The Neanderthal in the karst: First dating, morphometric, and paleogenetic data on the fossil skeleton from Altamura (Italy).

In 1993, a fossil hominin skeleton was discovered in the karst caves of Lamalunga, near Altamura, in southern Italy. Despite the fact that this specimen represents one of the most extraordinary hominin specimens ever found in Europe, for the last two decades our knowledge of it has been based purely on the documented on-site observations. Recently, the retrieval from the cave of a fragment of bone (part of the right scapula) allowed the first dating of the individual, the quantitative analysis of a diagnostic morphological feature, and a preliminary paleogenetic characterization of this hominin skeleton from Altamura. Overall, the results concur in indicating that it belongs to the hypodigm of Homo neanderthalensis, with some phenetic peculiarities that appear consistent with a chronology ranging from 172 ± 15 ka to 130.1 ± 1.9 ka. Thus, the skeleton from Altamura represents the most ancient Neanderthal from which endogenous DNA has ever been extracted.

Genetic evidence for patrilocal mating behavior among Neandertal groups.

The remains of 12 Neandertal individuals have been found at the El Sidrón site (Asturias, Spain), consisting of six adults, three adolescents, two juveniles, and one infant. Archaeological, paleontological, and geological evidence indicates that these individuals represent all or part of a contemporaneous social group of Neandertals, who died at around the same time and later were buried together as a result of a collapse of an underground karst. We sequenced phylogenetically informative positions of mtDNA hypervariable regions 1 and 2 from each of the remains. Our results show that the 12 individuals stem from three different maternal lineages, accounting for seven, four, and one individual(s), respectively. Using a Y-chromosome assay to confirm the morphological determination of sex for each individual, we found that, although the three adult males carried the same mtDNA lineage, each of the three adult females carried different mtDNA lineages. These findings provide evidence to indicate that Neandertal groups not only were small and characterized by low genetic diversity but also were likely to have practiced patrilocal mating behavior.

Ancient DNA studies: new perspectives on old samples.

In spite of past controversies, the field of ancient DNA is now a reliable research area due to recent methodological improvements. A series of recent large-scale studies have revealed the true potential of ancient DNA samples to study the processes of evolution and to test models and assumptions commonly used to reconstruct patterns of evolution and to analyze population genetics and palaeoecological changes. Recent advances in DNA technologies, such as next-generation sequencing make it possible to recover DNA information from archaeological and paleontological remains allowing us to go back in time and study the genetic relationships between extinct organisms and their contemporary relatives. With the next-generation sequencing methodologies, DNA sequences can be retrieved even from samples (for example human remains) for which the technical pitfalls of classical methodologies required stringent criteria to guaranty the reliability of the results. In this paper, we review the methodologies applied to ancient DNA analysis and the perspectives that next-generation sequencing applications provide in this field.

Sequences from first settlers reveal rapid evolution in Icelandic mtDNA pool.

A major task in human genetics is to understand the nature of the evolutionary processes that have shaped the gene pools of contemporary populations. Ancient DNA studies have great potential to shed light on the evolution of populations because they provide the opportunity to sample from the same population at different points in time. Here, we show that a sample of mitochondrial DNA (mtDNA) control region sequences from 68 early medieval Icelandic skeletal remains is more closely related to sequences from contemporary inhabitants of Scotland, Ireland, and Scandinavia than to those from the modern Icelandic population. Due to a faster rate of genetic drift in the Icelandic mtDNA pool during the last 1,100 years, the sequences carried by the first settlers were better preserved in their ancestral gene pools than among their descendants in Iceland. These results demonstrate the inferential power gained in ancient DNA studies through the application of population genetics analyses to relatively large samples.

Long-acting Injectable Antipsychotics during the COVID-19 pandemic in schizophrenia: An observational study in a real-world clinical setting.

