Genetic characterization of the South American coastal and riverine dolphins Sotalia

Abstract

Dolphins of the genus Sotalia are some of the least studied cetaceans. They are distributed along the Caribbean and Atlantic coast of Central and South America and also have populations found exclusively in freshwater along the Amazon River and in most of its tributaries. The taxonomy of this genus has been controversial and little information is available at present regarding the number of coastal and riverine populations and their genetic structure, or if coastal and riverine forms should be considered separate species. At a higher taxonomic level Sotalia is currently considered a member of the subfamily Stenoninae of the family Delphinidae, along with the genera Steno and Sousa, but the evolutionary relationships among members of the cetacean family Delphinidae are still not well understood. This thesis uses molecular genetics to investigate the genetic diversity, population structure and population boundaries in coastal and riverine Sotalia', to test hypotheses regarding their taxonomic status, including recent suggestions to consider each subspecies as separate species, and to test hypotheses of their divergence and speciation as well as to investigate the phylogenetic relationship between the genus Sotalia and other members of the family Delphinidae. Phylogenetic analyses of two (mtDNA) mitochondrial genes six autosomal introns and four Y chromosome introns (nuclear DNA, nuDNA), positioned Sotalia within a clade containing other members of the Delphininae subfamily that include Sousa, yet separated from Steno. Results suggest that the genus Sotalia should be placed into the subfamily Delphininae rather than Stenoninae. Sousa resolved as sister taxon to Sotalia in all phylogenetic reconstructions using the nuclear dataset, a finding that is not consistent with current subfamily classification. Genetic evidence from ten nuclear (4,312 bp) and three mitochondrial loci (2,096 bp) supported the elevation of each Sotalia subspecies to the species level, Sotalia fluviatilis (riverine species) and Sotalia guianensis (coastal species), under the Genealogical/Lineage Concordance Species Concept and the criterion of irreversible divergence. We also review additional evidence for this taxonomical revision from previously published and unpublished genetic, morphological and ecological studies. A divergence time between Sotalia fluviatilis and Sotalia guianensis of at least 1 million years was estimated using a molecular clock based on the rate of nucleotide substitution on the mtDNA control region. This divergence event is suggested to have happened during the Plio-Pleistocene and it seems to be related to a series of geological changes in the South American continent. The phylogeography and population structure of each of the two Sotalia species was investigated based on samples (S. guianensis n = 55 and S. fluviatilis n = 21) collected across more than 9,000 km of their distribution. A combined analysis of two mitochondrial genes (cytochrome b and control region, 1,052 bp) revealed 31 distinct haplotypes and confirmed reciprocal monophyly of the maternal lineages, confirming phylogenetic analysis from a smaller sample size (above). Within the coastal species, a spatial analysis of molecular variance of the control region sequences showed significant regional population differentiation (Fsr = 0.4; 05T= 0.6; P <0.001). The highest mitochondrial diversity among costal population units was found along the Caribbean Coast of Colombia and Venezuela. The genetic distinctiveness of the Maracaibo Lake (Venezuela) population has conservation implications regarding the threats faced by the animals in this region, including oil exploitation. razilian populations of Sotalia showed the lowest mitochondrial diversity and differentiation among the coastal species warrants further investigation. The Amazonian populations showed the highest haplotype diversity overall and moderate nucleotide diversity, suggesting a surprisingly large effective population size (Nef) and relatively high gene flow throughout the sampled regions of the main river and its tributaries. Suggestions regarding conservation measures required for each species are presented. To further investigate functional diversity, sequence variation at two MHC class II loci (DQA and DQB) in South American coastal and riverine Sotalia was investigated using cloning and direct sequencing. Four DQA and four DQB alleles were resolved in 33 and 32 Sotalia samples, respectively. High average pairwise difference at the nucleotide level was detected among DQB alleles, similar to the findings described for other cetacean species. Positive selection was evidenced at the Peptide-Binding-Region of the DQB locus of Sotalia. The presence of common alleles at both loci in S. fluviatilis and S. guianensis was consistent with trans-species mode of evolution in other mammalian species. There was a lack of apparent reduction of DQA and DQB allelic variation, as well as the presence of one unique DQA allele and one unique DQB allele in the Brazilian Coast population unit by comparison to lower mtDNA diversity. This seems to indicate either present or past balancing selection acting to maintain MHC variation in this population unit. Lastly, future research directions on Sotalia conservation genetics, delphinid systematics and mechanism of adaptation of cetaceans to the freshwater environment are presented, followed by a general hypothesis on the origin of aquatic and terrestrial biodiversity in the Amazon during the Plio-Pleistocene.