A Molecular Assessment of Speciation and Evolutionary History of the Globally Distributed Spotted Eagle Ray (Aetobatus narinari)

The spotted eagle ray (Aetobatus narinari), a species of conservation concern (Near Threatened IUCN category) is commonly associated with coral reef ecosystems worldwide where it is likely to play an important predatory role. Currently described as a single, circumglobally distributed species, geographic differences in parasite diversity have led to suggestions that A. narinari may constitute a species complex. There has been no systematic evidence to support this suggestion, however. If multiple species of spotted eagle ray exist, each will likely posses decreased geographic ranges and population sizes, altering the impacts of threats and requiring separate assessments of conservation needs. We assessed the validity of A. narinari as a single cosmopolitan species using 1570bp of sequence data from two mitochondrial genes (cytochrome b and COI) and the nuclear ribosomal ITS2 locus. Individuals from four major geographic regions were examined: western north Atlantic, and western, central, and eastern Pacific. Phylogenies for each locus concordantly described three distinct lineages (western north Atlantic, western/central Pacific, and eastern Pacific) with no genetic exchange among regions. Genetic distances among the most divergent lineages were comparable to taxonomically uncontroversial batoid and teleost congener pairs. Using combined genealogical concordance and genetic distance results, we recommend 1) that the western/central Pacific lineage be recognized as a distinct species from the western north Atlantic and eastern Pacific lineages, and 2) the western north Atlantic and eastern Pacific lineages, separated by the Isthmus of Panama, be given subspecies status. Dramatically higher nucleotide diversity and sequence divergence coupled with a basal position in multiple phylogenetic analyses support an Indo-West Pacific origin for the A. narinari species complex, with subsequent migration into the Atlantic. Evolutionary relationships among lineages suggest a westerly migration around the southern tip of Africa, with intensification of the Benguela coldwater upwelling system a possible vicariant mechanism underlying speciation

Population Structure in The Brown Tube Sponge (agelas Conifera) in The Florida Reef Tract And Caribbean

Sponges are broadly distributed, occur in a wide range of habitats, and comprise a substantial amount of the biomass and macro-biodiversity on coral reefs. Despite their ubiquity in reef ecosystems, their dispersal dynamics are largely unknown. Here we report on gene flow and population structure for the common brown tube sponge, Agelas conifera (Demospongiae, Poecilosclerida) in the Florida reef tract and Caribbean. Over 300 samples collected from ten geographically distinct locations throughout Florida and the Caribbean are being analyzed for variation at eight microsatellite loci. Preliminary results from screening four loci in 295 individuals from eight locations indicated significant population structure overall (FST = 0.072; P = 0.0018) with a significant heterozygosity deficit (FIS = 0.310; P = 0.001). Lower FST values occurred within the Florida reef tract sites compared to those between Caribbean island locations suggesting relatively greater gene flow within the Florida reef system. Significant isolation by distance was not found to occur Caribbean-wide (r = 0.25; P = 0.14), nor within the Florida reef tract (r = 0.39; P = 0.10). Fine scale population structure occurred among sites separated by \u3c1 km around the islands of Utila (Honduras) and St. Croix (USVI) suggesting A. conifera recruitment in these areas is highly local source dependant. In contrast, no structure was observed among sites around Grand Cayman (Cayman Islands). Only populations within St. Croix showed a significant isolation by distance pattern (r = 0.74; P \u3c 0.001). The strong overall and in some instances even highly localized genetic structure observed is consistent with the typically short larval durations known for sponges generally; suggesting recruitment in these important reef biomass components may be driven mainly from local sources

Characterization of the Heart Transcriptome of the White Shark (Carcharodon carcharias)

