Tiziana Lai

Mitochondrial DNA reveals genetic structuring of Pinna nobilis across the Mediterranean Sea.

Pinna nobilis is the largest endemic Mediterranean marine bivalve. During past centuries, various human activities have promoted the regression of its populations. As a consequence of stringent standards of protection, demographic expansions are currently reported in many sites. The aim of this study was to provide the first large broad-scale insight into the genetic variability of P. nobilis in the area that encompasses the western Mediterranean, Ionian Sea, and Adriatic Sea marine ecoregions. To accomplish this objective twenty-five populations from this area were surveyed using two mitochondrial DNA markers (COI and 16S). Our dataset was then merged with those obtained in other studies for the Aegean and Tunisian populations (eastern Mediterranean), and statistical analyses (Bayesian model-based clustering, median-joining network, AMOVA, mismatch distribution, Tajima’s and Fu’s neutrality tests and Bayesian skyline plots) were performed. The results revealed genetic divergence among three distinguishable areas: (1) western Mediterranean and Ionian Sea; (2) Adriatic Sea; and (3) Aegean Sea and Tunisian coastal areas. From a conservational point of view, populations from the three genetically divergent groups found may be considered as different management units.

Population structure of the Monocelis lineata (Proseriata, Monocelididae) species complex assessed by phylogenetic analysis of the mitochondrial Cytochrome c Oxidase subunit I (COI) gene

Monocelis lineata consists of a complex of sibling species, widespread in the Mediterranean and Atlantic Ocean. Previous genetic analysis placed in evidence at least four sibling species. Nevertheless, this research was not conclusive enough to fully resolve the complex or to infer the phylogeny/phylogeography of the group. We designed specific primers aiming at obtaining partial sequences of the mtDNA gene Cytochrome c Oxidase subunit I (COI) of M. lineata, and have identified 25 different haplotypes in 32 analyzed individuals. The dendrogram generated by Neighbor-Joining analysis confirmed the differentiation between Atlantic and Mediterranean siblings, as well as the occurrence of at least two Mediterranean sibling species. Thus validated, the method here presented appears as a valuable tool in population genetics and biodiversity surveys on the Monocelis lineata complex.

Combined analysis of four mitochondrial regions allowed the detection of several matrilineal lineages of the lessepsian fish Fistularia commersonii in the Mediterranean Sea

The bluespotted cornetfish ( Fistularia commersonii ) is an Indo-Pacific species that in the last ten years colonized a large part of the Mediterranean basin. The aim of this study was to sequence some portions of the mitochondrial DNA (D-loop II, 16S, 12S and Cyt b) of this fish from different localities of the Mediterranean Sea, in order to evaluate the level of its genetic variability in this area. The genetic analysis performed on specimens from seven localities of Sardinia, Tunisia and Libya revealed the presence of at least five mitochondrial lineages. The results obtained, compared with previous studies, indicate that the use of a sufficient number of mitochondrial regions may allow a more accurate estimate of genetic variability in lessepsian invasions.

Taxonomic distinction of Ophelia barquii and O. bicornis (Annelida: Polychaeta) in the Mediterranean as revealed by ISSR markers and the number of nephridiopores

Ophelia bicornis sensu lato is a polychaete living in intertidal sandy habitats of Mediterranean and European Atlantic coasts, whose systematics have been strongly debated in the past few decades. In the present work the count of nephridiopores was coupled with genetic analysis carried out with DNA markers (inter simple sequence repeats) for a total of 30 individuals collected at six Italian beaches. Exact test, analysis of molecular variance, non-metric multidimensional scaling and assignment tests clearly separated individuals with five nephridiopore pairs from those with six pairs. This finding validated results of a recent allozyme study in which O. bicornis sensu lato was split into O. bicornis sensu stricto (six nephridiopore pairs) and O. barquii (five nephridiopore pairs). This paper represents a further contribution to the estimation of biodiversity within marine invertebrates.

