immunity. Finally, several of the SVs differentiated between wilds and domestics would not have been detected by screening only the differentiation of SNPs surrounding them, highlighting the complementarity of SVs and SNPs based approaches to detect signatures of selection.

Northwest-African sheep represent an ideal case-study for assessing the potential impact of genetic homogenization as a threat to the future of traditional breeds that are adapted to local conditions. We studied ten Algerian and Moroccan breeds of sheep, including three transboundary breeds, distributed over a large part of the Maghreb region, which represents a geographically and historically coherent unit. Our analysis of the dataset that involved carrying out Genome-wide SNP genotyping, revealed a high level of homogenization (ADMIXTURE, NetView, fineSTRUCTURE and IBD segments analyses), in such a way that some breeds from different origins appeared genetically undistinguished: by grouping the eight most admixed populations, we obtained a mean global FST value of 0.0024. The sPCA analysis revealed that the major part of Morocco and the Northern part of Algeria were affected by the phenomenon, including most of the breeds considered. Unsupervised cross-breeding with the popular Ouled-Djellal breed was identified as a proximate cause of this homogenization. The issue of transboundary breeds was investigated, and the Hamra breed in particular was examined via ROH fragments analysis. Genetic diversity was considered in the light of historical archives and anthropological works. All of these elements taken together suggest that homogenization as a factor affecting the Maghrebin sheep stock, has been particularly significant over the last few decades, although this process probably started much earlier. In particular, we have identified the policies set by the French administration during the colonial period of the region’s history as a causal factor that probably contributed significantly to this process. The genetic homogenization that we have observed calls into question the integrity of the farm animal genomic resources represented by these local breeds, whose conservation is of critical importance to the future of the livestock sector.

The male-specific part of the Y-chromosome is in mammalian and many other species the longest haplotype that is inherited without recombination. By its paternal transmission it has a small effective population size in species with dominant males. In several species, Y-chromosomal haplotypes are sensitive markers of population history and introgression. Previous studies have identified in domestic goats four major Y-chromosomal haplotypes Y1A, Y1B, Y2A and Y2B with a marked geographic differentiation and several regional variants. In this study we used published whole-genome sequences of 70 male goats from 16 modern breeds, 11 ancient-DNA samples and 29 samples from seven wild goat species. We identified single-copy male-specific SNPs in four scaffolds, containing SRY, ZFY, DBY with SSX3Y and UTY, and USP9Y with UMN2001, respectively. Phylogenetic analyses indicated haplogroups corresponding to the haplotypes Y1B, Y2A and Y2B, respectively, but Y1A was split into Y1AA and Y1AB. All haplogroups were detected in ancient DNA samples from southeast Europe and, with the exception of Y1AB, in the bezoar goat, which is the wild ancestor of the domestic goats. Combining these data with those of previous studies and with genotypes obtained by Sanger sequencing or the KASP assay yielded haplogroup distributions for 132 domestic breeds or populations. The phylogeographic differentiation indicated paternal population bottlenecks on all three continents. This possibly occurred during the Neolithic introductions of domestic goats to those continents with a particularly strong influence in Europe along the Danubian route. This study illustrates the power of the Y-chromosomal haplotype for the reconstructing the history of mammalian species with a wide geographic range.

International audience; The EVOSHEEP project combines archaeozoology, geometric morphometrics and genetics to study archaeo- logical sheep assemblages dating from the sixth to the first millennia BC in eastern Africa, the Levant, the Ana- tolian South Caucasus, the Iranian Plateau and Mesopotamia. The project aims to understand changes in the physical appearance and phenotypic characteristics of sheep and how these related to the appearance of new breeds and the demand for secondary products to supply the textile industry.

In a context of climate change, identifying the genes underlying adaptations to extreme environments is essential. Small ruminants are adapted to a wide variety of habitats and thus, are promising study models. Here, we considered 17 goat and 25 sheep local Mediterranean breeds, in Italy, France and Spain. We proposed, and empirically tested a new methodology to highlight selection signatures in relation to the environments. Based on historical archives, we selected the breeds potentially most linked to a territory and defined their original geographical cradle, including transhumant pastoral areas. For both species, the cradles were arranged along latitudinal gradient of aridity and altitude. Then we used the programs PCAdapt and LFMM. Considering cradles, instead of current GPS coordinates, markedly improved the sensitivity of the LFMM analyses. All the results combined with a systematic literature review, revealed a set of genes with potentially key adaptive roles. Some of these genes, have been found implicated in lipid metabolism (SUCLG2, BMP2), hypoxia/heat stress (UBE2R2/UBAP2), lung function (BMPR2), seasonal patterns (SOX2, DPH6) or neuronal function (TRPC4, TRPC6). For the intergenic region between PCDH9 and KLH1, as well as NBEA, identified in both species, the adaptive role could be strong. We found for RXFP2, associated with sheep horn development, and MSRB3 and SLC26A4, both associated with hearing of sheep, a strong association with the environment gradient. We conclude that our new methodology, which considers breeds in their historical, environmental and anthropological context, provides novel and essential information on local breeds and their unique adaptations.

