DOI: 10.5061/dryad.fj6q573vf
title: Reevaluating claims of ecological speciation in Halichoeres bivittatus

description: Allopatry has traditionally been viewed as the primary driver of
 speciation in marine taxa, but the geography of the marine environment and
 the larval dispersal capabilities of many marine organisms render this
 view somewhat questionable. In marine fishes, one of the earliest and most
 highly cited empirical examples of ecological speciation with gene flow is
 the slippery dick wrasse, Halichoeres bivittatus. Evidence for this
 cryptic or incipient speciation event was primarily in the form of a deep
 divergence in a single mitochondrial locus between the northern and
 southern Gulf of Mexico, combined with a finding that these two haplotypes
 were associated with different habitat types (“tropical” vs.
 “subtropical”) in the Florida Keys and Bermuda, where they overlap. Here
 we examine habitat assortment in the Florida Keys using a broader sampling
 of populations and habitat types than were available for the original
 study. We find no evidence to support the claim that haplotype frequencies
 differ between habitat types, and little evidence to support any
 differences between populations in the Keys. These results undermine
 claims of ecological speciation with gene flow in Halichoeres bivittatus.
 Future claims of this type should be supported by multiple lines of
 evidence that illuminate potential mechanisms and allow researchers to
 rule out alternative explanations for spatial patterns of genetic
 differences.
description: We sampled eight populations in the Florida Keys including four
 populations on the edge of the continental shelf (Sombrero Light, 11 Foot
 Mound, XMuta, and Tennessee Reef), two populations on patch reefs in the
 inshore channel (East Washerwoman and East Turtle Shoal), and two grass
 beds located directly offshore in water &lt; 2m in depth (near mile
 marker 62 on Long Key and behind Keys Marine Lab (KML) on Vaca Key).  For
 broader geographic context we also sampled fishes from two sites further
 north on the gulf coast of Florida, two sites in the Bahamas, and one site
 from Belize.  Florida and Bahamas specimens were collected in 2005 and
 2006, and Belize specimens were collected in 2006.  In addition to
 comparing fore reef and inshore patch reef, we included the grass bed
 habitat as it experiences even greater seasonal and diurnal fluctuations
 in temperature than the inshore patch reef and as such provides an
 additional test of the proposed habitat segregation.
 All animal handling procedures were approved by the University of
 California, Davis Institutional Animal Care and Use Committee.  Fish were
 caught using a combination of hand nets, barrier nets, and otter trawls. 
 Specimens were euthanized using MS-222 dissolved in seawater, and samples
 were taken from muscle tissue and preserved in 95% ethanol.  We extracted
 DNA using DNeasy™ (Qiagen) columns and PCR amplified a fragment of the 
 mitochondrial cytochrome B gene using the L14768 and H15496 primers from
 Rocha et al. (2005).  PCR products were cleaned using ExoSap-IT (USB
 Corp.). Purified templates were dye labeled using BigDye (ABI) and
 sequenced on an ABI 3077 automated DNA Sanger sequencer.  Sequences were
 aligned using ClustalW (Thompson, Gibson, and Higgins 2002).
 Rocha, Luiz A., D. Ross Robertson, Joe Roman, and Brian W. Bowen. 2005. “Ecological Speciation in Tropical Reef Fishes.” <i>Proceedings. Biological Sciences / The Royal Society</i> 272 (1563): 573–79. Thompson, Julie D., Toby J. Gibson, and Des G. Higgins. 2002. “Multiple Sequence Alignment Using ClustalW and ClustalX.” <i>Current Protocols in Bioinformatics / Editoral Board, Andreas D. Baxevanis ... [et Al.]</i> Chapter 2 (August): Unit 2.3.