Comparative Transcriptome Analysis of Development in Salamandra infraimmaculata

For amphibians that have biphasic life cycles (aquatic larval stage and terrestrial juvenile and adult stages), habitat conditions during the larval/aquatic stage can have a great impact on their fitness. Ecological transcriptomics approaches are useful and increasingly affordable methods for research at the interface of ecology and evolution. We used microarray analysis to identify genes involved in developmental changes and/or local adaptation in Salamandra infraimmaculata larvae from a permanent (Tel Dan) and an ephemeral (Manof) habitat type in Israel. Gene expression patterns over time can shed light on the developmental process occurring in the larvae, since visual morphological changes occur just before the end of the metamorphic process. We designed custom microarray based on a de-novo RNA sequencing of S. infraimmaculata larvae, which were used for testing RNA from non-lethal samples of Salamandra tail clips. Principle component analysis (PCA) results showed that PC1, which distributed the different samples by time of development (e.g. time to metamorphosis), explained 61% of the variance of the gene expression, and PC2, which divided the samples by populations, explained additional 16% of the variance. From the most influential genes in PC1, 89% significantly were correlated between Manof and Tel-Dan and from the most influential genes on PC2, 12.6% were significantly different between populations Manof and Tel-Dan. Our microarray results identified candidate genes involved in different stages of development (e.g. beginning of metamorphosis) and candidate genes that can differentiate between populations or habitat type, which may be involved in local adaptation. Our results demonstrate the high potential of comparative transcriptomics in evolutionary and ecological studies.