Aspects of ancient Greek trade re-evaluated with amphora DNA evidence

Ancient DNA trapped in the matrices of ceramic transport jars from Mediterranean shipwrecks can reveal the goods traded in the earliest markets. Scholars generally assume that the amphora cargoes of 5th-3rd century B.C. Greek shipwrecks contained wine, or to a much lesser extent olive oil. Remnant DNA inside empty amphoras allows us to test that assumption. We show that short ∼100 nucleotides of ancient DNA can be isolated and analyzed from inside the empty jars from either small amounts of physical scrapings or material captured with non-destructive swabs. Our study material is previously inaccessible Classical/Hellenistic Greek shipwreck amphoras archived at the Ministry of Culture and Tourism Ephorate of Underwater Antiquities in Athens, Greece. Collected DNA samples reveal various combinations of olive, grape, Lamiaceae herbs (mint, rosemary, thyme, oregano, sage), juniper, and terebinth/mastic (genus Pistacia). General DNA targeting analyses also reveal the presence of pine (Pinus), and DNA from Fabaceae (Legume family); Zingiberaceae (Ginger family); and Juglandaceae (Walnut family). Our results demonstrate that amphoras were much more than wine containers. DNA shows that these transport jars contained a wide range of goods, bringing into question long-standing assumptions about amphora use in ancient Greece. Ancient DNA investigations open new research avenues, and will allow accurate reconstruction of ancient diet, medicinal compounds, value-added products, goods brought to market, and food preservation methods.     

Aspartic acid racemization variability in ancient human remains: implications in the prediction of ancient DNA recovery

The extent of racemization of aspartic acid (Asp) – expressed as d/l ratio – has been used as a marker of biomolecular degradation in ancient remains. However, Asp racemization rate is highly variable, and depends on biochemical and geochemical factors. In this paper we aim to determine to which extent the fraction analyzed and the kind of sample used may influence the d/l Asp ratios. Other factors, such as burial site and sample preservation conditions, are also considered.     

Digging deeper into the limits of ancient DNA research on syphilis

The search for the origins of syphilis has a long history in the medical and anthropological literatures. If we know more about the emergence of the pathogen that causes the disease in humans we will understand its evolution through time and space as well as shed light on its current state in living populations. Ancient DNA techniques used to isolate Treponema pallidum subsp. pallidum DNA from archaeological human specimens provide direct evidence of its existence in the past. However to date, only Kolman et al. (1999) have been successful in this endeavour, while other attempts have failed (e.g., Barnes and Thomas, 2006; Bouwman and Brown, 2005). Why has there been little success? This paper serves to compliment and add relevant information to Bouwman and Brown's and Barnes and Thomas' discussion concerning our inability to apply ancient DNA techniques to study venereal syphilis in past human populations.Our approach utilized 15 different human specimens from different geographies and different temporal periods: eight samples come from medically diagnosed individuals archived during the American Civil War period; six originate from the United Kingdom and predate 1492 with four of these samples having been previously analyzed by Bouwman and Brown and one sample comes from historic Canada. Human mitochondrial and amelogenin DNA, as well as several genes from the Treponema organism were analyzed revealing the relatively good preservation of human multi-copy and single copy DNA but not treponemal DNA. This study also incorporates a unique molecular experiment using rabbits infected with venereal syphilis to help illustrate that treponemal DNA disseminates to bone early during the first stages of infection but is not present in later stages of the disease using the techniques presented in this study.     