The COVID-19 pandemic is having an important impact on the practice of mental health services and on schizophrenia patients, and heterogeneous and conflicting findings are being reported on the reduction of long-acting injectable (LAI) antipsychotics use. Aims of the study were to assess the total number of patients treated with LAI, the start of novel LAI and the discontinuation of LAI treatments, analyzing register data of the first year of the pandemic, 2020, compared to a pre-pandemic reference year, 2019. Data from two outpatient centers were retrieved, for a total of 236 participants in 2020: no significant differences were observed comparing 2020 and 2019 when considering the total number of patients on LAI treatment (p = 0.890) and the number of dropouts (p = 0.262); however, a significant reduction in the start of LAI was observed (p = 0.022). In 2020, second generation LAI were more prescribed than first generation LAI (p = 0.040) while no difference was observed in 2019 (p = 0.191). These findings attest the efficacy of measures adopted in mental health services to face the consequences of COVID-19 and shed further light on the impact of the pandemic on the clinical practice of mental health services and on the continuity of care of people with schizophrenia.

Effects of bilateral, bipolar-nonbalanced, frontal transcranial Direct Current Stimulation (tDCS) on negative symptoms and neurocognition in a sample of patients living with schizophrenia: Results of a randomized double-blind sham-controlled trial.

Negative symptoms (NS), conceived as Avolition-Apathy (AA) and Expressive Deficit (EXP) domains, and neurocognitive impairments represent unmet therapeutic needs for patients with schizophrenia. The present study investigated if bilateral bipolar-nonbalanced frontal transcranial Direct Current Stimulation (tDCS) could improve these psychopathological dimensions. This randomized, double-blind, sham-controlled study (active-tDCS versus sham-tDCS, both, n = 25) included 50 outpatients diagnosed with schizophrenia clinically stabilized. Patients received 20-min 2 mA active-tDCS or sham-tDCS (anode: left Dorsolateral Prefrontal Cortex; cathode: right orbitofrontal region). Primary outcomes included: PANSS-Negative subscale, Negative Factor (Neg-PANSS), AA and EXP domains; neurocognitive performance at Brief Assessment of Cognition in Schizophrenia. Secondary outcomes included: PANSS subscales and total score, Disorganized/Concrete (DiscC-PANSS) and Positive Factors, Clinical Global Impression (CGI) scores, clinical insight at Scale to Assess Unawareness of Mental Disorder (SUMD). Analysis of covariance (ANCOVA) was performed evaluating between-group changes over time. Significant improvements following active-tDCS were observed for all NS measures (all, p < 0.001; d > 0.8) and for working memory (p = 0.025, d = 0.31). Greater variations following to active treatment emerged also for PANSS-General Psychopathology subscale (p < 0.001; d = 0.54), PANSS total score (p < 0.001; d = 0.69), CGI indexes (all, p < 0.001; d > 0.6), DiscC-PANSS (p < 0.001; d = 0.80) and SUMD-general Unawareness index (p = 0.005; d = 0.15) but not for positive symptoms and others insight measures. Good safety/tolerability profiles were found. Bilateral bipolar-nonbalanced frontal-tDCS is a non-pharmacological approach in schizophrenia effectively improving NS, particularly the AA and EXP domains, probably acting by modulating dysfunctional cortical-subcortical networks. Preliminary results also suggest working memory improvements following tDCS. Further studies are needed to confirm the neurobiological basis of these results.

Bitter taste perception in Neanderthals through the analysis of the TAS2R38 gene.

The bitter taste perception (associated with the ability or inability to taste phenylthiocarbamide) is mediated by the TAS2R38 gene. Most of the variation in this gene is explained by three common amino-acid polymorphisms at positions 49 (encoding proline or alanine), 262 (alanine or valine) and 296 (valine or isoleucine) that determine two common isoforms: proline-alanine-valine (PAV) and alanine-valine-isoleucine (AVI). PAV is the major taster haplotype (heterozygote and homozygote) and AVI is the major non-taster haplotype (homozygote). Amino acid 49 has the major effect on the distinction between tasters and non-tasters of all three variants. The sense of bitter taste protects us from ingesting toxic substances, present in some vegetables, that can affect the thyroid when ingested in large quantities. Balancing selection has been used to explain the current high non-taster frequency, by maintaining divergent TAS2R38 alleles in humans. We have amplified and sequenced the TAS2R38 amino acid 49 in the virtually uncontaminated Neanderthal sample of El Sidrón 1253 and have determined that it was heterozygous. Thus, this Neanderthal was a taster individual, although probably slightly less than a PAV homozygote. This indicates that variation in bitter taste perception pre-dates the divergence of the lineages leading to Neanderthals and modern humans.

