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The genomic basis of convergent evolution in modern sloths (Xenarthra, Mammalia)

Periodic Reporting for period 1 - CONVGENOMS (The genomic basis of convergent evolution in modern sloths (Xenarthra, Mammalia))

Reporting period: 2017-10-01 to 2019-09-30

Tree-sloths are some of the most unusual of mammals. The two extant genera, two-toed Choloepus, and three-toed Bradypus spend their lives suspended in tree canopies primarily eating leaves in the forests of South and Central America. To maintain this lifestyle, tree-sloths underwent a number of major adaptations during their evolution: they present a lower-than-average muscle content, a porous bone structure and several anatomical forelimb bone adaptations that allow them to hang against gravity on the tree canopy with minimal energy expenditure. They are the only inverted quadrupeds in the world. In addition, sloths are unusually heterothermic, partially regulation their body temperature and metabolism in accordance with the ambient temperature. They are also the only survivors of a once very diverse group (~100 genera) where most of the species perished at the end of the last ice age (about 11,000 years ago). In contrast to their extinct relatives - that were mostly very large and terrestrial - Bradypus and Choloepus are the only ones to live exclusively on the tree canopies. Very recent studies have shown that their numerous shared superficial adaptations evolved in parallel, as Bradypus and Choloepus are only distantly-related, sharing a last common ancestor ~30 million years ago. These studies show that each clade of extant sloths is more closely related to distinct groups of large-ground sloths than they are to each other, making their obligatory suspensory locomotion one of the most impressive cases of convergent evolution among mammals. Very little was known about the genomics of sloths until this work was performed. In fact, the whole Superorder Xenarthra (sloths, anteaters and armadillos) is understudied from a molecular perspective, despite representing an entire clade of placental mammals with unique adaptations and life-styles. This project aimed to sequence genomes for tree-sloths and perform comparative genomic analysis to study their unique adaptations from the molecular perspective. We have done so by sequencing one chromosome-level genome for Choloepus didactylus and a draft genome for Bradypus tritdactylus. Detailed comparative analysis of these genomes is allowing us to unveil the molecular basis of their peculiar adaptations.
This project has assembled a chromosome-level genome for the two-toed sloth Choloepus didactylus. This genome was produced with four sequencing technologies (Pacbio long reads, Chromium 10X linked reads, Bionano Optical Maps and Arima Hi-C reads) and a state-of-the-art hierarchical genome assembly pipeline (VGP 1.6 Assembly Pipeline). The experienced researcher and main supervisor have joined the Vertebrate Genomes Project - an international initiative within the Genome 10K consortium testing, developing and implementing methods to assemble the final reference genomes of all vertebrates, starting with one representative of each of the ~260 vertebrate orders. The Choloepus didactylus genome assembled by this project is the first representative of the Pilosa and Xenarthra on the VGP Phase I. C. didactylus genome was assembled to a total of 3.6Gb in 281 scaffolds with a N50=161 Mb, making it one of the best mammalian genomes assembled to date. This genome is in its final stages of manual curation, but sexual chromosomes are already identified (X and Y) and the Hi-C heat map seems to indicate that the karyotype of this specimen of C. didactylus is 2n=65. Several entire chromosome syntenies were found between C. didactylus and human chromosomes, allowing the start of the investigation of genome architecture evolution. In addition, 46% of C. didactylus genome is composed of repeats, with LINE1 being the most abundant element. Six tissues were sequenced as RNA-Seq (lung, blood, brain, heart, spleen and liver) to allow a high-quality annotation of C. didactylus genome. A genome annotation performed with MAKER2, followed by a single-copy-orthologs comparative analysis of C. didactylus and 10 other protein sets predicted from whole genomes representing all described mammalian lineages indicate Afrotheria to be more basal than Xenarthra within Eutherian mammals. The mitochondrial genome of C. didactylus was also assembled, presenting 16499 bp, 13 coding genes, 22 tRNAs, and 2 rRNAS. Both genomes of C. didactylus, nuclear and mitochondrial, are available at Further, this project has sequenced Illumina short reads for the three-toed sloth Bradypus tridactylus, and has assembled a draft genome for it with DISCOVAR. The kmer composition of B. tridactylus indicated a genome size of 3.3Gb. Homology analysis of C. didactylus chromosomes with B. tridactylus DISCOVAR contigs show 70% shared homology between both genomes. The genomes assembled by this work representing both clades of tree-sloths, Bradypus and Choloepus, present smaller genomes than previously estimated for sloths and other Xenarthrans (>4Gb). Comparative analysis of the unique genomic features of sloth’s genomes are being carried out and will be presented in open-access scientific manuscripts in the near future.
This project has also made efforts to train the next generation of genome bioinformaticians: the Experienced Researcher (ER) and Main Supervisor started an initiative to train masters students to run the Vertebrate Genomes Project Assembly Pipeline. The ER has trained at least four students at the Berlin Center for Genomics and Biodiversity Research to integrate the four sequencing technologies and run the VGP Assembly Pipeline. These students have produced at least one chromosome-level genome each, getting high-quality training and greatly contributing to the international scientific community studying genome evolution. This training initiative is getting larger and is being followed by other senior researchers within the VGP which are now integrating students with the consortium.
Nothing in biology makes sense if not in the light of evolution. This project has generated high-quality genome information for a basal clade of placental mammals that were, until now, understudied from the molecular perspective. In addition to the study of the specific molecular basis of sloths adaptations, this work also adds to the understanding of the molecular basis of adaptations of all mammals. Just comparative genomics - coupled with experimental analysis - will allow us to ultimately understand the complete picture of mammal’s molecular biology and employ them to innovative applications in human health. Finally, this work presents a high-quality genome that can be used as a reference for groups studying the diversity of two-toed sloths populations in the amazon. This will then enable numerous studies on population connectivity, local adaptation, relatedness, and the conservation genetics of these species. Understanding our natural world, and getting the tools to help preserve it are some of the most important and meaningful tasks for science and for humanity. This study aids our understanding of this special and unique group of animals, and may help preserve them into the future.
Hi-C heat map of Choloepus didactylus chromosome-level genome assembly