Deficiencies and challenges in the study of ancient tuberculosis DNA

We explore the standards of research and reporting needed to justify the destructive analysis of archaeological human bone for biomolecular studies of ancient tuberculosis (TB). Acceptable standards in osteological interpretation have been met in some biomolecular papers, but there are also cases where insufficient care has been taken in distinguishing between pathognomonic lesions and those that are ‘consistent with’ a diagnosis of TB. Some biomolecular studies have failed to recognize that archaeological bones might be contaminated with environmental mycobacteria whose DNA could give rise to false positives in polymerase chain reactions directed at members of the Mycobacterium tuberculosis complex. The difficulties of applying spoligotyping to ancient DNA have also been underestimated and conclusions drawn from such analyses are often weakly supported. Assumptions that mycobacterial DNA preserves better than human DNA, and that contamination with modern DNA is less of a problem, has led in some cases to a laxity in research standards with insufficient attention paid to the need to authenticate ancient DNA results. We illustrate our concerns by reference to a recent paper reporting biomolecular detection of ancient TB DNA in skeletons from the eastern Mediterranean Neolithic settlement of Atlit-Yam. We are unconvinced that the skeletal evidence presented in this paper gives sufficient indication of TB to warrant destructive analysis, and we are concerned that during the biomolecular part of the project inadequate attention was paid to the possibility that results might be due to laboratory cross-contamination or to amplification of environmental mycobacterial DNA present in the bones.     

A new miniSTR heptaplex system for genetic fingerprinting of ancient DNA from archaeological human bone

The analysis of short tandem repeats (STRs) is a useful tool in various contexts of ancient DNA research. Main applications are the reconstruction of kinship, identification, and authentication. Here we describe a short amplicon autosomal short tandem repeat (miniSTR) heptaplex system for the amplification of D13S317, D21S11, D18S51, TH01, D5S818, FGA and Amelogenin from highly degraded DNA as an inexpensive alternative to commercially available kits. All primers were newly designed and the amplicon length of all systems is less than 200 bp, with the exception of some rare alleles in the FGA and D21S11 systems. To validate the suitability of this system for typing STRs from human specimens with low DNA preservation we systematically tested it on 20 skeletal samples from four archaeological sites representing different burial environments and time spans since death. Finally, to test the sensitivity of the heptaplex system, we analyzed serial dilutions of control DNA and ancient DNA extracts. Using the system we were able to reproducibly obtain full STR profiles, down to a concentration of 0.06 ng DNA. Even with 0.004 ng DNA partial profiles could be amplified. The accumulated power of discrimination for the six selected STR loci is 0.99999984, plus the option of genetic sex determination through Amelogenin. The tests conducted prove that the system presented is efficient and especially suited for cases where STRs have to be typed and sex has to be assessed from human specimens with highly degraded DNA.     

Accurate sex identification of ancient human remains using DNA shotgun sequencing

Accurate identification of the biological sex of ancient remains is vital for critically testing hypotheses about social structure in prehistoric societies. However, morphological methods are imprecise for juvenile individuals and fragmentary remains, and molecular methods that rely on particular sex-specific marker loci such as the amelogenin gene suffer from allelic dropout and sensitivity to modern contamination. Analyzing shotgun sequencing data from 14 present-day humans of known biological sex and 16 ancient individuals from a time span of 100 to ∼70,000 years ago, we show that even relatively sparse shotgun sequencing (about 100,000 human sequences) can be used to reliably identify chromosomal sex simply by considering the ratio of sequences aligning to the X and Y chromosomes, and highlight two examples where the genetic assignments indicate morphological misassignment. Furthermore, we show that accurate sex identification of highly degraded remains can be performed in the presence of substantial amounts of present-day contamination by utilizing the signature of cytosine deamination, a characteristic feature of ancient DNA.     

Ancient DNA provides new insights into the origin of the Chinese domestic horse

Domestic horses played a pivotal role in ancient China, but their exact origin remains controversial. To investigate the origin of Chinese domestic horses, we analyzed mitochondrial DNA (mtDNA) from 35 horse remains, aged between 4000 and 2000 years, excavated from nine archaeological sites in northern China. The Chinese ancient horses exhibited high matrilineal diversity, falling into all the seven haplogroups (A–G) observed in modern horses. These results suggest that several maternal lines were introduced into the gene pool of Chinese horses in the past. Haplogroups A and F were more prevalent in ancient horses than the other haplogroups. Interestingly, only haplogroups A and F were present in the samples older than 4000 years, while the more recent horses (between 2000 and 3000 years BP) fell into all seven haplogroups. Comparison with DNA data of present-day horses suggests that haplogroup F is like to be an ancient haplogroup of East Asian origin. These analyses also suggest that the origin of Chinese domestic horses is complex, and external mtDNA input occurred after initial domestication. Our results indicate that the Chinese ancient horses are more related to the modern Mongolian horses. Lastly, our results cannot support the previous hypothesis that early Chinese domestic horses were derived from the Przewalski horse.     

