Allozyme analyses of the genus Trisopterus: taxonomic status and population structure of the poor cod

Genetic analysis of the four Trisopterus (Gadidae) taxa suggests that the interrelationships of the two morphs of poor cod ( T. minutus minutus in the Atlantic and T. minutus capelanus in the Mediterranean) should be reconsidered. The Mediterranean poor cod T. m. capelanus is more closely related to bib T. luscus than to the Atlantic poor cod, so the population structure in the Atlantic and Mediterranean poor cod must be considered separately. Among 635 Atlantic individuals there was some evidence of poor cod population differentiation (allele frequency heterogeneity test P <0·0005; F _ST=0·0135, P ≤0·0005). Levels of genetic variation were similar to those reported for related gadoid species. Some differentiation was present on the Norwegian coast (samples from Trondheimsfjord) and between the Faeroe Islands (Faeroe Bank) and the adjacent European coastal location. In contrast no statistically significant population differentiation was evident in Mediterranean poor cod, but fewer samples and individuals were screened.     

Taxonomic review of the genus Trisopterus(Teleostei: Gadidae) with recognition of the capelan Trisopterus capelanus as a valid species

Trisopterus is demonstrated to be monophyletic, including four distinct species: T. capelanus, T. esmarkii, T. luscus and T. minutus. The capelan T. capelanus is resurrected from the synonymy of poor cod T. minutus, and is shown to be morphologically more similar to T. luscus, to which species it is also more closely related, indicated by a phylogenetic analysis presented here. A lectotype is designated for T. luscus. Trisopterus fasciatus, the type species of Trisopterus, is a junior synonym of T. luscus, and the lectotype of T. luscus is designated as the neotype of T. fasciatus. The lectotype of T. luscus is also designated as the neotype of Gadus barbatus. Gadus barbatus has priority over T. luscus but the name is suppressed by prevailing usage of T. luscus. A neotype is designated also for T. minutus. A phylogenetic analysis using mitochondrial cytochrome b, and a fragment of the nuclear rhodopsin gene, shows that T. capelanus and T. luscus are sister species, and in turn sister to a clade formed by T. minutus and T. esmarkii.     

Evolutionary history of the genus Trisopterus

The group of small poor cods and pouts from the genus Trisopterus, belonging to the Gadidae family, comprises four described benthopelagic species that occur across the North-eastern Atlantic, from the Baltic Sea to the coast of Morocco, and the Mediterranean. Here, we combined molecular data from mitochondrial (cytochrome b) and nuclear (rhodopsin) genes to confirm the taxonomic status of the described species and to disentangle the evolutionary history of the genus. Our analyses supported the monophyly of the genus Trisopterus and confirmed the recently described species Trisopterus capelanus. A relaxed molecular clock analysis estimated an Oligocene origin for the group (∼30 million years ago; mya) indicating this genus as one of the most ancestral within the Gadidae family. The closure and re-opening of the Strait of Gibraltar after the Messinian Salinity Crisis (MSC) probably triggered the speciation process that resulted in the recently described T. capelanus.     

Allozyme variation and genetic inter-relationships between seven flatfish species (Pleuronectiformes)

A total of 11 enzymes coded by 14 loci were assayed for each of 17 populations (from the north east Atlantic and Mediterranean) of seven flatfish species, including representatives of three of the four European families of the order Pleuronectiformes. Diagnostic alleles were observed for each species and there were fixed differences between the species at many loci. Thus all species were genetically distinct, although there were some common alleles. H_obs was higher than the average of a range of fish species and was also higher than that of vertebrate species as a whole. It seems that flatfish as a group may show higher levels of genetic variation than other fish. Values of genetic identity for all pairwise comparisons fell comfortably within the ranges expected. The data support the grouping of plaice and flounder into a single genus, Pleuronectes , but it is concluded that the retention of the dab in a separate genus, Limanda , is justified. A high level of genetic divergence was found between Dover and thickback soles. Genetic divergence data support the hypothesis that Pleuronectes flesus luscus in the Aegean Sea is a distinct subspecies of Pleuronectes flesus (flounder). The data show a clear separation of the Mediterranean Dover sole population from those in the Atlantic. Low genetic divergence was observed between the Aegean Sea and Atlantic brill populations. It is speculated that about 5 Mya the families Pleuronectidae and later Soleidae evolved from the ancestral Scophthalmidae.     

