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In an effort to characterize the people who composed the groups known as the Xiongnu, nuclear and whole mitochondrial DNA data were generated from the skeletal remains of 52 individuals excavated from the Tamir Ulaan Khoshuu (TUK) cemetery in Central Mongolia. This burial site, attributed to the Xiongnu period, was used from the first century BC to the first century AD. Kinship analyses were conducted using autosomal and Y-chromosomal DNA markers along with complete sequences of the mitochondrial genome. These analyses suggested close kin relationships between many individuals. Nineteen such individuals composed a large family spanning five generations. Within this family, we determined that a woman was of especially high status; this is a novel insight into the structure and hierarchy of societies from the Xiongnu period. Moreover, our findings confirmed that the Xiongnu had a strongly admixed mitochondrial and Y-chromosome gene pools and revealed a significant western component in the Xiongnu group studied. Using a fine-scale approach (haplotype instead of haplogroup-level information), we propose Scytho-Siberians as ancestors of the Xiongnu and Huns as their descendants.

The likelihood ratio (LR) method is commonly used to determine kinship in civil, criminal, or forensic cases. For the past 15 years, our research group has also applied LR to ancient STR data and obtained kinship results for collections of graves or necropolises. Although we were able to reconstruct large genealogies, some pairs of individuals showed ambiguous results. Second-degree relationships, half-sibling pairs for example, were often inconsistent with detected first-degree relationships, such as parent/child or brother/sister pairs. We therefore set about providing empirical estimations of the error rates for the LR method in living populations with STR allelic diversities comparable to that of the ancient populations we had previously studied. We collected biological samples in the field in North-Eastern Siberia and West Africa and studied more than 800 pairs of STR profiles from individuals with known relationships. Because commercial STR panels were constructed for specific regions (namely Europe and North America), their allelic makeup showed a significant deficit in diversity when compared to European populations, replicating a situation often faced in ancient DNA studies. We assessed the capacity of the LR method to confirm known relationships (effectiveness) and its capacity to detect those relationships (reliability). Concerns over the effectiveness of LR determinations are mostly an issue in forensic studies, while the reliability of the detection of kinship is an issue for the study of necropolises or other large gatherings of unidentified individuals, such as disaster victims or mass graves. We show that the application of LR to both test populations highlights specific issues (both false positives and false negatives) that prevent the confirmation of second-degree kinship or even full siblingship in small populations. Up to 29% of detected full sibling relationships were either overestimated half-sibling relationships or underestimated parent-offspring relationships. The error rate for detected half-sibling relationships was even higher, reaching 41%. Only parent-offspring pairs were reliably detected or confirmed. This implies that, in populations that are small, ill-defined, or for which the STR loci analyzed are inappropriate, an examiner might not be able to distinguish a pair of full siblings from a pair of half-siblings. Furthermore, half-sibling pairs might be overlooked altogether, an issue that is exacerbated by the common confusion, in many languages and cultures, between half-siblings and full siblings. Consequently, in the study of ancient populations, human remains of unknown origins, or poorly surveyed modern populations, we recommend a conservative approach to kinship determined by LR. Next-generation sequencing data should be used when possible, but the costs and technology involved might be prohibitive. Therefore, in potentially contentious situations or cases lacking sufficient external information, uniparental markers should be analyzed: ideally, complete mitochondrial genomes and Y-chromosome haplotypes (STR, SNP, and/or sequencing).

This article describes current matrimonial strategies and residence patterns in two communities in the Sakha Republic. In Tolon, a rural settlement in central Sakha, community exogamy is predominant and patrilocality is detectable in postnuptial residence patterns. In the sub-Arctic town of Khonuu no gendered residence patterns are observed. Khonuu has an airport and serves as a regional capital. In Khonuu matrimonial decisions follow the immigration of men and couples rather than traditional strategies connected with horse- and cattle-based subsistence. This article discusses the possible biological, historical, and cultural reasons that explain the observance or lack of observance of traditional marriage in the contemporary Sakha Republic.

Paleovirology, the study of ancient viruses, is a recent scientific development. It belongs to the domain of ancient DNA studies, paleogenetics. The analytical power of this discipline, that has many applications, is nevertheless limited by the poor conservation of DNA (and RNA) found in fossils and the subsequent methodological problems (amplification issues, post-mortem lesions, contaminations). These issues must be addressed in order to exploit the full potential of preserved nucleic acids within ancient remains. Once these difficulties have been overcome, it becomes possible to characterize ancient viruses and particular steps of their evolution or even adaptation to their hosts. Shedding light on key aspects of past epidemics allows for a better comprehension of present situations and for the anticipation of future situations where viral pathogens emerge or re-emerge. The examples described in this article illustrate these facts.

Seventeen years of archaeological and anthropological expeditions in North-Eastern Siberia (in the Sakha Republic, Yakutia) have permitted the genetic analysis of 150 ancient (15th-19th century) and 510 modern individuals. Almost all males were successfully analysed (Y-STR) and this allowed us to identify paternal lineages and their geographical expansion through time. This genetic data was confronted with mythological, historical and material evidence to establish the sequence of events that built the modern Yakut genetic diversity. We show that the ancient Yakuts recovered from this large collection of graves are not representative of an ancient population. Uncommonly, we were also able to demonstrate that the funerary preference observed here involved three specific male lineages, especially in the 18th century. Moreover, this dominance was likely caused by the Russian conquest of Siberia which allowed some male clans to rise to new levels of power. Finally, we give indications that some mythical and historical figures might have been the actors of those genetic changes. These results help us reconsider the genetic dynamics of colonization in some regions, question the distinction between fact and myth in national histories and provide a rare insight into a funerary ensemble by revealing the biased process of its composition., Analysis of human remains from 15th-19th Century North-Eastern Siberia, in combination with modern samples, indicate the drivers of present day Yakut genetic diversity, and enable archaeological interpretations of population dynamics at this time.

