Wild or domesticated? An ancient DNA approach to canid species identification in South Africa's Western Cape Province

The fragmentary remains typical of archaeological fauna can prove impossible to identify when confronted with closely related sympatric species. In southern Africa domesticated dogs (Canis familiaris) and black-backed jackals (Canis mesomelas) pose just such a problem, rendering difficult an accurate assessment of the timing of arrival of domesticated dogs in the region. Contextual evidence has suggested that canid remains from four Later Stone Age archaeological sites were likely to be domesticated dogs rather than jackals, and were subjected to ancient DNA analyses to determine species. Every tested specimen proved to derive from black-backed jackals. These data provide not only an unexpected window on southern African prehistory, but also highlight the value in applying ancient DNA techniques to archaeological species identification.     

Ancient DNA analysis of desiccated wheat grains excavated from a Bronze Age cemetery in Xinjiang

Wheat has been one of the most important crop in Eurasia since the Neolithic period. Understanding the spread of wheat cultivation is crucial to understanding the spread of agriculture as a whole and the interactions between prehistoric populations across the Eurasian continent. However, the routes by which wheat cultivation spread eastwards have been poorly understood to date, due to the scarcity of plant remains recovered from archaeological sites. Desiccated wheat grains excavated from the Xiaohe cemetery in Xinjiang, and dated to the early Bronze Age, show excellent DNA preservation. Here we present an ancient DNA (aDNA) analysis of wheat (Triticum sp.) grains excavated from Xiaohe and provide the first definitive evidence for bread wheat in China during the Bronze Age. The nuclear ribosomal DNA internal transcribed spacer regions (ITS1 and ITS2) and the intergenic spacer region (IGS) were amplified. The IGS region within the D genome of wheat has a 71 bp insertion that is absent from corresponding regions in the A and B genomes. The results showed that the Xiaohe wheat showed most sequence similarity to hexaploid bread wheat (Triticum aestivum), including the characteristic insertion into the D genome. The presence of bread wheat at the Xiaohe cemetery is discussed in relation to it having spread into Xinjiang by the Bronze Age, providing new insight into the origins of bread wheat in East Asia.     

Evolving ecosystems: ecological data from an Iron Age small mammal accumulation at Alorda Park (Catalonia,Spain)

Small mammals are considered to be good indicators of ancient environments. One particular layer in a fortification tower of a Mediterranean Iberian village, Alorda Park (4th century BC), provided a large amount of well-preserved small mammal bones (about 19,200 remains) probably accumulated by barn owls (Tyto alba). Such an accumulation is rarely available from the Iron Age. It provides not only palaeoenvironmental data, but also further information about other studies such as morphometrics, ancient DNA and zoogeography. This article focuses on the ecological data, and attempts to develop an accurate taphonomic study to assess the reliability and statistical significance of this archaeological sample.     

Ectopic paragonimiasis from 400-year-old female mummy of Korea

Of the parasite eggs discovered in ancient samples from Korean archaeological sites, Paragonimus spp. are of particular importance in that they are regarded as one of the most insidious trematode infection sources. Although their infection prevalence decreased rapidly in the early 20th century, archaeoparasitological studies on the species are still required, as historical knowledge of paragonimiasis remains far from comprehensive. Fortunately, we recently were given a chance to examine a medieval Korean mummy in a good state of preservation. Using morphological techniques, we discovered evidence of ectopic paragonimiasis in the sample from the 17th-century female. When the ITS 2 gene was extracted, amplified and sequenced from Paragonimus eggs, it showed 100% homology to the sequences of modern Paragonimus westermani reported from Korea and Japan, forming a cluster distinct from South Asian P. westermani. Our report is the first-ever analysis of ancient Paragonimus DNA from any archaeological field in the world. The ectopic paragonimiasis diagnosis made in this study, especially involving the liver sample, also is the first of its kind in archaeoparasitology.     

Tuberculosis among Iron Age individuals from Tyva,South Siberia: palaeopathological and biomolecular findings

The study focuses on the evidence for tuberculosis apparent in an Iron Age population recovered from the cemetery of Aymyrlyg, Tyva (Tuva), South Siberia. A recent wholly molecular study of five of the cases confirmed the presence of Mycobacterium tuberculosis (MTB) complex DNA in four of the individuals. In all cases the disease was caused by strains of Mycobacterium bovis rather than Mycobacterium tuberculosis and represents the first positive identification of the bovine form of the disease in archaeological human remains. Details of the palaeopathological characteristics of the cases are provided in the current paper, while the molecular observations are extended to include a quantitative evaluation of the surviving mycobacterial DNA using real-time PCR. The observation that bovine tuberculosis was the pathogen responsible is discussed in terms of current understanding of the evolution of the MTB complex as well as the implications for future ancient DNA studies in this area.     

