Ancient DNA Damage Analysis of Late Quaternary Mammalian Fossil Samples in Northeast China / 中国生物化学与分子生物学报

The advancement of the next generation sequencing (NGS) technology has promoted the development of ancient DNA research. Ancient DNA has made outstanding contributions in various fields such as human origin, animal evolution, etc. How to effectively extract and mine the genetic information from fossil and sub-fossil remains excavated from specific locations is a prerequisite for optimizing their important roles in many fields. In this study, we correlated the two main indicators of DNA damage (terminal base replacement rate, average fragment length) with the possible factors such as the burial time, geological epochs, tissue types, and sequencing library construction methods. The results show that the end base replacement rate of ancient DNA from Northeastern China is positively correlated with the water content of the environment and the ages of the samples. Among samples of different geological epochs, ancient DNA end base replacement rates have significant differences. On the contrary, different tissue types of the remains have no significant effects on the end base replacement rate of ancient DNA. The average fragment size of the molecules has no obvious correlation with the factors mentioned above. The results provide both solid data for investigating the characteristics of ancient DNA from specimens collected in Northeastern China, and valuable information for collecting appropriate samples from different geographical locations and the downstream storage before wet lab procedures after excavation.

Peopling of India: Ancient DNA perspectives

To reconstruct and explain patterns of genetic diversity of modern humans, understanding their past and present geneticprofile is crucial. While genomes of contemporary people can provide information about present day population structure,analysis of ancient genomes may provide unprecedented insights about the past demographic events that have shaped thecontemporary gene pool. Population genetics has recently witnessed an explosion in studies on ancient human populationhistories, primarily from Europe and America. South Asia has no representation in the ancient genomics literature, despitethe wealth of archaeological richness in the form of human skeletal remains that exist in collections all over the country.Representing one-fifth of present day humanity calls for understanding the demographic history of south Asia not merely asa prerequisite but as an urgent need to understand its genetic variations on a global scale. Although the overall picture istaking form, new archaeological and genetic information from the region has started to reveal a more complex scenario ofancient human migrations and admixtures than was ever known before. In this article, we discuss a meaningful insight onthe current status of ancient DNA (aDNA) research in India. We have also summarized a few but important aDNA studies,which have been successfully carried out in India. Furthermore, we have highlighted the potential opportunity of aDNAresearch in the Indian subcontinent.

Early ‘Aryans’ and their neighbors outside and inside India

Data from archaeology, linguistics, population genetics, and from early Vedic texts, which deal with religion, mythologyand rituals, have to be assembled and closely compared in order to gain a comprehensive picture of the early ‘Aryans’. Suchinterdisciplinary dialogue is necessary in order to establish areas of overlap of data. This paper attempts to indicate awestern Central Asian origin of the Indo-Aryan speakers, in the steppe belt near the Urals, from where they moved, via theInner Asian Mountain belt and Bactria, into India. Their gradual migration entailed acculturation with previous populations,their languages and cultures.

Investigating the West Eurasian ancestry of Pakistani Hazaras

The Hazaras are a distinct ethnic group from central Afghanistan and northwestern Pakistan of Mongoloid descent. Here, we sought to dissect the genetic admixture history of the Pakistani Hazaras and investigated their likely affiliation to ancient and extant West Eurasian populations. Our results indicated that the likely proportion of West Eurasian ancestry was low in the Hazaras and could be attributed putatively to a combination of Steppe populations from Early/Middle Bronze Age or Middle/Late Bronze Age and the Neolithic Iranians. Our results expand upon the current understanding and provide an improved resolution into the population history of the Pakistani Hazaras.

Analysis of COI and ITS2 regions of DNA obtained from Paragonimus westermani eggs in ancient coprolites on Joseon dynasty mummies

The genetic information of ancient Paragonimus westermani, the oriental lung fluke infecting over 20 million people worldwide, has not been thoroughly investigated thus far. We analysed genetic markers (COI and ITS2) of P. westermani from coprolite specimens (n = 6) obtained from 15th to 18th century Korean mummies. Our results indicated that all P. westermani sequences were generally distinct from the other species of the genus Paragonimus. The sequences were clustered into three groups: Group I for East Asia; Group II for South and Southeast Asia; and Group III for India and Sri Lanka. In this study, we found that ancient P. westermani sequences in Korea belong to Group I, adding invaluable information to the existing knowledge of Paragonimus paleogenetics.