Genomic Structural Variations (SVs) are known to impact the evolution of genomes and to have consequences on individual’s fitness. Nevertheless, they remain challenging to detect in whole genome re-sequencing (WGS) data. Lots of methods detecting SVs are described in the literature but they might be hard to install, have non-trivial settings, do not detect all SVs categories and have generally high levels of false positive.Here we introduce BAdabouM, a fast (C written) and easy to install SVs discovery tool. BAdabouM auto evaluates read length, library size and mean coverage to set thresholds specific to each experiment. BAdabouM interprets multiple SVs signatures (reads aligned with a split, non-concordant mapped pairs or uneven coverage) to detect insertions, deletions, copy number variations, inversions, and translocations at single-nucleotide resolution.When compared with two widely used methods on simulated and real datasets, BAdabouM was faster, exhibited a similar accuracy with a good concordance on SVs detected, and detected significantly more insertions. BAdabouM was more reproducible to detect independently SVs across individuals, which is a clear advantage when characterizing population polymorphism. Furthermore, BAdabouM demonstrated a superior ability to detect breakpoints with a base pair resolution.BAdabouM proved to be efficient, fast and accurate to detect SVs, and handle. BAdabouM is a complementary method to be used for a more comprehensive detection of SVs, and is especially suited for studying polymorphism for all types of SVs with a high accuracy.

Whole genome sequences (WGS) greatly increase our ability to precisely infer population genetic parameters, demographic processes, and selection signatures. However, WGS may still be not affordable for a representative number of individuals/populations. In this context, our goal was to assess the efficiency of several SNP genotyping strategies by testing their ability to accurately estimate parameters describing neutral diversity and to detect signatures of selection. We analysed 110 WGS at 12× coverage for four different species, i.e., sheep, goats and their wild counterparts. From these data we generated 946 data sets corresponding to random panels of 1K to 5M variants, commercial SNP chips and exome capture, for sample sizes of five to 48 individuals. We also extracted low-coverage genome resequencing of 1×, 2× and 5× by randomly subsampling reads from the 12× resequencing data. Globally, 5K to 10K random variants were enough for an accurate estimation of genome diversity. Conversely, commercial panels and exome capture displayed strong ascertainment biases. Besides the characterization of neutral diversity, the detection of the signature of selection and the accurate estimation of linkage disequilibrium (LD) required high-density panels of at least 1M variants. Finally, genotype likelihoods increased the quality of variant calling from low coverage resequencing but proportions of incorrect genotypes remained substantial, especially for heterozygote sites. Whole genome resequencing coverage of at least 5× appeared to be necessary for accurate assessment of genomic variations. These results have implications for studies seeking to deploy low-density SNP collections or genome scans across genetically diverse populations/species showing similar genetic characteristics and patterns of LD decay for a wide variety of purposes.

Sheep, the Jaagsiekte sheep retrovirus (JSRV) and its endogenous forms (enJSRVs) are a good model to study long-time relationships between retroviruses and their hosts. Taking advantage of 76 whole genome resequencing data of wild and domestic Ovis, we investigated the evolution of this relationship. An innovative analysis of re-sequencing data allowed characterizing 462 enJSRVs insertion sites (including 435 newly described insertions) in the Ovis genus. We focused our study on endogenous copies inserted in the q13 locus of chromosome 6 (6q13). Those copies are known to confer resistance against exogenous JSRV thanks to alleles bearing a mutation in the gag gene. We characterized (i) the distribution of protective and non-protective alleles across Ovis species and (ii) the copy number variation of the 6q13 locus. Our results challenged the previous hypothesis of fixation and amplification of the protective copies in relation with domestication, and allowed building a new model for the evolution of the 6q13 locus. JSRV would have integrated the 6q13 locus after the Ovis-Capra divergence (5-11 MYA) and before the Ovis diversification (2.4-5 MYA). The protective mutation in the enJSRV 6q13 copy appeared shortly after its insertion and was followed by genomic amplifications, after the divergence between Pachyform lineage on one side and the Argaliform and moufloniform lineages on the other (2.4-5 MYA). Considering the potential selective advantage of the protective mutation, its fixation in both sheep and its closest wild relative Ovis orientalis may be due to natural selection before domestication from O. orientalis populations.

Domestication fundamentally reshaped animal morphology, physiology and behaviour, offering the opportunity to investigate the molecular processes driving evolutionary change. Here we assess sheep domestication and artificial selection by comparing genome sequence from 43 modern breeds (Ovis aries) and their Asian mouflon ancestor (O. orientalis) to identify selection sweeps. Next, we provide a comparative functional annotation of the sheep genome, validated using experimental ChIP-Seq of sheep tissue. Using these annotations, we evaluate the impact of selection and domestication on regulatory sequences and find that sweeps are significantly enriched for protein coding genes, proximal regulatory elements of genes and genome features associated with active transcription. Finally, we find individual sites displaying strong allele frequency divergence are enriched for the same regulatory features. Our data demonstrate that remodelling of gene expression is likely to have been one of the evolutionary forces that drove phenotypic diversification of this common livestock species., M.N.-S. and Q.N. are funded by the CSIRO Science Excellence Research Office. M.P.-E. was funded by a CSIRO McMaster Visiting Fellowship. Mouflon genome sequences were generated by the NextGen Consortium, as part of the European Union’s Seventh Framework Programme (FP7/2010-2014 grant 244356).

