Bioarchaeological study of ancient Teotihuacans based on complete mitochondrial genome sequences and diet isotopes.

BACKGROUND: The Teotihuacan civilisation was the largest one in ancient Mesoamerica. The Teotihuacan city was born in the north-eastern Basin of Mexico around the second century BC, reached its peak in the fourth century AD, and had cultural influence throughout Mesoamerica. At its peak, the size of the city reached more than 20 km2, and the total population is estimated to have increased from 100,000 to 200,000. However, knowledge of the genetic background of the Teotihuacan people is still limited. AIM: We aimed to determine the mitogenome sequences of the Teotihuacan human remains and compare the ancient and present Mesoamericans. In addition, we aimed to identify the food habits of ancient Teotihuacans. SUBJECTS AND METHODS: We determined the mitogenome sequences of human remains dated to 250-636 cal AD using target enrichment-coupled next generation sequencing. We also performed stable isotope analysis. RESULTS: We successfully obtained nearly full-length sequences newly unearthed from a civilian dwelling in the Teotihuacan site. Teotihuacan mitochondrial DNA was classified into the haplogroups in present and ancient Mesoamericans. In addition, Teotihuacan individuals had a diet dependent on C4 plants such as maize. CONCLUSION: Genetic diversity varied among the Teotihuacans.

Diversity in matrilineages among the Jomon individuals of Japan.

BACKGROUND: The Jomon period of Japan is characterised by a unique combination of sedentary and hunting/gathering lifestyles, spanning for more than 10,000 years from the final Pleistocene to the Holocene. The transition from the preceding Palaeolithic period to the Jomon period is known to have begun with the appearance of pottery usage. However, knowledge of the genetic background of the Jomon people is still limited. AIM: We aimed to determine the population-scale complete mitogenome sequences of the Initial Jomon human remains and compare the occurrence of mitochondrial haplogroups in the Jomon period from temporal and regional perspectives. SUBJECTS AND METHODS: For human remains dated to 8200-8600 cal BP, we determined their complete mitogenome sequences using target enrichment-coupled next-generation sequencing. RESULTS: We successfully obtained the complete mitogenome sequences with high depth of coverage and high concordance on consensus sequences. These sequences differed by more than three bases each, except for two individuals having completely identical sequences. Co-existence of individuals with haplogroups N9b and M7a was first observed at the same archaeological site from the Initial Jomon period. CONCLUSION: The genetic diversity within the population was not found to be low even in the Initial Jomon period.

Corrigendum: Comparison of Periodontal Bacteria of Edo and Modern Periods Using Novel Diagnostic Approach for Periodontitis With Micro-CT.

[This corrects the article DOI: 10.3389/fcimb.2021.723821.].

Deficient maternal behavior in multiparous Pou3f2⊿ mice is associated with an impaired exploratory activity.

Mammalian adult females develop specialized body parts, namely mammary glands and uterus, and exhibit specialized maternal behavior, lactation/nursing and care for their offspring. As the brain plays an essential role in regulating related physiological functions in the body, the morphology or function of the mammalian brain has been modified to manage newly equipped structures and functions. However, this evolutionary process is largely unknown. Pou3f2/Brn2 is an evolutionarily remarkable gene as it contains mammal-specific base sequences encoding three stretches of homopolymeric amino acids (polyAAs): poly-glycine (polyG), poly-glutamine (polyQ), and poly-proline (polyP). Previously, we demonstrated that POU3F2 acquisition of mammal-specific polyAAs contributed to the establishment of behaviors characteristic of mammals. Here, we demonstrated that Pou3f2⊿ mice displayed basic features required for maternal care. However, Pou3f2⊿ mice exhibited deficits in the reproductive performance and maternal behavior, which were not fully improved by multiparas. Therefore, we extensively investigated pup retrieval behavior and discovered that the retrieval and the exploratory behaviors were impaired in Pou3f2⊿ female mice, but not in males. Altogether, our data suggest that POU3F2 acquisition of mammal-specific polyAAs contributes to the continuous awareness and curiosity needed for maternal interaction.

Comparison of Periodontal Bacteria of Edo and Modern Periods Using Novel Diagnostic Approach for Periodontitis With Micro-CT.

