Phylogeographic study of the dwarf oyster, Ostreola stentina, from Morocco, Portugal and Tunisia: evidence of a geographic disjunction with the closely related taxa, Ostrea aupouria and Ostreola equestris

Despite the economic importance of oysters due to the high aquaculture production of several species, the current knowledge of oyster phylogeny and systematics is still fragmentary. In Europe, Ostrea edulis, the European flat oyster, and Ostreola stentina, the Provence oyster or dwarf oyster, are both present along the European and African, Atlantic and Mediterranean, coasts. In order to document the relationship not only between O. stentina and O. edulis, but also with the other Ostrea and Ostreola species, we performed a sequence analysis of the 16S mitochondrial fragment (16S rDNA: the large subunit rRNA-coding gene) and the COI fragment (COI: cytochrome oxidase subunit I). Oysters were sampled from populations in Portugal (two populations), Tunisia (two populations) and Morocco (one population), identified as O. stentina on the basis of shell morphological characters. Our data supported a high degree of differentiation between O. stentina and O. edulis and a close relationship between O. stentina and both Ostrea aupouria (from New Zealand) and Ostreola equestris (from Mexico Gulf/Atlantic). The status of this geographic disjunction between these closely related species is discussed. Furthermore, although identified in a separate genus Ostreola by Harry (Veliger 28:121–158, 1985), our molecular data on O. stentina, together with those available for the other two putative congeneric species, O. equestris and Ostreola conchaphila, would favour incorporation of Ostreola in Ostrea. Finally, a PCR-RFLP approach allowed the rapid identification of O. edulis and O. stentina.     

Low-mitochondrial diversity and lack of structure in the velvet swimming crab <Emphasis Type="Italic">Necora puber</Emphasis> along the Galician coast

The velvet swimming crab Necora puber is a common species along the European Atlantic coasts. Due to its increasing commercial importance, many studies have been carried out to characterize its growth and reproduction, but no genetic assessment has ever been attempted at the population level. Here, we describe the genetic diversity and population structure of N. puber in northwestern Spain (Galicia), including additional samples from France, Portugal, and southern Spain. To do so, we analysed two mitochondrial fragments of the COI and 16S genes in 217 individuals collected from ten localities. Our results unveil low-genetic diversity and weak population structure along the studied range. A range expansion after the last glacial maximum, followed by ongoing gene flow, seems to be the most likely explanation for the observed genetic pattern.     

Mitochondrial evolution and phylogeography in the hydrozoan<Emphasis Type="Italic"> Obelia geniculata</Emphasis> (Cnidaria)

The distribution and genetic structure of many marine invertebrates in the North Atlantic have been influenced by the Pleistocene glaciation, which caused local extinctions followed by recolonization in warmer periods. Mitochondrial DNA markers are typically used to reconstruct species histories. Here, two mitochondrial markers [16S rDNA and cytochrome c oxidase I (COI)] were used to study the evolution of the widely distributed hydrozoan Obelia geniculata (Linnaeus, 1758) from the North Atlantic and the Pacific and, more specifically, in the context of North Atlantic phylogeography. Samples were collected from six geographic localities between 1998 and 2002. Hydroids from the North Atlantic, North Pacific (Japan), and South Pacific (New Zealand) are reciprocally monophyletic and may represent cryptic species. Using portions of the 16S rDNA and COI genes and the date of the last trans-Arctic interchange (3.1–4.1 million years ago), the first calibrated rate of nucleotide substitutions in hydrozoans is presented. Whereas extremely low substitution rates have been reported in other cnidarians, mainly based on anthozoans, substitution rates in O. geniculata are comparable to other invertebrates. Despite a life history that ostensibly permits substantial dispersal, there is apparently considerable genetic differentiation in O. geniculata. Divergence estimates and the presence of unique haplotypes provide evidence for glacial refugia in Iceland and New Brunswick, Canada. A population in Massachusetts, USA, appears to represent a relatively recent colonization event.Communicated by J.P. Grassle, New Brunswick     