How humans got their goatsLittle is known regarding the location and mode of the early domestication of animals such as goats for husbandry. To investigate the history of the goat, Dalyet al.sequenced mitochondrial and nuclear sequences from ancient specimens ranging from hundreds to thousands of years in age. Multiple wild populations contributed to the origin of modern goats during the Neolithic. Over time, one mitochondrial type spread and became dominant worldwide. However, at the whole-genome level, modern goat populations are a mix of goats from different sources and provide evidence for a multilocus process of domestication in the Near East. Furthermore, the patterns described support the idea of multiple dispersal routes out of the Fertile Crescent region by domesticated animals and their human counterparts.Science, this issue p.85

DNA barcoding approaches are used to describe biodiversity by analysing specimens or environmental samples in taxonomic, phylogenetic and ecological studies. While sharing data among these disciplines would be highly valuable, this remains difficult because of contradictory requirements. The properties making a DNA barcode efficient for specimen identification or species delimitation are hardly reconcilable with those required for a powerful analysis of degraded DNA from environmental samples. The use of next generation sequencing methods open up the way towards the development of new markers (e.g., multilocus barcodes) that would overcome such limitations. However, several challenges should be taken up for coordinating actions at the interface between taxonomy, ecology, molecular biology and bioinformatics in order to develop methods and protocols compatible with both taxonomic and ecological studies.

International audience; Although it is generally agreed that the Arctic flora is among the youngest and least diverse on Earth, the processes that shaped it are poorly understood. Here we present 50 thousand years (kyr) of Arctic vegetation history, derived from the first large-scale ancient DNA metabarcoding study of circumpolar plant diversity. For this interval we also explore nematode diversity as a proxy for modelling vegetation cover and soil quality, and diets of herbivorous megafaunal mammals, many of which became extinct around 10 kyr bp (before present). For much of the period investigated, Arctic vegetation consisted of dry steppe-tundra dominated by forbs (non-graminoid herbaceous vascular plants). During the Last Glacial Maximum (25-15 kyr bp), diversity declined markedly, although forbs remained dominant. Much changed after 10 kyr bp, with the appearance of moist tundra dominated by woody plants and graminoids. Our analyses indicate that both graminoids and forbs would have featured in megafaunal diets. As such, our findings question the predominance of a Late Quaternary graminoid-dominated Arctic mammoth steppe.

The Podismini are melanopline grasshoppers with a Holarctic distribution and well represented in the Eurasian fauna. To investigate their controversial taxonomy and evolutionary history, we studied 86%, 78% and 33% respectively of the Eurasian, European and Asian Palaearctic genera (Otte, 1995; Eades et al., 2013). We reconstructed parsimony, maximum likelihood and Bayesian phylogenies using fragments of four genes (ITS1, 16S, 12S, CO2). We applied a Bayesian molecular clock to estimate the times of species divergence, and the event-based parsimony method to depict the biogeographic framework of the diversification. Our results suggest that the selected Eurasian Podismini constitute a monophyletic group inside the Melanoplinae, provided it includes the North American genus Phaulotettix. The clades proposed by the present study inside the Podismini do not fit the older morphological or cytological classifications, but are in agreement with more recent proposals. Furthermore, our results can be explained by a plausible biogeographic history in which the present geographical distribution of the Eurasian Podismini resulted from known changes, to the Cenozoic climate and vegetation, induced by major geological events including the genesis of high mountain chains (e.g., Himalayas, Altay, Alps) and large deserts (e.g., Gobi, Karakoum, Taklamakan), and the opening of marginal seas (e.g., Bering, Japanese and Yellow Seas).

Ecosystems across the globe are threatened by climate change and human activities. New rapid survey approaches for monitoring biodiversity would greatly advance assessment and understanding of these threats. Taking advantage of next-generation DNA sequencing, we tested an approach we call metabarcoding: high-throughput and simultaneous taxa identification based on a very short (usually

Virtually all empirical ecological studies require species identification during data collection. DNA metabarcoding refers to the automated identification of multiple species from a single bulk sample containing entire organisms or from a single environmental sample containing degraded DNA (soil, water, faeces, etc.). It can be implemented for both modern and ancient environmental samples. The availability of next-generation sequencing platforms and the ecologists' need for high-throughput taxon identification have facilitated the emergence of DNA metabarcoding. The potential power of DNA metabarcoding as it is implemented today is limited mainly by its dependency on PCR and by the considerable investment needed to build comprehensive taxonomic reference libraries. Further developments associated with the impressive progress in DNA sequencing will eliminate the currently required DNA amplification step, and comprehensive taxonomic reference libraries composed of whole organellar genomes and repetitive ribosomal nuclear DNA can be built based on the well-curated DNA extract collections maintained by standardized barcoding initiatives. The near-term future of DNA metabarcoding has an enormous potential to boost data acquisition in biodiversity research.

