Tasmanian devils have spawned two transmissible cancer lineages, named devil facial tumor 1 (DFT1) and devil facial tumor 2 (DFT2). We investigated the genetic diversity and evolution of these clones by analyzing 78 DFT1 and 41 DFT2 genomes relative to a newly assembled, chromosome-level reference. Time-resolved phylogenetic trees reveal that DFT1 first emerged in 1986 (1982 to 1989) and DFT2 in 2011 (2009 to 2012). Subclone analysis documents transmission of heterogeneous cell populations. DFT2 has faster mutation rates than DFT1 across all variant classes, including substitutions, indels, rearrangements, transposable element insertions, and copy number alterations, and we identify a hypermutated DFT1 lineage with defective DNA mismatch repair. Several loci show plausible evidence of positive selection in DFT1 or DFT2, including loss of chromosome Y and inactivation of MGA , but none are common to both cancers. This study reveals the parallel long-term evolution of two transmissible cancers inhabiting a common niche in Tasmanian devils.

Betta splendens, also known as Siamese fighting fish or “betta,” is a freshwater fish species renowned for its astonishing morphological diversity and extreme aggressive behavior. Despite recent advances in our understanding of the genetics and neurobiology of betta, the lack of tools to manipulate their genome has hindered progress at functional and mechanistic levels. In this study, we outline the use of three genetic manipulation technologies, which we have optimized for use in betta: CRISPR/Cas9-mediated knockout, CRISPR/Cas9-mediated knockin, and Tol2-mediated transgenesis. We knocked out three genes: alkal2l, bco1l, and mitfa, and analyzed their effects on viability and pigmentation. Furthermore, we knocked in a fluorescent protein into the mitfa locus, a proof-of-principle experiment of this powerful technology in betta. Finally, we used Tol2-mediated transgenesis to create fish with ubiquitous expression of GFP, and then developed a bicistronic plasmid with heart-specific expression of a red fluorescent protein to serve as a visible marker of successful transgenesis. Our work highlights the potential for the genetic manipulation of betta, providing valuable resources for the effective use of genetic tools in this animal model.

Siamese fighting (betta) fish are among the most popular and morphologically diverse pet fish, but the genetic bases of their domestication and phenotypic diversification are largely unknown. We assembled de novo the genome of a wild Betta splendens and whole-genome sequenced 98 individuals across five closely related species. We find evidence of bidirectional hybridization between domesticated ornamental betta and other wild Betta species. We discover dmrt1 as the main sex determination gene in ornamental betta and that it has lower penetrance in wild B. splendens. Furthermore, we find genes with signatures of recent, strong selection that have large effects on color in specific parts of the body or on the shape of individual fins and that most are unlinked. Our results demonstrate how simple genetic architectures paired with anatomical modularity can lead to vast phenotypic diversity generated during animal domestication and launch betta as a powerful new system for evolutionary genetics.

Betta splendens, also called Siamese fighting fish or 'betta,' are a popular species in the fishkeeping hobby. Native to South- east Asia, betta have been selectively bred for their fighting ability for hundreds of years, which has resulted in the species' characteristic male aggression. More recently, betta have been bred for a number of ornamental traits such as coloration, fin morphology, and body size. Betta have unique characteristics and an evolutionary history that make them a useful model for studies in the fields of behavior, endocrinology, neurobiology, genetics, development, and evolution. However, standard laboratory procedures for raising and keeping these fish are not well established, which has limited their use. Here, we briefly review the past and present use of betta in research, with a focus on their utility in behavioral, neurobiological, and evolutionary studies. We then describe effective husbandry practices for maintaining betta as a research colony.