Ancient dental calculus, formed from dental plaque, is a rich source of ancient DNA and can provide information regarding the food and oral microbiology at that time. Genomic analysis of dental calculus from Neanderthals has revealed the difference in bacterial composition of oral microbiome between Neanderthals and modern humans. There are few reports investigating whether the pathogenic bacteria of periodontitis, a polymicrobial disease induced in response to the accumulation of dental plaque, were different between ancient and modern humans. This study aimed to compare the bacterial composition of the oral microbiome in ancient and modern human samples and to investigate whether lifestyle differences depending on the era have altered the bacterial composition of the oral microbiome and the causative bacteria of periodontitis. Additionally, we introduce a novel diagnostic approach for periodontitis in ancient skeletons using micro-computed tomography. Ancient 16S rDNA sequences were obtained from 12 samples at the Unko-in site (18th-19th century) of the Edo era (1603-1867), a characteristic period in Japan when immigrants were not accepted. Furthermore, modern 16S rDNA data from 53 samples were obtained from a database to compare the modern and ancient microbiome. The microbial co-occurrence network was analyzed based on 16S rDNA read abundance. Eubacterium species, Mollicutes species, and Treponema socranskii were the core species in the Edo co-occurrence network. The co-occurrence relationship between Actinomyces oricola and Eggerthella lenta appeared to have played a key role in causing periodontitis in the Edo era. However, Porphyromonas gingivalis, Fusobacterium nucleatum subsp. vincentii, and Prevotella pleuritidis were the core and highly abundant species in the co-occurrence network of modern samples. These results suggest the possibility of differences in the pathogens causing periodontitis during different eras in history.

Population dynamics in the Japanese Archipelago since the Pleistocene revealed by the complete mitochondrial genome sequences.

The Japanese Archipelago is widely covered with acidic soil made of volcanic ash, an environment which is detrimental to the preservation of ancient biomolecules. More than 10,000 Palaeolithic and Neolithic sites have been discovered nationwide, but few skeletal remains exist and preservation of DNA is poor. Despite these challenging circumstances, we succeeded in obtaining a complete mitogenome (mitochondrial genome) sequence from Palaeolithic human remains. We also obtained those of Neolithic (the hunting-gathering Jomon and the farming Yayoi cultures) remains, and over 2,000 present-day Japanese. The Palaeolithic mitogenome sequence was not found to be a direct ancestor of any of Jomon, Yayoi, and present-day Japanese people. However, it was an ancestral type of haplogroup M, a basal group of the haplogroup M. Therefore, our results indicate continuity in the maternal gene pool from the Palaeolithic to present-day Japanese. We also found that a vast increase of population size happened and has continued since the Yayoi period, characterized with paddy rice farming. It means that the cultural transition, i.e. rice agriculture, had significant impact on the demographic history of Japanese population.

The number of SNPs required for distinguishing Japanese from other East Asians.

In some cases, it is necessary to estimate the national origin of an unknown subject in forensic medicine. The use of single nucleotide polymorphism (SNP) markers appears to be very effective for this purpose, since genome-wide SNP genotype data of many human populations are publicly available. In this study, we examined the number of SNPs that could objectively and accurately distinguish Japanese subjects (1KG-JPT) from the other East Asians (1KG-CDX, -CHB, -CHS, and -KHV) using the combination of principal component analysis and hierarchical cluster analysis. A computer simulation showed that approximately 3000 randomly selected SNPs were enough for the discrimination. Our results suggest that at least a 0.024% coverage is needed in the next generation sequencing experiment to objectively determine whether an unknown person is Japanese or not if the amount of DNA sample from him/her is insufficient or the quality is low.

A biomolecular anthropological investigation of William Adams, the first SAMURAI from England.