DNA metabarcoding refers to the DNA-based identification of multiple species from a single complex and degraded environmental sample. We developed new sampling and extraction protocols suitable for DNA metabarcoding analyses targeting soil extracellular DNA. The proposed sampling protocol has been designed to reduce, as much as possible, the influence of local heterogeneity by processing a large amount of soil resulting from the mixing of many different cores. The DNA extraction is based on the use of saturated phosphate buffer. The sampling and extraction protocols were validated first by analysing plant DNA from a set of 12 plots corresponding to four plant communities in alpine meadows, and, second, by conducting pilot experiments on fungi and earthworms. The results of the validation experiments clearly demonstrated that sound biological information can be retrieved when following these sampling and extraction procedures. Such a protocol can be implemented at any time of the year without any preliminary knowledge of specific types of organisms during the sampling. It offers the opportunity to analyse all groups of organisms using a single sampling/extraction procedure and opens the possibility to fully standardize biodiversity surveys.

Background The current extensive use of the domestic goat (Capra hircus) is the result of its medium size and high adaptability as multiple breeds. The extent to which its genetic variability was influenced by early domestication practices is largely unknown. A common standard by which to analyze maternally-inherited variability of livestock species is through complete sequencing of the entire mitogenome (mitochondrial DNA, mtDNA). Results We present the first extensive survey of goat mitogenomic variability based on 84 complete sequences selected from an initial collection of 758 samples that represent 60 different breeds of C. hircus, as well as its wild sister species, bezoar (Capra aegagrus) from Iran. Our phylogenetic analyses dated the most recent common ancestor of C. hircus to ~460,000 years (ka) ago and identified five distinctive domestic haplogroups (A, B1, C1a, D1 and G). More than 90 % of goats examined were in haplogroup A. These domestic lineages are predominantly nested within C. aegagrus branches, diverged concomitantly at the interface between the Epipaleolithic and early Neolithic periods, and underwent a dramatic expansion starting from ~12–10 ka ago. Conclusions Domestic goat mitogenomes descended from a small number of founding haplotypes that underwent domestication after surviving the last glacial maximum in the Near Eastern refuges. All modern haplotypes A probably descended from a single (or at most a few closely related) female C. aegagrus. Zooarchaelogical data indicate that domestication first occurred in Southeastern Anatolia. Goats accompanying the first Neolithic migration waves into the Mediterranean were already characterized by two ancestral A and C variants. The ancient separation of the C branch (~130 ka ago) suggests a genetically distinct population that could have been involved in a second event of domestication. The novel diagnostic mutational motifs defined here, which distinguish wild and domestic haplogroups, could be used to understand phylogenetic relationships among modern breeds and ancient remains and to evaluate whether selection differentially affected mitochondrial genome variants during the development of economically important breeds. Electronic supplementary material The online version of this article (doi:10.1186/s12864-015-2342-2) contains supplementary material, which is available to authorized users.

The analysis of food webs and their dynamics facilitates understanding of the mechanistic processes behind community ecology and ecosystem functions. Having accurate techniques for determining dietary ranges and components is critical for this endeavour. While visual analyses and early molecular approaches are highly labour intensive and often lack resolution, recent DNA-based approaches potentially provide more accurate methods for dietary studies. A suite of approaches have been used based on the identification of consumed species by characterization of DNA present in gut or faecal samples. In one approach, a standardized DNA region (DNA barcode) is PCR amplified, amplicons are sequenced and then compared to a reference database for identification. Initially, this involved sequencing clones from PCR products, and studies were limited in scale because of the costs and effort required. The recent development of next generation sequencing (NGS) has made this approach much more powerful, by allowing the direct characterization of dozens of samples with several thousand sequences per PCR product, and has the potential to reveal many consumed species simultaneously (DNA metabarcoding). Continual improvement of NGS technologies, on-going decreases in costs and current massive expansion of reference databases make this approach promising. Here we review the power and pitfalls of NGS diet methods. We present the critical factors to take into account when choosing or designing a suitable barcode. Then, we consider both technical and analytical aspects of NGS diet studies. Finally, we discuss the validation of data accuracy including the viability of producing quantitative data.

The present study characterizes the mitochondrial DNA diversity of Moroccan goats. 150 goats of different phenotypic entities were sampled over four geographic regions covering most of the Moroccan territory and the HVI segment of their mitochondrial DNA (mtDNA) control region was sequenced. The 150 Moroccan goats represented 97 haplotypes for this mtDNA fragment. Most of this diversity was present within phenotypic entities and within geographic regions. This weak genetic structure may result from the fact that all haplotypes were already mixed in the populations that colonized Morocco and/or the existence of recurrent gene flows from Mediterranean routes. Comparing the Moroccan haplotype diversity to that of 21 haplotypes representative of the worldwide diversity showed that all the Moroccan goats studied belonged to the A haplogroup that is preponderant in the world. The haplotypes of the Northern region appeared to be less diverse, what would probably reflect a stronger founder effect in this region.

