Olivier Hanotte

Genome-Wide Survey of SNP Variation Uncovers the Genetic Structure of Cattle Breeds

A survey of genetic diversity of cattle suggests two domestication events in Asia and selection by husbandry. Not Just Dinner on Legs Several thousand years ago, human beings realized the virtues of domesticating wild animals as easy meat. Soon other possibilities became apparent, and as revealed in a series of papers in this issue, early pastoralists became selective about breeding for wool, leather, milk, and muscle power. In two papers, Gibbs et al. report on the bovine genome sequence (p. 522; see the cover, the Perspective by Lewin, and the Policy Forum by Roberts) and trace the diversity and genetic history of cattle (p. 528), while Chessa et al. (p. 532) survey the occurrence of endogenous retroviruses in sheep and map their distribution to historical waves of human selection and dispersal across Europe. Finally, Ludwig et al. (p. 485) note the origins of variation in the coat-color of horses and suggest that it is most likely to have been selected for by humans in need of good-looking transport. The imprints of domestication and breed development on the genomes of livestock likely differ from those of companion animals. A deep draft sequence assembly of shotgun reads from a single Hereford female and comparative sequences sampled from six additional breeds were used to develop probes to interrogate 37,470 single-nucleotide polymorphisms (SNPs) in 497 cattle from 19 geographically and biologically diverse breeds. These data show that cattle have undergone a rapid recent decrease in effective population size from a very large ancestral population, possibly due to bottlenecks associated with domestication, selection, and breed formation. Domestication and artificial selection appear to have left detectable signatures of selection within the cattle genome, yet the current levels of diversity within breeds are at least as great as exists within humans.

Noninvasive genetic tracking of the endangered Pyrenean brown bear population

Pyrenean brown bears Ursus arctos are threatened with extinction. Management efforts to preserve this population require a comprehensive knowledge of the number and sex of the remaining individuals and their respective home ranges. This goal has been achieved using a combination of noninvasive genetic sampling of hair and faeces collected in the field and corresponding track size data. Genotypic data were collected at 24 micrdsatellite loci using a rigorous multiple‐tubes approach to avoid genotyping errors associated with low quantities of DNA. Based on field and genetic data, the Pyrenean population was shown to be composed at least of one yearling, three adult males, and one adult female. These data indicate that extinction of the Pyrenean brown bear population is imminent without population augmentation. To preserve the remaining Pyrenean gene pool and increase genetic diversity, we suggest that managers consider population augmentation using only females. This study demonstrates that comprehensive knowledge of endangered small populations of mammals can be obtained using noninvasive genetic sampling.

Genome 10K: A Proposal to Obtain Whole-Genome Sequence for 10 000 Vertebrate Species

The human genome project has been recently complemented by whole-genome assessment sequence of 32 mammals and 24 nonmammalian vertebrate species suitable for comparative genomic analyses. Here we anticipate a precipitous drop in costs and increase in sequencing efficiency, with concomitant development of improved annotation technology and, therefore, propose to create a collection of tissue and DNA specimens for 10,000 vertebrate species specifically designated for whole-genome sequencing in the very near future. For this purpose, we, the Genome 10K Community of Scientists (G10KCOS), will assemble and allocate a biospecimen collection of some 16,203 representative vertebrate species spanning evolutionary diversity across living mammals, birds, nonavian reptiles, amphibians, and fishes (ca. 60,000 living species). In this proposal, we present precise counts for these 16,203 individual species with specimens presently tagged and stipulated for DNA sequencing by the G10KCOS. DNA sequencing has ushered in a new era of investigation in the biological sciences, allowing us to embark for the first time on a truly comprehensive study of vertebrate evolution, the results of which will touch nearly every aspect of vertebrate biological enquiry.

Geographic distribution and frequency of a taurine Bos taurus and an indicine Bos indicus Y specific allele amongst sub‐Saharan African cattle breeds

We report for the first time, and for the whole of sub‐Saharan Africa, the geographical distribution and the frequency of an indicine and a taurine Y specific allele amongst African cattle breeds. A total of 984 males from 69 indigenous African populations from 22 countries were analysed at the microsatellite locus INRA 124. The taurine allele is probably the oldest one on the continent. However, the taurine and the indicine alleles were present in 291 males (30%), and 693 males (70%), respectively. More particularly, 96% of zebu males (n = 470), 50% of taurine males (n = 263), 29% of sanga males (crossbreed Bos taurus × Bos indicus, n = 263) and 95% of zebu × sanga crossbred males (n = 56) had the indicine allele. The Borgou, a breed classified as zebu × taurine cross showed only the zebu allele (n = 12). The indicine allele dominates today in the Abyssinian region, a large part of the Lake Victoria region and the sahelian belt of West Africa. All the sanga males (n = 64) but only one from the Abyssinian region had the indicine allele. The taurine allele is the commonest only among the sanga breeds of the southern African region and the trypanotolerant taurine breeds of West Africa. In West Africa and in the southern Africa regions, zones of introgression were detected with breeds showing both Y chromosome alleles. Our data also reveal a pattern of male zebu introgression in Mozambique and Zimbabwe, probably originating from the Mozambique coast. The sanga cattle from the Lake Victoria region and the Kuri cattle of Lake Chad, cattle populations surrounded by zebu breeds were, surprisingly, completely devoid of the indicine allele. Human migration, phenotypic preferences by the pastoralists, adaptation to specific habitats and to specific diseases are the main factors explaining the present‐day distribution of the alleles in sub‐Saharan Africa.

