Development of a real-time PCR assay for detection of Mytilus species specific alleles: Application to a sampling survey in Scotland

Shellfish aquaculture is a growing industry in Scotland, dominated by the production of the mussel Mytilus edulis, the native species. Recently the discovery of Mytilus galloprovincialis and Mytilus trossulus together with M. edulis and all 3 hybrids in cultivation in some Scottish sea lochs led to questions regarding the distribution of mussel species in Scotland. The establishment of an extensive sampling survey, involving the collection of mussels at 34 intertidal sites and 10 marinas around Scotland, motivated the development of a high-throughput method for identification of Mytilus alleles from samples. Three Taqman®-MGB probes and one set of primers were designed, based on the previously described Me 15/16 primers targeting the adhesive protein gene sequence, and samples were screened for the presence of M. edulis, M. galloprovincialis and M. trossulus alleles using real-time PCR. Mytilus edulis alleles were identified in samples from all 44 sites. Mytilus galloprovincialis alleles were found together with M. edulis alleles extensively in northern parts of the west and east coasts. Mytilus trossulus alleles were identified in samples from 6 sites in the west and south-west of Scotland. Because M. trossulus is generally undesirable in cultivation and therefore preventing the geographical spread of this species across Scotland is considered beneficial by the shellfish aquaculture industry, these 6 samples were further analysed for genotype frequencies using conventional PCR. Although distribution of the non-native species M. galloprovincialis and M. trossulus have proven to be more widespread than previously thought, there is no evidence from our study of either M. trossulus or M. galloprovincialis acting as an invasive species in Scotland. The real-time PCR method developed in this study has proven to be a rapid and effective tool for the identification of M. edulis, M. galloprovincialis and M. trossulus alleles from samples and should prove useful in future surveys, ecological or aquaculture management related studies in both unispecific and mixed species areas of these species.     

COMPARATIVE ANALYSIS OF KARYOTYPES AND NUCLEOLAR ORGANIZER REGIONS IN DIFFERENT POPULATIONS OF MYTILUS TROSSULUS, MYTILUS EDULIS AND MYTILUS GALLOPROVINCIALIS

The cylotaxonomical characteristics of Mytilus tros-sulus, M.edulis and M. galloprovincialis were studied using karyometricanalysis and silver staining. The karyotype of M. trossulus,reported here for the first time, consists of six metacentricand eight submetacentric-subtelocentric chromosome pairs. Theoccurrence of five metacentric chromosome pairs in M. galloprovincialisdifferentiates this taxon from M. trossulus and M. edulis whichboth have six meta-centrics. The number of submetacentric tosubtelo-centric chromosome pairs was variable between populationsand between taxa. We suggest that this variability has arisenfrom differential chromosome condensation rather than from structuralchange. Intercomparison of chromosome relative length showedstrong similarity among the three taxa. Chromosomal nucleolarorganizer regions (NORs) were identically located on two submetacentric-subtelocentricchromosome pairs in all three taxa. However, M. trossulus showeda characteristic difference in the occurrence of NORs on onemetacentric pair, which was absent in M. galloprovincialis andM. edulis. Variability in the number of NORs per cell was observedin all populations studied. We conclude that the three taxaof Mytilus studied here cannot be differentiated by asinglekaryological character, but that a combination of karyologicalcharacters is virtually diagnostic ^* To whom correspondence should be sent (Received 24 January 1994; accepted 9 March 1994)     

