Genetic and morphological discrimination of species within the nominal Brachidontes exustus (Mollusca: Bivalvia) cryptic species complex from the Florida Keys

The discovery of sibling and cryptic species complexes in the oceans has dramatically increased the estimated number of extant marine species. However, most cryptic species complexes remain taxonomically obscure, lacking descriptions of the morphological or ecological differences defining the species. The distributions and morphologies of species in the nominal Brachidontes exustus complex in the Florida Keys were investigated utilizing molecular and multivariate statistical techniques.;DNA barcoding, a method of comparing newly generated sequences of the mitochondrial cytocrome c oxidase I gene (COI) from specimens of unknown species to a database of known sequences from voucher specimens, identified two cryptic species on Long Key, Florida Keys. Two differing habitats, which were <5 km apart, had single-species populations, even though both locations were within the dispersal range of larval recruits from the other location. This was the first record from the Florida Keys for these species to be encountered as single species populations.;Tests for pseudo-crypsis among three species of the B. exustus morphospecies complex collected throughout the Florida Keys were performed with multivariate morphometics. Specimens were assigned to species using RFLP-based molecular methods. A discriminant function was constructed that, based on shell morphology, assigns individual mussels to a certain species with a high confidence (95%). Morphological differences among the species were sufficient to create robust statistical methods of resolving species using shell morphology alone. The suite of functions will facilitate future manipulation experiments with live specimens. The morphologies of the two most common species, provisionally called Bahamian and Gulf, were more similar in locations of coexistence than in locations of exclusivity.;An improved molecular-based technique for determining species, a multiplex PCR with species-specific forward oligonucleotides, was designed and tested. The method discriminates species by visualization of PCR products after electrophoresis on an agarose gel stained with ethidium brominde. This is a low cost, high throughput method that can effectively screen large numbers of specimens from the entire geographic range of the nominal species. This method can be used to identify species using larvae or juveniles which are unlikely to have the shell differences that can be used in the multivariate morphometric approach.