The oral cavity is a heterogeneous environment, varying in factors such as pH, oxygen levels, and salivary flow. These factors affect the microbial community composition and distribution of species in dental plaque, but it is not known how well these patterns are reflected in archaeological dental calculus. In most archaeological studies, a single sample of dental calculus is studied per individual and is assumed to represent the entire oral cavity. However, it is not known if this sampling strategy introduces biases into studies of the ancient oral microbiome. Here, we present the results of a shotgun metagenomic study of a dense sampling of dental calculus from four Chalcolithic individuals from the southeast Iberian peninsula (ca. 4500–5000 BP). Inter-individual differences in microbial composition are found to be much larger than intra-individual differences, indicating that a single sample can indeed represent an individual in most cases. However, there are minor spatial patterns in species distribution within the oral cavity that should be taken into account when designing a study or interpreting results. Finally, we show that plant DNA identified in the samples is likely of postmortem origin, demonstrating the importance of including environmental controls or additional lines of biomolecular evidence in dietary interpretations.
Systematic excavation and multidisciplinary research undertaken over three decades have deepened our understanding of the early Palaeolithic archaeology at Cueva Negra del Estrecho del Río Quípar (Caravaca de la Cruz, Murcia, Spain). New results from biochronology and combined ESR and U-series dating corroborate previous magnetostratigraphy, placing the entire excavated sequence between the Jaramillo sub-chron and the Matuyama-Brunhes boundary (i.e., ca. 990-772 thousand years ago [ka]); palaeontological and palynological findings reflect temperate environmental conditions. A bifacially-flaked limestone hand-axe was excavated one metre below the top of the Pleistocene sequence. The Equus cf. altidens tooth that provided the ESR estimate was excavated one metre below the hand-axe. Throughout its five-metre-deep sedimentary sequence, small nodules, fragments, and struck flakes make up the bulk of the Palaeolithic assemblage. Stratigraphical analysis points to undisturbed continuous sedimentary deposition above a layer of ashy sediment, encountered 4.5 m below the top of the Pleistocene sequence, which contained thermallyaltered bone and heat-shattered chert cores and flakes. Cueva Negra is among the earliest European sites with firm evidence of combustion..Introduction Context Geographical locationCueva Negra del Estrecho del Río Quípar is a large, north-facing, rock-shelter (ca. 10x10 m in area), in Upper Miocene (Tortonian) biocalcarenite rock, lying at 740 m above sea level, a.s.l. (38° 02' 12.5" N; 1° 53' 5.8" W) on the right bank 40 m above the Quípar River where it flows northwards from a one-kilometre-long gorge (the Estrecho, i.e., "the Narrows") below the hamlet of La Encarnación in Caravaca de la Cruz municipality (Murcia, Spain) (Figure 1). The Quípar is a tributary of the Segura River that reaches the Mediterranean Sea 110 km E of Cueva Negra, even though the cave lies only 70 km N of the Murcian coast. Important geological faults determine the alignment of tributaries in the Segura drainage basin. The Estrecho follows the sinistral reverse Quípar Fault, active since the Late Miocene (Messinian). Activity caused uplift of the right bank of the river, thereby saving from riverine erosion the fine-grained fluviatile sediments that had accumulated in Cueva Negra under conditions of low transport energy by intermittent overflow of an erstwhile swampy lake fed by the Quípar during the late Early Pleistocene. The Quípar enters the gorge at 725 m a.s.l. and leaves it at 690 m a.s.l.. The height of the land above sea level during the Early Pleistocene is unknown. The Upper Miocene (Tortonian) calcarenite formed under the Tethys Sea, arising to become a shoreline surface in the Upper Pliocene. Cueva Negra is a vestige of a trapezoidal endokarstic cavity that likely developed in relation to low-lying lagoons or lakes. LithostratigraphyThe fluviatile sediments inside Cueva Negra include clasts eroded from the cave roof and walls. Palaeolithic and faunal remains are present throughout the five-metre depth, implying...
Throughout the Pleistocene, early humans and carnivores frequented caves and large rock-shelters, usually generating bone accumulations. The well-preserved late Early Pleistocene sedimentary sequence at Cueva Negra del Estrecho del Río Quípar (CNERQ) has provided substantial evidence concerning the behavioural and adaptive skills of early humans in Western Europe, such as butchery practices, lithic technology or tending fire, whilst also bearing witness to the bone-altering activities of carnivores. Recent fieldwork has allowed the re-examination of the spatial and taphonomical nature of the macrofaunal assemblage from the upper layers of Complex 2. These layers are somewhat different from most of the underlying sequence, in showing quite a high representation of cranial and post-cranial bones of large mammals, including several Megaloceroscarthaginiensis antlers. The presence of Crocuta sp. at Cueva Negra represents one of the earliest instances of this genus in Western Eurasia. Identification of several juvenile Crocuta sp. remains alongside coprolites and bones with carnivore damage, indicates sporadical hyaenid denning activity. Furthermore, the presence of bones with percussion and cut-marks near to several hammerstones suggests a clear albeit limited anthropogenic input. We interpret the available taphonomical and spatial evidence from these layers as reflecting a multi-patterned palimpsest, likely representing the non-simultaneous and short-lived co-existence of hyaenas, humans, and other small carnivores in the Cueva Negra palaeolandscape during the final phase of sedimentation preserved at the site.
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