要特别重视古村落的保护——南漳漫云古村落的现状及保护探讨

南漳漫云古村落是全国第三次文物普查的新成果,其独特的景观价值、文化价值引起社会的广泛关注.其设计理念,建造方式体现了“天人合一”核心思想,具有浓厚的地域特色,是鄂西北建筑的代表.本文以南漳漫云古村落为例,对古村落的发展演变、结构布局、建筑特点以及保护现状进行分析和研究.并根据古村落的发展特点,探讨当前形势下南漳漫云古村落发展与保护.     

Ancient DNA,a Neolithic legging from the Swiss Alps and the early history of goat

Ancient DNA from a Neolithic legging (1st half of the 3rd millennium BC) found at Lenk, Schnidejoch (2750 m a.sl.) in the Swiss Alps has demonstrated, that modern distribution of genetic variation does not reflect past spatio-temporal signatures. The legging was made from the skin of a domestic goat (Capra hircus), belonging to the caprine haplogroup B1, which is marginal in Europe today, but represents a third highly diverse goat haplogroup entering Europe already in the Neolithic. Population expansion of lineage B therefore happened more than 4500 years ago, but their members were at some point almost completely replaced by goats of today's common A and C haplogroups.     

Ancient Greek Mercenaries: Facts,Theories and New Perspectives

This article presents an overview of the historiography of Greek mercenaries and the proliferation of the phenomenon during the fourth century BC. It evaluates theoretical approaches to the political, economic and military roles that mercenaries played in Classical Greece during that century. In doing so, it considers the ways that interstate relations between the poleis shaped the development and recruitment of this form of soldier. The article disentangles the mercenary from a rich body of scholarship in economic history, demonstrating that analysis of the roles and functions of this figure can shed light on other dimensions of the Classical period, including international relations. Finally, it signals paths that the study of Greek mercenaries might take in the coming years.     

Sex identification of ancient DNA samples using a microfluidic device

A microfluidic device has been developed for the sex identification of ancient DNA samples and works by manipulating liquids within an environment of micrometer dimensions. In this work a range of microfluidic DNA extraction methods were evaluated for their compatibility with ancient DNA samples, and the use of streptavidin-coated super paramagnetic particles to isolate biotin-labeled abasic sites within damaged DNA was shown to be the most reproducible. Polymerase chain reaction-based DNA amplification was possible on the microfluidic device when less than 50 pg of template DNA was added. As a proof-of-principle, powdered bone samples were analysed using the integrated methodology developed. Following conventional capillary gel electrophoresis, two out of the three samples produced positive amplification results and were successfully identified as female. These sex identifications were corroborated by independent Amelogenin, anthropological and Y chromosome analysis. The work reported here is the first step in the development of a complete miniaturized microfluidic system that would enable on-site ancient DNA analysis.     