Evidence for restricted gene flow over small spatial scales in a marine snapping shrimp <Emphasis Type="Italic">Alpheus angulosus</Emphasis>

Despite the apparent absence of geographic barriers, connectivity among marine populations may be restricted by, for example, ecological or behavioral mechanisms. In such cases, populations may show genetic differentiation even over relatively small spatial scales. Here, mitochondrial sequence data from the cytochrome oxidase I (COI) gene and seven polymorphic microsatellite markers were used to investigate fine geographic scale population genetic structure in the snapping shrimp Alpheus angulosus, a member of the A. armillatus species complex, from collections in Florida, Jamaica, and Puerto Rico carried out from 1999 to 2005. The COI data showed a deep divergence that separated these samples into two mitochondrial clades, but this divergence was not supported by the microsatellite data. The COI data reflect past population divergence not reflected in extant population structure on the whole genome level. The microsatellite data also revealed evidence for moderate population structure between populations as close as ∼10 km, and no evidence for isolation by distance, as divergences between near populations were at least as strong as those between more broadly separated populations. Overall, these data suggest a role for restricted gene flow between populations, though the mechanisms that reduce gene flow in this taxon remain unknown.     

Mitochondrial DNA sequence variation in spiny lobsters: population expansion,panmixia, and divergence

To investigate population differentiation in a comparative and historical context, segments of the mitochondrial cytochrome c oxidase subunit I gene and the control region were sequenced in Panulirus argus from nine sites along approximately 1,500 km of the Northern Caribbean Sea (n = 326) and analyzed with respect to available panulirid data. A mismatch analysis and Fu’s F _S test uncovered a signature of historical population expansion around the time of the Last Glacial Maximum. Significant population structure was not detected in the area. The data supported a hypothesis of panmixia resulting from ongoing larval transport by ocean currents and historical population expansion. Despite high intraspecific divergence levels at COI within Panulirus argus and several other Panulirus species, genetic species identification through DNA barcoding was feasible using either a modified distance threshold or a character-based approach.     

Genome-size variation in bivalve molluscs determined by flow cytometry

Six of the nine described species of the mole crab genus Emerita are distributed in the Americas, two [E. analoga (Stimpson, 1857) and E. rathbunae Schmitt, 1935] on the west coast, and four [E. benedicti Schmitt, 1935, E. brasiliensis Schmitt, 1935, E. portoricensis Schmitt, 1935 and E. talpoida (Say, 1817)] on the east. The presence of an extended planktonic larval stage in all Emerita species suggests high dispersal potential and the possibility of extensive gene flow among conspecific populations. Two taxa were sampled to study the extent of gene flow between widely separated conspecific populations: E. analoga (California and Chile) and E. talpoida (Massachusetts, South Carolina, and the west coast of Florida), while all other taxa were characterized from a single location. Portions of two mitochondrial genes, cytochrome oxidase I (COI) and 16S ribosomal RNA (16S rRNA) were sequenced. For data analysis, approximately 500 bp (COI) and 400 bp (16S rRNA) were examined. Estimated genetic divergence of 5.41% in COI between E. talpoida populations sampled from the Gulf of Mexico and the Atlantic coast, and 3.47% between E. analoga sampled in Chile and California, indicates that in both cases there has been no recent gene flow between disjunct populations. Additional molecular and morphological studies are necessary to decide whether disjunct populations should be accorded specific status. We predict that many marine invertebrates with antitropical distributions similar to E. analoga may consist of sibling species. In contrast to relationships inferred earlier from distribution patterns, parsimony analyses of both COI and 16S rRNA data yield similar phylogenetic trees in which E. analoga is separated from a clade composed of other species in the Americas; a bootstrap value (67%) in the COI inferred tree marginally supports the separation, but the same tree topology with a higher bootstrap value (84%) is obtained with 16S rRNA sequence data. Genetic divergence among the taxa indicates that the Emerita species constitute an old group and that distribution of species has been modified by past climatic and geological events.     