Cytochrome C Oxidase Subunit 1, Internal Transcribed Spacer 1, Nicotinamide Adenine Dinucleotide Hydrogen Dehydrogenase Subunits 2 and 5 of Clonorchis sinensis Ancient DNA Retrieved from Joseon Dynasty Mummy Specimens

We analyzed Clonorchis sinensis ancient DNA (aDNA) acquired from the specimens of the Joseon mummies. The target regions were cytochrome C oxidase subunit 1 (CO1), internal transcribed spacer 1 (ITS1), nicotinamide adenine dinucleotide hydrogen (NADH) dehydrogenase subunits 2 (NAD2) and 5 (NAD5). The sequences of C. sinensis aDNA was completely or almost identical to modern C. sinensis sequences in GenBank. We also found that ITS1, NAD2 and NAD5 could be good markers for molecular diagnosis between C. sinensis and the other trematode parasite species. The current result could improve our knowledge about genetic history of C. sinensis.

Mitochondrial DNA Analysis of a Sample of Pre-Columbian Acient Remains from Norte de Santander, Colombia (Chitarera Cultural Area) / Análisis de adn mitocondrial en una muestra de restos óseos precolombinos de norte de santander, colombia (área cultural chitarera)

RESUMEN Los análisis de ADN antiguo (ADNa) han incrementado en los últimos años permitiendo conocer la diversidad genética de las poblaciones precolombinas. En Colombia, existen pocos registros arqueológicos de la población prehispánica del Norte de Santander habitada en el siglo XVI por el grupo Chitarero. Por este motivo, nos propusimos analizar la diversidad genética a partir de secuencias de la región HVRI del ADNmt y determinar sus posibles relaciones con otras comunidades tanto antiguas como contemporáneas. Se analizaron siete individuos precolombinos asociados a este grupo prehispánico, recuperados en los municipios de Cácota y Silos en el departamento de Norte de Santander de los Andes Orientales colombianos, siguiendo criterios estrictos de autenticidad para el ADNa. En todos los individuos se logró identificar el haplogrupo B caracterizado por el polimorfismo en la posición 16217C, siendo éste uno de los más frecuentes en comunidades precolombinas y contemporáneas de los Andes Suramericanos. Este hallazgo indica que este grupo poblacional se encuentra estrechamente emparentado por línea materna, con posibles índices de endogamia, con una probable densidad demográfica baja y una baja diversidad genética, similares a lo observado en comunidades pertenecientes a periodos anteriores como el Formativo. Este grupo precolombino exhibe una de las diversidades genéticas más bajas reportadas en las poblaciones pertenecientes a la familia lingüistica Chibcha. Estos resultados genéticos coinciden con los planteamientos sobre el grupo Chitarero de pertenecer a comunidades pequeñas independientes, con asentamientos dispersos, apartados unos de otros.

ABSTRACT In the last few years there has been an increase in ancient DNA (aDNA) analyses that has allowed shedding light on the diversity of pre-Columbian populations. In Colombia, there are few archaeological records belonging to the prehispanic population from Norte de Santander inhabited in the XVI century by the Chitarero. For this reason, we performed a genetic diversity analysis of the HVRI region of mtDNA in order to determine their possible relationships with other communities both ancient and contemporary. We analyzed seven pre-Columbian individuals belonging to this pre-Hispanic group, recovered from the municipalities of Cácota and Silos in the department of Norte de Santander located at the Colombian Andes, following strict authenticity criteria for aDNA. All individuals were identified as belonging to haplogroup B, characterized by the polymorphism found at position 16217C which is one of the most frequent haplogroups in pre-Columbian and contemporary communities of the South American Andes. This finding suggests that this population group was closely related through its maternal lineage, with possible inbreeding indexes, low population density and therefore low genetic diversity, similar to what is observed in communities belonging to previous periods such as the Formative period. This pre-Columbian group exhibits one of the lowest genetic diversities reported in populations belonging to the Chibcha linguistic family. These genetic results coincide with the views on the Chitarero group as belonging to small independent communities, with dispersed settlements separated among them.