Genetic and morphological divergence between populations of the flatfish Platichthys flesus (L.) (Pleuronectidae)

Populations of the flounder, Platichthys flesus , were screened for electrophoretically detectable protein variation at up to 37 loci. Atlantic and North Sea populations (subspecies flesus ) were genetically very similar to one another ( I >0.99) but different from Adriatic (subspecies italicus ) and Black Sea (subspecies luscus ) populations. The values for genetic identity between subspecies were around 0.9. Diagnostic loci enabled specimens of flesus, italicus and luscus to be differentiated from one another. Samples of flesus showed two to three times the heterozygosity levels of italicus or luscus , consistent with the greater population size of flesus . Morphological comparisons enabled populations to be categorized to subspecies. Platichthys flesus italicus and luscus are valid subspecies: the terms are not synonyms. The genetic distance data provide an estimate of divergence time of the three subspecies at about 2 My ago, an estimate that seems reasonable in the light of present knowledge concerning the recent geological histories of the Mediterranean and Black Sea basins.     

Sex-specific somatic–otolith growth relationship in two Gadidae

Otolith and somatic mass of two Gadidae ( Merlangius merlangus and Trisopterus minutus ) were compared in order to analyse the sex-specific relationship between otolith and somatic growth at age. In the present study, otolith mass appeared a reliable indicator of age in both species. Otolith growth reflected somatic growth, but the relationship between these characters varied and differed between species and sexes.     

New host and locality records for two protozoan (Myxosporea, Coccidia) parasites of marine fishes

This is the first report of the myxosporean Ortholinea orientalis from Atlantic herring Clupea harrengus. It infects the kidney tubules and previously was known from Pacific herring Clupea pallasii and navaga Eleginus navaga in the White Sea and North Pacific Ocean. This is also the first report of the coccidian Eimeria raibauti from Norway pout Trisopterus esmarkii. It infects the epithelium of the pyloric ceca and previously was known only from poorcod Trisopterus minutus in the Mediterranean Sea. The new records are both from the northern North Sea.     

Discord in the family Sparidae (Teleostei): divergent phylogeographical patterns across the Atlantic-Mediterranean divide

The Strait of Gibraltar has been proposed to be the divide between two marine biogeographical regions, the Mediterranean Sea and the Northeast Atlantic. Intraspecific studies have shown, for several of the examined species, a reduction of gene flow between the two basins. The present study examines genetic variation at nuclear and mitochondrial loci in five marine teleost species belonging to the family Sparidae. Four samples for each species were analysed spanning the Northeast Atlantic and the Mediterranean. For all individuals 17 allozyme loci were scored and a combined single strand conformation polymorphism-sequencing approach was used to survey approximately 190 bp of the mitochondrial DNA (mtDNA) D-loop region. All five species share similar biological features. For three species, namely Lithognathus mormyrus, Spondyliosoma cantharus, and Dentex dentex, large mtDNA divergence was observed between Atlantic and Mediterranean samples. Little or no mtDNA differentiation was found in the other two species, Pagrus pagrus and Pagellus bogaraveo. Allozyme data revealed strong differentiation when comparing Atlantic and Mediterranean samples of L. mormyrus and D. dentex, moderate for P. pagrus, and no differentiation for P. bogaraveo and S. cantharus. These results provide evidence for a sharp phylogeographical break (sensu Avise) between the Atlantic and the Mediterranean for two (or possibly three) sparid species of the five investigated. At the same time, the obtained results for the other two species raise the question on which ecological/historical factors might have caused the observed discrepancy in the geographical distribution of genetic variation among otherwise biologically similar species.     

The Atlantic-Mediterranean transition: discordant genetic patterns in two seabream species, Diplodus puntazzo (Cetti) and Diplodus sargus (L.)

