Mitochondrial DNA phylogeography reveals the existence of an Evolutionarily Significant Unit of the sand goby Pomatoschistus minutus in the Adriatic (Eastern Mediterranean)

The sand goby Pomatoschistus minutus is a major component of marine shelf and estuarine food webs and an important study organism in behavioural research. Yet, despite the sand goby's significance, its past and present patterns of migration and gene flow are poorly understood. Here we use the mtDNA control region and parts of the flanking tRNA genes of 63 fish from six localities in the Adriatic (Eastern Mediterranean), Western Mediterranean, Atlantic, and North Sea to investigate the phylogeography of this gobiid. Phylogenetic analyses and population genetics statistics reveal the existence of an Evolutionarily Significant Unit, sensu Moritz (1994), in the Adriatic and another in the Western Mediterranean, Atlantic, and North Sea. A possible biogeographical scenario for the separation of the ancestral population is that sand gobies in the Adriatic and Western Mediterranean split between 10,000 and 5000 years ago when due to the rise in sea temperature they migrated northwards and were bisected by the Italian peninsula. A testable prediction of this scenario is that sand gobies from the Western Mediterranean, Adriatic, and Aegean form three reciprocally monophyletic groups which are the descendants of a three-way diversification event.     

A new West Balkanian sand-goby (Teleostei: Gobiidae)

A new species of West Balkanian freshwater sand-goby Pomatoschistus montenegrensis sp. nov. (Teleostei: Gobiidae) is described from the Morača River, southern Montenegro, and shown to be related to the euryhaline Adriatic Pomatoschistus canestrinii . The generic status of these two species is discussed with reference to the limits of Pomatoschistus (type Gobius minutus ) and the status of a subgenus [ Ninnigobius (type-species Gobius canestrini )]. The habitat and the local distribution of the new species are described.     

Mitochondrial DNA phylogeography reveals the existence of an Evolutionarily Significant Unit of the sand goby Pomatoschistus minutus in the Adriatic (Eastern Mediterranean)

The sand goby Pomatoschistus minutus is a major component of marine shelf and estuarine food webs and an important study organism in behavioural research. Yet, despite the sand goby’s significance, its past and present patterns of migration and gene flow are poorly understood. Here we use the mtDNA control region and parts of the flanking tRNA genes of 63 fish from six localities in the Adriatic (Eastern Mediterranean), Western Mediterranean, Atlantic, and North Sea to investigate the phylogeography of this gobiid. Phylogenetic analyses and population genetics statistics reveal the existence of an Evolutionarily Significant Unit, sensu Moritz (1994), in the Adriatic and another in the Western Mediterranean, Atlantic, and North Sea. A possible biogeographical scenario for the separation of the ancestral population is that sand gobies in the Adriatic and Western Mediterranean split between 10,000 and 5000 years ago when due to the rise in sea temperature they migrated northwards and were bisected by the Italian peninsula. A testable prediction of this scenario is that sand gobies from the Western Mediterranean, Adriatic, and Aegean form three reciprocally monophyletic groups which are the descendants of a three-way diversification event.     

First record of the Norway goby in the Adriatic Sea

One male and four female Norway gobies Pomatoschistus norvegicus are reported from the Adriatic Sea.     

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.     

Messinian salinity crisis and the origin of freshwater lifestyle in western Mediterranean gobies

The present paper reports on a molecular study based on 12S rRNA and 16S rRNA mitochondrial genes partly sequenced in 13 species of western Mediterranean gobies, three of which are strictly freshwater-dwelling. A total of 867 bp were aligned and used for the phylogenetic reconstruction. Two major lineages were identified, one clustering the sand gobies in a monophyletic clade. Relationships among taxa based on sequence analysis only partly match those based on morphological criteria, suggesting that the latter are somehow insufficient to correctly establish phylogenetic relationships within this family. The results provide evidence for a multiple independent evolution of the freshwater lifestyle in Knipowitschia and Padogobius lineages. On the basis of the present results, it is uncertain whether the freshwater preference within the genus Padogobius originated twice independently in P. nigricans and P. martensii or only once in their common ancestor. Estimation of the ages of the two major lineages of this group of fish with a molecular clock (in combination with the construction of a linearized tree) suggests that they are much older (at least 40 Myr) than previously thought. Thus, there should be no correlation between their diversification and the Miocene-Pliocene geological events, including the so-called Messinian salinity crisis, which occurred about 10 MYA and is believed to have played a role in their evolution. Alternatively, these gobies would have an evolutionary rate at least fourfold faster than those of other vertebrates.     