Principles of Archaeogenetics and the Current Trends of Ancient Genome Studies / 체질인류학회지

Archaeogenetics is an academic discipline that aims to establish scientific facts of human history by integrating ancient DNA analyses with archaeological and anthropological evidence. After ancient DNA research was initiated about 30 years ago, it has been innovated so rapidly that the range of analysis has been extended toward the whole genome sequence of ancient genomes in recent 10 years. By this development, researchers have been able to study in detail the origins and migration patterns of hominin species and ancient human populations by approaches of evolutionary genetics. This study has reviewed main principles of the archaeogenetic analysis and the current trends of ancient genome studies with recent achievements. While sampling techniques and statistical analyses have been improved, typical research methods have been established by the findings on hominins and ancient western Eurasia populations. Recently, archaeogenecists have been applying the methods to studying those in other geographical areas. Nonetheless, there is still the lack of ancient genome research about populations in Eastern Asia including the Korean peninsula. This review ultimately aims to predict possibilities and promise of future ancient genome studies of ancient Korean populations.

Ancient Mitochondrial DNA Analyses of Ascaris Eggs Discovered in Coprolites from Joseon Tomb

Analysis of ancient DNA (aDNA) extracted from Ascaris is very important for understanding the phylogenetic lineage of the parasite species. When aDNAs obtained from a Joseon tomb (SN2-19-1) coprolite in which Ascaris eggs were identified were amplified with primers for cytochrome b (cyt b) and 18S small subunit ribosomal RNA (18S rRNA) gene, the outcome exhibited Ascaris specific amplicon bands. By cloning, sequencing, and analysis of the amplified DNA, we obtained information valuable for comprehending genetic lineage of Ascaris prevalent among pre-modern Joseon peoples.

Insights from paleomicrobiology into the indigenous peoples of pre-colonial America - A Review

This review investigates ancient infectious diseases in the Americas dated to the pre-colonial period and considers what these findings can tell us about the history of the indigenous peoples of the Americas. It gives an overview, but focuses on four microbial pathogens from this period: Helicobacter pylori, Mycobacterium tuberculosis, Trypanosoma cruzi and Coccidioides immitis, which cause stomach ulceration and gastric cancer, tuberculosis, Chagas disease and valley fever, respectively. These pathogens were selected as H. pylori can give insight into ancient human migrations into the Americas, M. tuberculosis is associated with population density and urban development, T. cruzi can elucidate human living conditions and C. immitis can indicate agricultural development. A range of methods are used to diagnose infectious disease in ancient human remains, with DNA analysis by polymerase chain reaction one of the most reliable, provided strict precautions are taken against cross contamination. The review concludes with a brief summary of the changes that took place after European exploration and colonisation.

Chagas disease in prehistory

The classical hypothesis proposes that Chagas disease has been originated in the Andean region among prehistoric people when they started domesticating animals, changing to sedentary habits, and adopting agriculture. These changes in their way of life happened nearly 6,000 years ago. However, paleoparasitological data based on molecular tools showed that Trypanosoma cruzi infection and Chagas disease were commonly found both in South and North American prehistoric populations long before that time, suggesting that Chagas disease may be as old as the human presence in the American continent. The study of the origin and dispersion of Trypanosoma cruzi infection among prehistoric human populations may help in the comprehension of the clinical and epidemiological questions on Chagas disease that still remain unanswered.

A hipótese clássica sobre a origem da doença de Chagas propõe que tenha surgido entre as populações pré-históricas dos Andes quando começaram a domesticar animais, mudaram para hábitos sedentários e adotaram a agricultura. Estas mudanças em seus hábitos de vida aconteceram há aproximadamente 6.000 anos. Entretanto, os dados da paleoparasitologia, baseados na biologia molecular, mostraram que a infecção por Trypanosoma cruzi e a doença de Chagas eram comuns tanto em populações pré-históricas da América do Sul e América do Norte muito antes deste período. De acordo com os dados paleoparasitológicos, a doença de Chagas pode ser tão antiga quanto a presença humana no continente americano. O estudo sobre a origem e dispersão da infecção por Trypanosoma cruzi entre populações humanas pré-históricas pode auxiliar na compreensão de questões clínicas e epidemiológicas sobre a doença de Chagas que ainda permanecem sem resposta.