Sparids are a group of demersal perciform fish of high commercial value, which have experienced an extensive radiation, particularly in the Mediterranean, where they occupy a variety of different niches. The present study focuses on two species: Diplodus sargus and D. puntazzo, presenting a wide distribution from the Mediterranean to the eastern Atlantic coasts. They display similar ecological behaviour and are evolutionary closely related. Both are highly appreciated in fisheries and D. puntazzo is currently under domestication process. However, little is know on their population structure and it is an open question whether any genetic differentiation exists at the geographic level. To address this issue we examined sequence variation of a portion of the mitochondrial DNA (mtDNA) control region in population samples of each of the two species collected over a wide geographic range. In addition to the mtDNA, analysis of nuclear loci (allozymes) was included in the study to compare patterns revealed by nuclear and mitochondrial markers. The studied samples covered an area from the eastern Mediterranean to the Portuguese coasts immediately outside the Gibraltar Strait. The two species revealed a level of sequence polymorphism remarkably different for the control region with the D. puntazzo and D. sargus showing 111 and 28 haplotypes, respectively. Such a difference was not detected with allozyme markers. The two species also showed large differences in their population structure. While D. puntazzo presented a marked genetic divergence between the Atlantic and Mediterranean samples, D. sargus showed little intraspecific differentiation. These results were supported using both mtDNA and allozyme markers, and were interpreted as the consequence of differences in the history of the two species such as fluctuations in the effective population size due to bottlenecks and expansions, possibly combined with present-day differences in levels of gene flow.     

The genetic population structure of Thunnus thynnus (Linnaeus, 1758) in the Mediterranean Sea,a controversial issue

The Atlantic bluefin tuna (ABFT), Thunnus thynnus (Linnaeus, 1758), is an important commercial species managed as two different stocks, western and eastern Atlantic, with their spawning grounds in the Gulf of Mexico and in the Mediterranean Sea, respectively. The eastern Atlantic stock has been overexploited in the last decades, leading to the application of specific management measures introduced by the International Commission for the Atlantic Tuna (ICCAT). A clear understanding of the genetic structure of ABFT Mediterranean population should be pursued in order to support management decisions. To date the genetic studies on the Mediterranean ABFT, carried out with different molecular markers and sampling procedures, have produced unclear results. Here, we analysed ABFT samples from central and western Mediterranean Sea with mitochondrial sequences and 11 microsatellite loci to investigate, among the others, the area of the Strait of Messina, where environmental conditions seem to support a resident population of ABFT. Furthermore, genetic analyses of mitochondrial sequences were carried out including nucleotide sequences of Adriatic ABFT wild larvae retrieved from GenBank. Among the investigated areas a genetic differentiation was detected between the Strait of Messina and the Tyrrhenian Sea with microsatellite loci according to the exact G test, but not to the Bayesian analyses carried out with STRUCTURE. The analyses with mitochondrial sequences do not reveal any differentiation among sampled areas, however, a highly significant genetic divergence was observed between the Adriatic mitochondrial sequences retrieved from GenBank and the central‐western Mediterranean sequences obtained in the present work. Our results provide some evidence of population structure of Mediterranean ABFT adding pieces to a still unclear picture.     

Cryptic speciation and genetic structure of widely distributed brittle stars (Ophiuroidea) in Europe

