Transcriptome innovations in primates revealed by single-molecule long-read sequencing.

Transcriptomic diversity greatly contributes to the fundamentals of disease, lineage-specific biology, and environmental adaptation. However, much of the actual isoform repertoire contributing to shaping primate evolution remains unknown. Here, we combined deep long- and short-read sequencing complemented with mass spectrometry proteomics in a panel of lymphoblastoid cell lines (LCLs) from human, three other great apes, and rhesus macaque, producing the largest full-length isoform catalog in primates to date. Around half of the captured isoforms are not annotated in their reference genomes, significantly expanding the gene models in primates. Furthermore, our comparative analyses unveil hundreds of transcriptomic innovations and isoform usage changes related to immune function and immunological disorders. The confluence of these evolutionary innovations with signals of positive selection and their limited impact in the proteome points to changes in alternative splicing in genes involved in immune response as an important target of recent regulatory divergence in primates.

Local genetic adaptation to habitat in wild chimpanzees.

How populations adapt to their environment is a fundamental question in biology. Yet we know surprisingly little about this process, especially for endangered species such as non-human great apes. Chimpanzees, our closest living relatives, are particularly interesting because they inhabit diverse habitats, from rainforest to woodland-savannah. Whether genetic adaptation facilitates such habitat diversity remains unknown, despite having wide implications for evolutionary biology and conservation. Using 828 newly generated exomes from wild chimpanzees, we find evidence of fine-scale genetic adaptation to habitat. Notably, adaptation to malaria in forest chimpanzees is mediated by the same genes underlying adaptation to malaria in humans. This work demonstrates the power of non-invasive samples to reveal genetic adaptations in endangered populations and highlights the importance of adaptive genetic diversity for chimpanzees.

Past Connectivity but Recent Inbreeding in Cross River Gorillas Determined Using Whole Genomes from Single Hairs.

The critically endangered western gorillas (Gorilla gorilla) are divided into two subspecies: the western lowland (G. g. gorilla) and the Cross River (G. g. diehli) gorilla. Given the difficulty in sampling wild great ape populations and the small estimated size of the Cross River gorilla population, only one whole genome of a Cross River gorilla has been sequenced to date, hindering the study of this subspecies at the population level. In this study, we expand the number of whole genomes available for wild western gorillas, generating 41 new genomes (25 belonging to Cross River gorillas) using single shed hairs collected from gorilla nests. By combining these genomes with publicly available wild gorilla genomes, we confirm that Cross River gorillas form three population clusters. We also found little variation in genome-wide heterozygosity among them. Our analyses reveal long runs of homozygosity (>10 Mb), indicating recent inbreeding in Cross River gorillas. This is similar to that seen in mountain gorillas but with a much more recent bottleneck. We also detect past gene flow between two Cross River sites, Afi Mountain Wildlife Sanctuary and the Mbe Mountains. Furthermore, we observe past allele sharing between Cross River gorillas and the northern western lowland gorilla sites, as well as with the eastern gorilla species. This is the first study using single shed hairs from a wild species for whole genome sequencing to date. Taken together, our results highlight the importance of implementing conservation measures to increase connectivity among Cross River gorilla sites.

Ghost admixture in eastern gorillas.

Archaic admixture has had a substantial impact on human evolution with multiple events across different clades, including from extinct hominins such as Neanderthals and Denisovans into modern humans. In great apes, archaic admixture has been identified in chimpanzees and bonobos but the possibility of such events has not been explored in other species. Here, we address this question using high-coverage whole-genome sequences from all four extant gorilla subspecies, including six newly sequenced eastern gorillas from previously unsampled geographic regions. Using approximate Bayesian computation with neural networks to model the demographic history of gorillas, we find a signature of admixture from an archaic 'ghost' lineage into the common ancestor of eastern gorillas but not western gorillas. We infer that up to 3% of the genome of these individuals is introgressed from an archaic lineage that diverged more than 3 million years ago from the common ancestor of all extant gorillas. This introgression event took place before the split of mountain and eastern lowland gorillas, probably more than 40 thousand years ago and may have influenced perception of bitter taste in eastern gorillas. When comparing the introgression landscapes of gorillas, humans and bonobos, we find a consistent depletion of introgressed fragments on the X chromosome across these species. However, depletion in protein-coding content is not detectable in eastern gorillas, possibly as a consequence of stronger genetic drift in this species.

Insights from the rescue and breeding management of Cuvier's gazelle (Gazella cuvieri) through whole-genome sequencing.

Captive breeding programmes represent the most intensive type of ex situ population management for threatened species. One example is the Cuvier's gazelle programme that started in 1975 with only four founding individuals, and after more than four decades of management in captivity, a reintroduction effort was undertaken in Tunisia in 2016, to establish a population in an area historically included within its range. Here, we aim to determine the genetic consequences of this reintroduction event by assessing the genetic diversity of the founder stock as well as of their descendants. We present the first whole-genome sequencing dataset of 30 Cuvier's gazelles including captive-bred animals, animals born in Tunisia after a reintroduction and individuals from a genetically unrelated Moroccan population. Our analyses revealed no difference between the founder and the offspring cohorts in genome-wide heterozygosity and inbreeding levels, and in the amount and length of runs of homozygosity. The captive but unmanaged Moroccan gazelles have the lowest genetic diversity of all genomes analysed. Our findings demonstrate that the Cuvier's gazelle captive breeding programme can serve as source populations for future reintroductions of this species. We believe that this study can serve as a starting point for global applications of genomics to the conservation plan of this species.