Ancient DNA of the Pacific rat (Rattus exulans) from Rapa Nui (Easter Island)

We report analysis of ancient mitochondrial DNA sequences from nine archaeological specimens (8 femura and 1 incissor) of Rattus exulans excavated from Anakena Beach Dune on Rapa Nui. Sequence of a 239-base-pair fragment of the hypervariable mitochondrial control region reveals a single mitochondrial DNA (mtDNA) sequence of all samples corresponding to the R9 haplotype prevalent in East Polynesia. This suggests a single or very limited introduction of Rattus exulans to the island. Rapa Nui, like other remote islands of Polynesia, remained effectively isolated following colonization.     

Geoarchaeological insights from a Roman age incineration feature (ustrinum) at Enconsta de Sant'Ana (Lisbon,Portugal)

Ustrina are incineration funerary structures that are relatively common in Roman age cemeteries. Salvage excavations at Encosta de Sant'Ana (Lisbon, Portugal) in 2002 brought to light a part of the necropolis of the Roman town of Olisipo, including some ustrina. One of them, designated Burial 1 during fieldwork, is analyzed here from a geoarchaeological viewpoint to understand the formation processes of such an archaeological feature, namely its construction technique, use and function. The study of site context and subsurface pedofeatures, and the application of archaeological soil micromorphology, revealed that the ustrinum was constructed digging a hollow in bedrock and building a mud-brick ridge around it, with raw material taken from the local bedrock. The structure was used at various times and remodelled at least once, and its base was not always thoroughly cleaned – according to the custom called “pars pro toto” by Roman authors – and probably left open when not in use.     

Amplification and sequencing of Trichuris trichiura ancient DNA extracted from archaeological sediments

Despite high prevalence of Trichuris trichiura infection, PCR-based analysis on T. trichiura from archaeological samples has not been established so far. In the present study, we sought to perform PCR-based amplification of T. trichiura aDNA using the sediments from medieval tomb of Korea. The presence of Trichuris eggs were first detected by microscopic observation; then confirmed by PCR-based aDNA analysis. Obtained sequence showed 100% homology to that of the small subunit ribosomal RNA (SSUrRNA) gene of T. trichiura but distinct from that of other Trichuris species. PCR-based aDNA analysis in this study can serve as effective method to confirm the presence of T. trichiura eggs in the soils or coprolites collected from archaeological sites.     

The first archaeometric data on polychrome Iron Age glass from sites located in northern Italy

A large sample set of transparent and opaque glass artefacts recovered from Etruscan contexts in northern Italy (Bologna and Spina (FE) necropoleis) and dated to a period between the 6th and 4th century BC are analysed in this paper. Samples of highly decorated beads, spindle whorls and vessels of the ‘Mediterranean Group I’ (Alabastron, Oinochoes, Amphoriskos) were selected in order to determine whether these different glass artefacts were produced at the same manufacturing site. While the vessels almost certainly originate from Greece, the beads could derive from a more ancient local production ascertained at the site of Frattesina (Rovigo, Italy) and dated to the Bronze Age.     

Clarifying prehistoric parasitism from a complementary morphological and molecular approach

This paper reports an approach to the identification of prehistoric parasitic infection, which integrates traditional morphological methods with molecular methods. The approach includes the strengths of each method while mitigating the limitations. Demonstrating the efficacy of this approach, we provide a case study from a 1400 year old desiccated fecal sample from La Cueva de los Muertos Chiquitos, archaeological site, near Rio Zape, Durango, Mexico. Traditionally prepared microscope slides were processed via microscopy and tentative ascarids were identified. Information regarding the parasites' developmental stage was recorded. DNA was then extracted directly from the slide material. From this DNA extract, a small segment of the 18S ribosomal RNA gene variant that is specific to Ascaris, and its phylogenetically close relatives, was targeted for PCR amplification and sequencing. Phylogenetic analysis of the DNA sequence best matched a member of physalopterids, rather than ascarids, with a single exception of a match to Contracaecum spiculigerum. Subsequent extractions, amplifications and sequencing of the original rehydrated coprolite material confirmed these results. The C. spiculigerum sequence represented a phylogenetic anomaly and subsequent analysis determined the sequence was an error in the BLAST database, likely attributable to misidentification of juvenile specimens prior to sequencing and submission. Physaloptera are a difficult genus to identify morphologically and can carry major health burdens. They may be underreported in humans, in part, because of morphological similarities to the more common human parasites belonging to ascarids. We conclude that integrating traditional morphological methods with molecular methods can help resolve this issue, in both contemporary and prehistoric populations.     