According to historical sources, ancient Hungarians were made up of seven allied tribes and the fragmented tribes that split off from the Khazars, and they arrived from the Eastern European steppes to conquer the Carpathian Basin at the end of the ninth century AD. Differentiating between the tribes is not possible based on archaeology or history, because the Hungarian Conqueror artifacts show uniformity in attire, weaponry, and warcraft. We used Y-STR and SNP analyses on male Hungarian Conqueror remains to determine the genetic source, composition of tribes, and kin of ancient Hungarians. The 19 male individuals paternally belong to 16 independent haplotypes and 7 haplogroups (C2, G2a, I2, J1, N3a, R1a, and R1b). The presence of the N3a haplogroup is interesting because it rarely appears among modern Hungarians (unlike in other Finno-Ugric-speaking peoples) but was found in 37.5% of the Hungarian Conquerors. This suggests that a part of the ancient Hungarians was of Ugric descent and that a significant portion spoke Hungarian. We compared our results with public databases and discovered that the Hungarian Conquerors originated from three distant territories of the Eurasian steppes, where different ethnicities joined them: Lake Baikal-Altai Mountains (Huns/Turkic peoples), Western Siberia-Southern Urals (Finno-Ugric peoples), and the Black Sea-Northern Caucasus (Caucasian and Eastern European peoples). As such, the ancient Hungarians conquered their homeland as an alliance of tribes, and they were the genetic relatives of Asiatic Huns, Finno-Ugric peoples, Caucasian peoples, and Slavs from the Eastern European steppes.

A palaeogenetic analysis has been undertaken on the pre-Hispanic settlement of Los Amarillos (Regional Development Period, Jujuy Province, Argentina) to reconstruct kin relationship between individuals buried in two domestic areas. The aim of this study was first to genetically characterize the relationships between the individuals buried within the same funerary structure and, secondly, to correlate these genetic data with archaeo-anthropological data to discuss the burial practices and social organization of the Los Amarillos community. An analysis of both uniparental (mtDNA and Y-chromosome) and biparental (autosomal STRs) genetic markers was conducted on eighteen individuals recovered from three different burial structures. The very good DNA preservation contributed to characterize 13 mitochondrial haplotypes, 5 Y-chromosomal haplotypes and 11 complete autosomal STR profiles. The kinship analysis revealed that the domestic areas were used as family graves. Furthermore, they reveal that a maternal lineage is shared by a majority of the studied individuals from different sectors, suggesting matrilocal practices. Fil: Mendisco, Fanny. Université Paul Sabatier; Francia. Université de Strasbourg; Francia Fil: Keyser, Christine. Université de Strasbourg; Francia. Université Paul Sabatier; Francia Fil: Seldes, Verónica. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad de Buenos Aires. Facultad de Filosofía y Letras. Instituto Interdisciplinario Tilcara; Argentina Fil: Nielsen, Axel Emil. Secretaría de Cultura de la Nación. Dirección Nacional de Cultura y Museos. Instituto Nacional de Antropología y Pensamiento Latinoamericano; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Russo, Maria Gabriela. Universidad Maimónides. Área de Investigaciones Biomédicas y Biotecnológicas. Centro de Estudios Biomédicos, Biotecnológicos, Ambientales y de Diagnóstico; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Crubézy, Eric. Université Paul Sabatier; Francia Fil: Ludes, Bertrand. Université de Strasbourg; Francia. Université Paul Sabatier; Francia

Over the last few years, palaeogenomic studies of the petrous bone (the densest part of the temporal bone) have shown that it is a source of DNA in both larger quantities and of better quality than other bones. This dense bone around the otic capsule has therefore been called the choice substrate in palaeogenomics. Because the practice of forensic genetics responds to different imperatives, we implemented a study aimed at (i) understanding how and why the petrous bone is an advantageous substrate in ancient DNA studies and (ii) establishing whether it is advantageous in forensic STR typing. We selected 50 individual skeletal remains and extracted DNA from one tooth and one petrous bone from each. We then amplified 24 STR markers commonly used in forensic identification and compared the quality of that amplification using the RFU intensities of the signal as read on the STR profiles. We also performed histological analyses to compare (i) the microscopic structure of a petrous bone and of a tooth and (ii) the microscopic structure of fresh petrous bone and of an archaeological or forensic sample. We show that the RFU intensities read on STR profiles are systematically higher in experiments using DNA extracted from petrous bones rather than teeth. For this reason, we were more likely to obtain a complete STR profile from petrous bone material, increasing the chance of identification in a forensic setting. Histological analyses revealed peculiar microstructural characteristics (tissue organization), unique to the petrous bone, that might explain the good preservation of DNA in that substrate. Therefore, it appears that despite the necessity of analysing longer fragments in forensic STR typing compared to NGS palaeogenomics, the use of petrous bones in forensic genetics could prove valuable, especially in cases involving infants, toothless individuals or very degraded skeletal remains.