Pérez‐Portela, R., Almada, V. & Turon, X. (2012). Cryptic speciation and genetic structure of widely distributed brittle stars (Ophiuroidea) in Europe. —Zoologica Scripta, 00, 000–000. The development of molecular techniques has led to the detection of numerous cases of cryptic speciation within widely distributed marine invertebrate species and important taxonomic revisions in all the major marine taxa. In this study, we analysed a controversial marine species complex in the genus Ophiothrix, a widespread taxon in European waters traditionally assigned to two nominal species, Ophiothrix fragilis and O. quinquemaculata. These species are important components of the rocky shores and deep marine benthos along the North Atlantic and Mediterranean littoral. Their status (including variants of both species) has remained contentious due to overlapping variability in morphological characters. In this study, we analysed the genetic and morphological differences of Ophiothrix lineages along the Atlantic and Mediterranean coasts. We also assessed population genetic structure in the Atlantic and Mediterranean basins by sequencing two mitochondrial genes, the 16S rRNA gene and COI gene, of 221 specimens from 13 locations. Phylogenetic analyses demonstrated the existence of two genetically distinct lineages, attributable to two different species although unrelated to previous taxonomic distinctions. Morphological differences could also be detected between these lineages. Samples from the Northeast Atlantic and one from the deep Mediterranean grouped within Lineage I, whereas Lineage II pooled together the southern Atlantic and rocky shallow Mediterranean samples. In the northern region of the Iberian Peninsula and at a deep locality in the Mediterranean, both lineages overlap. Speciation processes likely happened during the Mio–Pliocene transition (about 4.8–7.5 million years ago), when marine‐level oscillations led to the blockage of major marine corridors in Europe and promoted genetic isolation by vicariance. Secondary contact between lineages following sea‐level increases and recolonization during the refilling of the Mediterranean after the Miocene salinity crisis could explain the present‐day distribution of genetic variability. No barriers to gene flow along the Atlanto‐Mediterranean area were detected for Lineage II, and the lack of genetic structure could be caused by a mixture of several factors, such as wide dispersal potential, recent demographic expansion and large population size.     

Genetic structure and environmental heterogeneity in the European hake (Merluccius merluccius)

This study aimed at assessing the genetic structure and the state of the stocks of the European hake (Merluccius merluccius). To this end, 15 samples were taken from the whole range of the species and analysed using allozymes. Since 11 samples were taken from the poorly studied Mediterranean Sea, the results obtained provided a complete picture of the hake's genetic structure and an initial insight into its relationships with environmental features. Atlantic and Mediterranean hake populations are separated by the Almeria-Oran front. This area has been proved to be the boundary between Atlantic and Mediterranean stocks of many marine organisms, but some doubt exists concerning the efficaciousness of the local gyres as barriers to the gene flow. Our data have evidenced a latitudinal cline at loci Gapdh and Gpi-2 within the Mediterranean Sea, with a further steep change across the Almeria-Oran front. The genetic pattern showed a strong correlation with the values of the salinity both at the surface and at -320 m and of the salinity + temperature at the surface, suggesting a role for these parameters in maintaining the genetic differentiation among the two population groups through selective processes. Finally, the levels of genetic variability were found to be slightly lower in the depleted Atlantic stock than in the Mediterranean one.     

Origin, diversification, and historical biogeography of the genus Trachurus (Perciformes: Carangidae)

We addressed phylogenetic relationships in the genus Trachurus using cytochrome b gene and D-loop sequences. The trees showed five groups: (1) the Southwest Pacific species (T. japonicus, T. novaezelandiae, and T. declivis); (2) The Mediterranean Sea and Eastern Atlantic species (T. mediterraneus); (3) The Atlantic Ocean species (T. lathami and T. trecae); (4) Eastern Atlantic species (T. trachurus and T. capensis); and (5) a group of highly mobile pelagic species, two from the Eastern Pacific (T. symmetricus and T. murphyi) and one from the Eastern Atlantic (T. picturatus). The phylogeny based on Cyt b, supports the molecular clock hypothesis and our results agree with the reported fossil indicating that the origin of this genus occur when the Thetys Sea closed (around 18.4 MYA). In addition, a very slow neutral substitution rate is reported identified only two periods of maximum diversification: the first occurring between 18.4 and 15.0 MYA and the second between 8.4 MYA and present day.     

