Increasingly, large phylogenomic data sets include transcriptomic data from nonmodel organisms. This not only has allowed controversial and unexplored evolutionary relationships in the tree of life to be addressed but also increases the risk of inadvertent inclusion of paralogs in the analysis. Although this may be expected to result in decreased phylogenetic support, it is not clear if it could also drive highly supported artifactual relationships. Many groups, including the hyperdiverse Lissamphibia, are especially susceptible to these issues due to ancient gene duplication events and small numbers of sequenced genomes and because transcriptomes are increasingly applied to resolve historically conflicting taxonomic hypotheses. We tested the potential impact of paralog inclusion on the topologies and timetree estimates of the Lissamphibia using published and de novo sequencing data including 18 amphibian species, from which 2,656 single-copy gene families were identified. A novel paralog filtering approach resulted in four differently curated data sets, which were used for phylogenetic reconstructions using Bayesian inference, maximum likelihood, and quartet-based supertrees. We found that paralogs drive strongly supported conflicting hypotheses within the Lissamphibia (Batrachia and Procera) and older divergence time estimates even within groups where no variation in topology was observed. All investigated methods, except Bayesian inference with the CAT-GTR model, were found to be sensitive to paralogs, but with filtering convergence to the same answer (Batrachia) was observed. This is the first large-scale study to address the impact of orthology selection using transcriptomic data and emphasizes the importance of quality over quantity particularly for understanding relationships of poorly sampled taxa.

RNA sequencing (RNA-seq) has become one of the most powerful tools to unravel the genomic basis of biological adaptation and diversity. Although challenging, RNA-seq is particularly promising for research on non-model, secretive species that cannot be observed in nature easily and therefore remain comparatively understudied. Among such animals, the caecilians (order Gymnophiona) likely constitute the least known group of vertebrates, despite being an old and remarkably distinct lineage of amphibians. Here, we characterize multi-tissue transcriptomes for five species of caecilians that represent a broad level of diversity across the order. We identified vertebrate homologous elements of caecilian functional genes of varying tissue specificity that reveal a great number of unclassified gene families, especially for the skin. We annotated several protein domains for those unknown candidate gene families to investigate their function. We also conducted supertree analyses of a phylogenomic dataset of 1,955 candidate orthologous genes among five caecilian species and other major lineages of vertebrates, with the inferred tree being in agreement with current views of vertebrate evolution and systematics. Our study provides insights into the evolution of vertebrate protein-coding genes, and a basis for future research on the molecular elements underlying the particular biology and adaptations of caecilian amphibians.

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Background The amphibian skin microbiome is an important mediator of host health and serves as a potential source of undiscovered scientifically significant compounds. However, the underlying modalities of how amphibian hosts obtain their initial skin-associated microbiome remains unclear. Here, we explore microbial transmission patterns in foam-nest breeding tree frogs from Southeast Asia (Genus: Polypedates) whose specialized breeding strategy allows for better delineation between vertically and environmentally derived microbes. To facilitate this, we analyzed samples associated with adult frog pairs taken after mating—including adults of each sex, their foam nests, environments, and tadpoles before and after environmental interaction—for the bacterial communities using DNA metabarcoding data (16S rRNA). Samples were collected from frogs in-situ in Brunei, Borneo, a previously unsampled region for amphibian-related microbial diversity. Results Adult frogs differed in skin bacterial communities among species, but tadpoles did not differ among species. Foam nests had varying bacterial community composition, most notably in the nests’ moist interior. Nest interior bacterial communities were discrete for each nest and overall displayed a narrower diversity compared to the nest exteriors. Tadpoles sampled directly from the foam nest displayed a bacterial composition less like the nest interior and more similar to that of the adults and nest exterior. After one week of pond water interaction the tadpole skin microbiome shifted towards the tadpole skin and pond water microbial communities being more tightly coupled than between tadpoles and the internal nest environment, but not to the extent that the skin microbiome mirrored the pond bacterial community. Conclusions Both vertical influence and environmental interaction play a role in shaping the tadpole cutaneous microbiome. Interestingly, the interior of the foam nest had a distinct bacterial community from the tadpoles suggesting a limited environmental effect on tadpole cutaneous bacterial selection at initial stages of life. The shift in the tadpole microbiome after environmental interaction indicates an interplay between underlying host and ecological mechanisms that drive community formation. This survey serves as a baseline for further research into the ecology of microbial transmission in aquatic animals.

