Ecological Transcriptomics in Salamandra salamandra and S. infraimmaculata, a Test Case for Applying Non-Lethal Sampling Techniques

Ecological transcriptomics approaches are useful for studying the ecology-evolution interface. Because transcriptome analyses often involve sacrificing whole individuals, which can be ethically and legally problematic especially for endangered species, we assessed the feasibility of using non-lethally sampled tail-fin clips for the transcriptome analysis of fire salamander larvae. A custom gene expression microarray including 22,145 transcript specific probes was designed based on De novo transcriptome sequencing of fire salamander larvae. We used the array to characterize the thermal response of fire salamander larvae and for comparing results obtained through the analysis of non-lethally sampled tail-clips vs. entire larvae. We found that transcriptome analysis of tail clips is a valid alternative to the entire larvae; 51% of the candidate genes derived from the analysis of whole larvae, could be inferred from analysis of tail-clips. While candidate genes were not identical, their functional categorization gave highly comparable results for the inference of likely phenotypes involved in larval thermal response (e.g. an overexpression of certain mitochondrial transcripts or an increase mitochondrial volume density), which match known thermal acclimatization responses of ectotherms. We also used the microarrays to identify developmental gene expression profiles, from birth to metamorphosis, in tail-clips sampled from Salamandra Infraimmaculata larvae. This analysis revealed transcriptome changes associated with initiation and preparation of metamorphosis. In conclusion, we recommend ecological transcriptomics based on non-lethal peripheral tissue samples like tail-clips as a valid alternative to lethal tissue sampling.