William Adams (Miura Anjin) was an English navigator who sailed with a Dutch trading fleet to the far East and landed in Japan in 1600. He became a vassal under the Shogun, Tokugawa Ieyasu, was bestowed with a title, lands and swords, and became the first SAMURAI from England. "Miura" comes from the name of the territory given to him and "Anjin" means "pilot". He lived out the rest of his life in Japan and died in Hirado, Nagasaki Prefecture, in 1620, where he was reportedly laid to rest. Shortly after his death, graveyards designated for foreigners were destroyed during a period of Christian repression, but Miura Anjin's bones were supposedly taken, protected, and reburied. Archaeological investigations in 1931 uncovered human skeletal remains and it was proposed that they were those of Miura Anjin. However, this could not be confirmed from the evidence at the time and the remains were reburied. In 2017, excavations found skeletal remains matching the description of those reinterred in 1931. We analyzed these remains from various aspects, including genetic background, dietary habits, and burial style, utilizing modern scientific techniques to investigate whether they do indeed belong to the first English SAMURAI.

A study of 8,300-year-old Jomon human remains in Japan using complete mitogenome sequences obtained by next-generation sequencing.

Ancient human remains have been assigned to their mitochondrial DNA (mtDNA) haplogroups. To obtain efficiently deep and reliable nucleotide sequences of ancient DNA of interest, we achieved target enrichment followed by next-generation sequencing (NGS). Complete mitochondrial genome (mitogenome) sequences were obtained for three human remains from the Iyai rock-shelter site of the Initial Jomon Period in Japan. All the Jomon mitogenomes belong to haplogroup N9b, but no sequences among them were identical. High genetic diversity was clarified even among the Jomon human remains belonging to haplogroup N9b, which has been described as a haplogroup representing the Jomon people.

Ancient DNA analysis of food remains in human dental calculus from the Edo period, Japan.

Although there are many methods for reconstructing diets of the past, detailed taxon identification is still challenging, and most plants hardly remain at a site. In this study, we applied DNA metabarcoding to dental calculus of premodern Japan for the taxonomic identification of food items. DNA was extracted from 13 human dental calculi from the Unko-in site (18th-19th century) of the Edo period, Japan. Polymerase chain reaction (PCR) and sequencing were performed using a primer set specific to the genus Oryza because rice (Oryza sativa) was a staple food and this was the only member of this genus present in Japan at that time. DNA metabarcoding targeting plants, animals (meat and fish), and fungi were also carried out to investigate dietary diversity. We detected amplified products of the genus Oryza from more than half of the samples using PCR and Sanger sequencing. DNA metabarcoding enabled us to identify taxa of plants and fungi, although taxa of animals were not detected, except human. Most of the plant taxonomic groups (family/genus level) are present in Japan and include candidate species consumed as food at that time, as confirmed by historical literature. The other groups featured in the lifestyle of Edo people, such as for medicinal purposes and tobacco. The results indicate that plant DNA analysis from calculus provides information about food diversity and lifestyle habits from the past and can complement other analytical methods such as microparticle analysis and stable isotope analysis.

Reduced home cage and social activity in Pou3f2⊿ mice.

Pou3f2/Brn2 is a transcription factor that helps to determine the cellular identity of neocortical or hypothalamic neurons. Mammalian Pou3f2 contains three homopolymeric amino acids that are not present in amphibian Pou3f2. These amino acids contribute to monoamine function, which may play specific roles in mammalian development and behavior. Previous work has indicated that Pou3f2⊿ mice, which lack the homopolymeric amino acids, exhibited declined maternal activity and impaired object and spatial recognition. The current study, analyzed weight gain, brain development, home cage activity, social interaction, and response to novel objects in Pou3f2⊿ mice to determine which aspects of behavior were affected by monoamine dysregulation. Compared to their wild type counterparts, Pou3f2⊿ mice showed decreased social interaction and reduced home cage activity during their active phase. However, they showed normal weight gain, brain development, and responses to novelty. These results indicate that monoamine dysregulation in Pou3f2⊿ mice may specifically affect basal activity and social development, without altering non-social motivation.

MitoIMP: A Computational Framework for Imputation of Missing Data in Low-Coverage Human Mitochondrial Genome.