On living in cold water: K-strategies in antarctic benthos

Comparison of the life-histories of two species pairs of caridean decapods, each pair containing a polar and a more temperate-water species (Chorismus antarcticus (Pfeffer, 1887)/Pandalus montagui Leach, 1814 and Notocrangon antarcticus (Pfeffer, 1887)/ crangon crangon (Linnaeus, 1758), suggested that in each case the polar species was more of a K-strategist than was the temperate species. In particular there were striking differences in brood size, egg weight and maturity of the newly hatched larvae. Measurements of individual annual reproductive effort, RE, as g fresh weight eggs per g fresh weight female indicated that in both species pairs the RE of the polar K-strategist was significantly less than that of the comparable temperate water r-strategist. Results expressing RE as g total egg lipid per g fresh weight female were equivocal. These data are discussed in relation to available autecological information for Anterctic marine invertebrates and it is concluded that many features of the polar benthos are explicable in terms of a general evolution of typical K-strategies. The role of low temperature in the widespread evolution of K-strategies may be crucial; consideration of this leads to a re-appraisal of cold-adaptation.     

Phylogeographic patterns of the mysid Mesopodopsis slabberi (Crustacea, Mysida) in Western Europe: evidence for high molecular diversity and cryptic speciation

The phylogeographic patterns among populations of Mesopodopsis slabberi (Crustacea, Mysida), an ecological important mysid species of marine and estuarine habitats, were analysed by means of DNA sequencing of a fragment of the mitochondrial cytochrome c oxidase subunit I (COI) and the 16S ribosomal RNA genes. Samples of M. slabberi collected from five Atlantic and two Western Mediterranean populations were investigated. Very high levels of within-population molecular diversity were observed in all samples (mean h=0.807 and π=0.0083), with exception of the Mediterranean Ebro population which contained only one haplotype. Differentiation among populations was high, and a clear phylogeographic break was observed between the Atlantic and Mediterranean populations. Moreover, a strong differentiation was detected between both populations in the Western Mediterranean Sea (Alicante and Ebro delta), while two divergent lineages occurred in sympatry within the Atlantic Mondego estuary. The high congruence between both the COI and 16S rRNA sequence data, the reciprocal monophyly of the different mitochondrial clades and the levels of nucleotide divergence between them suggest the presence of a complex of cryptic species within M. slabberi. Estimations of divergence time between the different mitochondrial lineages indicate that a split occurred during the late Miocene/early Pliocene. Such a divergence could be concordant with vicariant events during sea-level drops within the Mediterranean region at that time. However, within the Mediterranean Sea, the potential of divergence through ecological diversification cannot be ruled out.     

Mitochondrial cytochrome b variation in sleeper sharks (Squaliformes: Somniosidae)

Sleeper sharks are a poorly studied group of deep-sea sharks. The subgenus, Somniosus, contains three morphologically similar species: S. microcephalus found in the Arctic and North Atlantic; S. pacificus in the North Pacific; and S. antarcticus in the Southern Ocean. These sharks have been reported mainly in temperate to polar waters and occasionally in subtropical locations. They have not been recorded in tropical waters. This study investigates the relationships among the accepted species of Somniosus through analysis of mitochondrial cytochrome b sequence variation. Seventy-five samples were examined from four sampling locations in the North Pacific, Southern Ocean and North Atlantic. Twenty-one haplotypes were found. A minimum spanning parsimony network separated these haplotypes into two divergent clades, an S. microcephalus and an S. pacificus/antarcticus clade, strongly supporting the distinction of S. microcephalus as a separate species from the Pacific sleeper shark species. Analysis of genetic structure within the S. pacificus/antarcticus clade (analysis of molecular variance, allele frequency comparisons, and a nested clade analysis) showed limited or no differences amongst three populations. Further examination of genetic variation at more variable mtDNA and nuclear markers is needed to examine the species status of S. pacificus and S. antarcticus.     