Scythians are known from written sources as horse-riding nomadic peoples who dominated the Eurasian steppe throughout the Iron Age. However, their origins and the exact nature of their social organization remain debated. Three hypotheses prevail regarding their origins that can be summarized as a "western origin", an "eastern origin" and a "multi-regional origin". In this work, we first aimed to address the question of the familial and social organization of some Scythian groups (Scytho-Siberians) by testing genetic kinship and, second, to add new elements on their origins through phylogeographical analyses. Twenty-eight Scythian individuals from 5 archeological sites in the Tuva Republic (Russia) were analyzed using autosomal Short Tandem Repeats (STR), Y-STR and Y-SNP typing as well as whole mitochondrial (mtDNA) genome sequencing. Familial relationships were assessed using the Likelihood Ratio (LR) method. Thirteen of the 28 individuals tested were linked by first-degree relationships. When related, the individuals were buried together, except for one adult woman, buried separately from her mother and young sister. Y-chromosome analysis revealed a burial pattern linked to paternal lineages, with men bearing closely related Y-haplotypes buried on the same sites. Inversely, various mtDNA lineages can be found on each site. Y-chromosomal and mtDNA haplogroups were almost equally distributed between Western and Eastern Eurasian haplogroups. These results suggest that Siberian Scythians were organized in patrilocal and patrilineal societies with burial practices linked to both kinship and paternal lineages. It also appears that the group analyzed shared a greater genetic link with Asian populations than Western Scythians did.

The European DNA profiling group (EDNAP) organized a sixth collaborative exercise on RNA/DNA co-analysis for body fluid/tissue identification and STR profiling. The task was to identify skin samples/contact traces using specific RNA biomarkers and test three housekeeping genes for their suitability as reference genes. Eight stains, a skin RNA dilution series and, optionally, bona fide or mock casework samples of human or non-human origin were analyzed by 22 participating laboratories using RNA extraction or RNA/DNA co-extraction methods. Two sets of previously described skin-specific markers were used: skin1 pentaplex (LCE1C, LCE1D, LCE2D, IL1F7 and CCL27) and skin2 triplex (LOR, KRT9 and CDSN) in conjunction with a housekeeping gene, HKG, triplex (B2M, UBC and UCE). The laboratories used different chemistries and instrumentation. All laboratories were able to successfully isolate and detect mRNA in contact traces (e.g., human skin, palm-, hand- and fingerprints, clothing, car interiors, computer accessories and electronic devices). The simultaneous extraction of RNA and DNA provides an opportunity for positive identification of the tissue source of origin by mRNA profiling as well as a simultaneous identification of the body fluid donor by STR profiling. The skin markers LCE1C and LOR and the housekeeping gene marker B2M were detected in the majority of contact traces. Detection of the other markers was inconsistent, possibly due to the low amounts and/or poor quality of the genetic material present in shed skin cells. The results of this and the previous collaborative RNA exercises support RNA profiling as a reliable body fluid/tissue identification method that can easily be combined with current STR typing technology.

Objectives This work focuses on the populations of South Siberia during the Eneolithic and Bronze Age and specifically on the contribution of uniparental lineage and phenotypical data to the question of the genetic affinities and discontinuities between western and eastern populations. Materials and methods We performed molecular analyses on the remains of 28 ancient humans (10 Afanasievo (3600-2500 BC) and 18 Okunevo (2500-1800 BC) individuals). For each sample, two uniparentally inherited systems (mitochondrial DNA and Y-chromosome DNA) were studied, in order to trace back maternal and paternal lineages. Phenotype-informative SNPs (Single Nucleotide Polymorphisms) were also analyzed, along with autosomal STRs (Short Tandem Repeats). Results Most of the Afanasievo men submitted to analysis belonged to a single sub-haplogroup, R1b1a1a, which reveals the predominance of this haplogroup in these early Bronze Age populations. Conversely, Okunevo individuals carried more diverse paternal lineages that mostly belonged to Asian/Siberian haplogroups. These differences are also apparent, although less strongly, in mitochondrial lineage composition and phenotype marker variant frequencies. Discussion This study provides new elements that contribute to our understanding of the genetic interactions between populations in Eneolithic and Bronze Age southern Siberia. Our results support the hypothesis of a genetic link between Afanasievo and Yamnaya (in western Eurasia), as suggested by previous studies of other markers. However, we found no Y-chromosome lineage evidence of a possible Afanasievo migration to the Tarim Basin. Moreover, the presence of Y-haplogroup Q in Okunevo individuals links them to Native American populations, as was suggested by whole-genome sequencing.

Ancient genomics of horse domestication The domestication of the horse was a seminal event in human cultural evolution. Librado et al. obtained genome sequences from 14 horses from the Bronze and Iron Ages, about 2000 to 4000 years ago, soon after domestication. They identified variants determining coat color and genes selected during the domestication process. They could also see evidence of admixture with archaic horses and the demography of the domestication process, which included the accumulation of deleterious variants. The horse appears to have undergone a different type of domestication process than animals that were domesticated simply for food. Science , this issue p. 442

The Altai Mountains have been a long-term boundary zone between the Eurasian Steppe populations and South and East Asian populations. To disentangle some of the historical population movements in this area, 14 ancient human specimens excavated in the westernmost part of the Mongolian Altai were studied. Thirteen of them were dated from the Middle to the End of the Bronze Age and one of them to the Eneolithic period. The environmental conditions encountered in this region led to the good preservation of DNA in the human remains. Therefore, a multi-markers approach was adopted for the genetic analysis of identity, ancestry and phenotype markers. Mitochondrial DNA analyses revealed that the ancient Altaians studied carried both Western (H, U, T) and Eastern (A, C, D) Eurasian lineages. In the same way, the patrilineal gene pool revealed the presence of different haplogroups (Q1a2a1-L54, R1a1a1b2-Z93 and C), probably marking different origins for the male paternal lineages. To go further in the search of the origin of these ancient specimens, phenotypical characters (i.e. hair and eye color) were determined. For this purpose, we adapted the HIrisPlex assay recently described to MALDI-TOF mass spectrometry. In addition, some ancestry informative markers were analyzed with this assay. The results revealed mixed phenotypes among this group confirming the probable admixed ancestry of the studied Altaian population at the Middle Bronze Age. The good results obtained from ancient DNA samples suggest that this approach might be relevant for forensic casework too.