Ancient DNA reveals a major genetic change during the transition from hunting economy to reindeer husbandry in northern Scandinavia

The timing and origin of reindeer domestication has been highly debated. Recent molecular analyses show several mitochondrial lineages of domestic reindeer across Eurasia suggesting different origins of Fennoscandian and Siberian domestic reindeer. In order to investigate the origin of domestic Fennoscandian reindeer, we sequenced a mitochondrial control region fragment of 68 ancient reindeer remains from archaeological sites in Finnmark, the major county for extant reindeer husbandry in Norway, spanning from ca. BC 3400 to AD 1800. The majority of the Stone and Iron Age reindeer assemblages in Finnmark are from settlements in the eastern part of the county, in the Varangerfjord area. The reindeer remains from these settlements show affiliation to the large and complex Beringian haplotype cluster, found in extant reindeer from the Kola Peninsula to north-eastern Russia. A distinct haplotype shift is observed in the late medieval period, when the typical haplotype signatures of extant domestic Fennoscandian reindeer appeared in coastal regions of both eastern and western Finnmark. These haplotypes were not found among the Stone and Iron Age wild reindeer samples of Finnmark, suggesting that this population was not ancestral to extant domestic reindeer of Fennoscandia.     

Ancient DNA typing of archaeological pig remains corroborates historical records

The recent increase in both the abundance and taxonomic range of DNA sequence data in public repositories makes it possible to determine the maternal origin of lineages of faunal archaeological material by characterizing its mitochondrial DNA. Among the most commonly represented taxa are domesticated animals, for which extensive genetic characterization has revealed high levels of genetic diversity and (in at least some cases) strong phylogeographic clustering. Such information has significant implications not only for characterizing important aspects of the occupation history of archaeological sites, but also in providing novel insights into colonisation history and the scale and scope of trade and exchange networks. This can be done through studying the origins and dispersal of proxy organisms such as commensal and domesticated animals, as well as economically important wild fauna. To illustrate this approach, we compare historical records of maritime movement of people and pigs from two sites on Lord Howe Island, Australia, to phylogeographic results of DNA extracted from pig bones.     

略论昆明古城的历史沿革、文化特色及其底蕴

区,降及元初,虽“昆明”的族称在史籍中已逐渐被所谓“乌蛮”、“罗罗”取代,但很有可能在民间照常沿用而被官方采纳用作地名。其二是与唐初“昆州”、南诏“昆川”有明显的传承演进关系。对此,《元史·地理志》说:“昆明,中,倚郭。唐置(昆州) ,元宪宗四年,分其地立(昆明)千户二。至元十二年改鄯州,领(昆明)县。二十一年,州革,县名故。”从中不难发现,“昆州”与“昆明县”之间确有内在联系,而夹在其间的过渡性地名“鄯州”,则显然是从先前的“鄯阐府”转化而来。然而尽管这些解释已足够充分,且在边疆民族地区以当地少数民族的传统称谓用作…     

A flock of sheep,goats and cattle: ancient DNA analysis reveals complexities of historical parchment manufacture

Parchments comprise one of the most common and valuable sources of archaeological and historical data. Previous studies have shown that parchment also preserves genetic data. These data could be valuable for population studies, to understand past animal husbandry, the development of breeds and varieties and to comment on the provenance of parchments. To improve our understanding of DNA contained in parchments, we analysed genetic data, including both mitochondrial and autosomal loci, from 18th to 19th century English parchments which stable isotope analysis had indicated were well-preserved. DNA results were unexpected. All but one of the parchments produced multiple sequences matching several different species. Ion beam analysis ruled out surface treatments of the parchments (including ink and animal glues) as the origin of these multiple sequences. Our results suggest that the DNA content of parchment is more complex than previous research has suggested and that multiple stages of parchment manufacture, treatment and storage are preserved in parchment DNA extracts.     

Kinship between burials from Grave Circle B at Mycenae revealed by ancient DNA typing

The richness of the burials in Grave Circle B at Mycenae, Greece indicates that the 35 people interred there held elite status during their lifetimes 3500 years ago. It has been speculated that the groups of burials represent different dynasties or branches of the same family. To test this hypothesis, we carried out an exhaustive ancient DNA (aDNA) study of 22 of the skeletons. We were unable to identify nuclear aDNA in any specimen, but we obtained authentic mitochondrial aDNA sequences for four individuals. The results were compared with facial reconstructions and interpreted within the archaeological context represented by the organisation of the graves and the positions of the burials within the graves. We conclude that the contemporaneous male Γ55 and female Γ58 skeletons, which both possess the UK mitochondrial haplogroup, were brother and sister. The implication is that Γ58 was buried in Grave Circle B not because of a marital connection but because she held a position of authority by right of birth. The results illustrate the difficulty in using aDNA to study kinship relationships between archaeological specimens, but also show that aDNA can advance understanding of kinship when used to test hypotheses constructed from other evidence.     