New insights for the systematic and evolution of the wild sheep are provided by molecular phylogenies inferred from Maximum parsimony, Bayesian, Maximum likelihood, and Neighbor-Joining methods. The phylogeny of the wild sheep was based on cytochrome b sequences of 290 samples representative of most of the sub-species described in the genus Ovis. The result was confirmed by a combined tree based on cytochrome b and nuclear sequences for 79 Ovis samples representative of the robust clades established with mitochondrial data. Urial and mouflon, which are either considered as a single or two separate species, form two monophyletic groups (O. orientalis and O. vignei). Their hybrids appear in one or the other group, independently from their geographic origin. The European mouflon O. musimon is clearly in the O. orientalis clade. The others species, O. dalli, O. canadensis, O. nivicola, and O. ammon are monophyletic. The results support an Asiatic origin of the genus Ovis, followed by a migration to North America through North-Eastern Asia and the Bering Strait and a diversification of the genus in Eurasia less than 3 million years ago. Our results show that the evolution of the genus Ovis is a striking example of successive speciation events occurring along the migration routes propagating from the ancestral area.

The development of DNA barcoding (species identification using a standardized DNA sequence), and the availability of recent DNA sequencing techniques offer new possibilities in diet analysis. DNA fragments shorter than 100-150 bp remain in a much higher proportion in degraded DNA samples and can be recovered from faeces. As a consequence, by using universal primers that amplify a very short but informative DNA fragment, it is possible to reliably identify the plant taxon that has been eaten. According to our experience and using this identification system, about 50% of the taxa can be identified to species using the trnL approach, that is, using the P6 loop of the chloroplast trnL (UAA) intron. We demonstrated that this new method is fast, simple to implement, and very robust. It can be applied for diet analyses of a wide range of phytophagous species at large scales. We also demonstrated that our approach is efficient for mammals, birds, insects and molluscs. This method opens new perspectives in ecology, not only by allowing large-scale studies on diet, but also by enhancing studies on resource partitioning among competing species, and describing food webs in ecosystems.

International audience; For about 10 000 years, farmers have been managing cattle, sheep, and goats in a sustainable way, leading to animals that are well adapted to the local conditions. About 200 years ago, the situation started to change dramatically, with the rise of the concept of breed. All animals from the same breed began to be selected for the same phenotypic characteristics, and reproduction among breeds was seriously reduced. This corresponded to a strong fragmentation of the initial populations. A few decades ago, the selection pressures were increased again in order to further improve productivity, without enough emphasis on the preservation of the overall genetic diversity. The efficiency of modern selection methods successfully increased the production, but with a dramatic loss of genetic variability. Many industrial breeds now suffer from inbreeding, with effective population sizes falling below 50. With the development of these industrial breeds came economic pressure on farmers to abandon their traditional breeds, and many of these have recently become extinct as a result. This means that genetic resources in cattle, sheep, and goats are highly endangered, particularly in developed countries. It is therefore important to take measures that promote a sustainable management of these genetic resources; first, by in situ preservation of endangered breeds; second, by using selection programmes to restore the genetic diversity of industrial breeds; and finally, by protecting the wild relatives that might provide useful genetic resources.

International audience; Fungal communities are key components of soil, but the study of their ecological significance is limited by a lack of appropriated methods. For instance, the assessment of fungi occurrence and spatio-temporal variation in soil requires the analysis of a large number of samples. The molecular signature methods provide a useful tool to monitor these microbial communities and can be easily adapted to capillary electrophoresis (CE) allowing high-throughput studies. Here we assess the suitability of CE-FLA (Fragment Length Polymorphism, denaturing conditions) and CE-SSCP (Single-Stranded Conformation Polymorphism, native conditions) applied to environmental studies since they require a short molecular marker and no post-PCR treatments. We amplified the ITSI region from 22 fungal strains isolated from an alpine ecosystem and from total genomic DNA of alpine and infiltration basin soils. The CE-FLA and CE-SSCP separated 17 and 15 peaks respectively from a mixture of 19 strains. For the alpine soil-metagenomic DNA, the FLA displayed more peaks than the SSCP and the converse result was found for infiltration basin sediments. We concluded that CE-FLA and CE-SSCP of ITS I region provided complementary information. In order to improve CE-SSCP sensitivity. we tested its resolution according to migration temperature and found 32 degrees C to be optimal. Because of their simplicity, quickness and reproducibility. we found that these two methods were promising for high-throughput studies of soil fungal communities. (c) 2007 Elsevier B.V All rights reserved.