Summary This palaeogenetic study focused on the analysis of a late prehispanic Argentinean group from the Humahuaca valley, with the main aim of reconstructing its (micro)evolutionary history. The Humahuaca valley, a natural passageway from the eastern plains to the highlands, was the living environment of Andean societies whose cultural but especially biological diversity is still poorly understood. We analyzed the DNA extracted from 39 individuals who populated this upper valley during the Regional Development period (RDP) (between the 11th and 15th centuries CE), to determine their maternal and paternal genetic ancestry. Some mitochondrial and Y-chromosomal haplotypes specific to the Andean region are consistent with an origin in the highlands of Central Andes. On the other hand, a significant genetic affinity with contemporary admixed communities of the Chaco area was detected. Expectedly, recent demographic events, such as the expansion of the Inca Empire or the European colonization, have changed the original mitochondrial gene pool of the ancient Humahuaca Valley community. Finally, we identified a particular geographical organization of the prehispanic populations of Northwestern Argentina. Our results suggest that the communities of the region were divided between two different spheres of interaction, which is consistent with assumptions made by means of craniometric traits.

International audience; The extraction of DNA from skeletal remains is a major step in archeological or forensic contexts. However,diagenesis of mineralized tissues often compromises this task although bones and teeth may represent preservation niches allowing DNA to persist over a wide timescale. This exceptional persistence is not only explained on the basis of complex organo-mineral interactions through DNA adsorption on apatite crystals composing the mineral part of bones and teeth but is also linked to environmental factors such as low temperatures and/or a dry environment. The preservation of the apatite phase itself, as an adsorption substrate, is another crucial factor susceptible to significantly impact the retrieval of DNA.With the view to bring physicochemical evidence of the preservation or alteration of diagenetic biominerals, we developed here an analytical approach on various skeletal specimens (ranging from ancient archeological samples to recent forensic specimens), allowing us to highlight several diagenetic indices so as to better apprehend the complexity of bone diagenesis. Based on complementary techniques (X-ray diffraction (XRD), Fourier transform infrared (FTIR), calcium and phosphate titrations, SEM-EDX, and gravimetry), we have identified specific indices that allow differentiating 11 biological samples, primarily according to the crystallinity and maturation state of the apatite phase. A good correlation was found between FTIR results from the analysis of the v3(PO4) and v4(PO4) vibrational domains and XRD-based crystallinity features. A maximal amount of information has been sought from this analytical approach, by way of optimized posttreatment of the data (spectral subtraction and enhancement of curve-fitting parameters). The good overall agreement found between all techniques leads to a rather complete picture of the diagenetic changes undergone by these 11 skeletal specimens. Although the heterogeneity and scarcity of the studied samples did not allow us to seek direct correlations with DNA persistence, the physicochemical parameters described in this work permit a fine differentiation of key properties of apatite crystals among postmortem samples.As a perspective, this analytical approach could be extended to more numerous sets of specimens so as to draw statistical relationships between mineral and molecular conservation.

Les auteurs exposent les possibilites de retracer l’histoire des populations humaines du passe. A l’aide de marqueurs moleculaires de l’ADN nucleaire (autosomes et chromosome Y) et de l’ADN mitochondrial, ils ont notamment retrace les migrations des premiers peuples des steppes eurasiennes. L’analyse de 26 echantillons humains, dates du milieu du iie millenaire avant Jesus-Christ jusqu’au ive siecle apres Jesus-Christ et provenant de la region de Krasnoyarsk (sud de la Siberie), a revele que les populations europeennes y etaient majoritaires a l’âge du bronze. La repartition de l’haplogroupe R1a1a-M17 du chromosome Y suggere une migration vers l’est des peuples des Kourganes a travers la steppe russo-kazakhe. Grâce a l’etude de polymorphismes ponctuels de sequence (SNP), situes au sein des genes impliques dans la pigmentation, les auteurs ont montre que les habitants du sud de la Siberie, a l’âge du bronze et a l’âge du fer, devaient avoir les yeux, la peau et les cheveux clairs.

Human remains found in aquatic contexts are frequently recovered incomplete and badly decomposed, and therefore present a challenge for medico-legal institutes as their possibilities of analysis for identification and investigation of cause and manner of death are limited. This article aims to demonstrate the potential of forensic examination and analyses (DNA, toxicology, diatoms and entomology) on a set of bones recovered from a river in Strasbourg and the possibility to trigger identification of the victim and circumstances of death despite the state of decomposition and incompleteness of remains.

For fifteen years, part of the work of our research team has been focused on the study of parental links between individuals living hundreds or thousands of years ago, whose remains have been found in single graves or large funerary complexes. These studies have been undertaken using methods developed by forensic genetics to identify individuals, mainly based on the genotyping of autosomal STR (Short Tandem Repeats). Issues arose from this work, namely the limits of studying small numbers of subjects, originating from groups of finite sizes where kinships cannot be inferred a priori and for which reference allelic frequencies do not exist. Although ideal human populations are rare when undertaking such studies, the Yakuts of Eastern Siberia constitute a very advantageous model, with large numbers of small pastoral communities and well-preserved archaeological material. The study of kinship in the ancient Yakuts allowed us to highlight the difficulties in analysing genetic data from small ancient human groups and to develop a strategy to improve the accuracy of statistical computations. This work describes this strategy and possible solutions to the study of populations outside of the frame of reference of global meta-populations, due either to isolation, remoteness or antiquity.