In many species, vocal communication is essential for coordinating social behaviors including courtship, mating, parenting, rivalry, and alarm signaling. Effective communication requires accurate production, detection, and classification of signals, as well as selection of socially appropriate responses. Understanding how signals are generated and how acoustic signals are perceived is key to understanding the neurobiology of social behaviors. Here we review our long-standing research program focused on Xenopus, a frog genus which has provided valuable insights into the mechanisms and evolution of vertebrate social behaviors. In Xenopus laevis, vocal signals differ between the sexes, through development, and across the genus, reflecting evolutionary divergence in sensory and motor circuits that can be interrogated mechanistically. Using two ex vivo preparations, the isolated brain and vocal organ, we have identified essential components of the vocal production system: the sexually differentiated larynx at the periphery, and the hindbrain vocal central pattern generator (CPG) centrally, that produce sex- and species-characteristic sound pulse frequencies and temporal patterns, respectively. Within the hindbrain, we have described how intrinsic membrane properties of neurons in the vocal CPG generate species-specific vocal patterns, how vocal nuclei are connected to generate vocal patterns, as well as the roles of neurotransmitters and neuromodulators in activating the circuit. For sensorimotor integration, we identified a key forebrain node that links auditory and vocal production circuits to match socially appropriate vocal responses to acoustic features of male and female calls. The availability of a well supported phylogeny as well as reference genomes from several species now support analysis of the genetic architecture and the evolutionary divergence of neural circuits for vocal communication. Xenopus thus provides a vertebrate model in which to study vocal communication at many levels, from physiology, to behavior, and from development to evolution. As one of the most comprehensively studied phylogenetic groups within vertebrate vocal communication systems, Xenopus provides insights that can inform social communication across phyla.

Devil facial tumour 1 (DFT1) is a transmissible cancer clone endangering the Tasmanian devil. The expansion of DFT1 across Tasmania has been documented, but little is known of its evolutionary history. We analysed genomes of 648 DFT1 tumours collected throughout the disease range between 2003 and 2018. DFT1 diverged early into five clades, three spreading widely and two failing to persist. One clade has replaced others at several sites, and rates of DFT1 coinfection are high. DFT1 gradually accumulates copy number variants (CNVs), and its telomere lengths are short but constant. Recurrent CNVs reveal genes under positive selection, sites of genome instability, and repeated loss of a small derived chromosome. Cultured DFT1 cell lines have increased CNV frequency and undergo highly reproducible convergent evolution. Overall, DFT1 is a remarkably stable lineage whose genome illustrates how cancer cells adapt to diverse environments and persist in a parasitic niche.

Emerging infectious diseases are rising globally and understanding host-pathogen interactions during the initial stages of disease emergence is essential for assessing potential evolutionary dynamics and designing novel management strategies. Tasmanian devils (Sarcophilus harrisii) are endangered due to a transmissible cancer-devil facial tumour disease (DFTD)-that since its emergence in the 1990s, has affected most populations throughout Tasmania. Recent studies suggest that devils are adapting to the DFTD epidemic and that disease-induced extinction is unlikely. However, in 2014, a second and independently evolved transmissible cancer-devil facial tumour 2 (DFT2)-was discovered at the d'Entrecasteaux peninsula, in south-east Tasmania, suggesting that the species is prone to transmissible cancers. To date, there is little information about the distribution, epidemiology and effects of DFT2 and its interaction with DFTD. Here, we use data from monitoring surveys and roadkills found within and adjacent to the d'Entrecasteaux peninsula to determine the distribution of both cancers and to compare their epidemiological patterns. Since 2012, a total of 51 DFTD tumours have been confirmed among 26 individuals inside the peninsula and its surroundings, while 40 DFT2 tumours have been confirmed among 23 individuals, and two individuals co-infected with both tumours. All devils with DFT2 were found within the d'Entrecasteaux peninsula, suggesting that this new transmissible cancer is geographically confined to this area. We found significant differences in tumour bodily location in DFTD and DFT2, with non-facial tumours more commonly found in DFT2. There was a significant sex bias in DFT2, with most cases reported in males, suggesting that since DFT2 originated from a male host, females might be less susceptible to this cancer. We discuss the implications of our results for understanding the epidemiological and evolutionary interactions of these two contemporary transmissible cancers and evaluating the effectiveness of potential management strategies.