Estuarine habitats protect hybrid mussels from selection

The marine mussels, Mytilus edulis and Mytilus galloprovincialis, form an extensive hybrid zone in Europe where F₂ hybrids and mussels of mixed genetic ancestry are often locally abundant. Hybrid zones are maintained by the interplay of dispersal and selection on hybrid genotypes but there has been vigorous debate on the form of selection that may occur in these systems. Tension zone models argue that selection is against hybrids because of developmental misregulation and is independent of the external environment. Exogenous selection models argue that selection is habitat-dependent and the structure of the hybrid zone is due to the distribution of habitat patches that vary in selection intensity. We test this prediction by comparing the genetic structure of mussel populations in open-coast habitats, where selection on hybrids is strong, to those within two independent estuaries. We show that mussels within these estuaries are protected from selection and thus selection is strongly dependent on habitat, which supports the exogenous selection hypothesis. Hybrid mussel populations on the open-coast experience strong selection against M. edulis-like genotypes, which has been postulated to be the result of differential dislodgment by waves. This hypothesis is supported by our results since mussels within sheltered habitats are protected from selection. There was, despite previous suggestions, no evidence of selection in favor of M. edulis-like mussels within either estuary.     

Preliminary data on the genetic differentiation of Mytilus desolationis Lamy 1936 and Aulacomya ater regia Powell 1957 (Bivalvia,mytilidae) in the Kerguelen Islands (Terres Australes et Antarctiques Françaises)

Summary Mussel populations in the Kerguelen Islands constitute an excellent model in which to study genetic differentiation, because of the exceptional geographic position of the islands with consequent very low external gene flow, the variety of biotopes that they occupy and their abundance there. Enzyme polymorphism and the electrophoretic variability of some loci have been studied in four populations of Mytilus desolationis Lamy 1936 and in four other populations of Aulacomya ater regia Powell 1957. In both species 67% of the loci were polymorphic. The electrophoretic variability of ACP-2, LAP-1, LAP-2, PGI and PGM in M. desolationis populations, and of ACP-2, LAP-1, LAP-2 and MDH in A. ater regia populations was analyzed by correspondence analysis. For M. desolationis the four populations studied were found to be heterogeneous; four loci contribute clearly to this differentiation. The populations from sheltered and exposed areas were distinguishable on the basis of the allelic frequencies of LAP-1 and LAP-2. The allelic frequencies of PGI and PGM allow further differentiation of the populations. For A. ater regia the four populations studied were distinguishable on the basis of the allele frequencies of ACP-2 and LAP-1 and the differences could not actually be related to their environmental conditions.     

Cadmium induction of metallothionein synthesis in Mytilus galloprovincialis

1. The accumulation and subcellular distribution of cadmium and resulting induction of metallothionein has been studied in mussels, Mytilus galloprovincialis, exposed to 400 μg Cd 1⁻¹.2. Cadmium accumulated by mussels was primarily associated with metallothionein: a maximum four-fold increase in concentrations of the metal-binding protein (3 mg g⁻¹ to 12 mg g⁻¹) was observed after 30 days.3. Results for M. galloprovincialis confirm the importance of metallothionein in the detoxification of intracellular cadmium and indicate that induction rates are identical to those observed in the closely-related M. edulis.4. Intercomposition of data between these species may therefore be valid when employing metallothionein assays to determine sub-lethal response to cadmium contamination.     

Competition and facilitation between the alien mussel Mytilus galloprovincialis and indigenous species: Moderation by wave action

As the alien species that most dominates space along the South African coast, the Mediterranean mussel Mytilus galloprovincialis has radically altered community composition on invaded shores. We experimentally assessed interspecific interactions between this invasive species and dominant indigenous species in conjunction with considering how wave action moderates such interactions. The density of both M. galloprovincialis and the limpet Scutellastra granularis increased with wave action. Conversely, the tube-building polycheate Gunnarea capensis was negatively affected by wave exposure, being most abundant on sheltered shores. The influence of wave action on the indigenous mussel Aulacomya ater, however, remains unclear. M. galloprovincialis outcompeted both G. capensis and A. ater at moderate to high exposure levels, whereas it had both positive and negative effects on S. granularis. It outcompeted adult limpets on primary rock space on semi-exposed and exposed shores, reducing densities of this portion of the population. However, recruitment of S. granularis was facilitated by M. galloprovincialis, as greater numbers recruited to the secondary substratum offered by mussel shells. Again this interaction intensified with wave action. Due to the extremely high density of recruits on secondary space, the net effect of M. galloprovincialis on S. granularis was positive. Thus, wave action not only influences the abundance of individual species, but also mediates both positive and negative interspecific interactions in rocky shore communities, including the impact of alien species such as M. galloprovincialis.     