The incompleteness of partial human mitochondrial genome sequences makes it difficult to perform relevant comparisons among multiple resources. To deal with this issue, we propose a computational framework for deducing missing nucleotides in the human mitochondrial genome. We applied it to worldwide mitochondrial haplogroup lineages and assessed its performance. Our approach can deduce the missing nucleotides with a precision of 0.99 or higher in most human mitochondrial DNA lineages. Furthermore, although low-coverage mitochondrial genome sequences often lead to a blurred relationship in the multidimensional scaling analysis, our approach can correct this positional arrangement according to the corresponding mitochondrial DNA lineages. Therefore, our framework will provide a practical solution to compensate for the lack of genome coverage in partial and fragmented human mitochondrial genome sequences. In this study, we developed an open-source computer program, MitoIMP, implementing our imputation procedure. MitoIMP is freely available from https://github.com/omics-tools/mitoimp.

Proteomic profiling of archaeological human bone.

Ancient protein analysis provides clues to human life and diseases from ancient times. Here, we performed shotgun proteomics of human archeological bones for the first time, using rib bones from the Hitotsubashi site (AD 1657-1683) in Tokyo, called Edo in ancient times. The output data obtained were analysed using Gene Ontology and label-free quantification. We detected leucocyte-derived proteins, possibly originating from the bone marrow of the rib. Particularly prevalent and relatively high expression of eosinophil peroxidase suggests the influence of infectious diseases. This scenario is plausible, considering the overcrowding and unhygienic living conditions of the Edo city described in the historical literature. We also observed age-dependent differences in proteome profiles, particularly for proteins involved in developmental processes. Among them, alpha-2-HS-glycoprotein demonstrated a strong negative correlation with age. These results suggest that analysis of ancient proteins could provide a useful indicator of stress, disease, starvation, obesity and other kinds of physiological and pathological information.

Characterization of complete mitochondrial genomes of indigenous Mayans in Mexico.

BACKGROUND: The authors have previously published the complete mitochondrial genome (mitogenome) sequences of two indigenous Mesoamerican populations, Mazahua (n = 25) and Zapotec (n = 88). METHODS: This study determined the complete mitogenome sequences of nine unrelated individuals from the indigenous Maya population living in Mexico. RESULTS: Their mitogenome sequences could be classified into either of the haplogroups A2 and C1. Surprisingly, there were no mitogenome sequences (haplotypes) that the Maya, Mazahua, and Zapotec people share in common. CONCLUSIONS: This indicates that no genetic exchange, at least matrilineally, has occurred among them.

MitoSuite: a graphical tool for human mitochondrial genome profiling in massive parallel sequencing.

Recent rapid advances in high-throughput, next-generation sequencing (NGS) technologies have promoted mitochondrial genome studies in the fields of human evolution, medical genetics, and forensic casework. However, scientists unfamiliar with computer programming often find it difficult to handle the massive volumes of data that are generated by NGS. To address this limitation, we developed MitoSuite, a user-friendly graphical tool for analysis of data from high-throughput sequencing of the human mitochondrial genome. MitoSuite generates a visual report on NGS data with simple mouse operations. Moreover, it analyzes high-coverage sequencing data but runs on a stand-alone computer, without the need for file upload. Therefore, MitoSuite offers outstanding usability for handling massive NGS data, and is ideal for evolutionary, clinical, and forensic studies on the human mitochondrial genome variations. It is freely available for download from the website https://mitosuite.com.

Imputation approach for deducing a complete mitogenome sequence from low-depth-coverage next-generation sequencing data: application to ancient remains from the Moon Pyramid, Mexico.

It is considered that more than 15 depths of coverage are necessary for next-generation sequencing (NGS) data to obtain reliable complete nucleotide sequences of the mitogenome. However, it is difficult to satisfy this requirement for all nucleotide positions because of problems obtaining a uniform depth of coverage for poorly preserved materials. Thus, we propose an imputation approach that allows a complete mitogenome sequence to be deduced from low-depth-coverage NGS data. We used different types of mitogenome data files as panels for imputation: a worldwide panel comprising all the major haplogroups, a worldwide panel comprising sequences belonging to the estimated haplogroup alone, a panel comprising sequences from the population most closely related to an individual under investigation, and a panel comprising sequences belonging to the estimated haplogroup from the population most closely related to an individual under investigation. The number of missing nucleotides was drastically reduced in all the panels, but the contents obtained by imputation were quite different among the panels. The efficiency of the imputation method differed according to the panels used. The missing nucleotides were most credibly imputed using sequences of the estimated haplogroup from the population most closely related to the individual under investigation as a panel.