Genetic diversity of Nymphon (Arthropoda: Pycnogonida: Nymphonidae) along the Antarctic Peninsula with a focus on Nymphon australe Hodgson 1902

Sea spiders are conspicuous, and often abundant, members of the Antarctic benthic community. Nymphonidae (Pycnogonida) in Southern Ocean waters comprise over 240 species which are often difficult to assign due to their intraspecific ‘highly variable’ morphology. In particular, Nymphon australe, the numerically dominant species in Antarctic waters is known to have a high level of phenotypic variation in external morphology and is also reported to have a circumpolar distribution. Circumpolarity seems contradictory to the pycnogonid’s brooding lifestyle and presumably limited dispersal. Here we examine the genetic diversity of several Nymphon species collected in the Antarctic Peninsular region. Concomitantly, we assess the genetic structure of N. australe to gain insight into Nymphon dispersal capacity. Cytochrome c oxidase subunit I (COI) and 16S ribosomal gene data suggest a recent common history and/or recent gene-flow of N. australe populations across nearly 800 km of the Antarctic Peninsula. Furthermore, these data support that the Antarctic Peninsula region may hold two previously unrecognized species of Nymphon. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Assimilation efficiency of the antarctic marine isopod Glyptonotus antarcticus

When fed shrimp every 2 days, the isopod Glyptonotus antarcticus Eights assimilated over 90% of the ingested food. Errors in this estimate due to loss of food material during maceration by G. antarcticus were investigated and allowed for. These results are discussed in relation to data for other isopods and marine invertebrates.     

Reproductive isolation among morphotypes of the Atlantic seastar species<Emphasis Type="Italic"> Zoroaster fulgens</Emphasis> (Asteroidea: Echinodermata)

Zoroaster fulgens is a slope-dwelling seastar species that is distributed throughout the Atlantic Ocean. Studies into the population structure and systematics of marine animals have increasingly found that species with a reported cosmopolitan distribution are, in fact, collections of closely related cryptic or sibling species. In the Porcupine Seabight, three morphotypes of Z. fulgens can be found that have a distribution that is stratified by depth. This study investigates the genetic divergence between these morphotypes using sections of the cytochrome oxidase 1 (COI) and 16S regions of the mitochondrial genome. Bathymetrically separated morphotypes of Z. fulgens are reproductively isolated over distances of approximately 1 km while gene flow occurs among morphotypes, along isobaths, over distances of approximately 900 km. Reproductive isolation on the continental slope may have occurred as a result of selection exerted by gradients of depth-correlated physical factors, such as pressure and temperature. However, allopatric speciation with subsequent range expansion may also explain the observed patterns of genetic divergence. Further investigation of radiation within this group may provide important information on the evolution of slope species. Taxonomic revision of the genus is required.Communicated by J.P. Thorpe, Port Erin     

Molecular phylogeny of vestimentiferans collected around Japan, revealed by the nucleotide sequences of mitochondrial DNA

The phylogenetic relationships among local populations of undescribed vestimentiferan species that belong to the genera Lamellibrachia and Escarpia and had been collected at six sites around Japan were analyzed on the basis of the partial (1023 bp) nucleotide sequences of the mitochondrial gene for cytochrome oxidase I (COI), using a pogonophoran and a polychaete as outgroups. The identical amino acid sequence was deduced from the nucleotide sequence obtained from each of the vestimentiferans analyzed. The strong similarity among deduced amino acid sequences of COI suggested a close relationship between vestimentiferans and pogonophorans. On the basis of the phylogenetic relationships, the analyzed vestimentiferan populations were classified tentatively as five species. The genetic differentiation of vestimentiferans was suggested to occur bathymetrically as well as being a consequence of horizontal segregation. Two of these tentatively identified species inhabit both a hydrothermal area and a cold seep area, as is the case for some species of bivalves that belong to the genera Bathymodiolus and Calyptogena. Received: 28 August 1996 / Accepted: 2 October 1996     

RFLP analysis of the mtDNA control region in white shrimp (Litopenaeus vannamei) populations from the eastern Pacific

