Developing genomic resources for the common bottlenose dolphin (Tursiops truncatus): isolation and characterization of 153 single nucleotide polymorphisms and 53 genotyping assays.

Although single nucleotide polymorphisms (SNPs) are commonly used in human genetics, they have only recently been incorporated into genetic studies of non-model organisms, including cetaceans. SNPs have several advantages over other molecular markers for studies of population genetics: they are quicker and more straightforward to score, cross-laboratory comparisons of data are less complicated, and they can be used successfully with low-quality DNA. We screened portions of the genome of one of the most abundant cetaceans in U.S. waters, the common bottlenose dolphin (Tursiops truncatus), and identified 153 SNPs resulting in an overall average of one SNP every 463 base pairs. Custom TaqMan(®) Assays were designed for 53 of these SNPs, and their performance was tested by genotyping a set of bottlenose dolphin samples, including some with low-quality DNA. We found that in 19% of the loci examined, the minor allele frequency (MAF) estimated during initial SNP ascertainment using a DNA pool of 10 individuals differed significantly from the final MAF after genotyping over 100 individuals, suggesting caution when making inferences about MAF values based on small data sets. For two assays, we also characterized the basis for unusual clustering patterns to determine whether their data could still be utilized for further genetic studies. Overall results support the use of these SNPs for accurate analysis of both poor and good-quality DNA. We report the first SNP markers and genotyping assays for use in population and conservation genetic studies of bottlenose dolphins.

Restricted dispersal in a continuously distributed marine species: common bottlenose dolphins Tursiops truncatus in coastal waters of the western North Atlantic.

The marine environment provides an opportunity to examine population structure in species with high dispersal capabilities and often no obvious barriers to genetic exchange. In coastal waters of the western North Atlantic, common bottlenose dolphins, Tursiops truncatus, are a highly mobile species with a continuous distribution from New York to Florida. We examine if the highly mobile nature coupled with no obvious geographic barriers to movement in this region result in a large panmictic population. Mitochondrial control region sequences and 18 microsatellite loci indicate dolphins are partitioning the habitat both latitudinally and longitudinally. A minimum of five genetically differentiated populations were identified among 404 samples collected in the range of New Jersey to northern Florida using both genetic marker types, some inhabiting nearshore coastal waters and others utilizing inshore estuarine waters. The genetic results reject the hypothesis of a single stock of coastal bottlenose dolphins put forth after the 1987-1988 epizootic that caused a large-scale die-off of dolphins and suggest instead the disease vector was transferred from one population to the next as a result of seasonal migratory movements of some populations. These coastal Atlantic populations also differ significantly from bottlenose dolphin samples collected in coastal waters of the northern Gulf of Mexico, implying a long-term barrier to movement between the two basins.

Genetic differentiation among recently diverged delphinid taxa determined using AFLP markers.

In the mid-1990s, a new common dolphin species (Delphinus capensis) was defined in the northeast Pacific using morphological characters and mitochondrial DNA (mtDNA) sequences. This species is sympatric with a second species, Delphinus delphis; morphological differences between the two are slight and it is clear they are closely related. Does the phenotypic distinction result from only a few important genes or from large differences between their nuclear genomes? We used amplified fragment length polymorphism (AFLP) markers to broadly survey the nuclear genomes of these two species to examine the levels of nuclear divergence and genetic diversity between them. Furthermore, to create an evolutionary context in which to compare the level of interspecific divergence found between the two Delphinus taxa, we also examined two distinct morphotypes of the bottlenose dolphin (Tursiops truncatus). A nonmetric multidimensional scaling analysis clearly differentiated both Delphinus species, indicating that significant nuclear genetic differentiation has arisen between the species despite their morphological similarity. However, the AFLP data indicated that the two T. truncatus morphotypes exhibit greater divergence than D. capensis and D. delphis, suggesting that they too should be considered different species.

Genetic structure of harbour porpoise Phocoena phocoena populations in the northwest Atlantic based on mitochondrial and nuclear markers.

The harbour porpoise, Phocoena phocoena, experiences high levels of nonnatural mortality owing to interactions with commercial fisheries throughout its range. To accurately evaluate the significance of this bycatch, information on population structure is required. We have examined the population structure of this species in the northwest Atlantic Ocean using mitochondrial DNA (mtDNA) sequence and nuclear microsatellite data. Samples from four previously proposed summer breeding populations--the Gulf of Maine, eastern Newfoundland, the Gulf of St Lawrence and West Greenland--were analysed. Control-region sequences revealed a significant partitioning of genetic variation among most of these summer populations, indicating that northwest Atlantic harbour porpoises should not be considered one panmictic population. Analysis of females alone yielded the highest levels of population subdivision, suggesting that females are more philopatric than males. At least three management units may be defined for harbour porpoises in the northwest Atlantic based on these data. Analysis of six microsatellite loci failed to detect significant population subdivision. Male-mediated gene flow may maintain homogeneity among nuclear loci, while female philopatry is sufficient to produce a signal of population subdivision in the maternally inherited mtDNA genome. mtDNA analyses also indicate that winter aggregations of harbour porpoises along the US mid-Atlantic states comprise animals from more than one summer breeding population.

DNA sequence variation of mitochondrial large-subunit rRNA provides support for a two-subclass organization of the Anthozoa (Cnidaria).

We have sequenced a portion of the mitochondrial 16S rRNA gene from 29 species of Anthozoa, representing six orders of the subclasses Ceriantipatharia, Hexacorallia, and Octocorallia, with the focus on deep-seamount corals (> 500-m depth). We have detected significant length variation in the gene, with homologous gene fragments ranging from 545 bp in a shallow-water scleractinian coral to 911 bp in a deep-sea antipatharian black coral. The aligned sequences were divided into five regions: three high-identity sequence blocks (HSBs) and two highly variable blocks of insertions/deletions (INDELs). Most of the length variation among species occurred as varying numbers of nucleotides in the two INDELs. Little or no intraspecific sequence variation was detected over spatial scales of up to approximately 150 km. Interspecific sequence variation was lowest among the octocorals and greatest among the ceriantipatharians. Our data indicate that the orders Ceriantharia and Antipatharia are highly divergent, and a phylogenetic reconstruction provides support for the two-subclass system of the class Anthozoa (Hexacorallia and Octocorallia).

Phylogenetic relationships among the true porpoises (Cetacea:Phocoenidae).

Portions of the cytochrome b gene and control region of the mitochondrial DNA molecule were sequenced to investigate systematic relationships among the six extant species of true porpoises, (Cetacea: Phocoenidae). Phylogenetic analyses of mitochondrial cytochrome b sequences support a close relationship between Burmeister's porpoise, Phocoena spinipinnis, and the vaquita, Phocoena sinus, and the association of these two species with the spectacled porpoise, Australophocaena dioptrica. The latter result is not in concordance with a recent morphological reclassification which groups A. dioptrica with Dall's porpoise, Phocoenoides dalli, in the subfamily Phocoenoidinae. The molecular analysis found no support for this grouping. A. dioptrica was originally described as a member of the genus Phocoena, and our results support returning it to that genus at this time. Finally, the data suggest that the tropical species Neophocaena phocaenoides, the finless porpoise, may represent the most basal member of the family. The control region sequences corroborated the relationships among the closely related taxa P. sinus, P. spinipinnis, and A. dioptrica, but were unable to resolve the deeper branches of the tree, probably as a result of a high level of saturation of these sequences.