Rastreo genético del streptococcus mutans / Streptococcus mutans genetic tracking

Las técnicas moleculares para recuperar DNA antiguo brindan la posibilidad de comparar la evolución molecular a través del tiempo, ya que constituye una herramienta para aclarar el diagnóstico de posibles enfermedades infecciosas del pasado. Aislar y secuenciar un fragmento de DNA de Streptococcus mutans fosilizado, considerado el principal agente infeccioso implicado en la formación de la placa bacteriana y el desarrollo de la caries dental, utilizando la reacción en cadena de polimerasa (PCR). La muestra estuvo conformada por caries y tártaro dental proveniente de dientes de diferentes colecciones de México, Barcelona e Islas Baleares. La metodología fue adaptada a las condiciones de conservación de este tipo de muestra para obtener DNA y los primers fueron específicos para la amplificación de un fragmento de 124 pb del gen de la Dextranasa del S. mutans. De las 24 muestras analizadas, 9 resultaron positivas para la amplificación y en 6 se lograron las secuencias correspondientes. Para la alineación de las secuencias obtenidas, se empleó la base de datos BLAST encontrándose una homología del 95% con el genoma del S. mutans UA159. Este estudio demuestra la primera evidencia de obtención de la secuencia de un fragmento de DNA de Streptococcus mutans recuperados a partir de caries y cálculo dental de restos humanos antiguos, presentando un 95% de homología con el DNA de S. mutans de la subespecie UA159 moderno

The molecular techniques for ancient DNA recovering, offers the possibility to compare the molecular evolution through time as these are tools which make clear possible infectious diseases from the ancient times Objective: To isolate and sequence fossilized Streptococcus mutans DNA fragments, considered the infectious agent involved with dental caries and plaque formation and development, by using the polymerase chain reaction (PCR). Dental caries and tartar samples of teeth collections from Mexico, Barcelona and Balearic Islands. The methodology was adapted to the conservation conditions of this type of DNA samples, and primers were specific to amplify a fragment of 124 bp of S. mutans dextranase gene. Results: 24 samples were analyzed, 9 were positive for amplification and 6 were obtained with its corresponding sequences. To alignment the sequences obtained, we used the BLAST database, giving us the 95% homology with the S. mutans UA159 genome. This study shows us the first evidence of Streptococcus mutans DNA sequence fragment recovered from dental caries and tartar from ancient human remains, presenting a 95% homology with S. mutans UA159 modern subspecies DNA

Paleoparasitologic, paleogenetic and paleobotanic analysis of XVIII century coprolites from the church La Concepción in Santa Cruz de Tenerife, Canary Islands, Spain

We present the results of a paleoparasitologic, paleogenetic and paleobotanic analysis of coprolites recovered during the excavation of the church La Concepción in Santa Cruz de Tenerife. Coprolites (n = 4) were rehydrated and a multidisciplinary analysis was conducted. The paleobotanic analysis showed numerous silicates, seeds and fruits of the family Moraceae. In the paleoparasitologic study, Ascaris sp. eggs (n = 344) were identified. The paleogenetic results confirmed the Ascaris sp. infection as well as the European origin of human remains. These findings contribute to our knowledge of ancient helminthes infections and are the first paleoparasitological record of Ascaris sp. infection in Spain.

Paleoparasitological report on Ascaris aDNA from an ancient East Asian sample

In this study, Ascaris DNA was extracted and sequenced from a medieval archaeological sample in Korea. While Ascaris eggs were confirmed to be of human origin by archaeological evidence, it was not possible to pinpoint the exact species due to close genetic relationships among them. Despite this shortcoming, this is the first Ascaris ancient DNA (aDNA) report from a medieval Asian country and thus will expand the scope of Ascaris aDNA research.