White shrimp (Litopenaeus vannamei) population genetic structure from the eastern Pacific was determined by restriction fragment length polymorphism analysis of the mitochondrial DNA control region. Four localities were surveyed with four endonucleases (Alu I, Taq I, Spe I, Ssp I) yielding 48 composite haplotypes. White shrimp showed high average within-locality haplotype (0.823) and nucleotide (5.41%) diversities and also high average nucleotide divergence between all pairs of localities (0.73%). A mismatch analysis of pairwise differences between haplotypes indicated that white shrimp does not fit the sudden population expansion model. An analysis of molecular variance showed significant geographic variation in the frequencies of haplotypes (Φ_ST=0.1382, P<0.0001). Population differentiation may be maintained by a combination of physical, oceanographic, and biological factors acting as barriers to gene flow among localities. Because of its high polymorphism, the control region might be useful as a genetic marker for monitoring genetic diversity in aquaculture stocks.     

Genetic evidence for the existence of cryptic species in an endangered clam <Emphasis Type="Italic">Coelomactra antiquata</Emphasis>

Coelomactra antiquata is a commercially important bivalve species, but has been suffering from severe population decline due to over-exploitation and the deterioration of environmental conditions. Previous genetic survey of C. antiquata conducted with allozymes combined with morphology revealed high levels of genetic differentiation between northern and southern populations which suggests a cryptic species might exist in C. antiquata. To test this hypothesis, amplified fragment length polymorphisms (AFLPs) and 16S rRNA gene sequence were used to re-evaluate the spatial genetic structure of six populations of C. antiquata along the coast of China. Both genetic markers display a sharp genetic break between the four northern populations (northern lineage) and two southern population (southern lineage). Large numbers of private alleles (AFLP) were found within the northern or southern populations and a deep divergence of about 6.5% in 16S rRNA gene sequence between the northern and southern lineages suggests the occurrence of potential cryptic or sibling species of C. antiquata. Applying previously published rates of mutation, divergence between the two lineages is estimated to have occurred approximately 3 million years ago and may be due to allopatric isolation during the middle Pliocene times. While no genetic differentiation was found within the northern or southern populations in both AFLP and 16S mtDNA markers, the results indicate that the northern and southern lineage should be managed separately and any translocation between the two areas should be avoided.     

Diversity of free-living marine nematodes (Enoplida) from Baja California assessed by integrative taxonomy

We used morphological and molecular approaches to evaluate the diversity of free-living marine nematodes (order Enoplida) at four coastal sites in the Gulf of California and three on the Pacific coast of Baja California, Mexico. We identified 22 morphological species belonging to six families, of which Thoracostomopsidae and Oncholaimidae were the most diverse. The genus Mesacanthion (Thoracostomopsidae) was the most widespread and diverse. Five allopatric species, genetically and morphologically differentiated, were found in two localities in the Gulf of California (M. sp1 and M. sp2) and three in the Pacific coast (M. sp3, M. sp4 and M. sp5). Overall, we produced 19 and 20 sequences for the 18S and 28S genes, respectively. Neither gene displayed intraspecific polymorphisms, which allowed us to establish that some morphological variation was likely either ontogenetic or due to phenotypic plasticity. Although 18S and 28S phylogenies were topologically congruent (incongruence length difference test, P > 0.05), divergences between species were much higher in the 28S gene. Moreover, this gene possessed a stronger phylogenetic signal to resolve relationships involving Rhabdodemania and Bathylaimus. On the other hand, the close relationship of Pareurystomina (Enchilidiidae) with oncholaimids warrants further study. The 28S sequences (D2D3 domain) may be better suited for DNA barcoding of marine nematodes than those from the 18S rDNA, particularly for differentiating closely related or cryptic species. Finally, our results underline the relevance of adopting an integrative approach encompassing morphological and molecular analyses to improve the assessment of marine nematode diversity and advance their taxonomy.     