Influence of genotype and geography on shell shape and morphometric trait variation among North Atlantic blue mussel (Mytilus spp.) populations

The influence of geography and genotype on shell shape (outline) and trait (morphometric) variation among North Atlantic blue mussels and their hybrids has been examined. Shape differences among reference taxa (Mytilus trossulus, Mytilus edulis and Mytilus galloprovincialis) were consistent with an association between taxon‐specific genes and shape genes. Newfoundland M. edulis × M. trossulus populations and northern Quebec M. trossulus populations exhibited an uncoupling of taxon‐specific genes from shape genes, whereas Nova Scotia M. trossulus populations and SW England M. edulis × M. galloprovincialis populations exhibited an association between taxon‐specific genes and shape genes. We found no evidence of a geographic effect (NE versus NW Atlantic) for shape variation, indicating that the genotype effect is stronger than any geographic effect at macrogeographic scales. Pronounced differences were observed in trait variability consistent with an association between taxon‐specific genes and trait genes in European populations, and trait divergence of New York M. edulis from all European mussels. Trait variability in mussels from Newfoundland, Nova Scotia and northern Quebec indicated an uncoupling of taxon genes from trait genes, whereas trait variability in SW England M. edulis × M. galloprovincialis populations was consistent with background genotype, indicating a strong association between taxon genes and trait genes. A pronounced macrogeographic split (NE versus NW Atlantic) regardless of taxonomy was observed, indicating that geography exerts a greater influence than genotype on trait variation at the macrogeographic scale. This is consistent with pronounced within‐taxon genetic divergence, indicative of different selection regimes or more likely of different evolutionary histories of mussels on either side of the North Atlantic. © 2009 The Linnean Society of London, Biological Journal of the Linnean Society, 2009, 96 , 875–897.     

Larval settlement can explain the adult distribution of Mytilus californianus Conrad but not of M. galloprovincialis Lamarck or M. trossulus Gould in Moss Landing, central California: Evidence from genetic identification of spat

We investigated the spatial distribution of adult and newly settled mussels (Mytilus galloprovincialis Lamarck, Mytilus trossulus Gould and Mytilus californianus Conrad) on the shore at Moss Landing, California to test the hypothesis that adult distributions are a result of settlement patterns. Adult M. californianus were most abundant on a wave-exposed rocky jetty and adults of Blue mussels (M. trossulus and M. galloprovincialis) were more abundant inside the protected Moss Landing harbor. Using taxon-specific polymerase chain reactions, we monitored recruitment during continuous 1-2 week intervals on fibrous scrubbing pads for 12 months in 2002-2003. All mussel species settled in greatest numbers on the exposed jetty, and Blue mussels settled in greater numbers there than did M. californianus. Because Blue mussels settled abundantly where their adults were rare, post-settlement mortality appeared to be the strongest influence on adult distribution. In contrast, M. californianus settled mostly in their adult habitat.     

ASYMMETRIC INTROGRESSION OF MITOCHONDRIAL DNA AMONG EUROPEAN POPULATIONS OF BLUE MUSSELS (MYTILUS SPP.)

