Summary Horse domestication revolutionized warfare and accelerated travel, trade, and the geographic expansion of languages. Here, we present the largest DNA time series for a non-human organism to date, including genome-scale data from 149 ancient animals and 129 ancient genomes (≥1-fold coverage), 87 of which are new. This extensive dataset allows us to assess the modern legacy of past equestrian civilizations. We find that two extinct horse lineages existed during early domestication, one at the far western (Iberia) and the other at the far eastern range (Siberia) of Eurasia. None of these contributed significantly to modern diversity. We show that the influence of Persian-related horse lineages increased following the Islamic conquests in Europe and Asia. Multiple alleles associated with elite-racing, including at the MSTN “speed gene,” only rose in popularity within the last millennium. Finally, the development of modern breeding impacted genetic diversity more dramatically than the previous millennia of human management.

The Eneolithic Botai culture of the Central Asian steppes provides the earliest archaeological evidence for horse husbandry, ~5500 years ago, but the exact nature of early horse domestication remains controversial. We generated 42 ancient-horse genomes, including 20 from Botai. Compared to 46 published ancient- and modern-horse genomes, our data indicate that Przewalski's horses are the feral descendants of horses herded at Botai and not truly wild horses. All domestic horses dated from ~4000 years ago to present only show ~2.7% of Botai-related ancestry. This indicates that a massive genomic turnover underpins the expansion of the horse stock that gave rise to modern domesticates, which coincides with large-scale human population expansions during the Early Bronze Age.

The genomic changes underlying both early and late stages of horse domestication remain largely unknown. We examined the genomes of 14 early domestic horses from the Bronze and Iron Ages, dating to between ~4.1 and 2.3 thousand years before present. We find early domestication selection patterns supporting the neural crest hypothesis, which provides a unified developmental origin for common domestic traits. Within the past 2.3 thousand years, horses lost genetic diversity and archaic DNA tracts introgressed from a now-extinct lineage. They accumulated deleterious mutations later than expected under the cost-of-domestication hypothesis, probably because of breeding from limited numbers of stallions. We also reveal that Iron Age Scythian steppe nomads implemented breeding strategies involving no detectable inbreeding and selection for coat-color variation and robust forelimbs.

Domestication of horses fundamentally transformed long-range mobility and warfare1. However, modern domesticated breeds do not descend from the earliest domestic horse lineage associated with archaeological evidence of bridling, milking and corralling2–4 at Botai, Central Asia around 3500 bc3. Other longstanding candidate regions for horse domestication, such as Iberia5 and Anatolia6, have also recently been challenged. Thus, the genetic, geographic and temporal origins of modern domestic horses have remained unknown. Here we pinpoint the Western Eurasian steppes, especially the lower Volga-Don region, as the homeland of modern domestic horses. Furthermore, we map the population changes accompanying domestication from 273 ancient horse genomes. This reveals that modern domestic horses ultimately replaced almost all other local populations as they expanded rapidly across Eurasia from about 2000 bc, synchronously with equestrian material culture, including Sintashta spoke-wheeled chariots. We find that equestrianism involved strong selection for critical locomotor and behavioural adaptations at the GSDMC and ZFPM1 genes. Our results reject the commonly held association7 between horseback riding and the massive expansion of Yamnaya steppe pastoralists into Europe around 3000 bc8,9 driving the spread of Indo-European languages10. This contrasts with the scenario in Asia where Indo-Iranian languages, chariots and horses spread together, following the early second millennium bc Sintashta culture11,12.

The DNA molecules that can be extracted from archaeological and palaeontological remains are often degraded and massively contaminated with environmental microbial material. This reduces the efficacy of shotgun approaches for sequencing ancient genomes, despite the decreasing sequencing costs of high-throughput sequencing (HTS). Improving the recovery of endogenous molecules from the DNA extraction and purification steps could, thus, help advance the characterization of ancient genomes. Here, we apply the three most commonly used DNA extraction methods to five ancient bone samples spanning a ~30 thousand year temporal range and originating from a diversity of environments, from South America to Alaska. We show that methods based on the purification of DNA fragments using silica columns are more advantageous than in solution methods and increase not only the total amount of DNA molecules retrieved but also the relative importance of endogenous DNA fragments and their molecular diversity. Therefore, these methods provide a cost-effective solution for downstream applications, including DNA sequencing on HTS platforms.