Population dynamics and genetic connectivity in recent chimpanzee history.

Knowledge on the population history of endangered species is critical for conservation, but whole-genome data on chimpanzees (Pan troglodytes) is geographically sparse. Here, we produced the first non-invasive geolocalized catalog of genomic diversity by capturing chromosome 21 from 828 non-invasive samples collected at 48 sampling sites across Africa. The four recognized subspecies show clear genetic differentiation correlating with known barriers, while previously undescribed genetic exchange suggests that these have been permeable on a local scale. We obtained a detailed reconstruction of population stratification and fine-scale patterns of isolation, migration, and connectivity, including a comprehensive picture of admixture with bonobos (Pan paniscus). Unlike humans, chimpanzees did not experience extended episodes of long-distance migrations, which might have limited cultural transmission. Finally, based on local rare variation, we implement a fine-grained geolocalization approach demonstrating improved precision in determining the origin of confiscated chimpanzees.

HuConTest: Testing Human Contamination in Great Ape Samples.

Modern human contamination is a common problem in ancient DNA studies. We provide evidence that this issue is also present in studies in great apes, which are our closest living relatives, for example in noninvasive samples. Here, we present a simple method to detect human contamination in short-read sequencing data from different species: HuConTest. We demonstrate its feasibility using blood and tissue samples from these species. This test is particularly useful for more complex samples (such as museum and noninvasive samples) which have smaller amounts of endogenous DNA, as we show here.

Maximizing the acquisition of unique reads in noninvasive capture sequencing experiments.

Noninvasive samples as a source of DNA are gaining interest in genomic studies of endangered species. However, their complex nature and low endogenous DNA content hamper the recovery of good quality data. Target capture has become a productive method to enrich the endogenous fraction of noninvasive samples, such as faeces, but its sensitivity has not yet been extensively studied. Coping with faecal samples with an endogenous DNA content below 1% is a common problem when prior selection of samples from a large collection is not possible. However, samples classified as unfavourable for target capture sequencing might be the only representatives of unique specific geographical locations, or to answer the question of interest. To explore how library complexity may be increased without repeating DNA extractions and generating new libraries, in this study we captured the exome of 60 chimpanzees (Pan troglodytes) using faecal samples with very low proportions of endogenous content (<1%). Our results indicate that by performing additional hybridizations of the same libraries, the molecular complexity can be maintained to achieve higher coverage. Also, whenever possible, the starting DNA material for capture should be increased. Finally, we specifically calculated the sequencing effort needed to avoid exhausting the library complexity of enriched faecal samples with low endogenous DNA content. This study provides guidelines, schemes and tools for laboratories facing the challenges of working with noninvasive samples containing extremely low amounts of endogenous DNA.

Metagenomic analysis of a blood stain from the French revolutionary Jean-Paul Marat (1743-1793).

The French revolutionary Jean-Paul Marat (1743-1793) was assassinated in 1793 in his bathtub, where he was trying to find relief from the debilitating skin disease he was suffering from. At the time of his death, Marat was annotating newspapers, which got stained with his blood and were subsequently preserved by his sister. We extracted and sequenced DNA from the blood stain and also from another section of the newspaper, which we used for comparison. Results from the human DNA sequence analyses were compatible with a heterogeneous ancestry of Marat, with his mother being of French origin and his father born in Sardinia. Metagenomic analyses of the non-human reads uncovered the presence of fungal, bacterial and low levels of viral DNA. Relying on the presence/absence of microbial species in the samples, we could cast doubt on several putative infectious agents that have been previously hypothesised as the cause of his condition but for which we detect not a single sequencing read. Conversely, some of the species we detect are uncommon as environmental contaminants and may represent plausible infective agents. Based on all the available evidence, we hypothesize that Marat may have suffered from a fungal infection (seborrheic dermatitis), possibly superinfected with bacterial opportunistic pathogens.

Selective single molecule sequencing and assembly of a human Y chromosome of African origin.

Mammalian Y chromosomes are often neglected from genomic analysis. Due to their inherent assembly difficulties, high repeat content, and large ampliconic regions, only a handful of species have their Y chromosome properly characterized. To date, just a single human reference quality Y chromosome, of European ancestry, is available due to a lack of accessible methodology. To facilitate the assembly of such complicated genomic territory, we developed a novel strategy to sequence native, unamplified flow sorted DNA on a MinION nanopore sequencing device. Our approach yields a highly continuous assembly of the first human Y chromosome of African origin. It constitutes a significant improvement over comparable previous methods, increasing continuity by more than 800%. Sequencing native DNA also allows to take advantage of the nanopore signal data to detect epigenetic modifications in situ. This approach is in theory generalizable to any species simplifying the assembly of extremely large and repetitive genomes.