The canine transmissible venereal tumor (CTVT) is a cancer lineage that arose several millennia ago and survives by “metastasizing” between hosts through cell transfer. The somatic mutations in this cancer record its phylogeography and evolutionary history. We constructed a time-resolved phylogeny from 546 CTVT exomes and describe the lineage’s worldwide expansion. Examining variation in mutational exposure, we identify a highly context-specific mutational process that operated early in the cancer’s evolution but subsequently vanished, correlate ultraviolet-light mutagenesis with tumor latitude, and describe tumors with heritable hyperactivity of an endogenous mutational process. CTVT displays little evidence of ongoing positive selection, and negative selection is detectable only in essential genes. We illustrate how long-lived clonal organisms capture changing mutagenic environments, and reveal that neutral genetic drift is the dominant feature of long-term cancer evolution.

Transmissible cancers are clonal lineages that spread through populations via contagious cancer cells. Although rare in nature, two facial tumor clones affect Tasmanian devils. Here we perform comparative genetic and functional characterization of these lineages. The two cancers have similar patterns of mutation and show no evidence of exposure to exogenous mutagens or viruses. Genes encoding PDGF receptors have copy number gains and are present on extrachromosomal double minutes. Drug screening indicates causative roles for receptor tyrosine kinases and sensitivity to inhibitors of DNA repair. Y chromosome loss from a male clone infecting a female host suggests immunoediting. These results imply that Tasmanian devils may have inherent susceptibility to transmissible cancers and present a suite of therapeutic compounds for use in conservation.

Dogs were present in the Americas before the arrival of European colonists, but the origin and fate of these precontact dogs are largely unknown. We sequenced 71 mitochondrial and 7 nuclear genomes from ancient North American and Siberian dogs from time frames spanning ~9000 years. Our analysis indicates that American dogs were not derived from North American wolves. Instead, American dogs form a monophyletic lineage that likely originated in Siberia and dispersed into the Americas alongside people. After the arrival of Europeans, native American dogs almost completely disappeared, leaving a minimal genetic legacy in modern dog populations. The closest detectable extant lineage to precontact American dogs is the canine transmissible venereal tumor, a contagious cancer clone derived from an individual dog that lived up to 8000 years ago.

Tasmanian devils have spawned two transmissible cancer clones, known as devil facial tumour 1 (DFT1) and devil facial tumour 2 (DFT2). DFT1 and DFT2 are transmitted between animals by the transfer of allogeneic contagious cancer cells by biting, and both cause facial tumours. DFT1 and DFT2 tumours are grossly indistinguishable, but can be differentiated using histopathology, cytogenetics or genotyping of polymorphic markers. However, standard diagnostic methods require specialist skills and equipment and entail long processing times. Here, we describe Tasman-PCR: a simple polymerase chain reaction (PCR)-based diagnostic assay that identifies and distinguishes DFT1 and DFT2 by amplification of DNA spanning tumour-specific interchromosomal translocations. We demonstrate the high sensitivity and specificity of this assay by testing DNA from 546 tumours and 804 normal devils. A temporal–spatial screen confirmed the reported geographic ranges of DFT1 and DFT2 and did not provide evidence of additional DFT clones. DFT2 affects disproportionately more males than females, and devils can be co-infected with DFT1 and DFT2. Overall, we present a PCR-based assay that delivers rapid, accurate and high-throughput diagnosis of DFT1 and DFT2. This tool provides an additional resource for devil disease management and may assist with ongoing conservation efforts.

Devil Facial Tumour 2 (DFT2) is a recently discovered contagious cancer circulating in the Tasmanian devil (Sarcophilus harrisii), a species which already harbours a more widespread contagious cancer, Devil Facial Tumour 1 (DFT1). Here we show that in contrast to DFT1, DFT2 cells express major histocompatibility complex (MHC) class I molecules, demonstrating that loss of MHC is not necessary for the emergence of a contagious cancer. However, the most highly expressed MHC class I alleles in DFT2 cells are common among host devils or non-polymorphic, reducing immunogenicity in a population sharing these alleles. In parallel, MHC class I loss is emerging in vivo, thus DFT2 may be mimicking the evolutionary trajectory of DFT1. Based on these results we propose that contagious cancers may exploit partial histocompatibility between the tumour and host, but that loss of allogeneic antigens could facilitate widespread transmission of DFT2.