Times Cited: 1; International audience; In conservation biology genetic diversity is recognized as an important criterion to consider when prioritizing populations for protection. Today, population genomics offers the opportunity to evaluate both neutral and adaptive components of genetic diversity directly at the genome level with molecular tools. By screening the genome with many genetic markers, it is possible to detect loci supposedly under natural selection and thus of adaptive significance. We devised a new diversity index, the population adaptive index (PAI), which accounts for the adaptive value of the population it refers to. To estimate this index, we performed a genome scan with amplified fragment length polymorphism markers to identify neutral and selected loci in several populations of a widespread amphibian (common frog, Rana temporaria) and a threatened plant (Austrian dragonhead, Dracocephalum austriacum L.). We then investigated four different conservation strategies aimed at protecting the maximum amount of genetic diversity (neutral or selected). In particular we explored the relevance of the principle of complementarity, usually applied to the protection of species, in the management of intraspecific diversity. This principle advocates the conservation of sets of units that together maximize the species' orgenetic diversity, which is in opposition to the traditional approach of targeting populations that are the most diverse individually. Four major conclusions emerged from these results. First, the PAI seemed to be a valuable index to evaluate the adaptive diversities within populations. Second, in the two species, the neutral and adaptive diversities within and among populations were not correlated, so conservation strategies based on the neutral and adaptive indexes would not select the same populations for protection. Third, because of its efficiency in conserving genetic diversity, the principle of complementarity deserves to be used more often for this purpose. Fourth, when neutral and adaptive results conflict, additional arguments (e.g., demography, ecology, and geographic proximity) should be considered together with levels of genetic diversity to determine a conservation strategy.

International audience; Life history and spatio-temporal patterns of resource utilisation were characterised in four Chiastocheta (Diptera: Anthomyiidae) species, whose larvae compete as seed predators on Trollius europaeus fruits. Interspecific co-occurrence was observed in 80% of the resource patches (= Trollius fruits) in the two communities studied. Isolated larvae from all species had a similar food intake, but differed in development time and size at emergence Different species exhibit contrasting resource exploitation strategies with specific mining patterns and a partial temporal shift. Two species exhibited particularly singular strategies. C. rotundiventris escaped from strong interactions with other species because it was the first species to develop and the only one to exploit the central pith of Trollius fruits. The key role of this species as the main pollinator of the host-plant appears to be a by-product of constraints imposed by occupying a restricted niche. Although the resource is ephemeral due to seed dispersal, C. dentifera, the last species to oviposit, is not disadvantaged because it has a short development time and rapid food intake. The different patterns can partly explain the stability of Chiastocheta communities, but do not prevent competition to occur at high larval densities. (c) 2006 Elsevier SAS. All rights reserved.

Genotyping errors occur when the genotype determined after molecular analysis does not correspond to the real genotype of the individual under consideration. Virtually every genetic data set includes some erroneous genotypes, but genotyping errors remain a taboo subject in population genetics, even though they might greatly bias the final conclusions, especially for studies based on individual identification. Here, we consider four case studies representing a large variety of population genetics investigations differing in their sampling strategies (noninvasive or traditional), in the type of organism studied (plant or animal) and the molecular markers used [microsatellites or amplified fragment length polymorphisms (AFLPs)]. In these data sets, the estimated genotyping error rate ranges from 0.8% for microsatellite loci from bear tissues to 2.6% for AFLP loci from dwarf birch leaves. Main sources of errors were allelic dropouts for microsatellites and differences in peak intensities for AFLPs, but in both cases human factors were non-negligible error generators. Therefore, tracking genotyping errors and identifying their causes are necessary to clean up the data sets and validate the final results according to the precision required. In addition, we propose the outline of a protocol designed to limit and quantify genotyping errors at each step of the genotyping process. In particular, we recommend (i) several efficient precautions to prevent contaminations and technical artefacts; (ii) systematic use of blind samples and automation; (iii) experience and rigor for laboratory work and scoring; and (iv) systematic reporting of the error rate in population genetics studies.

Environmental DNA (eDNA) metabarcoding is increasingly used to study the present and past biodiversity. eDNA analyses often rely on amplification of very small quantities or degraded DNA. To avoid missing detection of taxa that are actually present (false negatives), multiple extractions and amplifications of the same samples are often performed. However, the level of replication needed for reliable estimates of the presence/absence patterns remains an unaddressed topic. Furthermore, degraded DNA and PCR/sequencing errors might produce false positives. We used simulations and empirical data to evaluate the level of replication required for accurate detection of targeted taxa in different contexts and to assess the performance of methods used to reduce the risk of false detections. Furthermore, we evaluated whether statistical approaches developed to estimate occupancy in the presence of observational errors can successfully estimate true prevalence, detection probability and false-positive rates. Replications reduced the rate of false negatives; the optimal level of replication was strongly dependent on the detection probability of taxa. Occupancy models successfully estimated true prevalence, detection probability and false-positive rates, but their performance increased with the number of replicates. At least eight PCR replicates should be performed if detection probability is not high, such as in ancient DNA studies. Multiple DNA extractions from the same sample yielded consistent results; in some cases, collecting multiple samples from the same locality allowed detecting more species. The optimal level of replication for accurate species detection strongly varies among studies and could be explicitly estimated to improve the reliability of results.

