Translocation, the movement of organisms for conservation purposes, can result in unintended introgression if genetic material flows between populations in new ways. The Bluemask Darter Etheostoma akatulo is a federally endangered species of freshwater fish inhabiting the Caney Fork River system and three of its tributaries (Collins River, Rocky River, and Cane Creek) in Tennessee. The current conservation strategy for Bluemas­k Darters involves translocating the progeny of broodstock from the Collins River, in the west, to the Calfkiller River, in the east, where the species had been extirpated. In this study, we use ddRAD sequence data from across the extant range to assess this translocation strategy in light of population structure, phylogeny, and demography. We also include museum specimen data to assess morphological variation among extant and extirpated populations. Our analyses reveal substantial genetic and phenotypic disparities between a western population in the Collins River and an eastern population encompassing the Rocky River, Cane Creek, and upper Caney Fork, the two of which shared common ancestry more than 100,000 years ago. Furthermore, morphological analyses classify 12 of 13 Calfkiller River specimens with phenotypes consistent with the eastern population. These results suggest that current translocations perturb the evolutionary boundaries between two delimited populations. Instead, we suggest that repopulating the Calfkiller River using juveniles from the Rocky River could balance conflicting signatures of demography, diversity, and divergence. Beyond conservation, the microgeographic structure of Bluemask Darter populations adds another puzzle to the phylogeography of the hyper diverse freshwater fishes in eastern North America.

Raw ddRAD sequence data (Data/fastqs/*):

We sampled specimens of Etheostoma akatulo from sites among the four tributaries of the Caney Fork River system that span the complete extant range of the species. Samples (collected by J.W.S. under permit from the TVA) were small tissue biopsies from the upper lobe of the caudal fin. To minimize injury and mortality, tissue biopsies were only sampled from individuals with a total length > 30 mm and all specimens were released at their collection site. We also included several previously collected specimens of other species closely related to E. akatulo to serve as outgroups for phylogenetic and molecular divergence time analyses: two specimens of E. stigmaeum (from Crooked Creek, Jefferson County, AL, USA), two specimens of E. meadiae (Copper Creek, Scott County, VA, USA), two specimens of E. jessiae (McNair Creek, Franklin County, KY, USA), and one specimen of E. chlorosoma (Wildcat Creek, Calloway County, KY, USA). We extracted genomic DNA from our ethanol-preserved tissue specimens using the Qiagen DNeasy Blood and Tissue Kit following the manufacturer's protocol (Qiagen, Valencia, CA, USA). After ddRAD library preparation, libraries were sequenced for 100 bp, single-end reads on an Illumina HiSeq2000 at the Genomics & Cell Characterization Core Facility, University of Oregon (Eugene, OR, USA). Specimen metadata is available in the Data/akatulo_genetic_metadata.tsv file.

Morphological data (Data/akatulo_morphology_data.tsv):

Courtesy of Dr. Steve Layman and the Auburn University Museum of Natural History. See Layman, S. R., & Mayden, R. L. (2009). A New Species of the Darter Subgenus Doration (Percidae: Etheostoma) from the Caney Fork River System, Tennessee. Copeia, 2009(1), 157–170.

Instructions on using these files and replicating analyses are available in the README.md file. Each script contains a documented header and in-line documentation.

NOTE that the precise site info, latitude, and longtidue columns have been masked in accordance with Dryad submission protocols for imperiled species (see README_MASKING in Data/ folder).