Correction: Insight into the introduction of domestic cattle and the process of Neolithization to the Spanish region Galicia by genetic evidence.

[This corrects the article DOI: 10.1371/journal.pone.0249537.].

Equine diet during protohistoric times in the Northeast of the Iberian Peninsula: Stable isotope data (C, N) from bone collagen.

The analysis of stable isotopes in bone collagen allows us to infer the diet of the animals studied. This dataset consists of isotopic signatures (δ13C and δ15N) obtained by isotope ratio mass spectrometry from the skeletal remains of 42 equines (horse, ass and their hybrids) from the Can Roqueta site (Sabadell, Northeast Iberian Peninsula). Their chronology spans from Late Bronze Age to Late Roman Period, with particular emphasis on the Early Iron Age. These animals were found in storage silos and graves and were probably sacrificed as ritual offerings. The isotopic values are accompanied by data to assess the quality of the collagen analyzed. This fills a gap in equine isotopic values for this region and chronology, which may be of use to archaeologists interested in the study of livestock management or palaeodiet.

Insight into the introduction of domestic cattle and the process of Neolithization to the Spanish region Galicia by genetic evidence.

Domestic cattle were brought to Spain by early settlers and agricultural societies. Due to missing Neolithic sites in the Spanish region of Galicia, very little is known about this process in this region. We sampled 18 cattle subfossils from different ages and different mountain caves in Galicia, of which 11 were subject to sequencing of the mitochondrial genome and phylogenetic analysis, to provide insight into the introduction of cattle to this region. We detected high similarity between samples from different time periods and were able to compare the time frame of the first domesticated cattle in Galicia to data from the connecting region of Cantabria to show a plausible connection between the Neolithization of these two regions. Our data shows a close relationship of the early domesticated cattle of Galicia and modern cow breeds and gives a general insight into cattle phylogeny. We conclude that settlers migrated to this region of Spain from Europe and introduced common European breeds to Galicia.

Biomechanical simulations reveal a trade-off between adaptation to glacial climate and dietary niche versatility in European cave bears.

The cave bear is one of the best known extinct large mammals that inhabited Europe during the "Ice Age," becoming extinct ≈24,000 years ago along with other members of the Pleistocene megafauna. Long-standing hypotheses speculate that many cave bears died during their long hibernation periods, which were necessary to overcome the severe and prolonged winters of the Last Glacial. Here, we investigate how long hibernation periods in cave bears would have directly affected their feeding biomechanics using CT-based biomechanical simulations of skulls of cave and extant bears. Our results demonstrate that although large paranasal sinuses were necessary for, and consistent with, long hibernation periods, trade-offs in sinus-associated cranial biomechanical traits restricted cave bears to feed exclusively on low energetic vegetal resources during the predormancy period. This biomechanical trade-off constitutes a new key factor to mechanistically explain the demise of this dominant Pleistocene megafaunal species as a direct consequence of climate cooling.

Partial genomic survival of cave bears in living brown bears.

Although many large mammal species went extinct at the end of the Pleistocene epoch, their DNA may persist due to past episodes of interspecies admixture. However, direct empirical evidence of the persistence of ancient alleles remains scarce. Here, we present multifold coverage genomic data from four Late Pleistocene cave bears (Ursus spelaeus complex) and show that cave bears hybridized with brown bears (Ursus arctos) during the Pleistocene. We develop an approach to assess both the directionality and relative timing of gene flow. We find that segments of cave bear DNA still persist in the genomes of living brown bears, with cave bears contributing 0.9 to 2.4% of the genomes of all brown bears investigated. Our results show that even though extinction is typically considered as absolute, following admixture, fragments of the gene pool of extinct species can survive for tens of thousands of years in the genomes of extant recipient species.

Paleogenomic Evidence for Multi-generational Mixing between Neolithic Farmers and Mesolithic Hunter-Gatherers in the Lower Danube Basin.

The transition from hunting and gathering to farming involved profound cultural and technological changes. In Western and Central Europe, these changes occurred rapidly and synchronously after the arrival of early farmers of Anatolian origin [1-3], who largely replaced the local Mesolithic hunter-gatherers [1, 4-6]. Further east, in the Baltic region, the transition was gradual, with little or no genetic input from incoming farmers [7]. Here we use ancient DNA to investigate the relationship between hunter-gatherers and farmers in the Lower Danube basin, a geographically intermediate area that is characterized by a rapid Neolithic transition but also by the presence of archaeological evidence that points to cultural exchange, and thus possible admixture, between hunter-gatherers and farmers. We recovered four human paleogenomes (1.1× to 4.1× coverage) from Romania spanning a time transect between 8.8 thousand years ago (kya) and 5.4 kya and supplemented them with two Mesolithic genomes (1.7× and 5.3×) from Spain to provide further context on the genetic background of Mesolithic Europe. Our results show major Western hunter-gatherer (WHG) ancestry in a Romanian Eneolithic sample with a minor, but sizeable, contribution from Anatolian farmers, suggesting multiple admixture events between hunter-gatherers and farmers. Dietary stable-isotope analysis of this sample suggests a mixed terrestrial/aquatic diet. Our results provide support for complex interactions among hunter-gatherers and farmers in the Danube basin, demonstrating that in some regions, demic and cultural diffusion were not mutually exclusive, but merely the ends of a continuum for the process of Neolithization.

