Zoonoses are infectious diseases transmitted from animals to humans. Bats have been suggested to harbour more zoonotic viruses than any other mammalian order¹. Infections in bats are largely asymptomatic^2,3, indicating limited tissue-damaging inflammation and immunopathology. To investigate the genomic basis of disease resistance, the Bat1K project generated reference-quality genomes of ten bat species, including potential viral reservoirs. Here we describe a systematic analysis covering 115 mammalian genomes that revealed that signatures of selection in immune genes are more prevalent in bats than in other mammalian orders. We found an excess of immune gene adaptations in the ancestral chiropteran branch and in many descending bat lineages, highlighting viral entry and detection factors, and regulators of antiviral and inflammatory responses. ISG15, which is an antiviral gene contributing to hyperinflammation during COVID-19 (refs. ^4,5), exhibits key residue changes in rhinolophid and hipposiderid bats. Cellular infection experiments show species-specific antiviral differences and an essential role of protein conjugation in antiviral function of bat ISG15, separate from its role in secretion and inflammation in humans. Furthermore, in contrast to humans, ISG15 in most rhinolophid and hipposiderid bats has strong anti-SARS-CoV-2 activity. Our work reveals molecular mechanisms that contribute to viral tolerance and disease resistance in bats.

High-quality and complete reference genome assemblies are fundamental for the application of genomics to biology, disease, and biodiversity conservation. However, such assemblies are only available for a few non-microbial species ^1–4 . To address this issue, the international Genome 10K (G10K) consortium ^5,6 has worked over a five-year period to evaluate and develop cost-effective methods for assembling the most accurate and complete reference genomes to date. Here we summarize these developments, introduce a set of quality standards, and present lessons learned from sequencing and assembling 16 species representing major vertebrate lineages (mammals, birds, reptiles, amphibians, teleost fishes and cartilaginous fishes). We confirm that long-read sequencing technologies are essential for maximizing genome quality and that unresolved complex repeats and haplotype heterozygosity are major sources of error in assemblies. Our new assemblies identify and correct substantial errors in some of the best historical reference genomes. Adopting these lessons, we have embarked on the Vertebrate Genomes Project (VGP), an effort to generate high-quality, complete reference genomes for all ~70,000 extant vertebrate species and help enable a new era of discovery across the life sciences.

Background: Studies in vertebrate genomics require sampling from a broad range of tissue types, taxa, and localities. Recent advancements in long-read and long-range genome sequencing have made it possible to produce high-quality chromosome-level genome assemblies for almost any organism. However, adequate tissue preservation for the requisite ultra-high molecular weight DNA (uHMW DNA) remains a major challenge. Here we present a comparative study of preservation methods for field and laboratory tissue sampling, across vertebrate classes and different tissue types. Results: We find that storage temperature was the strongest predictor of uHMW fragment lengths. While immediate flash-freezing remains the sample preservation gold standard, samples preserved in 95% EtOH or 20–25% DMSO-EDTA showed little fragment length degradation when stored at 4°C for 6 hours. Samples in 95% EtOH or 20–25% DMSO-EDTA kept at 4°C for 1 week after dissection still yielded adequate amounts of uHMW DNA for most applications. Tissue type was a significant predictor of total DNA yield but not fragment length. Preservation solution had a smaller but significant influence on both fragment length and DNA yield. Conclusion: We provide sample preservation guidelines that ensure sufficient DNA integrity and amount required for use with long-read and long-range sequencing technologies across vertebrates. Our best practices generated the uHMW DNA needed for the high-quality reference genomes for phase 1 of the Vertebrate Genomes Project, whose ultimate mission is to generate chromosome-level reference genome assemblies of all ∼70,000 extant vertebrate species.

Summary Paragraph: The genus Myotis is one of the largest clades of bats, and exhibits some of the most extreme variation in lifespans among mammals alongside unique adaptations to viral tolerance and immune defense ^1–3 . To study the evolution of these phenotypes we generated cell lines and near-complete genome assemblies for eight closely related Myotis species. Using genome-wide screens of positive selection, analyses of structural variation, and functional experiments in primary cells, we identify patterns of adaptation contributing to longevity, cancer resistance, and viral interactions. We demonstrate distinct modes of adaptation to DNA and RNA viruses compared to all other mammals, with bats exhibiting genome-wide overrepresentation of positive selection for DNA virus-interacting proteins and elevated rates of copy number variation for RNA virus-interacting proteins. Characterization of Myotis- specific duplications of the key immune factor protein kinase R ( PKR ) reveals multiple ancient segregating trans-species copy number polymorphisms. We show that the recurrent evolution of longevity seen in Myotis is associated with positive selection in cancer pathways, and demonstrate a unique response to DNA damage in primary cells of the long-lived M. lucifugus . Together, our results suggest that bats’ remarkable longevity and immunity are linked through pleiotropic adaptations to viruses and aging-related disease.