The Basque population inhabits the Franco-Cantabrian region in southwest Europe where Palaeolithic human groups took refuge during the Last Glacial Maximum. Basques have been an isolated population, largely considered as one of the most ancient European populations and it is possible that they maintained some pre-Neolithic genetic characteristics. This work shows the results of mitochondrial DNA analysis of seven ancient human remains from the Cave of Santimamine in the Basque Country dated from Mesolithic to the Late Roman period. In addition, we compared these data with those obtained from a modern sample of Basque population, 158 individuals that nowadays inhabits next to the cave. The results support the hypothesis that Iberians might have been less affected by the Neolithic mitochondrial lineages carried from the Near East than populations of Central Europe and revealed the unexpected presence of prehistoric maternal lineages such as U5a2a and U3a in the Basque region. Comparison between ancient and current population samples upholds the hypothesis of continuity of the maternal lineages in the area of the Franco-Cantabrian region.

In the absence of any other conclusive forensic evidence, DNA profiling is the method of choice for body identification. This study focuses on the case of a carbonized corpse whose complete autosomal short tandem repeat (STR) profile could not lead to direct identification by the investigators. To assist in the progress of investigation, we endeavoured to determine the biogeographical origin and eye colour of the deceased individual. Along with Y chromosome and mitochondrial DNA analyses, we applied a next-generation sequencing (NGS) approach to the study of ancestry informative markers (AIMs) using the HID-Ion AmpliSeq™ Ancestry Panel launched by Thermo Fisher Scientific. This work gave us the opportunity to test this new technology in a real forensic case. Although this study highlights the benefits of such a combined approach, as it markedly improves the specificity of the biogeographical profile, it also underlines the need for the accurate characterization of a larger collection of reference populations and the necessity of caution in data interpretation.

Understanding the peopling history of Europe is crucial to comprehend the origins of modern populations. Of course, the analysis of current genetic data offers several explanations about human migration patterns which occurred on this continent, but it fails to explain precisely the impact of each demographic event. In this context, direct access to the DNA of ancient specimens allows the overcoming of recent demographic phenomena, which probably highly modified the constitution of the current European gene pool. In recent years, several DNA studies have been successfully conducted from ancient human remains thanks to the improvement of molecular techniques. They have brought new fundamental information on the peopling of Europe and allowed us to refine our understanding of European prehistory. In this review, we will detail all the ancient DNA studies performed to date on ancient European DNA from the Middle Paleolithic to the beginning of the protohistoric period.

High-resolution melting (HRM) analysis is a recently developed molecular technique proved to be applicable for detection of genetic variation, notably in sudden cardiac death. In certain circumstances, especially in postmortem genetic investigations, the formalin-fixed and paraffin-embedded (FFPE) tissues are the only DNA source available. The present study aimed to develop HRM assays, optimized for the analysis of FFPE tissues, to detect sequence variations in KCNQ1 exons in a prospective population-based series of postmortem negative sudden death and to compare the results between the paired freshly frozen and FFPE tissue samples simultaneously obtained from the same case. The analyses were conducted in each case of sudden death involving cases younger than 35 years with no significant morphological anomalies particularly with no cardiac structural disease and with negatives toxicological investigations. HRM analysis was successfully optimized for 13 of the 16 exons of the KCNQ1 gene. All mutated samples were correctly identified by HRM whatever the type of tissue tested. However, for FFPE samples, HRM indicated more positive samples than classical sequencing, used in parallel, due to the degradation of DNA by formalin fixation. This is the first postmortem study of KCNQ1 mutation detection with HRM on DNA extracted from FFPE samples with adapted protocol. Despite the false-positive detection, we concluded that the use of HRM as a screening method with FFPE samples to analyze KCNQ1 mutations can reduce the number of sequencing reactions and, thus, results in substantial time and cost savings.

Objective Mechanisms of sepsis-induced myocardial and diaphragmatic alteration are multiple and remain largely unknown, particularly in humans. In the present study, we compared the inducible nitric oxide synthase (NOS-2) expression, the peroxynitrite production and the expression and activation of the ubiquitin proteolytic pathway in the wall of the 4 cardiac chambers, in the diaphragm, and in the rectus abdominis. Patients Seven patients who died from septic shock associated with a myocardial depression and 5 nonseptic (control) patients. Measurements and results We evaluated protein expression by Western blot. Nitrotyrosin and ubiquitin residues were localized by immunofluorescence. NOS-2, nitrated proteins, free ubiquitin, and ubiquitinated proteins are overexpressed in the wall of the four cardiac cavities, in the diaphragm and in the rectus abdominis of septic patients at a similar level. Ubiquitinated proteins with a molecular mass of 50, 35, 30, and 25 kD were consistently detected in heart, diaphragm, and rectus abdominis of septic shock patients but lacking in nonseptic patients. In situ immunolabelling of ubiquitin showed a colocalisation with nitrotyrosine residues at the sarcomeric level of cardiac myocytes in septic patients. Conclusions This study showed the first evidence for the activation of the proteolytic ubiquitin–proteasome pathway in human heart and diaphragm in septic shock.

Formalin-fixed and paraffin-embedded tissue (FF-PET) is an invaluable resource for retrospective molecular genetic studies, but the extraction of high-quality genomic DNA from FF-PET is still a problematic issue. Despite the range of DNA extraction methods currently in use, the association of phenol-chloroform extraction and silica-based purification protocols, reported in ancient DNA studies on archaeological bones, has, to our knowledge, not been used for DNA extraction from FF-PET yet. The present study compared the efficiency of three DNA extraction and purification protocols from two different FF-PET substrates, heart and liver, by using quantitative PCR and multiplex amplification. We showed that the method, using phenol-chloroform and the QIAamp DNA mini Kit (Qiagen), was the most effective DNA extraction and purification method and that the DNA quantity extracted from liver is statistically more important than that extracted from heart. Autosomal STR typing by multiplex amplifications gave partial allelic profiles with only small size products (less than 300 bases) amplified, suggesting that DNA extracted from FF-PET was degraded. In conclusion, the protocol presented here, previously described in studies on ancient bones, should find application in different molecular studies involving FF-PET material.