International audience; Roads are of major concern in conservation biology, as they are known to restrict animal movements through landscape fragmentation, and may therefore impact genetic patterns in native terrestrial organisms. We assessed the effect of two large-scale transportation infrastructures (LTIs), a 40-year-old highway and a 30 year-old high-speed railway, on the spatial genetic structure of the alpine newt Ichthyosaura alpestris, a highly nomadic amphibian. Genetic data were gathered following a targeted individual-based sampling scheme and analysed using both overlay and correlative methods. While simulations suggested that the highway may be old enough for a significant barrier effect to be detected, LTIs were never detected as barriers to gene flow: inferred genetic boundaries rather coincided with transition zones between major landscape entities. Furthermore, spatial principal component analysis, a method designed to reveal cryptic genetic spatial patterns in high gene flow species, counter-intuitively suggested that the highway may act as a potential dispersal corridor in low-quality habitats, thus challenging traditional hypotheses on road impacts in amphibians. Our study showed that considering local interactions between species, infrastructures and landscape-specific characteristics is essential for better understanding the potential impacts of roads on movement patterns in terrestrial organisms.

International audience; The interactions between botanical composition of pasture, quality of herbage grazed, performances of dairy cows and sensory and nutritional properties of dairy products were investigated using an integrated system approach. Two contrasting grazing systems were evaluated from May to September in two years. The treatments included a continuous grazing system (DIV) managed at a lenient stocking rate (1.0 LU ha−1) on a botanically-rich permanent pasture, and a rotational grazing system (PROD) set up at a higher stocking rate (1.7 LU ha−1) on a former temporary grassland presenting moderate biodiversity. DIV aimed to maximize biodiversity and obtain high sensory and nutritional quality cheese, whereas PROD was oriented towards milk production and herbage quality. In each system, 12 non-feed-supplemented Montbéliarde cows were used. The DIV system led to higher milk production per cow in the early grazing season than the PROD system (22.2 vs. 19.9 kg d−1). At the beginning of summer, this milk production pattern was inverted following a decrease in grass nutritive value in the DIV system. In parallel, DIV cows showed a more marked loss of body condition than PROD cows over the season. In terms of milk fatty acid profile, the DIV system proved very interesting early in the grazing season but lost its value over time as the herbage matured. Cheese sensory properties differed between systems only after a long ripening period (6 months). Regarding the ecological performances, the DIV plot showed greater botanical and entomological biodiversity than the PROD plot. This study provides evidence that the balance between animal performances, dairy product quality and biodiversity in dairy systems is more complex than previously thought, since the expected benefits of each system vary markedly over periods. The evolution of herbage vegetation stage during the grazing season combined with the botanical composition of the pasture is a key component for understanding these variations.

Summary 1. In obligate plant/seed parasite–pollinator mutualisms, the plant is exclusively pollinated by an insect whose larvae are specific seed predators. Hence, outcomes of the interaction for the plant can vary with the number of eggs laid and the number of seeds eaten per larva. 2. In the work reported here, predation by Chiastocheta larvae on seeds of Trollius europaeus was analysed as a function of the number of eggs laid on the flower. Flowers with an increasing number of eggs were bagged in three populations and seeds were counted after the end of larval predation, in order to assess whether there was competition among larvae. 3. Seed predation on single-egg flowers was high and variable (mean per population ranging from 15 to 40% of the developed seeds). Seed predation increased weakly with increasing egg load and was lower than gross seed production (always

Biological control agents such as entomophagus insects (e.g., Trichogramma sp.) are important components of integrated pest management (IPM) programs. Because pesticides are also an important component of IPM programs, it is essential to understand how biological control agents are affected by pesticides and how these effects may affect the effectiveness of the biological control agents. Contrary to most insect species that use volatile pheromones for mate location, T. brassicae is one of the few species using a substrate-borne sex pheromone for mating. In this work we determined the effects of an organophosphorus insecticide, chlorpyrifos, on sex pheromone reception by males and emission by virgin females. The insects used in the pheromone tests were survivors from an acute toxicity test in which individuals were exposed to a dose of chlorpyrifos equivalent to a 20% lethal dose. Males that survived the insecticide exposure spent much less time on the area marked with the female pheromone than control males (mean choice indexes, 0.46 [0.34 SD] vs 0.70 [0.25 SD], p < 0.001). The kinetics of the response of males to marking by control and treated females indicated that chlorpyrifos decreased the emission of sex pheromone by females surviving the insecticide. Therefore, chlorpyrifos inhibited pheromone reception by males and decreased pheromone emission by virgin females. These results are interpreted in the context of sex pheromone regulation by the nervous system of insects and according to the mode of action of chlorpyrifos. Extensions of these results on actual work on sex pheromone, the use of pheromone traps, and the population biology of Trichogramma sp. are discussed.