Abstract.—Mytilus edulis and M. galloprovincialis are two blue mussel species that coexist in western Europe. Previously, we reported that M. galloprovincialis populations contain female and male haplotypes that are fixed in M. edulis populations as well as unique haplotypes. This study assesses whether paraphyly for these species is due to introgression or incomplete lineage extinction. The lineage extinction hypothesis predicts that the shared mtDNA haplotypes in M. galloprovincialis will be significantly diverged from those in M. edulis and form distinct sequence clades. In contrast, the introgression hypothesis proposes that M. edulis haplotypes have only recently been introduced into M. galloprovincialis through hybridization with relatively little divergence accumulating between the shared RFLP haplotypes. We examined 80 mtl6S gene sequences for both the maternal and paternal mtDNA lineages from mussels sampled from various European populations and found strong support for the introgression hypothesis. In addition, we found that M. edulis mtDNA haplotypes appear to be introgressing into mussel populations in the Baltic Sea, which have predominantly M. trossulus nuclear genotypes, indicating that introgressive hybridization is prevalent among European mussel populations.     

Aspects of the population genetics of Mytilus (Mytilidae; Mollusca) in the British Isles

Starch gel electrophoresis was used to study variation at 11 loci in mussels sampled mainly from British coastal sites. Two types of mussel were identified, Mytilus edulis, the common mussel and its southern relative Mytilus galloprovincialis. Several partially diagnostic loci were used to map the distribution of the two forms. Mytilus edulis was present at all sites sampled in Britain and Ireland but was at low frequency in SW England; M. galloprovincialis was detected in SW England, the south and west of Ireland. Scotland and NE England, but was absent from south Wales, the Irish sea coasts of Wales and Ireland, and SE England. Apart from the occurrence of M. galloprovincialis in NE England, this distribution conforms with the results of studies using morphological characters and parallels the distribution of many other southern species in Britain. At the microgeographical level, M. edulis was found to prefer more sheltered and estuarine conditions than M. galloprovincialis. Analysis using the best diagnostic loci showed that hybridization is occurring between M. edulis and M. galloprovincialis at all localities where they occur sympatrically but that the extent of hybridization varies considerably between localities. The distribution of localities having high proportions of hybrid individuals is best interpreted by assuming that hybrids have higher fitness than parental types at these localities. A study was made of variation within and between those localities where only M. edulis individuals were observed. Little significant geographic variation in allele frequency was detected, but significant deficits of heterozygotes compared with Hardy-Weinberg expectations were seen for most loci. Analysis suggests that the Wahlund effect is not involved and that the most likely cause of the deficit is low frequencies of null alleles. In M. edulis no differences in phenotypic variance in shell height and width were observed between samples of multiply heterozygous and multiply homozygous individuals and no genetic differences were found between juveniles and adults. Overall little evidence was found that balancing selection is responsible for maintenance of the polymorphisms studied in M. edulis. The pattern of geographic variation in gene frequencies in Mytilus in the British Isles is discussed in relation to variation in the south and north of Europe and North America. It is concluded that steep clines in gene frequencies in M. edulis observed by other workers in the Baltic and in Long Island Sound cannot be attributed to the presence of M. galloprovincialis.     

Bioinvasion threatens the genetic integrity of native diversity and a natural hybrid zone: smooth‐shelled blue mussels (Mytilus spp.) in the Strait of Magellan

Smooth‐shelled blue mussels of the Mytilus edulis species complex are widely distributed bivalve molluscs whose introductions threaten native marine biodiversity (non‐indigenous species – NIS). The aim of the present study was to identify the species and hybrids of Mytilus present in the Magellan Region (southern Chile). Results indicate that three mussel species of the Mytilus edulis complex are found in the region – M. edulis, M. chilensis (or the Southern Hemisphere lineage of Mytilus galloprovincialis), and M. galloprovincialis of Northern Hemisphere origin. For the first time, alleles of the introduced M. trossulus are reported from the Southern Hemisphere. In the Strait of Magellan the native Pacific blue mussel, Mytilus chilensis and the native Atlantic blue mussel, Mytilus edulis, meet and mix at a natural hybrid zone (about 125 km in length). This is the first record of a natural Mytilus hybrid zone in the Southern Hemisphere and is also the first record of the co‐occurrence of genes from all four Mytilus species in any one region. These results contribute to the knowledge of the biodiversity and delimitation of mussel species in southern South America, and highlight how introduced species may threaten the genetic integrity of native species through hybridization and introgression.     