A partial nuclear genome of the Jomons who lived 3000 years ago in Fukushima, Japan.

The Jomon period of the Japanese Archipelago, characterized by cord-marked 'jomon' potteries, has yielded abundant human skeletal remains. However, the genetic origins of the Jomon people and their relationships with modern populations have not been clarified. We determined a total of 115 million base pair nuclear genome sequences from two Jomon individuals (male and female each) from the Sanganji Shell Mound (dated 3000 years before present) with the Jomon-characteristic mitochondrial DNA haplogroup N9b, and compared these nuclear genome sequences with those of worldwide populations. We found that the Jomon population lineage is best considered to have diverged before diversification of present-day East Eurasian populations, with no evidence of gene flow events between the Jomon and other continental populations. This suggests that the Sanganji Jomon people descended from an early phase of population dispersals in East Asia. We also estimated that the modern mainland Japanese inherited <20% of Jomon peoples' genomes. Our findings, based on the first analysis of Jomon nuclear genome sequence data, firmly demonstrate that the modern mainland Japanese resulted from genetic admixture of the indigenous Jomon people and later migrants.

Rice Varieties in Archaic East Asia: Reduction of Its Diversity from Past to Present Times.

The Asian cultivated rice, Oryza sativa, is one of the most important crops feeding more than a third of global population. In spite of the studies for several decades, the origin and domestication history of rice varietal groups, japonica and indica, have not been fully unveiled. Genetic information of ancient rice remains is essential for direct and exclusive insight into the domestication history of rice. We performed ancient DNA analysis of 950- to 2,800-year-old rice remains excavated from Japan and Korea. We found the presence of both japonica- and indica-type varieties in the Yayoi period and the middle ages of Japan and the middle part of Korea Peninsula 2,000 years ago. It is popularly considered that japonica has been exclusively cultivated in northern part of East Asia including Japan and Korea. Our result disclosed unexpectedly wide diversity of rice varieties in archaic East Asia. The present results from ancient rice DNA reveal an exclusive insight for the domestication history of rice which is not provided as far as contemporary rice.

mtDNA diversity of the Zapotec in Mexico suggests a population decline long before the first contact with Europeans.

The New World is the last continent colonized by anatomically modern humans, Homo sapiens. The first migrants entered the New World from Asia through Beringia. It is suggested that there were three streams of Asian gene flow, one major and two additional minor gene flows. The first major migrants took a Pacific coastal route and began spreading to the American continent before the opening of the ice-free corridor. We investigated the diversity of full-length mitochondrial DNA genomes of the Zapotec population, residing in the Mesoamerican region, and reconstructed their demographic history using Bayesian Skyline Plots. We estimated the initial date of gene flow into the New World by Zapotec ancestors at around 17 000­19 000 years ago,which is highly concordant with previous studies. We also show a population decline after the initial expansion. This decline started 4000 years ago, long before European contact with Native Americans. This indicates that other factors including climatec hange should be considered to explain the observed demographic pattern.

Complete mitogenome analysis of indigenous populations in Mexico: its relevance for the origin of Mesoamericans.

Mesoamerica has an important role in the expansion of Paleoamericans as the route to South America. In this study, we determined complete mitogenome sequences of 113 unrelated individuals from two indigenous populations of Mesoamerica, Mazahua and Zapotec. All newly sequenced mitogenomes could be classified into haplogroups A2, B2, C1 and D1, but one sequence in Mazahua was D4h3a, a subclade of haplogroup D4. This haplogroup has been mostly found in South America along the Pacific coast. Haplogroup X2a was not found in either population. Genetic similarity obtained using phylogenetic tree construction and principal component analysis showed that these two populations are distantly related to each other. Actually, the Mazahua and the Zapotec shared no sequences (haplotypes) in common, while each also showed a number of unique subclades. Surprisingly, Zapotec formed a cluster with indigenous populations living in an area from central Mesoamerica to Central America. By contrast, the Mazahua formed a group with indigenous populations living in external areas, including southwestern North America and South America. This intriguing genetic relationship suggests the presence of two paleo-Mesoamerican groups, invoking a scenario in which one group had expanded into South America and the other resided in Mesoamerica.