Ancient DNA typing shows that a Bronze Age mummy is a composite of different skeletons

Excavations at Cladh Hallan, a Bronze Age-Iron Age settlement on South Uist in the Outer Hebrides off the west coast of Scotland, revealed the skeletons of two adults, a sub-adult and a child buried beneath the foundations of three roundhouses. Osteological and isotopic evidence has shown that the male adult skeleton is a composite made up of parts of at least three different individuals. To test the hypothesis that the female skeleton was also a composite we examined ancient DNA from four of its components: the skull, mandible, right humerus and right femur. Seven polymerase chain reactions (PCRs) were attempted, these covering positions 15,996–16,420, 16–132 and 232–368 of the mitochondrial DNA hypervariable I and II regions. Three PCRs were successful for each sample and a total of 55 sequences were obtained from the cloned products. After exclusion of possible contaminating sequences, the remaining 34 were compared. It was concluded that the mandible, humerus and femur come from different individuals. Insufficient data were obtained to draw conclusions regarding the origin of the skull. The presence of two composite skeletons at Cladh Hallan indicates that the merging of identities may have been a deliberate act, perhaps designed to amalgamate different ancestries into a single lineage.     

Ancient DNA in human bones from Neolithic and Bronze Age sites in Greece and Crete

Attempts were made to detect ancient DNA (aDNA) in samples of 88 human skeletons from eight Neolithic and Bronze Age sites in Greece and Crete. Ancient DNA was absent in specimens from Nea Nikomedia, Lerna, Karaviádena (Zakro), Antron Grave Circle A and Mycenae Grave Circle A. For each of three skeletons from Antron Grave Circle B that were sampled, polymerase chain reactions (PCRs) gave products for nuclear but not mitochondrial DNA, but amplicon yield was low and inconsistent with replicate PCRs failing to give reproducible results. With specimens from Mycenae Grave Circle B, evidence for mitochondrial aDNA was obtained for four of the 22 skeletons that were studied, and at Kouphovouno evidence for mitochondrial and/or nuclear aDNA was obtained with eight of the 20 skeletons that were examined. We conclude that, although aDNA might be present in some Eastern Mediterranean skeletons from later centuries of the Bronze Age, it is not commonly found in material from this period and is likely to be absent from older material.     

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.     

Fossil dogs and wolves from Palaeolithic sites in Belgium,the Ukraine and Russia: osteometry,ancient DNA and stable isotopes

Using multivariate techniques, several skulls of fossil large canids from sites in Belgium, Ukraine and Russia were examined to look for possible evidence of the presence of Palaeolithic dogs. Reference groups constituted of prehistoric dogs, and recent wolves and dogs. The fossil large canid from Goyet (Belgium), dated at c. 31,700 BP is clearly different from the recent wolves, resembling most closely the prehistoric dogs. Thus it is identified as a Palaeolithic dog, suggesting that dog domestication had already started during the Aurignacian. The Epigravettian Mezin 5490 (Ukraine) and Mezhirich (Ukraine) skulls are also identified as being Palaeolithic dogs. Selected Belgian specimens were analyzed for mtDNA and stable isotopes. All fossil samples yielded unique DNA sequences, indicating that the ancient Belgian large canids carried a substantial amount of genetic diversity. Furthermore, there is little evidence for phylogeographic structure in the Pleistocene large canids, as they do not form a homogenous genetic group. Although considerable variation occurs in the fossil canid isotope signatures between sites, the Belgian fossil large canids preyed in general on horse and large bovids.