A toxic environment selects for specialist microbiome in poison frogs.

Shifts in microbiome community composition can have large effects on host health. It is therefore important to understand how perturbations, like those caused by the introduction of exogenous chemicals, modulate microbiome community composition. In poison frogs within the family Dendrobatidae, the skin microbiome is exposed to the alkaloids that the frogs sequester from their diet and use for defense. Given the demonstrated antimicrobial effects of these poison frog alkaloids, these compounds may be structuring the skin microbial community. To test this, we first characterized microbial communities from chemically defended and closely related non-defended frogs from Ecuador. Then we conducted a laboratory experiment to monitor the effect of the alkaloid decahydroquinoline (DHQ) on the microbiome of a single frog species. In both the field and lab experiments, we found that alkaloid-exposed microbiomes are more species rich and phylogenetically diverse, with an increase in rare taxa. To better understand the strain-specific behavior in response to alkaloids, we cultured microbial strains from poison frog skin and found the majority of strains exhibited either enhanced growth or were not impacted by the addition of DHQ. Additionally, stable isotope tracing coupled to nanoSIMS suggests that some of these strains are able to metabolize DHQ. Taken together, these data suggest that poison frog chemical defenses open new niches for skin-associated microbes with specific adaptations, including the likely metabolism of alkaloids, that enable their survival in this toxic environment. This work helps expand our understanding of how exposure to exogenous compounds like alkaloids can impact host microbiomes.

Tissue-specific in vivo transformation of plasmid DNA in Neotropical tadpoles using electroporation.

Electroporation is an increasingly common technique used for exogenous gene expression in live animals, but protocols are largely limited to traditional laboratory organisms. The goal of this protocol is to test in vivo electroporation techniques in a diverse array of tadpole species. We explore electroporation efficiency in tissue-specific cells of five species from across three families of tropical frogs: poison frogs (Dendrobatidae), cryptic forest/poison frogs (Aromobatidae), and glassfrogs (Centrolenidae). These species are well known for their diverse social behaviors and intriguing physiologies that coordinate chemical defenses, aposematism, and/or tissue transparency. Specifically, we examine the effects of electrical pulse and injection parameters on species- and tissue-specific transfection of plasmid DNA in tadpoles. After electroporation of a plasmid encoding green fluorescent protein (GFP), we found strong GFP fluorescence within brain and muscle cells that increased with the amount of DNA injected and electrical pulse number. We discuss species-related challenges, troubleshooting, and outline ideas for improvement. Extending in vivo electroporation to non-model amphibian species could provide new opportunities for exploring topics in genetics, behavior, and organismal biology.