Tardigrada, a phylum of meiofaunal organisms, have been at the center of discussions of the evolution of Metazoa, the biology of survival in extreme environments, and the role of horizontal gene transfer in animal evolution. Tardigrada are placed as sisters to Arthropoda and Onychophora (velvet worms) in the superphylum Panarthropoda by morphological analyses, but many molecular phylogenies fail to recover this relationship. This tension between molecular and morphological understanding may be very revealing of the mode and patterns of evolution of major groups. Limnoterrestrial tardigrades display extreme cryptobiotic abilities, including anhydrobiosis and cryobiosis, as do bdelloid rotifers, nematodes, and other animals of the water film. These extremophile behaviors challenge understanding of normal, aqueous physiology: how does a multicellular organism avoid lethal cellular collapse in the absence of liquid water? Meiofaunal species have been reported to have elevated levels of horizontal gene transfer (HGT) events, but how important this is in evolution, and particularly in the evolution of extremophile physiology, is unclear. To address these questions, we resequenced and reassembled the genome of H. dujardini, a limnoterrestrial tardigrade that can undergo anhydrobiosis only after extensive pre-exposure to drying conditions, and compared it to the genome of R. varieornatus, a related species with tolerance to rapid desiccation. The 2 species had contrasting gene expression responses to anhydrobiosis, with major transcriptional change in H. dujardini but limited regulation in R. varieornatus. We identified few horizontally transferred genes, but some of these were shown to be involved in entry into anhydrobiosis. Whole-genome molecular phylogenies supported a Tardigrada+Nematoda relationship over Tardigrada+Arthropoda, but rare genomic changes tended to support Tardigrada+Arthropoda.

Across diverse taxa, selfing species have evolved independently from outcrossing species thousands of times. The transition from outcrossing to selfing significantly decreases the effective population size, effective recombination rate, and heterozygosity within a species. These changes lead to a reduction in genetic diversity, and therefore adaptive potential, by intensifying the effects of random genetic drift and linked selection. Within the nematode genus Caenorhabditis , selfing has evolved at least three times and all three species, including in the model organism Caenorhabditis elegans , show substantially reduced genetic diversity relative to outcrossing species. Selfing and outcrossing Caenorhabditis species are often found in the same niches, but we still do not know how selfing species with limited genetic diversity can adapt to these environments. Here, we examine the whole-genome sequences from 609 wild C. elegans strains isolated worldwide and show that genetic variation is concentrated in punctuated hyper-divergent regions that cover 20% of the C. elegans reference genome. These regions are enriched in environmental response genes that mediate sensory perception, pathogen response, and xenobiotic stress response. Population genomic evidence suggests that genetic diversity in these regions has been maintained by long-term balancing selection. Using long- read genome assemblies for 15 wild strains, we show that hyper-divergent haplotypes contain unique sets of genes and show levels of divergence comparable to levels found between Caenorhabditis species that diverged millions of years ago. These results provide an example for how species can avoid the evolutionary “dead end” associated with selfing.

Tardigrades are meiofaunal ecdysozoans that are key to understanding the origins of Arthropoda. Many species of Tardigrada can survive extreme conditions through cryptobiosis. In a recent paper [Boothby TC, et al. (2015) Proc Natl Acad Sci USA 112(52):15976-15981], the authors concluded that the tardigrade Hypsibius dujardini had an unprecedented proportion (17%) of genes originating through functional horizontal gene transfer (fHGT) and speculated that fHGT was likely formative in the evolution of cryptobiosis. We independently sequenced the genome of H. dujardini. As expected from whole-organism DNA sampling, our raw data contained reads from nontarget genomes. Filtering using metagenomics approaches generated a draft H. dujardini genome assembly of 135 Mb with superior assembly metrics to the previously published assembly. Additional microbial contamination likely remains. We found no support for extensive fHGT. Among 23,021 gene predictions we identified 0.2% strong candidates for fHGT from bacteria and 0.2% strong candidates for fHGT from nonmetazoan eukaryotes. Cross-comparison of assemblies showed that the overwhelming majority of HGT candidates in the Boothby et al. genome derived from contaminants. We conclude that fHGT into H. dujardini accounts for at most 1-2% of genes and that the proposal that onesixth of tardigrade genes originate from functional HGT events is an artifact of undetected contamination.tardigrade | blobtools | contamination | metagenomics | horizontal gene transfer

The nematode Caenorhabditis elegans has been central to the understanding of metazoan biology. However, C. elegans is but one species among millions and the significance of this important model organism will only be fully revealed if it is placed in a rich evolutionary context. Global sampling efforts have led to the discovery of over 50 putative species from the genus Caenorhabditis , many of which await formal species description. Here, we present species descriptions for 10 new Caenorhabditis species. We also present draft genome sequences for nine of these new species, along with a transcriptome assembly for one. We exploit these whole‐genome data to reconstruct the Caenorhabditis phylogeny and use this phylogenetic tree to dissect the evolution of morphology in the genus. We reveal extensive variation in genome size and investigate the molecular processes that underlie this variation. We show unexpected complexity in the evolutionary history of key developmental pathway genes. These new species and the associated genomic resources will be essential in our attempts to understand the evolutionary origins of the C. elegans model.

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