Mitochondrial DNA of pre-last glacial maximum red deer from NW Spain suggests a more complex phylogeographical history for the species.

The major climatic oscillations that characterized the Quaternary had a great influence on the evolution and distribution of several species. During cold periods, the distribution of temperate-adapted species became fragmented with many surviving in southern refugia (Iberian, Italian, and Balkan Peninsulas). Red deer was one of the species that contracted its original range to southern refugia. Currently, two main lineages have been described for the species: western and eastern. We have analyzed fossils pre-dating the last glacial maximum (LGM) from Liñares cave (NW Spain) that belongs to the peripheral range of the western clade, and fossils from the Danish Holocene belonging to the central part of the same clade. Phylogenetic analyses place our samples in the western clade. However, some specimens from Liñares represent an early split in the tree along with other pre-LGM western samples from previous studies. Despite low bootstrap values in the Bayesian phylogenies, haplotype networks connect these foreign haplotypes to the eastern clade. We suggest a mixed phylogeographical model to explain this pattern with range expansions from the east during the expansion phase after the cold periods in marine isotope stage 3. We find slight isolation by distance in post-LGM populations that could be a consequence of the recolonization from southern refugia after the LGM.

Ancient DNA reveals differences in behaviour and sociality between brown bears and extinct cave bears.

Ancient DNA studies have revolutionized the study of extinct species and populations, providing insights on phylogeny, phylogeography, admixture and demographic history. However, inferences on behaviour and sociality have been far less frequent. Here, we investigate the complete mitochondrial genomes of extinct Late Pleistocene cave bears and middle Holocene brown bears that each inhabited multiple geographically proximate caves in northern Spain. In cave bears, we find that, although most caves were occupied simultaneously, each cave almost exclusively contains a unique lineage of closely related haplotypes. This remarkable pattern suggests extreme fidelity to their birth site in cave bears, best described as homing behaviour, and that cave bears formed stable maternal social groups at least for hibernation. In contrast, brown bears do not show any strong association of mitochondrial lineage and cave, suggesting that these two closely related species differed in aspects of their behaviour and sociality. This difference is likely to have contributed to cave bear extinction, which occurred at a time in which competition for caves between bears and humans was likely intense and the ability to rapidly colonize new hibernation sites would have been crucial for the survival of a species so dependent on caves for hibernation as cave bears. Our study demonstrates the potential of ancient DNA to uncover patterns of behaviour and sociality in ancient species and populations, even those that went extinct many tens of thousands of years ago.

Pitfalls in comparing modern hair and fossil bone collagen C and N isotopic data to reconstruct ancient diets: a case study with cave bears (Ursus spelaeus).

Stable isotope analyses provide one of the few means to evaluate diet of extinct taxa. However, interpreting isotope data from bone collagen of extinct animals based on isotopic patterns in different tissues of modern animal proxies is precarious. For example, three corrections are needed before making comparisons of recent hair and ancient bone collagen: calibration of carbon-13 variations in atmospheric CO2, different isotopic discrimination between diet-hair keratin and diet-bone collagen, and time averaging of bone collagen versus short-term record in hair keratin. Recently, Robu et al. [Isotopic evidence for dietary flexibility among European Late Pleistocene cave bears (Ursus spelaeus). Can J Zool. 2013;91:227-234] published an article comparing extant carbon (δ(13)C) and nitrogen (δ(15)N) stable isotopic data of European cave bear bone collagen with those of Yellowstone Park grizzly bear hair in order to test the prevailing assumption of a largely vegetarian diet among cave bears. The authors concluded that cave bears were carnivores. This work is unfortunately unfounded as the authors failed to consider the necessary corrections listed above. When these corrections are applied to the Romanian cave bears, these individuals can be then interpreted without involving consumption of high trophic-level food, and environmental changes are probably the reason for the unusual isotopic composition of these cave bears in comparison with other European cave bears, rather than a change of diet. We caution researchers to pay careful attention to these factors when interpreting feeding ecology of extinct fauna using stable isotope techniques.

Withering away--25,000 years of genetic decline preceded cave bear extinction.

The causes of the late Pleistocene megafaunal extinctions are still enigmatic. Although the fossil record can provide approximations for when a species went extinct, the timing of its disappearance alone cannot resolve the causes and mode of the decline preceding its extinction. However, ancient DNA analyses can reveal population size changes over time and narrow down potential causes of extinction. Here, we present an ancient DNA study comparing late Pleistocene population dynamics of two closely related species, cave and brown bears. We found that the decline of cave bears started approximately 25,000 years before their extinction, whereas brown bear population size remained constant. We conclude that neither the effects of climate change nor human hunting alone can be responsible for the decline of the cave bear and suggest that a complex of factors including human competition for cave sites lead to the cave bear's extinction.