Genetic characterization of the ABO blood group in Neandertals.

BACKGROUND: The high polymorphism rate in the human ABO blood group gene seems to be related to susceptibility to different pathogens. It has been estimated that all genetic variation underlying the human ABO alleles appeared along the human lineage, after the divergence from the chimpanzee lineage. A paleogenetic analysis of the ABO blood group gene in Neandertals allows us to directly test for the presence of the ABO alleles in these extinct humans. RESULTS: We have analysed two male Neandertals that were retrieved under controlled conditions at the El Sidron site in Asturias (Spain) and that appeared to be almost free of modern human DNA contamination. We find a human specific diagnostic deletion for blood group O (O01 haplotype) in both Neandertal individuals. CONCLUSION: These results suggest that the genetic change responsible for the O blood group in humans predates the human and Neandertal divergence. A potential selective event associated with the emergence of the O allele may have therefore occurred after humans separated from their common ancestor with chimpanzees and before the human-Neandertal population divergence.

Pleistocene Mitochondrial Genomes Suggest a Single Major Dispersal of Non-Africans and a Late Glacial Population Turnover in Europe.

How modern humans dispersed into Eurasia and Australasia, including the number of separate expansions and their timings, is highly debated [1, 2]. Two categories of models are proposed for the dispersal of non-Africans: (1) single dispersal, i.e., a single major diffusion of modern humans across Eurasia and Australasia [3-5]; and (2) multiple dispersal, i.e., additional earlier population expansions that may have contributed to the genetic diversity of some present-day humans outside of Africa [6-9]. Many variants of these models focus largely on Asia and Australasia, neglecting human dispersal into Europe, thus explaining only a subset of the entire colonization process outside of Africa [3-5, 8, 9]. The genetic diversity of the first modern humans who spread into Europe during the Late Pleistocene and the impact of subsequent climatic events on their demography are largely unknown. Here we analyze 55 complete human mitochondrial genomes (mtDNAs) of hunter-gatherers spanning ∼35,000 years of European prehistory. We unexpectedly find mtDNA lineage M in individuals prior to the Last Glacial Maximum (LGM). This lineage is absent in contemporary Europeans, although it is found at high frequency in modern Asians, Australasians, and Native Americans. Dating the most recent common ancestor of each of the modern non-African mtDNA clades reveals their single, late, and rapid dispersal less than 55,000 years ago. Demographic modeling not only indicates an LGM genetic bottleneck, but also provides surprising evidence of a major population turnover in Europe around 14,500 years ago during the Late Glacial, a period of climatic instability at the end of the Pleistocene.

Mitochondrial DNA from El Mirador cave (Atapuerca, Spain) reveals the heterogeneity of Chalcolithic populations.

Previous mitochondrial DNA analyses on ancient European remains have suggested that the current distribution of haplogroup H was modeled by the expansion of the Bell Beaker culture (ca 4,500-4,050 years BP) out of Iberia during the Chalcolithic period. However, little is known on the genetic composition of contemporaneous Iberian populations that do not carry the archaeological tool kit defining this culture. Here we have retrieved mitochondrial DNA (mtDNA) sequences from 19 individuals from a Chalcolithic sample from El Mirador cave in Spain, dated to 4,760-4,200 years BP and we have analyzed the haplogroup composition in the context of modern and ancient populations. Regarding extant African, Asian and European populations, El Mirador shows affinities with Near Eastern groups. In different analyses with other ancient samples, El Mirador clusters with Middle and Late Neolithic populations from Germany, belonging to the Rössen, the Salzmünde and the Baalberge archaeological cultures but not with contemporaneous Bell Beakers. Our analyses support the existence of a common genetic signal between Western and Central Europe during the Middle and Late Neolithic and points to a heterogeneous genetic landscape among Chalcolithic groups.