The horse was domesticated only 5.5 KYA, thousands of years after dogs, cattle, pigs, sheep, and goats. The horse nonetheless represents the domestic animal that most impacted human history; providing us with rapid transportation, which has considerably changed the speed and magnitude of the circulation of goods and people, as well as their cultures and diseases. By revolutionizing warfare and agriculture, horses also deeply influenced the politico-economic trajectory of human societies. Reciprocally, human activities have circled back on the recent evolution of the horse, by creating hundreds of domestic breeds through selective programs, while leading all wild populations to near extinction. Despite being tightly associated with humans, several aspects in the evolution of the domestic horse remain controversial. Here, we review recent advances in comparative genomics and paleogenomics that helped advance our understanding of the genetic foundation of domestic horses.

Enamel thickness continues to be an important morphological character in hominin systematics and is frequently invoked in dietary reconstructions of Plio-Pleistocene hominin taxa. However, to date, the majority of published data on molar enamel thickness of Pliocene and early Pleistocene hominins derive from naturally fractured random surfaces of a small number of specimens.In this study we systematically analyze enamel thickness in a large sample of Plio-Pleistocene fossil hominins (n = 99), extant hominoids (n=57), and modern humans (n=30). Based on analysis of 2D mesial planes of section derived from microtomography, we examine both average and relative enamel thickness, and the distribution of enamel across buccal, occlusal, and lingual components of mandibular molars. Our results confirm the trend for increasing enamel thickness during the Pliocene that culminates in the thick enamel of the robust Australopithecus species, and then decreases from early Homo to recent modern humans. All hominin taxa, and Pongo, share a regional average enamel thickness pattern of thick occlusal enamel and greater buccal than lingual enamel thickness. Pan is unique in exhibiting thinnest average enamel thickness in the occlusal basin. Statistical analysis indicates that among Pliocene hominins enamel thickness is a weak taxonomic discriminator. The data underlying these results are included as an appendix in the study.We thank the editorial staff and the reviewers for their critical and helpful comments. Please find below a point by point explanation of how we addressed these comments with our revision. Reviewers' comments:AE Comments: This is a very clean, easy to read manuscript that provides a thorough update to studies of enamel thickness in hominins. All three reviewers find the manuscript useful and praise its clarity, thoroughness, and brevity. However, the reviewers diverge in their opinions on how much revision is needed. Reviewer #3 finds the manuscript effectively acceptable as is, while the other two reviewers request some degree of revision. Reviewer #1 requests that the manuscript be revised with regard to "....inferences regarding the taxonomic valence and functional utility of enamel thickness..." Reviewer #2 focuses on the measurements of enamel thickness, and asks the authors especially to comment on why enamel is so thick in modern Homo, but not so in early Homo. The reviewer also offers some organizational suggestions.The suggestions of the reviewers are certainly in the spirit of constructive advice, and should be relatively easy to manage. Given that the changes may involve extra analysis and an alteration in the primary thrust of the manuscript, I recommend that the authors revise the manuscript and reply to the reviewers comments. Pending the nature of the revisions, I suggest returning the ms to reviewer #1 if necessary.Reviewer #1: Overall, Skinner et al. provide a thorough and much needed compendium of enamel thickness in a range of early hominins, including early Homo. As such, it provide...

High-throughput sequencing has dramatically fostered ancient DNA research in recent years. Shotgun sequencing, however, does not necessarily appear as the best-suited approach due to the extensive contamination of samples with exogenous environmental microbial DNA. DNA capture-enrichment methods represent cost-effective alternatives that increase the sequencing focus on the endogenous fraction, whether it is from mitochondrial or nuclear genomes, or parts thereof. Here, we explored experimental parameters that could impact the efficacy of MYbaits in-solution capture assays of ~5000 nuclear loci or the whole genome. We found that varying quantities of the starting probes had only moderate effect on capture outcomes. Starting DNA, probe tiling, the hybridization temperature and the proportion of endogenous DNA all affected the assay, however. Additionally, probe features such as their GC content, number of CpG dinucleotides, sequence complexity and entropy and self-annealing properties need to be carefully addressed during the design stage of the capture assay. The experimental conditions and probe molecular features identified in this study will improve the recovery of genetic information extracted from degraded and ancient remains.