First report of Gyrodactylus spp. (Platyhelminthes: Monogenea) in the western Mediterranean Sea: molecular and morphological descriptions

The Gyrodactylus spp. fauna on species of gobies, Pomatoschistus, Gobiusculus, and Knipowitschia (Gobiidae: Teleostei), from the western Mediterranean and Adriatic seas is strikingly similar to that found in the Baltic Sea and eastern Atlantic Ocean, both in morphology and in internal transcribed spacer (ITS) rRNA. The fauna consisted of Gyrodactylus branchialis, G. ostendicus, G. gondae, G. rugiensis, G. rugiensoides, and G. arcuatus. No new species have been found. A morphometric comparison between G. branchialis from the Mediterranean and Adriatic seas and its type locality in Ostend (Belgium) showed significant differences in ventral bar and marginal hook features. The morphometric variation was lower in G. rugiensis, whereas no significant differences were found in G. ostendicus. Gyrodactylus branchialis and G. ostendicus collected on P. microps were slightly different in the ITS rDNA (-0.6%) compared with specimens on the closely related P. marmoratus, probably reflecting ongoing speciation. A hybrid zone was identified in the Vaccarès lagoon complex (France) where both host species are sympatric. There was no clear geographic or host-related pattern in the variation found in the ITS2 rDNA in G. arcuatus sampled from P. microps, G. flavescens, Pungitius pungitius, K. panizzae, and its original host Gasterosteus aculeatus (2 polymorphic sites). Of all studied species, only G. arcuatus, G. rugiensis, and G. rugiensoides showed minor intraspecific variation in the ITS rDNA. Hence, the physical separation by a shoreline of more than 10,000 km is hardly reflected in the parasite ITS rDNA.     

Identification of a host-associated species complex using molecular and morphometric analyses,with the description of Gyrodactylus rugiensoides n. sp. (Gyrodactylidae,Monogenea)

Gyrodactylus rugiensis was originally described as a parasite occurring on the marine gobies Pomatoschistus minutus and Pomatoschistus microps. In our preliminary survey this species was also frequently found on Pomatoschistus pictus and Pomatoschistus lozanoi. Subsequent molecular analysis of the internal transcribed spacers rDNA region revealed that this parasite actually represents a complex of two apparently cryptic species, one restricted to P. microps and the other shared by P. minutus, P. lozanoi and P. pictus. Morphometric analyses were conducted on 17 features of the opisthaptoral hard parts of specimens collected from all four host species. Standard discriminant analysis showed a clear separation of both genotypes by significant differences in marginal hook and ventral bar features. Statistical classifiers (linear discriminant analysis and nearest neighbours) resulted in an estimated misclassification rate of 4.7 and 3.1%, respectively. Based on molecular, morphological and statistical analyses a new species, Gyrodactylus rugiensoides is described. This species seems to display a lower host-specificity than generally observed for Gyrodactylus species as it infects three sympatric host species.However, seasonal and host-dependent morphometric variation is shown for G. rugiensoides collected on P. pictus. Host-switching and gene flow might be important factors preventing speciation on closely related and sympatric host species. The presence of host associated species complexes in this Gyrodactylus-Pomatoschistus system is also confirmed by the presence of two host-dependent genotypes within G. micropsi found on P. minutus and P. lozanoi, and P. microps, respectively. By comparing host and parasite phylogeny, phylogenetic and ecological factors influencing host-specificity are discussed.     

Findings of new and rare fish species in the coastal zone of the Crimea (the Black Sea)

Characteristics of seven Atlantic-Mediterranean fish species are provided. Of them, four (gobies—yellow-headed Gobius xanthocephalus and red-mouthed G. cruentatus, incognito blenny Parablennius incognitus, and greater pipefish Syngnathus acus) were either first found in the Black Sea coastal zone of the Crimea, or their findings were doubted, and three species (thin-lipped grey mullet Liza ramada, European barracuda Sphyraena sphyraena, and Black Sea salmon Salmo trutta labrax) are very rare. Over the last decade, the authors recorded 15 fish species; of them, seven Atlantic-Mediterranean species have dispersed along the Crimean coasts by now, one endemic of the far eastern seas (chameleon goby Tridentiger trigonocephalus) has naturalized in Sevastopol Bay, four invading species, freshwater by origin, permanently occur in the estuary zones of the peninsula; the remaining three species are known from single findings and apparently belong to the category of accidental fish. An increase in the species composition of ichthyofauna off the Black Sea coasts of the Crimea results from the on-going process of mediterranization—the invasion in the estuary of freshwater tolerant species at the expense of accidental species that penetrated here either independently or with ballast waters.     