Intertidal zonation of prosobranch gastropods: Analysis of densities of four co-existing species

The densities of populations of Nerita atramentosa Reeve, Austrocochlea constricta Lamarck, Bembiciuin nanum (Lamarck), and Cellana tramoserica (Sowerby) were recorded in 0.25 m² quadrats on landward and seaward halves of a sheltered and a moderately exposed rock platform from June 1972 to June 1973. The data have been subjected to variance and multiple regression analysis. Differences between densities of each species on the two shores and in the two areas of each shore are discussed with respect to the regressions on littoral height. Densities of Nerita on the sheltered shore, Austrocochlea and Bembicium on both shores, and Cellana, on the exposed shore, increased with height. The density of Cellana on the sheltered shore decreased as height increased. The density differences in the landward and seaward areas on the two shores were attributable to the effect of height for Nerita and Austrocochlea. Height was the major variable affecting the density of these two species on the sheltered shore. Bembicium and Cellana densities on both shores, and Austrocochlea density on the exposed shore, are not primarily determined by height.On the sheltered shore, Nerita and Cellana increased in density with increasing cover of water in rock pools. On the exposed shore, Bembicium and Cellana became sparser with increasing water cover. Bembicium on both shores increased in density with increasing cover of the alga Peyssonelia, as did Cellana on the exposed shore. Cellana and Austrocochlea tended to increase in density with Nerita on the sheltered shore. Cellana tended to be sparse where Bembicium was dense on the exposed shore. The biological effects of substrata and other species on the densities of each species remain unexplained.Significant time effects were found for variations in the densities of Nerita and Cellana on the sheltered shore and of Austrocochlea and Cellana on the exposed shore. The seasonal change in density of Nerita was parabolic with a minimum in summer, and is consistent with the known period of recruitment of this species. The lack of significant time effects on density of Austrocochlea on the sheltered shore is consistent with the known continuous recruitment of this species. The declining density of Austrocochlea on the moderately exposed shore was found, by size-frequency analysis, to be due to reduced recruitment compared with surrounding shores. The density of Cellana on both shores showed seasonal changes, with a maximum in summer, which is consistent with the known period of recruitment in Cellana. The lack of seasonal changes in density of Bembicium is briefly discussed.This type of multifactor analysis identifies biological interactions affecting variation in density of these species. It also corroborates the results of other ecological investigations and determines the relative importance of a variety of physical and biological factors in the distribution of density of intertidal gastropods.     

Identification of Mytilus spp. and Pecten maximus in Irish Waters by Standard PCR of the 18S rDNA Gene and Multiplex PCR of the 16S rDNA Gene

Two molecular protocols for the identification of mussel and scallop have been developed using specific primers targeting the mitochondrial 16S ribosomal DNA gene and the nuclear 18S ribosomal DNA gene. Primers for the mitochondrial 16S ribosomal DNA gene in multiplex polymerase chain reaction (PCR) protocols yielded diagnostic DNA fragments for the mussels Mytilus edulis, Mytilus galloprovincialis, and the hybrid Mytilus edulis/galloprovincialis (335 bp), the king scallop Pecten maximus (382 bp) and the black scallop Mimachlamys varia (398 bp). DNA from the queen scallop Aequipecten opercularis showed no consistent PCR amplification of the 16S rDNA gene. Primers for the nuclear 18S rDNA gene in standard PCR protocols yielded similar-sized, diagnostic DNA fragments (approx. 190 bp) for the mussels Mytilus edulis, Mytilus galloprovincialis, and the hybrid Mytilus edulis/galloprovincialis, the king scallop Pecten maximus, the black scallop Mimachlamys varia, and the queen scallop Aequipecten opercularis. Both protocols have been tested with Mytilus spp., P. maximus, and 6 other bivalve species from a wide range of locations in Irish and European waters. Cross reaction of the specific primers with DNA template from any of the 6 other bivalve species was not observed. Rapid DNA extraction using FTA Card technology and the16S rDNA primers allowed for the detection of at least 10 mussel larvae in a subsample of natural plankton.     