Current and historic gene flow of the sand goby Pomatoschistus minutus on the European Continental Shelf and in the Mediterranean Sea

Phylogeographical patterns of the sand goby Pomatoschistus minutus (Gobiidae, Teleostei) were studied by means of sequence and single-stranded conformational polymorphism analysis of a 283-bp fragment of the cytochrome b locus of the mtDNA. A total of 228 individuals sampled at 13 sites throughout the species's distributional range revealed a moderate level of diversity and a low but significant level of overall genetic differentiation at all but one site. The goby sample from the Adriatic Sea differed in sequence by approximately 10% from the Atlantic P. minutus and is thought to belong to a cryptic species of the genus Pomatoschistus . Limited genetic differentiation with a weak pattern of isolation-by-distance was recorded throughout the distributional range of the typical P. minutus . Phylogeographical analysis suggested a contiguous range expansion in the Atlantic and Baltic basins during the Eemian and evidence for a glacial refugium in the southern North Sea during the Weichselian. In P. minutus from the western Mediterranean Sea a high number of endemic haplotypes as well as the most common Atlantic haplotype were recorded in appreciable frequencies. This might be explained by secondary contact between different mitochondrial lineages, which evolved in allopatry.   © 2004 The Linnean Society of London, Biological Journal of the Linnean Society , 2004, 83 , 561–576.     

Some aspects of the infection of bib, Trisopterus luscus (L.), by the parasitic copepod Lernaeocera lusci (Basset-Smith, 1896) in Portuguese waters

Winter samples totalling 2308 specimens of Trisopterus luscus (total length range: 13–36 cm) caught in northern Portugal were examined for infections with Lernaeocera lusci. Infection prevalence was of 42%, and infection intensity was 1.7 copepods per infected fish. Individual infection varied between 1 and 9 copepods per infected specimen; 30.6% of the fish had bilateral parasitosis. There was a significant correlation (P<0.01) between fish length and prevalence. Length-weight relationships of uninfected and infected fish were not significantly different (P > 0.05) and thus provide evidence that L. lusci has no effect on the condition of T. luscus in all the examined 1–cm size classes.     

Rapid radiation and cryptic speciation in mediterranean triplefin blennies (Pisces: Tripterygiidae) combining multiple genes

The genus Tripterygion is the unique genus of the family Tripterygiidae in the Mediterranean Sea and in the northeastern Atlantic coast. Three species and four subspecies had been described: Tripterygion tripteronotus and Tripterygion melanurus (T. m. melanurus and T. m. minor) are endemic of the Mediterranean, and T. delaisi (T. d. delaisi and T. d. xanthosoma) is found in both areas. We used five different genes (12S, 16S, tRNA-val, COI, and 18S) to elucidate their taxonomy status and their phylogenetic relationships. We employed different phylogenetic reconstructions that yielded different tree topologies. This discrepancy may be caused by the speciation process making difficult the reconstruction of a highly supported tree. All pair comparisons between these three species showed the same genetic divergence indicating that the speciation process could have been resolved by a rapid radiation event after the Messinian Salinity Crisis (5.2Mya) leading to a trichotomy. Our molecular data revealed two clearly supported clades within T. tripteronotus, whose divergence largely exceeded that found between other fish species, consequently these two groups should be considered two cryptic species diverging 2.75-3.32Mya along the Pliocene glaciations. On the contrary, none of the genes studied supported the existence of two subspecies of T. melanurus. Finally, the two subspecies of T. delaisi were validated and probably originated during the Quaternary climatic fluctuations (1.10-1.23Mya), however their distribution ranges should be redefined.     

Genetic diversity and spatial correlation patterns unravel the biogeographical history of the European sweet vernal grasses (Anthoxanthum L., Poaceae)

Different processes have contributed to shaping the present distribution of the European biotas. Up to three different tertiary- to quaternary-time-scale evolutionary scenarios have been proposed to interpret the divergence and genetic structuring of plant species in Europe. In the present study, the Amplified Fragment Length Polymorphisms technique has been used to unravel the species and regional phylogeography of the European sweet vernal grasses (Anthoxanthum L. Poaceae). Forty-six populations belonging to all seven European species of Anthoxanthum and covering a broad geographical and ecological range were selected. Different phylogeography and population genetics diversity and structure estimates indicated a clear divergence of old Messinian Mediterranean lineages, followed by a pre-Pliocene split between Mediterranean annuals and Eurosiberian perennials and a more recent Pleistocene differentiation of Arctic-Alpine, Atlantic and Submediterranean diploid to polyploid landraces. Regional and population correlation tests between geographical and genetic distances allowed to postulate distinct pre- and post-glacial colonization pathways across Europe for the taxa of this widespread genus.     