Background: The white shark (Carcharodon carcharias) is a globally distributed, apex predator possessing physical, physiological, and behavioral traits that have garnered it significant public attention. In addition to interest in the genetic basis of its form and function, as a representative of the oldest extant jawed vertebrate lineage, white sharks are also of conservation concern due to their small population size and threat from overfishing. Despite this, surprisingly little is known about the biology of white sharks, and genomic resources are unavailable. To address this deficit, we combined Roche-454 and Illumina sequencing technologies to characterize the first transcriptome of any tissue for this species. Results: From white shark heart cDNA we generated 665,399 Roche 454 reads (median length 387-bp) that were assembled into 141,626 contigs (mean length 503-bp). We also generated 78,566,588 Illumina reads, which we aligned to the 454 contigs producing 105,014 454/Illumina consensus sequences. To these, we added 3,432 non-singleton 454 contigs. By comparing these sequences to the UniProtKB/Swiss-Prot database we were able to annotate 21,019 translated open reading frames (ORFs) of ≥ 20 amino acids. Of these, 19,277 were additionally assigned Gene Ontology (GO) functional annotations. While acknowledging the limitations of our single tissue transcriptome, Fisher tests showed the white shark transcriptome to be significantly enriched for numerous metabolic GO terms compared to the zebra fish and human transcriptomes, with white shark showing more similarity to human than to zebra fish (i.e. fewer terms were significantly different). We also compared the transcriptome to other available elasmobranch sequences, for signatures of positive selection and identified several genes of putative adaptive significance on the white shark lineage. The white shark transcriptome also contained 8,404 microsatellites (dinucleotide, trinucleotide, or tetranucleotide motifs ≥ five perfect repeats). Detailed characterization of these microsatellites showed that ORFs with trinucleotide repeats, were significantly enriched for transcription regulatory roles and that trinucleotide frequency within ORFs was lower than for a wide range of taxonomic groups including other vertebrates. Conclusion: The white shark heart transcriptome represents a valuable resource for future elasmobranch functional and comparative genomic studies, as well as for population and other biological studies vital for effective conservation of this globally vulnerable species

Transcriptome Derived Microsatellites Demonstrate Strong Genetic Differentiation in Pacific White Sharks

Recent advances in genome-scale sequencing technology have allowed the development of high resolution genetic markers for the study of non-model taxa. In particular, transcriptome sequencing has proven to be highly useful in generating genomic markers for use in population genetic studies, allowing for insight into species connectivity, as well as local adaptive processes as many transcriptome-derived markers are found within or associated with functional genes. Herein, we developed a set of 30 microsatellite markers from a heart transcriptome for the white shark (Carcharodon carcharias), a widely distributed and globally vulnerable marine predator. Using these markers as well as ten published anonymous genomic microsatellite loci, we provide (i) the first nuclear genetic assessment of the cross-Pacific connectivity of white sharks, and (ii) a comparison of the levels of inferred differentiation across microsatellite marker sets (i.e., transcriptome versus anonymous) to assess their respective utility to elucidate the population genetic dynamics of white sharks. Significant (FST = 0.083, P = 0.05; G”ST = 0.200; P = 0.001) genetic differentiation was found between Southwestern Pacific (n = 19) and Northeastern Pacific (n = 20) white sharks, indicating restricted, cross Pacific gene flow in this species. Transcriptome-derived microsatellite marker sets identified much higher (up to 2X) levels of genetic differentiation than anonymous genomic markers, underscoring potential utility of transcriptome markers in identifying subtle population genetic differences within highly vagile, globally distributed marine species

Genetic Connectivity in the Florida Reef System: Comparative Phylogeography of Commensal Invertebrates With Contrasting Reproductive Strategies

Effective spatial management of coral reefs including design of marine protected areas requires an understanding of interpopulation genetic connectivity. We assessed gene flow along 355 km of the Florida reef system and between Florida and Belize in three commensal invertebrates occupying the same host sponge (Callyspongia vaginalis) but displaying contrasting reproductive dispersal strategies: the broadcast-spawning brittle star Ophiothrix lineata and two brooding amphipods Leucothoe kensleyi and Leucothoe ashleyae. Multiple analytical approaches to sequence variation in the mitochondrial COI gene demonstrated a high degree of overall connectivity for all three species along the Florida reef system. Ophiothrix lineata showed significant genetic structuring between Florida and Belize, and a pattern of isolation by distance but no significant genetic structuring along the Florida coastline. Bayesian estimates of migration detected a strong southerly dispersal bias for O. lineata along the Florida reef system, contrary to the general assumption of northerly gene flow in this region based on the direction of the Florida Current. Both amphipods, despite direct development, also showed high gene flow along the Florida reef system. Multiple inferences of long-distance dispersal from a nested clade analysis support the hypothesis that amphipod transport, possibly in detached sponge fragments, could generate the high levels of overall gene flow observed. However, this transport mechanism appears much less effective across deep water as connectivity between Florida and Belize (1072 km) is highly restricted

Contrasting Patterns of Population Structure and Dispersal for the Giant Barrel Sponge (Xestospongia muta) within the Florida Reef Tract and Caribbean