Paleoparasitology of Chagas disease: a review

One hundred years since the discovery of Chagas disease associated with Trypanosoma cruzi infection, growing attention has focused on understanding the evolution in parasite-human host interaction. This interest has featured studies and results from paleoparasitology, not only the description of lesions in mummified bodies, but also the recovery of genetic material from the parasite and the possibility of analyzing such material over time. The present study reviews the evidence of Chagas disease in organic remains excavated from archeological sites and discusses two findings in greater detail, both with lesions suggestive of chagasic megacolon and confirmed by molecular biology techniques. One of these sites is located in the United States, on the border between Texas and Mexico and the other in state of Minas Gerais, in the Brazilian cerrado (savannah). Dated prior to contact with Europeans, these results confirm that Chagas disease affected prehistoric human groups in other regions outside the Andean altiplanos and other transmission areas on the Pacific Coast, previously considered the origin of T. cruzi infection in the human host.

A Practical and Efficient Method for The Retrieval of Ancient DNA Sequence / 生物化学与生物物理进展

Retrieval of ancient DNA (aDNA) sequences from organism remains provide direct view of their evolutionary history. However, researches on aDNA have suffered from lots of technical problems. Specifically, discredited sequences were generated from damaged aDNA templates, and expensive and time-consuming methods were employed. Here, a method which could recover the endogenous aDNA as well as to reduce the cost and research period is described. This is achieved by improving the ancient DNA extraction method of isopropanol precipitation, and reevaluating the method of PCR after N-glycosylase (UNG) treatment, which could remove the damaged DNA from the aDNA extract. The efficiency of these methods were tested by comparing with traditional methods using ancient specimens of pig teeth aged between 4 300 years before present (BP) and 3 900 BP. The results showed that: firstly, the extraction efficiency of the improved method of isopropanol precipitation and current method with silica-based spin column were all 60%. Furthermore, the research period at least could be reduced by half with the application of the improved methods and the cost to 1/10 of the current method. Secondly, sequences obtained through the method of PCR after UNG treatment were 100% authentic. In contrast, 66%～ 88% sequences were authentic based on the results obtained with the method of multiple PCRs without UNG treatment. And the research cost and period needed by the method with UNG treatment were only half of the later one. These results demonstrate that the improved extraction method of isopropanol precipitation combined with the method of PCR after UNG treatment could increase the success rate of authentic DNA amplified and at least reduce the research cost and period by half. Therefore, this method can be applied in the large-scale detection of ancient specimens.

Experience on Sequencing of Mitochondrial DNA from 1200 Year Old Bone Using Cloning / 대한법의학회지

Ancient bones have undergone natural decomposition and have been exposed to external environment for long period. Ancient DNA from old bone is usually fragmented. In addition, various kinds of inhibitors are co-extracted. All these may inhibit proper sequencing reaction. Cloning is regarded as the standard method when sequencing aDNA. When cloning, each clone from the same sample may not be of same sequence, and to exact consensus sequence may be difficult. Here we present our experience on 1200 year old bone from Russia, Primorsky Kray area. We have tried to sequence for HV I, II region of mtDNA using modified mini-primer set, which consisted of 7 set to cover the HV I, II. We cloned the PCR product and sequenced all the clones. Amplification efficiency and subsequent success rates were different for each mini primer set. Loci of variation that differ from consensus sequences were rather frequent, and the pattern were variable depending on sample. Except major polymorphic sites that are important when haplogroup designation, 16129 was the most frequent site that was discarded when extracting haplogroup designation.

Pre-Columbian Chagas disease in Brazil: Trypanosoma cruzi I in the archaeological remains of a human in Peruaçu Valley, Minas Gerais, Brazil

We evaluated the presence and distribution of Trypanosoma cruzi DNA in a mummy presenting with megacolon that was dated as approximately 560 ± 40 years old. The mummy was from the Peruaçu Valley in the state of Minas Gerais, Brazil. All samples were positive for T. cruzi minicircle DNA, demonstrating the presence and broad dissemination of the parasite in this body. From one sample, a mini-exon gene fragment was recovered and characterized by sequencing and was found to belong to the T. cruzi I genotype. This finding suggests that T. cruzi I infected humans during the pre-Columbian times and that, in addition to T. cruzi infection, Chagas disease in Brazil most likely preceded European colonization.