Phylogeography of the planktonic chaetognath Sagitta setosa reveals isolation in European seas

Numerous planktonic species have disjunct distribution patterns in the world's oceans. However, it is unclear whether these are truly unconnected by gene flow, or whether they are composed of morphologically cryptic species. The marine planktonic chaetognath Sagitta setosa Müller has a discontinuous geographic distribution over the continental shelf in the northeastern Atlantic, Mediterranean Sea, and Black Sea. Morphological variation between these populations has been described, but overlaps and is therefore unsuitable to determine the degree of isolation between populations. To test whether disjunct populations are also genetically disjunct, we sequenced a 504-bp fragment of mitochondrial DNA comprising the cytochrome oxidase II region of 86 individuals. Sequences were highly variable; each represented a different haplotype. Within S. setosa, sequence divergence ranged from 0.2 to 8.1% and strong phylogeographic structure was found, with four main groups corresponding to the northeastern Atlantic, Mediterranean Sea (including Ligurian Sea, Tyrrhenian Sea and Gulf of Gabes), Adriatic Sea, and Black Sea. Two of these (Atlantic and Black Sea) were resolved as monophyletic clades, thus gene flow between disjunct populations of S. setosa has been extremely limited and lineage sorting has taken place. The deepest divergence was between Atlantic and Mediterranean/Black Sea populations followed by a split between Mediterranean and Black Sea populations. The Mediterranean/Black Sea clade comprised three groups, with the Adriatic Sea as the most likely sister clade of the Black Sea. These data are consistent with a colonization of the Black Sea from the Mediterranean. Furthermore, a possible cryptic species was found in the Black Sea with 23.1% sequence divergence from S. setosa. Two possibilities for the evolutionary origin of this species are proposed, namely, that it represents a relict species from the ancient Paratethys, or that it represents another chaetognath species that colonized the Black Sea more recently. Even though the exact timing of disjunction of S. setosa populations remains unclear, on the basis of the geological and paleoclimatic history of the European basins and our estimates of net nucleotide divergence, we suggest that disjunct populations arose through vicariance resulting from the cyclical changes in temperature and sea levels during the Pleistocene. We conclude that these populations have remained disjunct, not because of limited dispersal ability, but because of the inability to maintain viable populations in suboptimal, geographically intermediate areas.     

Parasites as indicator of fish behaviour

In the present study two species of gobies, Pomatoschistus microps and P. minutus from southwest Wales were studied and appearance of some unusual parasite of gobies, were used as an indicator for fish behaviour. The occurrence of pleurocercoids of Ligula intestinalis in the body cavity of these fish and Proteocephalus sp., excysted metacercariae and adult of Timoniella spp., excysted metacercariae of Bucephalus baeri , the excysted metacercariae of Podocotyle atomon and progenetic adult of Bucephaloides gracilescens in their gut indicate host anadromous, cannibalistic and scavenging behaviour.     

Molecular phylogeny of the three paedomorphic Mediterranean gobies (Perciformes: Gobiidae)

Pelagic gobies are considered of particular zoological interest because the acquisition of a pelagic lifestyle is achieved through the persistence of larval anatomical features. The Mediterranean Sea is inhabited by three goby species (Aphia minuta, Crystallogobius linearis and Pseudaphya ferreri) characterized by paedomorphic traits. Owing to the shared larval morphological features, these species have generally been considered as a monophyletic group. This study aimed at establishing the phylogenetic relationships of these paedomorphic species within the family Gobiidae to ascertain whether the pelagic lifestyle achieved through paedomorphosis is due to an event that took place in a common ancestor or not. For this purpose, we amplified by polymerase chain reaction and sequenced the mitochondrial 12S rDNA (complete sequence) and 16S rDNA (partial sequence) of 15 Mediterranean gobies. The phylogenetic analysis strongly supported the polyphyletic origin of the three paedomorphic gobies, indicating that the heterochronic change leading to the retention of larval features seems to have occurred independently in the ancestors of these species. Thus, the sharing of morphological traits can be considered homoplasious and the classification of these species in the same taxonomic group is rejected on the basis of molecular data.     