A new gene family of single fibrinogen domain lectins in Mytilus

In molluscs haemolymph lectins bearing ?brinogen-like domain (FREP) act as immune pattern-recognition receptors. A full-length cDNAs of MytFREP1 and MytFREP2 cloned from haemocytes of blue mussel Mytilus edulis encoded putative polypeptides of 230 and 241 amino acids. Both polypeptides consist of signal peptide and C-terminal fibrinogen-like domain. Immune functions of these molecules may be extrapolated from the close-related and functionally characterized lectin AiFREP from bay scallop, Argopecten irradians. However, immune challenge experiments with zymosan particles, Escherichia coli bacterium and cercariae of Himasthla elongata (Trematoda) failed to modulate MytFREP1 and MytFREP2 mRNA expression in M. edulis haemocytes. Hypothetically, it argues into rather high specificity of mechanisms triggering a differential expression of MytFREP genes. The search in the EST database revealed orthologous copies for described genes and portion of relatively similar genes from two close-related mytilids, Mytilus galloprovincialis and Mytilus californianus. We document the new multigene family of FREPs from bivalves of genus Mytilus. MytFREP family currently represented by 2 genes from M. edulis, 4 genes from M. californianus and 7 genes from M. galloprovincialis.     

Differential introgression from a sister species explains high FST outlier loci within a mussel species

Scanning genomes for loci with high levels of population differentiation has become a standard of population genetics. F_ST outlier loci are most often interpreted as signatures of local selection, but outliers might arise for many other reasons too often left unexplored. Here, we tried to identify further the history and genetic basis underlying strong differentiation at F_ST outlier loci in a marine mussel. A genome scan of genetic differentiation has been conducted between Atlantic and Mediterranean populations of Mytilus galloprovincialis. The differentiation was low overall (F_ST = 0.03), but seven loci (2%) were strong F_ST outliers. We then analysed DNA sequence polymorphism at two outlier loci. The genetic structure proved to be the consequence of differential introgression of alleles from the sister‐hybridizing species Mytilus edulis. Surprisingly, the Mediterranean population was the most introgressed at these two loci, although the contact zone between the two species is nowadays localized along the Atlantic coasts of France and the British Isles. A historical contact between M. edulis and Mediterranean M. galloprovincialis should have happened during glacial periods. It proved difficult to disentangle two hypotheses: (i) introgression was adaptive, implying edulis alleles have been favoured in Mediterranean populations, or (ii) the genetic architecture of the barrier to edulis gene flow is different between the two M. galloprovincialis backgrounds. Five of the seven outliers between M. galloprovincialis populations were also outliers between M. edulis and Atlantic M. galloprovincialis, which would support the latter hypothesis. Differential introgression across semi‐permeable barriers to gene flow is a neglected scenario to interpret outlying loci that may prove more widespread than anticipated.     