Resolving the evolution of extant and extinct ruminants with high-throughput phylogenomics

The Pecorans (higher ruminants) are believed to have rapidly speciated in the Mid-Eocene, resulting in five distinct extant families: Antilocapridae, Giraffidae, Moschidae, Cervidae, and Bovidae. Due to the rapid radiation, the Pecoran phylogeny has proven difficult to resolve, and 11 of the 15 possible rooted phylogenies describing ancestral relationships among the Antilocapridae, Giraffidae, Cervidae, and Bovidae have each been argued as representations of the true phylogeny. Here we demonstrate that a genome-wide single nucleotide polymorphism (SNP) genotyping platform designed for one species can be used to genotype ancient DNA from an extinct species and DNA from species diverged up to 29 million years ago and that the produced genotypes can be used to resolve the phylogeny for this rapidly radiated infraorder. We used a high-throughput assay with 54,693 SNP loci developed for Bos taurus taurus to rapidly genotype 678 individuals representing 61 Pecoran species. We produced a highly resolved phylogeny for this diverse group based upon 40,843 genome-wide SNP, which is five times as many informative characters as have previously been analyzed. We also establish a method to amplify and screen genomic information from extinct species, and place Bison priscus within the Bovidae. The quality of genotype calls and the placement of samples within a well-supported phylogeny may provide an important test for validating the fidelity and integrity of ancient samples. Finally, we constructed a phylogenomic network to accurately describe the relationships between 48 cattle breeds and facilitate inferences concerning the history of domestication and breed formation.

Worldwide Patterns of Ancestry, Divergence, and Admixture in Domesticated Cattle

The domestication and development of cattle has considerably impacted human societies, but the histories of cattle breeds and populations have been poorly understood especially for African, Asian, and American breeds. Using genotypes from 43,043 autosomal single nucleotide polymorphism markers scored in 1,543 animals, we evaluate the population structure of 134 domesticated bovid breeds. Regardless of the analytical method or sample subset, the three major groups of Asian indicine, Eurasian taurine, and African taurine were consistently observed. Patterns of geographic dispersal resulting from co-migration with humans and exportation are recognizable in phylogenetic networks. All analytical methods reveal patterns of hybridization which occurred after divergence. Using 19 breeds, we map the cline of indicine introgression into Africa. We infer that African taurine possess a large portion of wild African auroch ancestry, causing their divergence from Eurasian taurine. We detect exportation patterns in Asia and identify a cline of Eurasian taurine/indicine hybridization in Asia. We also identify the influence of species other than Bos taurus taurus and B. t. indicus in the formation of Asian breeds. We detect the pronounced influence of Shorthorn cattle in the formation of European breeds. Iberian and Italian cattle possess introgression from African taurine. American Criollo cattle originate from Iberia, and not directly from Africa with African ancestry inherited via Iberian ancestors. Indicine introgression into American cattle occurred in the Americas, and not Europe. We argue that cattle migration, movement and trading followed by admixture have been important forces in shaping modern bovine genomic variation.

Polymorphic microsatellites in the blue tit Parus caeruleus and their cross‐species utility in 20 songbird families

© 2000 Blackwell Science Ltd, Molecular Ecology, 9, 1919–1952 Using the Primer 3 computer program (Rozen & Skaletsky 1997), primers could be designed to microsatellite-flanking regions of 37 clones. PCR was performed in 25 μL volumes containing 1.6 mm MgCl2, 0.2 mm dCTP, dGTP and dTTP, 0.02 mm dATP, 0.06 μL [α32P]dATP, 75 mm Tris–HCl (pH 9), 20 mm (NH4)2SO4, 0.01% Tween-20, 5 pmol of each primer, 0.5 units Taq DNA polymerase (Eurogentec) and 10 ng of template DNA. After initial denaturation (95 °C, 20 s), PCR was run for 35 cycles (94 °C for 20 s, 53 °C for 25 s, 67 °C for 23 s) in a Perkin-Elmer 2400 thermocycler. Products were separated on 6% sequencing gels and autoradiographed (Sambrook et al. 1989). Primer functionality was initially tested on three A. rabiei isolates, and one isolate of A. fabae. Using an annealing temperature of 53 °C, single bands of the expected size were obtained for 26 loci. Twenty-four primer pairs also amplified one or more products from A. fabae. Whether these fragments contain microsatellites and/or are polymorphic within A. fabae remains to be examined. Twenty marker loci were selected to genotype 22 A. rabiei isolates from the USA, Pakistan, Syria, Turkey and Tunisia. Each of these constituted a unique clonal lineage as defined by its fingerprint haplotype (Geistlinger et al. 1997). The Popgene computer package (Yeh & Boyle 1997) was used to calculate allele frequencies and to determine Nei’s expected genetic diversity valves for haploid organisms (Nei 1987; Tenzer et al. 1999; Table 1). All loci were polymorphic, with 2–14 alleles and genetic diversity values of 0.17–0.90. With up to 14 alleles among 22 isolates, these microsatellites are more variable than other types of molecular markers applied to A. rabiei. Locus-specific primers not only ensure a high reproducibility of results, but also allow the analysis of mixed samples. Groppe & Boller (1997) have specifically amplified a microsatellite locus from the fungal endophyte Epichloe in the presence of contaminating host DNA. Following a similar strategy, microsatellite markers could be used to type A. rabiei isolates directly from individual lesions. The distribution of A. rabiei pathotypes in the field could then be monitored more efficiently, avoiding the lengthy procedures of single-sporing and culturing.