DNA methylation is mediated by a conserved family of DNA methyltransferases (Dnmts). The human genome encodes three active Dnmts (Dnmt1, Dnmt3a and Dnmt3b), the tRNA methyltransferase Dnmt2, and the regulatory protein Dnmt3L. Despite their high degree of conservation among different species, genes encoding Dnmts have been duplicated and/or lost in multiple lineages throughout evolution, indicating that the DNA methylation machinery has some potential to undergo evolutionary change. However, little is known about the extent to which this machinery, or the methylome, varies among vertebrates. Here, we study the molecular evolution of Dnmt1, the enzyme responsible for maintenance of DNA methylation patterns after replication, in 79 vertebrate species. Our analyses show that all studied species exhibit a single copy of the DNMT1 gene, with the exception of tilapia and marsupials (tammar wallaby, koala, Tasmanian devil and opossum), each of which displays two apparently functional DNMT1 copies. Our phylogenetic analyses indicate that DNMT1 duplicated before the radiation of major marsupial groups (i.e., at least ~75 million years ago), thus giving rise to two DNMT1 copies in marsupials (copy 1 and copy 2). In the opossum lineage, copy 2 was lost, and copy 1 recently duplicated again, generating three DNMT1 copies: two putatively functional genes (copy 1a and 1b) and one pseudogene (copy 1ψ). Both marsupial copies (DNMT1 copies 1 and 2) are under purifying selection, and copy 2 exhibits elevated rates of evolution and signatures of positive selection, suggesting a scenario of neofunctionalization. This gene duplication might have resulted in modifications in marsupial methylomes and their dynamics.

Symbiosis, defined as living together in physical contact between different species, greatly impacts many aspects of organismal biology. The pervasiveness of these systems in nature led Lynn Margulis to propose that symbiotic interactions are one of the major drivers of evolution (Margulis, 1998). Many characteristics of these associations can vary, including direction of cost/benefit effects, degree of dependence between partners and symbiont transmission traits. This variation profoundly affects ecological and evolutionary

Background: Gene expression profiles can provide insights into the molecular machinery behind tissue functions and, in turn, can further our understanding of environmental responses, and developmental and evolutionary processes. During vertebrate evolution, the skin has played a crucial role, displaying a wide diversity of essential functions. To unravel the molecular basis of skin specialisations and adaptations, we compared gene expression in the skin with eight other tissues in a phylogenetically and ecologically diverse species sample of one of the most neglected vertebrate groups, the caecilian amphibians (order Gymnophiona). Results: The skin of the five studied caecilian species showed a distinct gene expression profile reflecting its developmental origin and showing similarities to other epithelial tissues. We identified 59 sequences with conserved enhanced expression in the skin that might be associated with caecilian dermal specialisations. Some of the upregulated genes shared expression patterns with human skin and potentially are involved in skin functions across vertebrates. Variation trends in gene expression were detected between mid and posterior body skin suggesting different functions between body regions. Several candidate biologically active peptides were also annotated. Conclusions: Our study provides the first atlas of differentially expressed sequences in caecilian tissues and a baseline to explore the molecular basis of the skin functions in caecilian amphibians, and more broadly in vertebrates.

While some pathogens are limited to single species, others can colonize many hosts, likely contributing to the emergence of novel disease outbreaks. Despite this biodiversity threat, traits associated with host niche expansions are not well understood in multihost pathogens. Here, we aimed to uncover functional machinery driving multihost invasion by focusing on Batrachochytrium dendrobatidis (Bd), a pathogen that infects the skin of hundreds of amphibians worldwide. We performed a meta‐analysis of Bd gene expression using data from published infection experiments and newly generated profiles. We analysed Bd transcriptomic landscapes across the skin of 14 host species, reconstructed Bd isolates phylogenetic relationships, and inferred the origin and evolutionary history of differentially expressed genes under a phylogenetic framework comprising other 12 zoosporic fungi. Bd displayed plastic infection strategies when challenged by hosts with different disease susceptibility. Our analyses identified sets of differentially expressed genes under host environments with similar infection outcome. We stressed nutritional immunity and gene silencing as important processes required to overcome challenging skin environments in less susceptible hosts. Overall, Bd genes expressed during amphibian skin exploitation have arisen mainly via gene duplications with great family expansions, increasing the gene copy events previously described for this fungal species. Finally, we provide a comprehensive gene data set that can be used to further examine eco‐evolutionary hypotheses for this host‐pathogen system. Our study supports the idea that host environments exert contrasting selective pressures, such that gene expression plasticity could be one of the evolutionary keys leading to the success of multihost pathogens.