Mytilus galloprovincialis Lmk. in Southern Africa

The Mediterranean mussel, Mytilus galloprovincialis Lmk. has been identified on the west coast of southern Africa using morphological and biochemical-genetic comparisons with samples of “pure” M. edulis L. from Denmark and M. galloprovincialis from the Mediterranean coast of Spain. Our results show that the southern African Mytilus more closely resembles M. galloprovincialis in anterior adductor, posterior adductor and hinge size. It also has a more ventrally located maximal shell width like the Mediterranean mussel. The electrophoretic examination of 23 proteins showed that many alleles diagnostic for M. galloprovincialis appeared in the southern African Mytilus. Nei's genetic distance between the southern African Mytilus and M. galloprovincialis was 0.010 ± 0.004 and the average genetic distance between these samples and M. edulis was 0.162 ± 0.077.The presence of M. galloprovincialis in southern Africa could be explained by biogeographic dispersal along the west coast of Africa during Pleistocene cooling, or by a recent inadvertant or intentional introduction by man. To test these two hypotheses, we examined ≈750 mussel shells collected from Koi-san middens and ≈350 shells from a raised-beach deposit all located on the west coast of Cape Province, South Africa. The ages of these deposits range from 1500 yr to 120000 yr and pre-date the arrival of Europeans to the Cape. This examination did not reveal any shells of Mytilus. We, therefore, conclude that the introduction of M. galloprovincialis to southern Africa was a recent event. We suggest that, since the first confirmed reference to Mytilus in southern Africa was made from a 1972 collection, the introduction occurred within the last two decades.     

Hybridisations between Mytilus edulis and Mytilus galloprovincialis and performance of pure species and hybrid veliger larvae at different temperatures

The mussels Mytilus edulis L. and Mytilus galloprovincialis Lamark hybridise naturally in the wild along the Atlantic coast of Europe producing a patchwork of mixed pure species and hybrid populations. Individuals of both species were spawned in the laboratory and were hybridised in a series of reciprocal crosses. After 72 h, the proportion of eggs which developed into larvae (%yield) and the proportion of those larvae which had a normal veliger morphology (%normality) were estimated and compared between pure species and hybrid families. There were no significant differences in %yield or %normality between pure species and hybrids, but significant differences were evident between the offspring from different parents irrespective of whether they were hybrids or pure species. Therefore confirmation of hybrid heterosis in laboratory studies should not be based on a single, or a few reciprocal crosses. Hybrid and pure species veliger larvae were grown for approximately 4 weeks at 10, 14 or 20 °C. In all trials, pure M. galloprovincialis larvae grew significantly faster at 20 °C than either reciprocal hybrid or pure M. edulis larvae. Irrespective of temperature, in general, hybrid larvae grew slower than larvae of either pure species. Increased exposure to planktonic predation due to slow growth can be interpreted as selection against hybrids and this may play a role in the structure and distribution of mixed pure species and hybrid populations.     

Genetics and taxonomy of Chilean smooth-shelled mussels, Mytilus spp. (Bivalvia: Mytilidae)

It has been previously established that native smooth-shelled mussels in southern South America possess close evolutionary affinities with Northern-Hemisphere Mytilus edulis L. 1758 (McDonald et al. (1991) [5]). This result has since been challenged by authors claiming that Chilean mussels should be considered a local subspecies of M. galloprovincialis Lmk. 1819. Moreover, morphological, physiological, ecotoxicological and molecular genetic studies on Chilean smooth-shelled mussels still frequently refer to ‘M. chilensis’ Hupé 1854, even though the previous discovery of alien M. galloprovincialis and considerable heterogeneity in shell morphology among samples collected along the Chilean shores raise concerns that different Mytilus spp. species might have been included under ‘M. chilensis’. Here we reviewed the molecular and morphological data available on smooth-shelled mussels from Chile in an attempt to clarify both their genetic composition and their taxonomic status. Using multivariate analysis on sample × allozyme-frequency matrices, we confirmed the widespread occurrence of the Southern-Hemisphere form of M. edulis along the shores from the North Patagonia region of Chile to the southern tip of the South American continent. The populations sampled in southern central Chile showed some evidence of slight introgression from Southern-Hemisphere M. galloprovincialis. Morphological characterization of a sample from Dichato in southern central Chile was consistent with its previous genetic identification as Mediterranean M. galloprovincialis. The occurrence of Southern-Hemisphere M. galloprovincialis in Punta Arenas at the southern tip of the South American continent was also reported. Southern-Hemisphere M. edulis, including native Chilean smooth-shelled Mytilus, should be assigned subspecific rank and named M. edulis platensis d’Orbigny 1846.     

