The Middle to Upper Palaeolithic transition in Europe witnessed the replacement and partial absorption of local Neanderthal populations by Homo sapiens populations of African origin1. However, this process probably varied across regions and its details remain largely unknown. In particular, the duration of chronological overlap between the two groups is much debated, as are the implications of this overlap for the nature of the biological and cultural interactions between Neanderthals and H. sapiens. Here we report the discovery and direct dating of human remains found in association with Initial Upper Palaeolithic artefacts2, from excavations at Bacho Kiro Cave (Bulgaria). Morphological analysis of a tooth and mitochondrial DNA from several hominin bone fragments, identified through proteomic screening, assign these finds to H. sapiens and link the expansion of Initial Upper Palaeolithic technologies with the spread of H. sapiens into the mid-latitudes of Eurasia before 45 thousand years ago3. The excavations yielded a wealth of bone artefacts, including pendants manufactured from cave bear teeth that are reminiscent of those later produced by the last Neanderthals of western Europe4,5,6. These finds are consistent with models based on the arrival of multiple waves of H. sapiens into Europe coming into contact with declining Neanderthal populations7,8.
The data that support the findings of this study are available from the corresponding author upon reasonable request. Genetic sequence reads from all libraries and corresponding negative controls are deposited at European Nucleotide Archive under the study accession number PRJEB35466. The FASTA files of the mitochondrial genomes are deposited in GenBank with the accession numbers MN706602–MN706607. Details are as follows: Bacho Kiro AA7-738, MN706602; Bacho Kiro BB7-240, MN706603; Bacho Kiro BK-1653, MN706604; Bacho Kiro CC7-335, MN706605; Bacho Kiro CC7-2289, MN706606; and Bacho Kiro molar F6-620, MN706607.
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We thank the tourism association of Bacho Kiro Cave in the town of Dryanovo, the History museum – Dryanovo, the Regional History museum in the city of Gabrovo, Dryanovo town hall and V. Lafchiiski for their assistance with the fieldwork and in the laboratory; N. Spassov from the National Museum of Natural History in Sofia for cooperating and hosting researchers of our project; H. Temming and J. Honeyford for their technical assistance and S. Nagel, B. Nickel, B. Schellbach and A. Weihmann for their help with the ancient DNA laboratory procedures and sequencing. Field operations were funded by the Max Planck Society. AixMICADAS and its operation are funded by Collège de France and the EQUIPEX ASTER-CEREGE (principal investigator, E.B.). S.T. is funded by the European Research Council under the European Union’s Horizon 2020 Research and Innovation Programme (grant agreement no. 803147-951 RESOLUTION). The ancient DNA part of this study was funded by the Max Planck Society and the European Research Council (grant agreement no. 694707 to S.P.).
The authors declare no competing interests.
Peer review information Nature thanks William Banks, Richard G. Klein and the other, anonymous, reviewer(s) for their contribution to the peer review of this work.
Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
Extended data figures and tables
a, Plan view of the entrance and the excavated areas of the cave, with the grid system of our recent excavations (letters in the left column) and those of the 1971–1975 excavations (letters in the right column). b, Site location in southeastern Europe. c, Photograph of the entrance of the cave. The floor is artificially raised; the original entrance was several metres lower than shown in this photograph. d, Initial stratigraphic section drawing of the exposed profile from the Main sector in 2015 (codes for the archaeological layers are on the left, with the corresponding layers from the 1971–1975 excavations in parentheses). e, Frontal view of the Niche 1 sector and its stratigraphic subdivisions. f, Lower part of the stratigraphic section drawing of the Niche 1 sector, in 2018. Note the thickness and preservation of the lower deposits here in comparison with the Main sector profile. g, Photograph of the Main sector transversal section on the line between squares F5–F6 and squares G5–G6 before excavation in 2015. CF, combustion feature. h–n, Hominin remains identified by ZooMS with their IDs: BK-1653 (h) and F6-597 (j) from layer B, with h coming from the 1971–1975 excavations (dashed line); BB7-240 (k), CC7-2289 (l), CC7-335 (m) and AA7-738 (n) from layer N1-I. Continuous lines connect the fossils with their find locations. i, Second lower molar (F6-620) from layer J in the Main sector.
Geographical distribution of the main IUP sites of western and central Eurasia (black dots), directly dated early H. sapiens predating 37,000 cal. bp (empty black dots) and directly dated late Neanderthals associated with Châtelperronian assemblages (orange squares). Bacho Kiro Cave is represented by a red circle.
Pointed retouched blades and fragments (1–4, 6, 7) and piece with bifacial retouch (5). Photographs by V.S.-M. and T. Tsanova.
Pointed retouched blade with slightly oblique truncation and base modified by inverse retouch (1), pointed blade fragments (2 and 5, which has an oblique truncation and slight notch on the left edge, and was perhaps intentionally fragmented), pointed, small blades fragments (3, 7, 8 and 9), pointed blade fragment with opposing pseudo-burin blows on the apex and on the distal fracture edge (perhaps indicating use as a projectile) (4) and Levallois flake (6). Drawings by I.K. and T. Tsanova).
a, Mesial, buccal and distal views of the crown, root and pulp chamber (left) and occlusal views of the enamel and dentine crown (right). b, A principal component analysis of the shape of the enamel–dentine junction ridge and cervix places the Bacho Kiro Cave second lower molar (F6-620) represented by a red star within the samples of recent (n = 8) and Pleistocene (n = 9) H. sapiens, and outside the distribution of Neanderthals (n = 20) and H. erectus (n = 3).
Extended Data Fig. 6 MALDI–TOF MS spectra for the six bone specimens identified as hominins through ZooMS analysis.
a, B4-1653 (interface of layers 6a and 7). b, AA7-738 (layer N1-I). c, BB7-240 (layer N1-I). d, CC7-2289 (layer N1-I). e, CC7-335 (layer N1-I). f, F6-597 (layer B).
Extended Data Fig. 7 Frequency of nucleotide substitutions at the beginning and the ends of mtDNA alignments for the Bacho Kiro Cave specimens.
Only fragments of at least 35 base pairs in length that mapped to the revised Cambridge Reference Sequence with a mapping quality of at least 25 were used for this analysis. Solid lines in red depict all fragments and dashed lines depict the fragments that have a C-to-T substitution at the opposing end (‘conditional’ C-to-T substitutions). All other types of substitution are marked in grey.
Extended Data Fig. 8 Bayesian phylogenetic tree relating Bacho Kiro Cave mtDNA to 54 present-day humans, 10 directly radiocarbon dated ancient H. sapiens and the Vindija 33.16 Neanderthal.
The Bacho Kiro Cave specimens are in red. Other ancient H. sapiens used as calibration points to estimate the tip dates of Bacho Kiro Cave specimens are italicized. The posterior probabilities are denoted above the branches. The mtDNA of Vindija 33.16 was used to root the tree (not shown).