Evolutionary relationships of deep-sea mussels inferred by mitochondrial DNA sequences

In order to elucidate the evolutionary process of deep-sea Bathymodiolus mussels, we investigated the phylogenetic relationships of 16 species worldwide by analyzing nucleotide sequences of the mitochondrial COI and ND4 genes. Deep-sea mussels were clustered into three groups by basal trichotomous divergence. The first was composed of four species found in Japanese waters and one species from the Gulf of Mexico, which contain methanotrophic endosymbiotic bacteria. The second included nine species distributed in the West and East Pacific, Indian, and Atlantic Oceans. Members of the second group were trichotomously divided into the Indo-West Pacific, Atlantic, and East Pacific subclusters. The Indo-West Pacific subcluster was composed of three very closely related species with mutual genetic distances at the intraspecific level (av. 0.019 in COI and 0.009 in ND4 relative to av. 0.156 in COI and 0.265 in ND4 among Bathymodiolus species other than Cluster A species), suggesting some gene flow among these species. The third consisted of two West Pacific species. Species in the second and third groups contain mainly thioautotrophic endosymbionts, including some species harboring both methanotrophs and thioautotrophs.     

Low levels of global genetic differentiation and population expansion in the deep-sea teleost <Emphasis Type="Italic">Hoplostethus atlanticus</Emphasis> revealed by mitochondrial DNA sequences

The orange roughy Hoplostethus atlanticus is a well-known commercial species with a global distribution. There is no consensus about levels of connectivity among populations despite a range of techniques having been applied. We used cytochrome c oxidase subunit I (COI) and cytochrome b sequences to study genetic connectivity at a global scale. Pairwise Φ_ST analyses revealed a lack of significant differentiation among samples from New Zealand, Australia, Namibia, and Chile. However, low but significant differentiation (Φ_ST = 0.02–0.13, P < 0.05) was found between two Northeast Atlantic sites and all the other sites with COI. AMOVA and the haplotype genealogy confirmed these results. The prevalent lack of genetic differentiation is probably due to active adult dispersal under the stepping-stone model. Demographic analyses suggested the occurrence of two expansion events during the Pleistocene period.     

Reproductive biology in two species of deep-sea squids

Deep-sea squids, Moroteuthis ingens and Gonatus antarcticus, were collected in the slope waters off the Falkland Islands and their reproductive systems preserved and investigated onshore. Changes in oocyte length-frequencies at maturation and spawning, and their fecundity were studied. These squids, as well as many other species, are characterised by a synchronous oocyte growth and ovulation. Oviducts are not used for ripe egg accumulation and consequently the universal scale of Lipinski (1979) cannot be applied to assign female maturity. M. ingens spawns near the bottom; its fecundity is 168–297 thousand eggs. Maximum egg size is 1.8–2.7 mm. G. antarcticus spawns midwater; its fecundity is 10–25 thousand eggs. Egg size is 3.2–3.3 mm. In M. ingens spawning takes place in the austral autumn and winter, in G. antarcticus—in austral winter. Our data and the literature data show that the so-called “synchronous ovulation” probably occurs in all deepwater squids. This pattern is very rare among fish, but is quite common among benthic octopods that brood their egg masses.     

The trophic ecology of two abundant mesopredators in south-east coastal waters of Tasmania, Australia

Investigating predator–prey relationships is an important component for identifying and understanding the factors that influence the structure and function of ecosystems. Mesopredators, defined as mid-level predators, have a profound effect on ecosystem structure by contributing an important link between apex predators and lower trophic levels. The diet of two elasmobranch mesopredators, Squalus acanthias and Mustelus antarcticus, was investigated in three locations in south-east Tasmania. Squalus acanthias consumed predominantly pelagic teleosts and cephalopods, while M. antarcticus predominantly consumed benthic crustaceans. As a result, there was low dietary and niche overlap between the two species. There was however evidence of intra-specific dietary variations between locations for both the species. This study has contributed to a better understanding of the top-down dynamics of the food web in coastal Tasmania, by providing important dietary information of two abundant mesopredators. In addition, the similar dietary patterns for S. acanthias and other Mustelus species over much of their global range suggest they may be consistent in their trophic roles across systems, with limited competition between these two sympatric mesopredators to be expected.