Tracing the origin of the east-west population admixture in the Altai region (Central Asia).

A recent discovery of Iron Age burials (Pazyryk culture) in the Altai Mountains of Mongolia may shed light on the mode and tempo of the generation of the current genetic east-west population admixture in Central Asia. Studies on ancient mitochondrial DNA of this region suggest that the Altai Mountains played the role of a geographical barrier between West and East Eurasian lineages until the beginning of the Iron Age. After the 7th century BC, coinciding with Scythian expansion across the Eurasian steppes, a gradual influx of East Eurasian sequences in Western steppes is detected. However, the underlying events behind the genetic admixture in Altai during the Iron Age are still unresolved: 1) whether it was a result of migratory events (eastward firstly, westward secondly), or 2) whether it was a result of a local demographic expansion in a 'contact zone' between European and East Asian people. In the present work, we analyzed the mitochondrial DNA lineages in human remains from Bronze and Iron Age burials of Mongolian Altai. Here we present support to the hypothesis that the gene pool of Iron Age inhabitants of Mongolian Altai was similar to that of western Iron Age Altaians (Russia and Kazakhstan). Thus, this people not only shared the same culture (Pazyryk), but also shared the same genetic east-west population admixture. In turn, Pazyryks appear to have a similar gene pool that current Altaians. Our results further show that Iron Age Altaians displayed mitochondrial lineages already present around Altai region before the Iron Age. This would provide support for a demographic expansion of local people of Altai instead of westward or eastward migratory events, as the demographic event behind the high population genetic admixture and diversity in Central Asia.

Genetic analysis of the presumptive blood from Louis XVI, King of France.

A text on a pyrographically decorated gourd dated to 1793 explains that it contains a handkerchief dipped with the blood of Louis XVI, king of France, after his execution. Biochemical analyses confirmed that the material contained within the gourd was blood. The mitochondrial DNA (mtDNA) hypervariable region 1 (HVR1) and 2 (HVR2), the Y-chromosome STR profile, some autosomal STR markers and a SNP in HERC2 gene associated to blue eyes, were retrieved, and some results independently replicated in two different laboratories. The uncommon mtDNA sequence retrieved can be attributed to a N1b haplotype, while the novel Y-chromosome haplotype belongs to haplogroup G2a. The HERC2 gene showed that the subject analyzed was a heterozygote, which is compatible with a blue-eyed person, as king Louis XVI was. To confirm the identity of the subject, an analysis of the dried heart of his son, Louis XVII, could be undertaken.

Fragmentation of contaminant and endogenous DNA in ancient samples determined by shotgun sequencing; prospects for human palaeogenomics.

BACKGROUND: Despite the successful retrieval of genomes from past remains, the prospects for human palaeogenomics remain unclear because of the difficulty of distinguishing contaminant from endogenous DNA sequences. Previous sequence data generated on high-throughput sequencing platforms indicate that fragmentation of ancient DNA sequences is a characteristic trait primarily arising due to depurination processes that create abasic sites leading to DNA breaks. METHODOLOGY/PRINCIPALS FINDINGS: To investigate whether this pattern is present in ancient remains from a temperate environment, we have 454-FLX pyrosequenced different samples dated between 5,500 and 49,000 years ago: a bone from an extinct goat (Myotragus balearicus) that was treated with a depurinating agent (bleach), an Iberian lynx bone not subjected to any treatment, a human Neolithic sample from Barcelona (Spain), and a Neandertal sample from the El Sidrón site (Asturias, Spain). The efficiency of retrieval of endogenous sequences is below 1% in all cases. We have used the non-human samples to identify human sequences (0.35 and 1.4%, respectively), that we positively know are contaminants. CONCLUSIONS: We observed that bleach treatment appears to create a depurination-associated fragmentation pattern in resulting contaminant sequences that is indistinguishable from previously described endogenous sequences. Furthermore, the nucleotide composition pattern observed in 5' and 3' ends of contaminant sequences is much more complex than the flat pattern previously described in some Neandertal contaminants. Although much research on samples with known contaminant histories is needed, our results suggest that endogenous and contaminant sequences cannot be distinguished by the fragmentation pattern alone.