Patterns of spatial genetic structuring in the endangered limpet Patella ferruginea: implications for the conservation of a Mediterranean endemic

Patella ferruginea Gmelin, 1791 is an endangered marine gastropod endemic to the Western Mediterranean. Its range is restricted to the Sardinian-Corsican region (SCR), North Africa, a few scattered sites in Southern Spain, and Sicily. Inter-simple sequence repeat (ISSR) markers and three different mitochondrial DNA (mtDNA) regions, Cytochrome c Oxidase subunit I, 12S (small-subunit ribosomal RNA gene) and 16S (large-subunit ribosomal RNA gene), were used to investigate the presence of genetic population structuring. The mtDNA sequences showed very low levels of genetic differentiation. Conversely, ISSRs showed the presence of two main genetic groups, corresponding to Spain, North Africa and Sicily and the SCR. The SCR was further split into two subgroups. The ISSR results suggest that, on a regional scale, the genetic structure of P. ferruginea is mainly determined by the restriction of gene flow by dispersal barriers. On a more local scale human harvesting may play a crucial role in population structuring by increasing the effect of genetic drift.

PCR-RFLP: a practical method for the identification of specimens of Patella ulyssiponensis s.l. (Gastropoda: Patellidae )

Abstract Patella ulyssiponensis s.l. is widely distributed in the Mediterranean as well as in the north-eastern Atlantic, from southern Norway to north-western Africa, including Macaronesia. Throughout its range, P. ulyssiponensis s.l. shows a high degree of variability in the shape and colour pattern of the shell and in the colour pattern of the foot. In addition, the species demonstrates widely overlapping morphological parameters with the coexisting P. caerulea (in the Mediterranean), P. vulgata (in north-eastern Atlantic) and P. candei (in the Macaronesian islands). Thus P. ulyssiponensis s.l. identification proves to be difficult. So far, method to unequivocally identify uncertain specimens of P. ulyssiponensis s.l. from the coexisting species was using a molecular taxonomy approach based on the Cytochrome c Oxidase subunit I (COI) sequences analysis. The aim of the present study was to identify specific PCR-RFLP (Restriction Fragment Length Polymorphism PCR-based) markers on the COI sequences which could be used as an alternative and fast diagnostic tool for the discrimination of P. ulyssiponensis s.l. from P. caerulea, P. vulgata and P. candei, when the morphology of the coexisting individuals provided equivocal information. First, we sampled 65 Patella spp. and classified these by means of the COI sequencing. Then, we selected the TaqI endonuclease for the specific RFLP reaction, which produced a combination of RFLP electrophoretic bands, allowing an unequivocal identification of the individuals of P. ulyssiponensis s.l.

Patterns of spatial genetic variation in Patella ulyssiponensis: insights from the western Mediterranean marine ecoregion

Patterns of genetic variation in marine species reflect the interplay of species-specific traits, oceanographic features, historical processes and selection. In the Atlantic–Mediterranean regions, Patella ulyssiponensis (Mollusca: Gastropoda) was previously used as a model to investigate these patterns. Our study gained insight into the genetic patterns of P. ulyssiponensis in the western Mediterranean marine ecoregion (WME), by means of ISSRs and COI. We evaluated the genetic structure of the WME with respect to the Atlantic and eastern Mediterranean, as well as the occurrence of further structuring within this ecoregion. Both population- and individual-based analysis evidenced that WME does not appear to be sharply isolated from the adjacent regions. Within the WME, P. ulyssiponensis displays a pattern of genetic structure that may reflect a chaotic patchiness scenario: structuring is neither related to coastal distance nor to other factors that may constrain dispersal. Compared to the congener P. ferruginea, P. ulyssiponensis shows weaker spatial genetic structure, which may reflect a higher dispersal potential coupled with greater population effective size. Whilst other processes that may have influenced the genetic pattern in the WME remain to be cleared, a recent range expansion coupled with species-specific traits favouring larval dispersal may have contributed to the genetic structuring.