Developmental stability is not disrupted by extensive hybridization and introgression among populations of the marine bivalve molluscs Mytilus edulis (L.) and M. galloprovincialis (Lmk.) from south-west England

The effects of hybridization and introzgression were assessed among two naturally hybridizing bivalve molluscs (the mussels Mytilus edulis and M. galloprovincialis) from western Europe to estimate how disruptive these processes are to developmental stability (measured in terms of morphological variability). Ten shell traits were measured for 392 mussels from four allopatric populations (two each of At. edulis and At. galloprovincialis) and two hybrid populations. An index of variability (defined as Z_i= |y_i–y_i where y_; is the population mean of the length-standardized trait, and y_i is the individualcar;s length-standardized trait value) was constructed for each trait, and for the sum of the traits. The hybrid populations did not exhibit greater mean variability than the allopatric populations for any of the indices. Upon pooling, the hybrid populations had significantly lower variability than the pooled M. edulis populations and the pooled M. galloprovincialis populations in two analyses, and had similar means in the remaining nine analyses. Where significant differences existed, the pooled M. galloprovincialis had lower levels of mean trait variability than the pooled M. edulis. Among the two hybrid populations, the putative Fl hybrids and backcross individuals exhibited means of trait variability which were similar to those of the parental types. Thus, extensive hybridization and introgression do not adversely affect developmental stability among these mussel populations. There was a strong significant correlation between the ranking of indices (based on the amount of variability) across all six populations, indicating that a large genetic component determines the measured morphological variability. It is concluded that the genes or gene complexes which control morphological development in M. edulis and M. galloprovincialis arc very similar, providing further evidence of the close evolutionary relatedness of these mussel taxa.     

Species status and population structure of mussels (Mollusca: Bivalvia: Mytilus spp.) in the Wadden Sea of Lower Saxony (Germany)

Three species of mussel (genus Mytilus) occur in Europe: M. edulis (Linnaeus 1758), M. galloprovincialis (Lamarck 1819) and M. trossulus (Gould, Boston Society of Natural History 3: 343?C348, 1850). Although these species are indigenous to the North Sea, the Mediterranean and the Baltic Sea, respectively, they form an extended patchy species complex along the coasts of Europe (??the Mytilus edulis complex??) and are able to hybridize where their distributions overlap. Recent studies examining the taxonomic status and genetic composition of Mytilus populations in the Netherlands and the British Isles have revealed introgressive hybridization processes within this species complex, with hints of an invasion of nonindigenous M. galloprovincialis into the North Sea. Furthermore, an extensive international mussel fishery industry in Europe (i.e., Great Britain, the Netherlands, Denmark, and Germany) is also in discussion for a possibly anthropogenically induced bioinvasion of nonindigenous Mytilus traits into the Wadden Sea area. Although it is assumed that the Wadden Sea of Germany comprises M. edulis only, this has never been confirmed in a molecular genetic study. To assess the situation for the Wadden Sea of Lower Saxony, we conducted the first molecular study of the Mytilus genus in the region. Taxonomic identification of 504 mussels from 13 intertidal mussel banks using the nDNA marker Me15/16 revealed a population composition of 99% M. edulis and 1% M. edulis X M. galloprovincialis hybrids. Hence, the Wadden Sea population is unaffected by range expansion of nonindigenous Mytilus traits. The genetic structure of the M. edulis populations was investigated using the phylogenetic and population genetics analyses of the mitochondrial DNA cytochrome-c-oxidase subunit I (COI) and the first variable domain of the control region (VD1), which were sequenced for >120 female individuals. These results showed a heterogeneous, panmictic population due to unrestricted gene flow. This can be attributed to extensive larval dispersal linked to the tidal circulation system in the back barrier basins of the Wadden Sea.