An approach to the optimal allocation of conservation funds to minimize loss of genetic diversity between livestock breeds

About 30% of all farm animal breeds worldwide are at risk of extinction. To prevent this irreversible erosion of genetic diversity, the limited funds available for conservation need to be allocated in the most efficient way. Applying the diversity concept of Weitzman [Quart. J. Econ. CVII (1992) 363; Quart. J. Econ. CVIII (1993) 157] this paper presents a framework for the allocation of a given budget among a set of breeds such that the expected amount of between-breed diversity conserved is maximized. As a novel methodological contribution, a functional relationship between conservation funds spent in one population and the conservation effect in terms of reduced extinction probability is suggested. Based on arguments from population genetics, three different functions are derived, which may reflect the range of possible functions in typical conservation situations. The methodology is illustrated with an example of 23 African zebu and zenga cattle breeds. The results indicate that conservation funds should be spent on only three to nine of the 23 breeds, depending on the model used. Highest priority is given to breeds, for which the ‘conservation potential’, that is, the product of extinction probability and marginal diversity is maximum, and these are not necessarily the most endangered breeds. The methodology can be extended to the maximization of total utility, which incorporates diversity, as well as other direct use, and special value, characteristics. However, a number of essential input parameters such as extinction probabilities and economic values are lacking and realistic models for developing cost-efficient conservation strategies have to be derived. Given these lacking bits of information become available, the methodology suggested provides a breakthrough towards applicability of diversity-based approaches for decision taking in conservation programs. # 2003 Elsevier Science B.V. All rights reserved.

Hybridization of banteng (Bos javanicus) and zebu (Bos indicus) revealed by mitochondrial DNA, satellite DNA, AFLP and microsatellites

Hybridization between wild and domestic bovine species occurs worldwide either spontaneously or by organized crossing. We have analysed hybridization of banteng (Bos javanicus) and zebu (Bos indicus) in south-east Asian cattle using mitochondrial DNA (PCR-RFLP and sequencing), AFLP, satellite fragment length polymorphisms (SFLP or PCR-RFLP of satellite DNA) and microsatellite genotyping. The Indonesian Madura zebu breed is reputed to be of hybrid zebu–banteng origin, but this has never been documented and Bali cattle are considered to be a domesticated form of banteng. The banteng mitochondrial type was found in all animals sampled on the isle of Bali, Indonesia, but only in 35% of the animals from a Malaysian Bali-cattle population. The Madura animals also carried mitochondrial DNA of either zebu and banteng origin. In both populations, zebu introgression was confirmed by AFLP and SFLP. Microsatellite analysis of the Malaysian Bali population revealed for 12 out of 15 loci screened, Bali-cattle-specific alleles, several of which were also found in wild banteng animals. The tools we have described are suitable for the detection of species in introgression studies, which are essential for the genetic description of local breeds and the preservation of their economic and cultural value.

Extra-pair paternity and intraspecific brood parasitism in the European starling, Sturnus vulgaris: evidence from DNA fingerprinting

Abstract Abstract. The frequency of extra-pair paternity and intraspecific brood parasitism was determined, using multilocus DNA fingerprinting, in a single population of the European starling during two subsequent breeding seasons. The results were compared with behavioural data about the rates of extra-pair copulation and intraspecific brood parasitism. Fourteen families including 62 chicks were analysed. Behavioural observations suggested a low level of intraspecific brood parasitism and indeed only one case was detected by DNA fingerprinting. Extra-pair copulations were not observed, but the DNA fingerprint data revealed six offspring (9·7%) from four broods (28·6%) where the putative father of the chick was not the genetic father. One brood was fathered by three different males. Behavioural observations suggested that at least three cases of extra-pair paternity (three offspring from two broods) could have arisen through rapid mate replacement rather than through extra-pair copulations. There was no significant evidence that polygynous males suffer from a higher risk of being cuckolded than monogamous males.