Recovery of DNA from archaeological insect remains: first results,problems and potential

We report the recovery of short fragments of PCR amplifiable ancient DNA from exoskeletal fragments of the grain weevil Sitophilus granarius (L.) extracted from Roman and medieval deposits in Northern England. If DNA preservation in archaeological insect remains is widespread then many applications in the spheres of evolutionary studies and archaeology can be conceived, some of which are outlined.     

Short-chain n-alkanes (C16–20) in ancient soil are useful molecular markers for prehistoric biomass burning

The incorporation of plant biomass into soil usually leads to long-chain n-alkanes with a relative predominance of odd carbon numbered homologues. Contrastingly, an increase in short-chain even carbon numbered n-alkanes was found in charred biomass with progressing temperatures. We applied lipid analysis to buried ancient topsoils that contained charred organic matter and to corresponding control soils, which were characterized by a lighter color and lower contents of charred materials. Commonly, the proportion of lipid extracts was found to be lower in the ancient soil than in the control samples, which indicated an enhanced degradation of organic matter, e.g., by thermal degradation. All samples displayed a particular pattern of short-chain and even carbon numbered n-alkanes (maximum at n-C₁₆ or n-C₁₈). The ratios CPI (carbon preference index) and ACL (average chain length) for the investigated soil samples matched the ratios found for maize and rye straw charred at 400 °C or 500 °C, respectively. These molecular ratios indicate the presence of charred biomass. The predominance of short-chain and even carbon numbered n-alkanes was a result of thermal degradation of biomass. The degradation products were preserved in ancient soils and could be applied as molecular markers in archaeological or palaeoenvironmental research.     

Ancient Iberian horses: a method to recover DNA from archaeological samples buried under sub-optimal conditions for preservation

Existing methods to extract, amplify, and sequence ancient DNA (aDNA) from horse bone and teeth were optimized to recover DNA from a depositional environment of highly permeable acidic soil. DNA was successfully retrieved using 0.10g of bone powder from horse (Equus sp.) remains dating to 25 K years utilizing the methods optimized for this archaeological material. The genetic analyses were performed in a facility that is dedicated to ancient DNA research (Paleo-DNA Laboratory, Thunder Bay, ON, Canada) and has not been previously used to analyse modern or ancient horse DNA. Research was replicated to obtain reliable sequencing results for six samples from the Iberian Peninsula that were consistent with published sequences of Equus caballus. The archaeological sequence data obtained support hypotheses that promote the significance that the Iberian Peninsula has had to the multi-focal centres of origin for horse domestication and distribution of modern horse breeds. The data presented may provide evidence of the existence of an Iberian refugium for Equus during the last glacial period, 10 K years BP. Further molecular data analyses will enhance the ideas presented by this data and our understanding of horse domestication and phylogeny. The optimization of molecular techniques to successfully obtain DNA using minimally destructive, cosmetically sensitive techniques from archaeological remains endeavours to foster further cooperation between museums and researchers.     

Rice chaff in ceramics from the archaeological site of Sumhuram (Dhofar,Southern Oman)

The identification of the chaff used as temper in ancient ceramics represents a possible source of information about the area of origin of the pottery. This paper studies the occurrence of rice (Oryza sp.) chaff in potsherds from the archaeological site of Sumhuram (Dhofar, Sultanate of Oman). The information gathered at the site offers insight into commerce routes and networks to which the town belonged in the first centuries AD. The analysis of the plant material in the pottery and potsherds can also furnish interesting data on the ancient technologies of pottery production.     

Mineral weathering in forest soils and its relevance to the preservation of the buried archaeological resource

There is more woodland in Britain now than for many centuries and considering many international climate change mitigation policies, woodland cover, both in Britain and internationally, is being promoted. However, neither the management of existing woodlands nor their expansion should be at the expense of important archaeological evidence. Due to the large number of known archaeological sites, the large areas of land with uninvestigated archaeological potential and the expanding woodland cover, suitable mitigation strategies need to be developed to allow preservation in situ of important sites. An understanding of how woodland soils and the buried archaeological resource interact is, therefore, essential. This paper utilises ongoing environmental research into the mineral weathering rates in forest soils and considers its application to artefact preservation. The study concludes that soil water pH, its movement, and the saturation of dissolved ions in the soil solution are major factors determining both mineral and artefact longevity. A simple guide to artefact longevity based on these properties is proposed and a geochemical model for predicting loss is tested. These tools could be applied to any soil or individual horizon irrespective of land-use.     