Outlier SNP markers reveal fine‐scale genetic structuring across European hake populations (Merluccius merluccius)

Shallow population structure is generally reported for most marine fish and explained as a consequence of high dispersal, connectivity and large population size. Targeted gene analyses and more recently genome‐wide studies have challenged such view, suggesting that adaptive divergence might occur even when neutral markers provide genetic homogeneity across populations. Here, 381 SNPs located in transcribed regions were used to assess large‐ and fine‐scale population structure in the European hake (Merluccius merluccius), a widely distributed demersal species of high priority for the European fishery. Analysis of 850 individuals from 19 locations across the entire distribution range showed evidence for several outlier loci, with significantly higher resolving power. While 299 putatively neutral SNPs confirmed the genetic break between basins (F_CT = 0.016) and weak differentiation within basins, outlier loci revealed a dramatic divergence between Atlantic and Mediterranean populations (F_CT range 0.275–0.705) and fine‐scale significant population structure. Outlier loci separated North Sea and Northern Portugal populations from all other Atlantic samples and revealed a strong differentiation among Western, Central and Eastern Mediterranean geographical samples. Significant correlation of allele frequencies at outlier loci with seawater surface temperature and salinity supported the hypothesis that populations might be adapted to local conditions. Such evidence highlights the importance of integrating information from neutral and adaptive evolutionary patterns towards a better assessment of genetic diversity. Accordingly, the generated outlier SNP data could be used for tackling illegal practices in hake fishing and commercialization as well as to develop explicit spatial models for defining management units and stock boundaries.     

Microsatellites in Aureliana fasciculata var. fasciculata (Solanaceae), a shrub that inhabits the Atlantic Rainforest

? Premise of the study: Microsatellite primers were developed in Aureliana fasciculata var. fasciculata, a shrub that inhabits the Atlantic Rainforest, to investigate the intraspecific genetic structure of the species. ? Methods and Results: Nine pairs of microsatellite primers were designed that were based on a simple sequence repeat (SSR)-enriched library. Seven loci were amplified successfully. Two populations of A. fasciculata var. fasciculata were genotyped and all loci were polymorphic, with two to five alleles per locus per population detected. Cross-species amplification was achieved in other members of the genus and members of the sister genus Athenaea. ? Conclusions: The development of these microsatellite markers will contribute to future population genetic studies in A. fasciculata var. fasciculata. Cross-amplification among other species in the genus and in the sister genus Athenaea might be a useful tool to establish patterns of genetic variability within the Atlantic Rainforest.     

Population Structure and Dispersal Patterns within and between Atlantic and Mediterranean Populations of a Large-Range Pelagic Seabird

Dispersal is critically linked to the demographic and evolutionary trajectories of populations, but in most seabird species it may be difficult to estimate. Using molecular tools, we explored population structure and the spatial dispersal pattern of a highly pelagic but philopatric seabird, the Cory''s shearwater Calonectris diomedea. Microsatellite fragments were analysed from samples collected across almost the entire breeding range of the species. To help disentangle the taxonomic status of the two subspecies described, the Atlantic form C. d. borealis and the Mediterranean form C. d. diomedea, we analysed genetic divergence between subspecies and quantified both historical and recent migration rates between the Mediterranean and Atlantic basins. We also searched for evidence of isolation by distance (IBD) and addressed spatial patterns of gene flow. We found a low genetic structure in the Mediterranean basin. Conversely, strong genetic differentiation appeared in the Atlantic basin. Even if the species was mostly philopatric (97%), results suggest recent dispersal between basins, especially from the Atlantic to the Mediterranean (aprox. 10% of migrants/generation across the last two generations). Long-term gene flow analyses also suggested an historical exchange between basins (about 70 breeders/generation). Spatial analysis of genetic variation indicates that distance is not the main factor in shaping genetic structure in this species. Given our results we recommend gathering more data before concluded whether these taxa should be treated as two species or subspecies.