Fish communities along an oxygen-poor salinity gradient (Zeeschelde Estuary, Belgium)

Fish living in the tidal fresh and brackish water reaches of the Zeeschelde Estuary were studied in samples collected from the cooling-water inlets of three power stations. Between July 1994 and June 1995, 42 different species were recorded including 26 marine migrants, 14 freshwater species and two diadromous species. Species number as well as fish abundance were correlated significantly with salinity and oxygen concentration. The community structure was analysed with a correlation biplot based on principal component analysis of the root-root transformed fish abundance. Four separated communities could be distinguished since the first principal component expressed a salinity and the second a temperature gradient. During summer and fall Pomatoschistus microps, P. minutus, P. lozanoi and Syngnathus rostellatus were most abundant in the brackish-water reach. At this time, freshwater species seemed to avoid the freshwater area. During winter, Sprattus sprattus, Clupea harengus and Dicentrarchus labrax were the dominant species of the brackish-water zone while the freshwater reaches were dominated by Gasterosteus aculeatus . Migrating fish such as Pleuronectes flesus, Lampetra fluviatilis, Anguilla anguilla and Osmerus eperlanus were restricted to the brackish environment.     

Sublittoral abundance and food consumption of Baltic gobies

Two small demersal fishes, the sand goby Pomatoschistus minutus and the common goby Pomatoschistus microps , were quantified on soft bottoms at 20–40 m depth in the Baltic Sea, using a camera placed above the bottom. The largest numbers of gobies were seen following the settlement of young in late summer and autumn. Most recorded fishes were sand gobies. An annual average of 4·7 individuals m⁻²(0·24 g dry mass m⁻²) was recorded in 1983–1985 and 2·5 individuals m⁻²(0·13 g m⁻²) in 1997–1998. Using these densities, the annual goby food consumption was estimated to 100 kJ m⁻² in 1983–1985 and 50 kJ m⁻² in 1997–1998, corresponding to most of the annual macrobenthos production available to the gobies. The resulting goby production, assumed equal to 25% of the food consumed, must have been an important food source for the larger fishes occasionally recorded in the photographs.     

Cryptic speciation and paraphyly in the cosmopolitan bryozoan Electra pilosa--impact of the Tethys closing on species evolution

Cosmopolitan nature of the marine bryozoan Electra pilosa was studied to clarify geographic structure and to outline evolution and phylogeography of the species. Several local populations from the Northeast Atlantic (North Sea and Baltic Sea), Arctic (Barents Sea and White Sea) and Indo-West Pacific (New Zealand) were compared. In addition, we examined the closely related species E. posidoniae from the Mediterranean Sea. Phylogenetic analysis based on both 16S and 18S rDNA indicate that the Indo-West Pacific E. pilosa is a sister species to the Atlantic-Mediterranean clade, with the latter including the species E. posidoniae and the Atlantic population of E. pilosa. The topology of the phylogenetic tree leads us to conclude that E. pilosa is a paraphyletic species group relative to E. posidoniae, and a molecular dating of its divergence is consistent to geologic events associated with the closure of the Tethys Sea.     

Rhodopsin population genetics and local adaptation: variable dim-light vision in sand gobies is illuminated

The visual pigments of fish are thought to be adapted to the variable spectral qualities of aquatic light environments. Most research on the role of natural selection on the evolution of rhodopsins and dim-light vision in fish has focused on variation among species and higher taxa. In this issue, Larmuseau et al. reveal substantial intraspecific sequence variation in RH1 (the rhodopsin gene) in sand gobies ( Pomatoschistus minutus ). Using population genetics and molecular evolution approaches, they detect positive selection on RH1 and find evidence for adaptation to local light conditions.     