Molecular support for morphology‐based family‐rank taxa: The contrasting cases of two families of Proseriata (Platyhelminthes)

Representatives of the Meidiamidae and Otomesostomidae (Platyhelminthes: Proseriata) are seldom encountered, and the monophyly and phylogenetic relationships of these families have never been assessed on molecular basis. Here, we present the first exhaustive molecular study of Proseriata at the family level, including species belonging to the genera Meidiama and Yorknia (Meidiamidae), and Otomesostoma auditivum (Otomesostomidae), using 18S and 28S genes as markers. We performed phylogenetic analyses (Maximum Likelihood [ML] and Bayesian Inference [BI] methods) and species delimitation methods (Single/Multiple Threshold‐Generalized Mixed Yule Coalescent [ST/MT‐GMYC] and Poisson Tree Processes [PTP/bPTP]). The taxon Meidiamidae was not supported, since the type species (Meidiama lutheri) and Meidiama etrusca sp. n. are nested within the Archimonocelididae, formerly restricted to specialized cnidarian feeders. Species belonging to the genus Yorknia resulted genetically well separated from species of Meidiama and from the rest of Archimonocelididae. The new family‐level taxon Yorkniidae fam. n. is thus here introduced, to include the type species of Yorknia (Yorknia aprostatica), and six new species, five of which are formally described here. Otomesostoma auditivum, representative of Otomesostomidae, the only exclusively freshwater taxon of the Proseriata, is the sister taxon of the predominantly marine Apingospermata. This result is not conflictual with the family level attributed to Otomesostomidae on morphological grounds, but it raises speculations on the marine versus freshwater origin of Apingospermata.

Fistularia commersonii (Teleostea: Fistulariidae): walking through the Lessepsian paradox of mitochondrial DNA

Abstract The Mediterranean Lessepsian migrations excite the interest of biologists who are devoted to inferring the effects of selection on the genetic structure of immigrants. The bluespotted cornetfish Fistularia commersonii is an Indo-Pacific species that was first recorded in the Levantine cost of Mediterranean, and within a few years, it rapidly expanded throughout the entire basin. Studies on its genetic variability, performed via mitochondrial sequencing of the Mediterranean specimens, suggest that a limited number of mitochondrial lineages passed through the Suez Canal. However, nuclear markers provide a scenario, with a high genetic variability among the Mediterranean F. commersonii migrants, along with the occurrence of haplotype sharing between the Mediterranean and the Red Sea. The aim of this study was to enlarge the number of Mediterranean sites in order to evaluate if the rapid expansion and different patterns of spread of F. commersonii in the basin could have led to a genetic structuring. The analysis was carried out by sequencing mitochondrial D-loop I in individuals from Sardinia, Sicily, Tunisia, Lampedusa, Libya and Lebanon. Sequences available from previous studies were included in the data set, allowing us to obtain a data set that likely represents the entire distribution range of the species. Results suggest the possible occurrence of two mitochondrial lineages involved in the Mediterranean invasion of F. commersonii, a bottleneck may have caused a loss in the genetic variation, leading to the fixation of specific lineages as an adaptive response to the new environmental conditions.

COI sequencing as tool for the taxonomic attribution of Patella spp. (Gastropoda): the case of morphologically undistinguishable juveniles settled on a Patella ferruginea adult

Patella ferruginea is an endangered marine gastropod, distributed on the western Mediterranean coasts, whose range has progressively contracted, due to intense human exploitation. A genetic analysis was performed on two unidentified young individuals belonging to the genus Patella found attached to the shell of an adult of P. ferruginea , with a twofold aim: (i) to achieve their correct taxonomic attribution by means of the DNA barcoding; and (ii) to shed some light on the hypothesized larval philopatry and/or juveniles phoresis in P. ferruginea . The survey was carried out comparing the sequences of the cytochrome c oxidase subunit I (COI) Folmer region obtained for the two juveniles with those obtained for adults of P. ferruginea , P. caerulea , P. rustica , and P. ulyssiponensis , from different sites of the western Mediterranean, by means of maximum likelihood cluster analysis and a Bayesian-based assignment test. Results obtained evidenced that: (i) COI may be used with confidence as DNA barcoding in the genus Patella ; and (ii) the two juveniles studied are not conspecific: one belonged to P. ferruginea , the other to P. rustica . The latter finding raises doubts about the juvenile phoresis and about the occurrence of larval philopatry in P. ferruginea , suggesting that an extensive use of a molecular approach for a better evaluation of the recruitment features of this endangered species should be adopted.