Ancient DNA recovered from 21 individuals excavated from burial sites in the Pampa Grande (PG) region (Salta province) of North-Western Argentina (NWA) was analyzed using various genetic markers (mitochondrial DNA, autosomal STRs, and Y chromosomal STRs). The results were compared to ancient and modern DNA from various populations in the Andean and North Argentinean regions, with the aim of establishing their relationships with PG. The mitochondrial haplogroup frequencies described (11% A, 47% B, and 42% D) presented values comparable to those found for the ancient Andean populations from Peru and San Pedro de Atacama. On the other hand, mitochondrial and Y chromosomal haplotypes were specific to PG, as they did not match any other of the South American populations studied. The described genetic diversity indicates homogeneity in the genetic structure of the ancient Andean populations, which was probably facilitated by the intense exchange network in the Andean zone, in particular among Tiwanaku, San Pedro de Atacama, and NWA. The discovery of haplotypes unique to PG could be due to a loss of genetic diversity caused by recent events affecting the autochthonous populations (establishment of the Inca Empire in the region, colonization by the Europeans).

To help unravel some of the early Eurasian steppe migration movements, we determined the Y-chromosomal and mitochondrial haplotypes and haplogroups of 26 ancient human specimens from the Krasnoyarsk area dated from between the middle of the second millennium BC. to the fourth century AD. In order to go further in the search of the geographic origin and physical traits of these south Siberian specimens, we also typed phenotype-informative single nucleotide polymorphisms. Our autosomal, Y-chromosomal and mitochondrial DNA analyses reveal that whereas few specimens seem to be related matrilineally or patrilineally, nearly all subjects belong to haplogroup R1a1-M17 which is thought to mark the eastward migration of the early Indo-Europeans. Our results also confirm that at the Bronze and Iron Ages, south Siberia was a region of overwhelmingly predominant European settlement, suggesting an eastward migration of Kurgan people across the Russo-Kazakh steppe. Finally, our data indicate that at the Bronze and Iron Age timeframe, south Siberians were blue (or green)-eyed, fair-skinned and light-haired people and that they might have played a role in the early development of the Tarim Basin civilization. To the best of our knowledge, no equivalent molecular analysis has been undertaken so far.

International audience; The aim of this study was to demonstrate the presence of the A189G age-related point mutation on DNA extracted from bone. For this, a peptide nucleic acid (PNA)/DNA sequencing method which can determine an age threshold for the appearance of the mutation was used. Initially, work was done in muscle tissue in order to evaluate the sensitivity of the technique and afterwards in bone samples from the same individuals. This method was also applied to ancient bones from six well-preserved skeletal remains. The mutation was invariably found in muscle, and at a rate of up to 20% in individuals over 60 years old. In modern bones, the mutation was detected in individuals aged 38 years old or more, at a rate of up to 1%, but its occurrence was not systematic (only four out of ten of the individuals over 50 years old carried the heteroplasmy). For ancient bones, the mutation was also found in the oldest individuals according to osteologic markers. The study of this type of age-related mutation and a more complete understanding of its manifestation has potentially useful applications. Combined with traditional age markers, it could improve identification accuracy in forensic cases or in anthropological studies of ancient populations.

Οι « εκτελεσθείσες των Φουρνών ». Ιατροδικαστική και παλαιοπαθολογική επανεξέταση Οι δύο αποκεφαλισμένοι ανθρώπινοι σκελετοί που ανακαλύφθηκαν το 1960 σε ένα βόθρο, που συνδέεται με την αποχέτευση των αποχωρητηρίων της Οικίας των Φούρνων στη Δήλο, δημοσιεύτηκαν το 1973 (Études déliennes [ BCH Suppl. I], σελ. 173-181) και οδήγησαν σε διάφορες ερμηνείες. Οι σκελετοί αυτοί υφίστανται μια νέα ανθρωπολογική εξέταση, καθώς και, για πρώτη φορά, μια παλαιοπαθολογική εξέταση, που καταλήγει σε απρόσμενα συμπεράσματα. Πρώτον, και τα δύο θύματα ήταν γυναίκες. Επίσης, η χρονολόγηση του θανάτου τους, όπως προκύπτει από τη χρήση άνθρακα 14, είναι πολύ αρχαιότερη από αυτήν που είχε προταθεί με βάση τα ιστορικά συμφραζόμενα. Η εκτέλεση ακολούθησε μια περίοδο κατά την οποία τα θύματα κρατούνταν αλυσοδεμένα. Τέλος, τα πτώματα ρίφθηκαν σε ένα βόθρο, ο οποίος ήταν ακόμα σε λειτουργία. Δεν μπορούμε να συνδέσουμε το επεισόδιο αυτό με ένα συγκεκριμένο γεγονός ή κοινωνικό πλαίσιο. Ωστόσο, η θανάτωση αυτή θυμίζει περισσότερο τελετουργικό φόνο μιας θρησκευτικής ομάδας παρά εκτέλεση θανατικής ποινής., The "decapitated of Fourni". A medical-legal and palaeopathological reexamination The two decapitated human skeletons discovered in 1960 in a bothros connected to the clearing of the latrines of the Maison de Fourni at Delos were published in 1973 (Études déliennes [BCH Suppl. I], p. 173-181) and have since led to divergent interpretations. They are here subjected to a new anthropolical study and, for the first time, to a palaeopathological examination, which leads to unexpected conclusions. Firstly, both victimes would have been feminine. Secondly, the date of their beheading as provided by C-14 analysis is much higher than that proposed based on the historical context. The execution was preceded by a period of shackling and restraint. Finally, the bodies were thrown into a cesspool still in use. This episode cannot be connected to a precise event or social context. Nevertheless, this killing recalls more the ritual killing of a sect than a capital execution., Les deux squelettes humains décapités découverts en 1960 dans un bothros lié à l’évacuation des latrines de la Maison de Fourni à Délos ont été publiés en 1973 (Études déliennes [ BCH Suppl. I], p. 173-181) et ont depuis lors donné lieu à des interprétations divergentes. Elles font ici l’objet d’une nouvelle étude anthropologique et, pour la première fois, paléopathologique, qui aboutit à des conclusions inattendues. En premier lieu, les victimes seraient toutes deux de sexe féminin. D’autre part, la date de leur supplice telle qu’elle est donnée par le 14C est nettement plus haute que celle qu’on avait proposée en se fondant sur le contexte historique. L’exécution a été précédée d’une période d’entrave et de contention. Enfin, les corps ont été jetés dans une fosse d’aisance encore en activité. On ne peut rattacher cet épisode à un événement ou à un cadre social précis. Toutefois, cette mise à mort évoque plus le meurtre rituel de sectateurs qu’une exécution capitale., Charlier Philippe,Le Roy Christian,Keyser Christine. Les « suppliciées de Fourni ». Réexamen médico-légal et paléopathologie. In: Bulletin de correspondance hellénique. Volume 132, livraison 1, 2008. pp. 617-637.