Evolutionary distinctness of grayling {Thymallus thymallus) inhabiting the Adriatic river system, as based on mtDNA variation

The northern part of the peri-Adriatic river system is inhabited by a native grayling population. In comparison with the Danubian population most of the Adriatic population is regarded as morphologically distinct and therefore recognized as racially differentiated. DNA sequence analysis of the 394-bp fragment of the mtDNA control region and 363-bp fragment of the cytochrome b gene region from grayling specimens of the Adriatic, Danubian and Atlantic river systems revealed nine composite mtDNA haplotypes. The genetic divergence of the Adriatic and the Danubian-Atlantic haplotypes ranged from 2.52 to 3.10% and from 3.38 to 3.92% in the cytochrome b gene and control region, respectively. On the basis of sequence data an attempt was made to elucidate the phylogeography of the Adriatic population. The molecular data indicate an ancient monophyletic origin and also confirm its distinct position within the Thymallus thymallus species, in spite of the fact that stocking with the Danubian type occurred in the Adriatic river system. Therefore the Adriatic population deserves special attention. As a first step in its preservation, its formal distinction by using the name Adriatic grayling is proposed.     

Effect of salinity on cable bacteria species composition and diversity

Cable bacteria (CB) are Desulfobulbaceae that couple sulphide oxidation to oxygen reduction over centimetre distances by mediating electric currents. Recently, it was suggested that the CB clade is composed of two genera, Ca. Electronema and Ca. Electrothrix, with distinct freshwater and marine habitats respectively. However, only a few studies have reported CB from freshwater sediment, making this distinction uncertain. Here, we report novel data to show that salinity is a controlling factor for the diversity and the species composition within CB populations. CB sampled from a freshwater site (salinity 0.3) grouped into Ca. Electronema and could not grow under brackish conditions (salinity 21), whereas CB from a brackish site (salinity 21) grouped into Ca. Electrothrix and decreased by 93% in activity under freshwater conditions. On a regional scale (Baltic Sea), salinity significantly influenced species richness and composition. However, other environmental factors, such as temperature and quantity and quality of organic matter were also important to explain the observed variation. A global survey of 16S rRNA gene amplicon sequencing revealed that the two genera did not co-occur likely because of competitive exclusion and identified a possible third genus.     

Non-concerted ITS evolution, early origin and phylogenetic utility of ITS pseudogenes in Pyrus

Molecular studies of 19 species of the genus Pyrus L. revealed different degrees of intra-individual polymorphism of the internal transcribed spacer (ITS 1, 5.8S rDNA and ITS 2) region due to the existence of putative non-functional copies (pseudogenes), putative recombinants and non-concerted evolution among functional copies. Different types of ITS pseudogenes displaying lower GC content and unstable secondary structure were preferentially amplified under normal PCR conditions. Functional ITS copies were successfully obtained in all investigated accessions under the modified PCR conditions. All pseudogenes were highly divergent from their corresponding functional copies and formed a monophyletic group in the phylogenetic tree based on all paralogs, indicating they were of relatively early origin. Functional ITS copies led to confused and poorly resolved phylogeny as a result of low sequence divergence, existence of unidentified ancient recombinants and a high degree of intra-individual functional ITS polymorphism, while certain types of pseudogenes and some relict pseudogenes offered more credible clues for the evolutionary history of Pyrus species.     

Four new species of Gyrodactylus von Nordmann, 1832 (Monogenea,Gyrodactylidae) on gobiid fishes: combined DNA and morphological analyses

Four Gyrodactylus species parasitising four closely related gobiid species in European coastal waters were studied and compared with G. arcuatus Bychowsky sensu Bychowsky & Poljansky (1953) from Gasterosteus aculatus . These were G. gondae n. sp. from Pomatoschistus minutus and P. lozanoi , G. flavescensis n. sp. from Gobiusculus flavescens , G. arcuatoides n. sp. from P. minutus and G. branchialis n. sp. from P. microps. Combined molecular and morphological analyses, as well as morphometric and statistical methods, were used. The ssrRNA V4 region and the complete ITS rDNA region were sequenced. Genetically the four new species are clearly distinct from G. arcuatus . From a morphological point of view, the haptoral hard parts of G. gondae n. sp., G. flavescensis n. sp. and G. arcuatoides n. sp. are related to those of G. arcuatus, while these parts of G. branchialis n. sp. are different, but related to those of G. quadratidigitus Longshaw, Pursglove & Shinn, 2003. For the latter two species, a new species group is formed. The V4 and ITS sequence analyses, however, indicate a close relationship between G. branchialis and the three G. arcuatus-like species.