While the use of chemical cues in host location is well known in parasitoid insects, little is known about their role in sexual communication. Sex pheromones were investigated in the egg parasitoid Trichogramma brassicae, and the evidence for a substrate-borne pheromone was demonstrated. Mated and unmated males concentrated their walking in an area where virgin females (

La caractérisation du régime alimentaire des ruminants au pâturage en milieu diversifié (prairies d’association et permanentes) est un verrou méthodologique important dans les études portant sur les relations animal-plantes dans ce type de milieu. Le développement récent de techniques permettant l’identification des végétaux dans des substrats complexes ou dégradés (DNA metabarcoding), basées sur l’analyse de fragments résiduels d’ADN des plantes dans les produits de la digestion, pourrait permettre une innovation méthodologique complémentaire aux méthodes actuelles basées sur les examens micro-histologiques, l’extraction et le dosage des alcanes à longue chaîne contenus dans les cuticules des plantes, ou les méthodes spectrales (spectroscopie en proche Infrarouge), ces méthodes étant fastidieuses, peu précises et mal adaptées aux couverts végétaux complexes. L’objectif de ce travail était de tester l’aspect quantitatif de cette méthodologie, déjà sensible pour apprécier qualitativement la composition d’une ration (Farruggia et al., 2012). Des échantillons de produits de la digestion (contenus ruminaux et fèces) ont été prélevés lors d’expérimentations au cours desquelles des espèces fourragères ont été distribuées à des moutons à l’auge en différentes proportions bien définies (mélanges binaires ray-grass anglais (RGA)-légumineuses à l’INRA de Theix, en 2010 et 2011) ou à des vaches laitières au pâturage dans des proportions mesurées par la méthode des alcanes (mélanges de une à cinq espèces à l’INRA de Rennes en 2012). Ces échantillons ont été analysés par le LECA en utilisant un couple d’amorces universelles ciblant une région variable de l’intron du gène chloroplastique trnL(UAA) des végétaux, un fragment adéquat pour l’identification de l’ADN résiduel dans les produits de la digestion car la région amplifiée est à la fois très courte et très variable selon les espèces. Puis, après séquençage massif de ces fragments, le nombre relatif de séquences obtenues dans le produit PCR pour chaque espèce végétale a été comparé à la proportion relative de ces espèces dans l’alimentation des animaux. Les résultats obtenus à partir des mélanges binaires contrôlés montrent une très bonne quantification des proportions de RGA et de légumineuses ingérées (en %, 0:100, 25:75, 50:50, 75:25 et 100:0) lorsque les analyses de DNA metabarcoding ont été réalisées sur contenus ruminaux, et ceci lors des deux années d’étude (mélanges RGA-luzerne et RGA-trèfle blanc, respectivement) (Fig. 1). Les résultats étaient plus mitigés lorsque les analyses de DNA metabarcoding ont été réalisées sur les fèces, avec une quantification correcte avec le mélange RGA-luzerne, bien que la variabilité soit élevée, mais moins bonne avec le mélange RGA-trèfle blanc pour lequel les proportions de RGA étaient sous-estimées (Fig. 1). Il est encore difficile, à ce stade, de déterminer pourquoi la quantification est de moins bonne qualité avec les fèces qu’avec les contenus ruminaux, mais il semble que l’ADN des fèces soit moins efficacement récupéré. Ceci pourrait s’expliquer par un différentiel dans la digestibilité post-ruminale des espèces ingérées, produisant un biais en raison de la méthode d’extraction de l’ADN utilisée qui tend à favoriser la récupération de l’ADN intracellulaire. Une autre explication pourrait résider dans l'assignation des séquences aux espèces : un certain niveau d'erreur est toléré dans l'assignation pour prendre en compte les erreurs de PCR et séquençage. Les ADN étant plus dégradés dans les fèces, un biais d'assignation pourrait être présent qui n'existe pas avec le contenu ruminal. Les échantillons prélevés en 2012 sur vaches laitières sont en cours d’analyse et complèteront les résultats déjà acquis.

International audience; Genetic data are increasingly used in landscape ecology for the indirect assessment of functional connectivity, that is, the permeability of landscape to movements of organ- isms. Among available tools, matrix correlation analyses (e.g. Mantel tests or mixed models) are commonly used to test for the relationship between pairwise genetic dis- tances and movement costs incurred by dispersing individuals. When organisms are spatially clustered, a population-based sampling scheme (PSS) is usually performed, so that a large number of genotypes can be used to compute pairwise genetic distances on the basis of allelic frequencies. Because of financial constraints, this kind of sam- pling scheme implies a drastic reduction in the number of sampled aggregates, thereby reducing sampling coverage at the landscape level. We used matrix correlation analy- ses on simulated and empirical genetic data sets to investigate the efficiency of an individual-based sampling scheme (ISS) in detecting isolation-by-distance and isola- tion-by-barrier patterns. Provided that pseudo-replication issues are taken into account (e.g. through restricted permutations in Mantel tests), we showed that the use of inter- individual measures of genotypic dissimilarity may efficiently replace interpopulation measures of genetic differentiation: the sampling of only three or four individuals per aggregate may be sufficient to efficiently detect specific genetic patterns in most situa- tions. The ISS proved to be a promising methodological alternative to the more con- ventional PSS, offering much flexibility in the spatial design of sampling schemes and ensuring an optimal representativeness of landscape heterogeneity in data, with few aggregates left unsampled. Each strategy offering specific advantages, a combined use of both sampling schemes is discussed.