How Long Does Evolution of the Troglomorphic Form Take? Estimating Divergence Times in Astyanax Mexicanus

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Features including colonization routes (stream capture) and the existence of both epigean and cave-adapted hypogean populations make Astyanax mexicanus an attractive system for investigating the subterranean evolutionary time necessary for acquisition of the troglomorphic form. Using published sequences, we have estimated divergence times for A. mexicanus using: 1) two different population-level mitochondrial datasets (cytochrome b and NADH dehydrogenase 2) with both strict and relaxed molecular clock methods, and 2) broad phylogenetic approaches combining fossil calibrations and with four nuclear (recombination activating gene, seven in absentia, forkhead, and α-tropomyosin) and two mitochondrial (16S rDNA and cytochrome b) genes. Using these datasets, we have estimated divergence times for three events in the evolutionary history of troglomorphic A. mexicanus populations. First, divergence among cave haplotypes occurred in the Pleistocene, possibly correlating with fluctuating water levels allowing the colonization and subsequent isolation of new subterranean habitats. Second, in one lineage, A. mexicanus cave populations experienced introgressive hybridization events with recent surface populations (0.26-2.0 Ma), possibly also correlated with Pleistocene events. Finally, using divergence times from surface populations in the lineage without evidence of introgression as an estimate, the acquisition of the troglomorphic form in A. mexicanus is younger than 2.2 (fossil calibration estimates) – 5.2 (cytb estimate) Ma (Pliocene).