Paleogenomics in a temperate environment: shotgun sequencing from an extinct Mediterranean caprine.

BACKGROUND: Numerous endemic mammals, including dwarf elephants, goats, hippos and deers, evolved in isolation in the Mediterranean islands during the Pliocene and Pleistocene. Most of them subsequently became extinct during the Holocene. Recently developed high-throughput sequencing technologies could provide a unique tool for retrieving genomic data from these extinct species, making it possible to study their evolutionary history and the genetic bases underlying their particular, sometimes unique, adaptations. METHODOLOGY/PRINCIPALS FINDINGS: A DNA extraction of a approximately 6,000 year-old bone sample from an extinct caprine (Myotragus balearicus) from the Balearic Islands in the Western Mediterranean, has been subjected to shotgun sequencing with the GS FLX 454 platform. Only 0.27% of the resulting sequences, identified from alignments with the cow genome and comprising 15,832 nucleotides, with an average length of 60 nucleotides, proved to be endogenous. CONCLUSIONS: A phylogenetic tree generated with Myotragus sequences and those from other artiodactyls displays an identical topology to that generated from mitochondrial DNA data. Despite being in an unfavourable thermal environment, which explains the low yield of endogenous sequences, our study demonstrates that it is possible to obtain genomic data from extinct species from temperate regions.

Statistical analysis of post mortem DNA damage-derived miscoding lesions in Neandertal mitochondrial DNA.

BACKGROUND: We have analysed the distribution of post mortem DNA damage derived miscoding lesions from the datasets of seven published Neandertal specimens that have extensive cloned sequence coverage over the mitochondrial DNA (mtDNA) hypervariable region 1 (HVS1). The analysis was restricted to C-->T and G-->A miscoding lesions (the predominant manifestation of post mortem damage) that are seen at a frequency of more than one clone among sequences from a single PCR, but do not represent the true endogenous sequence. FINDINGS: The data indicates an extreme bias towards C-->T over G-->A miscoding lesions (observed ratio of 67:2 compared to an expected ratio of 7:2), implying that the mtDNA Light strand molecule suffers proportionally more damage-derived miscoding lesions than the Heavy strand. CONCLUSION: The clustering of Cs in the Light strand as opposed to the singleton pattern of Cs in the Heavy strand could explain the observed bias, a phenomenon that could be further tested with non-PCR based approaches. The characterization of the HVS1 hotspots will be of use to future Neandertal mtDNA studies, with specific regards to assessing the authenticity of new positions previously unknown to be polymorphic.

Genetic analysis of the skeletal remains attributed to Francesco Petrarca.

We report on the mitochondrial DNA (mtDNA) analysis of the supposed remains of Francesco Petrarca exhumed in November 2003, from the S. Maria Assunta church, in Arquà Padua (Italy) where he died in 1374. The optimal preservation of the remains allowed the retrieval of sufficient mtDNA for genetic analysis. DNA was extracted from a rib and a tooth and mtDNA sequences were determined in multiple clones using the strictest criteria currently available for validation of ancient DNA sequences, including independent replication. MtDNA sequences from the tooth and rib were not identical, suggesting that they belonged to different individuals. Indeed, molecular gender determination showed that the postcranial remains belonged to a male while the skull belonged to a female. Historical records indicated that the remains were violated in 1630, possibly by thieves. These results are consistent with morphological investigations and confirm the importance of integrating molecular and morphological approaches in investigating historical remains.