In the present study, a set of 13 Y-chromosomal single nucleotide polymorphisms (Y-SNPs) selected for the identification of the most frequent Asian Y-haplogroups was included in an allele-specific primer extension assay. Single nucleotide polymorphism (SNP) genotyping was accomplished by co-amplification of these 13 DNA fragments within 2 multiplex PCRs followed by detection with 1 minisequencing reaction using the SNaPshottrade mark Multiplex kit and analysis of extension products by capillary electrophoresis. First developed on modern samples, the assay was optimized for the analysis of 11 ancient DNA (aDNA) samples from the Krasnoyarsk region (southern Siberia) that were dated from 5,500-1,800 years before present (YBP). SNP typing was successful for most of them, which were all assigned to Y-haplogroup R1a1 except one. These results show that SNPs are well-suited for the analysis of aged and degraded DNA samples. Moreover, we found that the SNaPshot minisequencing methodology is a convenient, robust, and efficient method for SNP typing. To our knowledge, this study reports the first successful investigation of Y-SNPs on aDNA samples. The potential use of Y-SNPs in both evolutionary and forensic fields is also discussed.

The allele frequency distribution of 15 short tandem repeats (STR) loci contained in the AmpFlSTR Identifiler™ PCR Amplification Kit (Applied Biosystems), was determined in two Berber populations from Asni and Bouhria, in Central and Eastern Morocco, respectively. A total of 209 individuals were typed. No deviations from the Hardy–Weinberg equilibrium were observed for Asni at the 15 STRs loci whereas for the Bouhria samples, two loci (D5S818 and TH01) showed significant departures from Hardy–Weinberg expectations (after Bonferroni's correction). All loci are highly polymorphic and population differentiation tests showed that the Moroccan samples from Asni and Bouhria have significant differences in 4 out of 15 loci (D21S11, D7S820, D16S539 and TPOX). The aim of the study was to obtain accurate allele frequencies relevant for forensic applications. Comparative analyses between our population data and other population samples gathered from the literature are also presented.

Genetic testing for cardiac channelopathies in sudden unexplained death (SUD) has developed substantially over the last years. The Next Generation Sequencing (NGS) technology provides an unprecedented opportunity to screen for genetic variations underlying arrhythmogenic genes in a short period of time at a low cost. The present study aimed to perform genetic testing with NGS technologies on the Ion Torrent Personal Genome Machine™ (Ion PGM™) sequencer, in targeting a total of 23 genes reported to be associated with inherited cardiac channelopathies in order to identify the possible cause of death in a cohort of post-mortem cases. The molecular analyses focused on 16 cases of SUD, aged less than 35 years old. In all cases, the cause of death could not be determined after a rigorous autopsy associated with histopathological and toxicological analyses according to the guidelines of the Association for European Cardiovascular Pathology. DNA was extracted from fresh frozen tissue. An average of 200 variants was identified per case. However, after the prioritization process using a new scoring program (VaRank) and after the conjunction of clinical data and molecular findings, four “likely pathogenic” variants (including two undescribed variants), were identified in three cases (18.75%) of our cohort in the genes KCNH2 , ANK2 , SCN5A and RYR2 . One case, who died during psychiatric hospitalization after administration of a QT prolonging drug, showed a double “likely pathogenic” variant in Long QT genes ( ANK2 and SCN5A ) which may have predisposed to drug-induced cardiac arrhythmias. Our study illustrates that the NGS approach based on AmpliSeq™ libraries and Ion Torrent PGM™ sequencing may be an efficient approach, integrated to post-mortem examination. Given the massive amount of information generated by NGS, a rigorous filtration strategy of variants coupled with multidisciplinary collaboration is crucial to determine the potential pathogenic role of identified variants in the cause of death.

We successfully extracted DNA from a bone sample of a Neolithic skeleton (dated 3,600 +/- 60 years BP) excavated in northeastern Yakutia (east Siberia). Ancient DNA was analyzed by autosomal STRs (short tandem repeats) and by sequencing of the hypervariable region I (HV1) of the mitochondrial DNA (mtDNA) control region. The STR profile, the mitochondrial haplotype, and the haplogroup determined were compared with those of modern Eurasian and Native American populations. The results showed the affinity of this ancient skeleton with both east Siberian/Asian and Native American populations.