Ancient DNA analysis on Clonorchis sinensis eggs remained in samples from medieval Korean mummy

In the present study, Clonorchis sinensis (C. sinensis) ancient DNA (aDNA) was successfully extracted from human remains discovered in a tomb dating to the medieval Joseon dynasty of Korea. The presence of C. sinensis eggs was confirmed by microscopic observation, after which a PCR-based aDNA analysis was performed using primer sets designed for the amplification of nuclear ribosomal internal transcribed spacer 1 (ITS1), 2 (ITS2) and mitochondrial cytochrome c oxidase 1 (CO1) genes. The sequences obtained were 100% homologous to some contemporary C. sinensis gene sequences reported from Korea and other East Asian countries. We believe that the results of our analysis expand the temporal and geographical scopes of research on the history of C. sinensis infection in different human populations.     

First ancient bovine DNA evidence from India: difficult but not impossible

Ancient DNA isolation from the tropical countries has been shown to be very difficult in the past. Here for the first time we have been successful in isolating ancient DNA from Indian cattle samples. We were able to obtain DNA and sequence the partial mitochondrial D-loop in 3 of the 15 bovine fossil samples ranging in age from 2000 BC to 1000 AD, and were able to further identify the most recent sample as being of Bos indicus origin. Our results on ancient DNA extraction from India will encourage other researchers in this field to carry out further studies of ancient DNA from Indian bovine samples. Our results represent the first successful extraction and amplification of bovine ancient DNA from India, and thus may pave the road for a better understanding of demographic and historical processes of cattle domestication that has taken place in this region.     

First dendroarchaeological dates of prehistoric contexts in South America: chullpas in the Central Andes

Despite the existence of long-lived tree species with excellent tree-ring characteristics, the use of dendrochronological techniques for dating archaeological contexts has been very limited in South America. Recent research in the Bolivian Altiplano of the Central Andes has yielded a network of Polylepis tarapacana chronologies that goes back to the 13th century. This species was regularly used by local populations since pre-Columbian times as raw material for beams, lintels, hooks and other architectural features in houses, storage chambers, and sepulchers. The aim of this study is to argue in favor of the potential of P. tarapacana for providing high resolution dates of significant archaeological events during the pre-Columbian era. The chronology of chullpas (burial towers and storage chambers) in the southern Andean Altiplano is used as a case study. Growth rings from P. tarapacana have provided several centennial-length dendrochronological records including a reference regional chronology covering the last 786 years in the Central Andean highlands. Based on this long reference chronology calendar years were assigned to woody pieces from pre-Hispanic chullpas. Dendroarchaeological results are consistent with contextual evidence and most radiocarbon dates from woody samples and related archaeological items. Our results indicate that P. tarapacana has a high potential for providing annually-resolved tree-ring dates for archaeological contexts in the Altiplano since the early 13th century.     

Starch grain characterization of Prosopis chilensis (Mol.) Stuntz and P. flexuosa DC,and the analysis of their archaeological remains in Andean South America

The fruit (pods) of Prosopis (Fabaceae) are frequently recovered from pre-Hispanic Argentinian archaeological sites, suggesting that this genus was of importance in ancient economies in this region. Yet it is only recently that archaeobotanists have begun to carry out systematic research into this genus. Therefore many questions remain to be addressed concerning the food value of Prosopis fruit, and its potential contribution. This paper examines starch from the pods of two species, Prosopis flexuosa and Prosopis chilensis, for the purposes of describing and classifying their morphological features and biometrical parameters. Pods of both species were gathered from two extremes (northern and southern) of the Hualfín Valley, Catamarca, Argentina. Starch abundance is estimated and compared with that of Zea mays, an economically important plant with high concentrations of starch. This paper reveals that Prosopis pods contain very low amounts of starch compared to starch-rich edible species. Despite this, the recovery of Prosopis starch grains can be successfully used to assess archaeological tool uses. Prosopis starch grains were found to be highly variable in shape. Grain size ranged between 10 and 20 μm. Granule irregularity and the high birefringence under polarized light are two of the most important diagnostic features. No significant statistical differences were found in the structure (morphology, size and hilum) of the starch of the same species from both localities. Finally, the implications for the role of this plant in past societies are discussed.     

A short note on starch and xylem of Colocasia esculenta (taro) in archaeological deposits from Pitcairn Island,southeast Polynesia

We present results of starch analysis of archaeological deposits from Pitcairn Island. High concentrations of starch grains preserved in cell membranes, and xylem tracheary elements, consistent with introduced Colocasia esculenta (taro) were found. Because of limited age control, we are uncertain if the microfossils are prehistoric. Problems associated with identifying taxa with small starch grains in extractions from weathered deposits are highlighted.