Sponges are one of the dominant fauna on Florida and Caribbean reefs, with species diversity often exceeding that of scleractinian corals. Despite their importance as structural components and habitat providers on reefs, their dispersal dynamics are little understood. We utilized eight microsatellite markers to study the population structure and migration patterns of the giant barrel sponge (Xestospongia muta), a widespread species throughout Florida and the Caribbean. Bayesian multilocus genotype analyses clustered 157 samples from the Bahamas, Honduras, and the US Virgin Islands into three distinct groups. 159 samples from nine locations within 284 km of the Florida reef tract (Key Largo to the Dry Tortugas) formed a fourth group. Population structure among the four groups was high (FST = 0.155; P = 0.001), with no recent migration among the groups. In contrast, high levels of migration were detected within the Florida reef tract. Reefs in the Upper Keys (Long Key) appear to be sources of larvae to reefs in the north (Key Largo) and also to reefs in the south (Key West and the Dry Tortugas). This pattern of migration closely matches current pathways within the South Florida recirculation system, suggesting that currents play an important role in dispersing X. muta larvae within the Florida reef tract. Although there was an overall lack of isolation by distance among the four groups, a significant correlation between genetic and geographic distance was found among the Florida sampling sites indicating that mating within the reef tract is not random. Asexual reproduction appears not to be the cause as only 1.3% of individuals in Florida shared the same genotype (1.6% overall). Rather, limited larval dispersal along the reef tract and among Caribbean locations has probably led to inbreeding within reefs, explaining the significant deficit in heterozygosity detected (FIS = 0.219; P = 0.001)

Genetic Connectivity in The Branching Vase Sponge (callyspongia Vaginalis) Across The Florida Reef Tract And Caribbean

The Porifera constitute a substantial fraction of the biomass on coral reefs and frequently have higher species diversity than corals and algae, making this phylum an important model for the investigation of reef connectivity. We examined genetic connectivity in the common branching vase sponge, Callyspongia vaginalis, by analyzing DNA sequence variation in 511 bp of the mitochondrial cytochrome oxidase I (COI) gene in 401 individuals sampled from 16 locations throughout the Florida reef tract and Caribbean. Populations of Callyspongia vaginalis were highly genetically structured over the study area (ΦST = 0.48, P \u3c 0.0001), including over distances as short as tens of kilometers within the Florida reef tract, and had a significant overall pattern of isolation by distance (P = 0.0002). However, nonsignificant pairwise ΦST values were also found between a few Florida sampling sites suggesting that long distance dispersal, perhaps by means of fragmentation, may occur over continuous, shallow coastlines. Indeed, sufficient gene flow appears to occur along the Florida reef tract to obscure a signal of isolation by distance (P = 0.164), but not to homogenize haplotype frequencies over 465 km from Palm Beach to the Dry Tortugas. Statistical parsimony analysis revealed two highly divergent haplotypes from Honduras suggestive of cryptic speciation. Inferences from a nested clade analysis supported the pattern of restricted gene flow and isolation by distance in the Caribbean, and suggested a northward range extension of C. vaginalis from a hypothesized Central American ancestral population into the Gulf of Mexico and Florida. The extensive genetic structuring in this common reef sponge is consistent with expectations based on typically short sponge larval durations, suggesting that sponge recruitment to coral reefs may be largely local source driven

Enantiomerically Pure [2.2]Paracyclophane-4-thiol: A Planar Chiral Sulfur-Based Building Block Readily Available by Resolution with an Amino Acid Chiral Auxiliary

Acyl chloride of N-phthaloyl-(S)-isoleucine is an efficient chiral auxiliary for the resolution of (+/-)-[2.2]paracyclophane-4-thiol. A preparative protocol, based on the conversion into diastereoisomeric thiolesters and separation by two fractional crystallizations and column chromatography, was developed. Deprotection with LiAlH4 allowed isolation of the individual thiol enantiomers in good yield (similar to 40%) and high enantiomeric purity (ee >93%). The absolute configurations were determined by comparison of the optical rotation value of the products with literature data and were confirmed by X-ray crystallography

Blastocystis Mitochondrial Genomes Appear to Show Multiple Independent Gains and Losses of Start and Stop Codons.

Complete mitochondrion-related organelle (MRO) genomes of several subtypes (STs) of the unicellular stramenopile Blastocystis are presented. Complete conservation of gene content and synteny in gene order is observed across all MRO genomes, comprising 27 protein coding genes, 2 ribosomal RNA genes, and 16 transfer RNA (tRNA) genes. Despite the synteny, differences in the degree of overlap between genes were observed between subtypes and also between isolates within the same subtype. Other notable features include unusual base-pairing mismatches in the predicted secondary structures of some tRNAs. Intriguingly, the rps4 gene in some MRO genomes is missing a start codon and, based on phylogenetic relationships among STs, this loss has happened twice independently. One unidentified open reading frame (orf160) is present in all MRO genomes. However, with the exception of ST4 where the feature has been lost secondarily, orf160 contains variously one or two in-frame stop codons. The overall evidence suggests that both the orf160 and rps4 genes are functional in all STs, but how they are expressed remains unclear