DNA markers used for individual identification in forensic sciences are based on repeat sequences in nuclear DNA and the mitochondrial DNA hypervariable regions 1 and 2. An alternative to these markers is the use of single nucleotide polymorphisms (SNPs). These have a particular advantage in the analysis of degraded or poor samples, which are often all that is available in forensics or anthropology. In order to study the potential of SNP analysis in these fields, 41 SNPs were selected on the basis of following criteria: conservation, lack of phenotypic expression, and frequency of occurrence in populations. Thirty-six autosomal SNPs were used for genotyping 21 inclusionary and 3 exclusionary paternity cases. The behavior of 5 X-chromosome SNPs was analyzed in a French representative population. Our approach to SNP typing is a multiplex PCR based amplification followed by simultaneous detection by primer extension (PEX) analyzed by Matrix Assisted Laser Desorption Ionization Time-of-Flight Mass Spectrometry (MALDI-TOF MS). The selected autosomal SNPs showed independent inheritance and gave clear results in paternity investigation. All X-SNPs were useful as both paternity and identification markers. PEX and MALDI-TOF MS, with their high sensitivity, precision and speed, gave a powerful method for forensic and anthropological exploitation of biallelic markers.

We extracted DNA from two skeletons belonging to the Sytho-Siberian population, which were excavated from the Sebystei site (dating back 2,500 years) in the Altai Republic (Central Asia). Ancient DNA was analyzed by autosomal short tandem repeats (STRs) and by the sequencing of the hypervariable region 1 (HV1) of the mitochondrial DNA (mtDNA) control region. The results showed that these two skeletons were not close relatives. Moreover, their haplogroups were characteristic of Asian populations. Comparison with the haplogroup of 3,523 Asian and American individuals linked one skeleton with a putative ancestral paleo-Asiatic population and the other with Chinese populations. It appears that the genetic study of ancient populations of Central Asia brings important elements to the understanding of human population movements in Asia.

Fifteen somatic (D8S1179, D21S11, D7S820, CSF1PO, D3S1358, TH01, D13S317, D16S539, D2S1338, D19S433, VWA, TPOX, D18S51, D5S818, FGA) and five Y-chromosome (DYS19, DYS390, DYS391, DYS392, DYS393) short tandem repeat (STR) system analyses were carried out in a Mongolian population in order to define the possible relationship between Mongolians and old Hungarian population. For STR data analysis the Microsoft Excel-PowerStats program was used. Inter-population data analysis was performed with Arlequin Software ver. 2.000. The somatic markers showed meaningful difference between Mongolians and old Hungarians but the distribution of the Y-chromosome STR systems refers to a closer relationship between the old Hungarian and oriental populations.

Genetic analysis of archaeological remains may provide information of prime importance to the understanding of human past history. In order to investigate the history of Mongolian populations, we have analyzed the skeletal remains of 56 specimens excavated from the Egyin Gol necropolis located in northern Mongolia. This burial site is linked to the Xiongnu period, which extended from the 3rd century B.C. to the 3rd century A.D. The environmental conditions at this site were cold and dry, and the DNA preservation was exceptional, allowing nuclear markers such as microsatellites to be studied. Molecular analyses were performed on long cortical bone samples with the AmplFlSTR Profiler Plus kit (Applied Biosystems). Amplifications were successful for 47 of the 56 specimens analysed. Molecular sexing confirmed archeological data based on morphometric parameters and allowed the gender of four juvenile skeletons to be determined. Short tandem repeat (STR) analysis showed close relationships between several specimens and provides additional background information concerning social organization within the necropolis as well as funeral practices linked to Xiongnu populations. D 2003 Elsevier Science B.V. All rights reserved.

After initially slow progress in identifying DNA polymorphisms on the non recombining part of the human Y chromosome, a large number of polymorphic markers are now available to define individual or population-specific haplotypes or haplogroups. Among them the Y-chromosomal STRs (Short Tandem Repeats) have been increasingly used over the past few years for the study of human evolution as well as for human identification in forensic case-work and paternity testing. After a brief summary of the features of such markers, this paper deals with some applications of the Y-STR haplotyping.

International audience; We present here the results of our inter-disciplinary examination of the mummified heart of Blessed Anne-Madeleine Remuzat (1696–1730). This organ has been examined in the context of a canonization process. This analysis is related to important aspects of the early history of anatomy in Europe, that of “Holy autopsies”, and to the relationship between anatomical investigations, Catholic theology, and religious/medical customs. According to anatomical, genetic, toxicological, and palynological analyses, it has been shown that this organ has not been naturally (“miraculously”) conserved but embalmed using myrtle, honey, and lime. Moreover, a right ventricle dilatation has been diagnosed, that may represent a post-tuberculosis condition and may have played a role in the cause of death of this religious figure.

International audience; In order to shed some light on DNA preservation over time in skeletal remains from a physicochemicalviewpoint, adsorption and desorption of DNA on a well characterized synthetic apatite mimicking boneand dentin biominerals were studied. Batch adsorption experiments have been carried out to determinethe effect of contact time (kinetics), DNA concentration (isotherms) and environmentally relevant factorssuch as temperature, ionic strength and pH on the adsorption behavior. The analogy of the nanocrystallinecarbonated apatite used in this work with biological apatite was first demonstrated by XRD, FTIR, andchemical analyses. Then, DNA adsorption kinetics was fitted with the pseudo-first order, pseudo-secondorder, Elovich, Ritchie and double exponential models. The best results were achieved with the Elovichkinetic model. The adsorption isotherms of partially sheared calf thymus DNA conformed satisfacto-rily to Temkin's equation which is often used to describe heterogeneous adsorption behavior involvingpolyelectrolytes. For the first time, the irreversibility of DNA adsorption toward dilution and significantphosphate-promoted DNA desorption were evidenced, suggesting that a concomitant ion exchange pro-cess between phosphate anionic groups of DNA backbone and labile non-apatitic hydrogenphosphateions potentially released from the hydrated layer of apatite crystals. This work should prove helpful fora better understanding of diagenetic processes related to DNA preservation in calcified tissues.