Initial Phylotranscriptomic Confirmation of Homoplastic Evolution of the Conspicuous Coloration and Bufoniform Morphology of Pumpkin-Toadlets in the Genus Brachycephalus.

Sven Künzel,Thais H Condez,Loïs Rancilhac,Mariana L Lyra,Eugenia Sanchez,Luís Felipe Toledo,Célio F B Haddad,Mirco Solé,Miguel Vences,Juliane P C Monteiro,Iker Irisarri,Omar Rojas-Padilla

doi:10.3390/toxins13110816

Abstract

The genus Brachycephalus is a fascinating group of miniaturized anurans from the Brazilian Atlantic Forest, comprising the conspicuous, brightly colored pumpkin-toadlets and the cryptic flea-toads. Pumpkin-toadlets are known to contain tetrodotoxins and therefore, their bright colors may perform an aposematic function. Previous studies based on a limited number of mitochondrial and nuclear-encoded markers supported the existence of two clades containing species of pumpkin-toadlet phenotype, but deep nodes remained largely unresolved or conflicting between data sets. We use new RNAseq data of 17 individuals from nine Brachycephalus species to infer their evolutionary relationships from a phylogenomic perspective. Analyses of almost 5300 nuclear-encoded ortholog protein-coding genes and full mitochondrial genomes confirmed the existence of two separate pumpkin-toadlet clades, suggesting the convergent evolution (or multiple reversals) of the bufoniform morphology, conspicuous coloration, and probably toxicity. In addition, the study of the mitochondrial gene order revealed that three species (B. hermogenesi, B. pitanga, and B. rotenbergae) display translocations of different tRNAs (NCY and CYA) from the WANCY tRNA cluster to a position between the genes ATP6 and COIII, showing a new mitochondrial gene order arrangement for vertebrates. The newly clarified phylogeny suggests that Brachycephalus has the potential to become a promising model taxon to understand the evolution of coloration, body plan and toxicity. Given that toxicity information is available for only few species of Brachycephalus, without data for any flea-toad species, we also emphasize the need for a wider screening of toxicity across species, together with more in-depth functional and ecological study of their phenotypes.

Highlights

The evolution of aposematism, i.e the co-occurrence of conspicuous coloration and defensive mechanisms, is fascinating and poorly understood, despite intensive research over many decades (e.g., [1,2,3,4,5])
Several pumpkin-toadlets are known to contain toxins: tetrodotoxin analogues (TTX) has been reported in B. ephippium, B. nodoterga, B. pernix, B. pitanga, and B. rotenbergae [15,21,22] and bradykinin-potentiating peptides (BPP) have been reported in B. rotenbergae ([23]—as B. ephippium)
The position of B. albolineatus within the B. pernix group was different between the datasets: phylotranscriptomic analyses recovered the species as sister to B. actaeus, whereas the mitogenomic analysis placed it sister to the B. pernix group

Summary

Introduction

The evolution of aposematism, i.e the co-occurrence of conspicuous coloration and defensive mechanisms, is fascinating and poorly understood, despite intensive research over many decades (e.g., [1,2,3,4,5]). Amphibians have long been a prime group for research on the evolution of aposematism and contain a large number of species that exhibit putative warning signals [14]. They are the only class of terrestrial vertebrates known to contain tetrodotoxin analogues (TTX; [15]). Flea-toads have an inconspicuous brown coloration [19,20] and their putative toxicity has so far not been assessed These characteristics make Brachycephalus an excellent model for studying the evolution of anti-predator defense mechanisms associated with toxins

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Conclusion