Evidence for recombination of mtDNA in the marine mussel Mytilus trossulus from the Baltic

A number of studies have claimed that recombination occurs in animal mtDNA, although this evidence is controversial. Ladoukakis and Zouros (2001) provided strong evidence for mtDNA recombination in the COIII gene in gonadal tissue in the marine mussel Mytilus galloprovincialis from the Black Sea. The recombinant molecules they reported had not however become established in the population from which experimental animals were sampled. In the present study, we provide further evidence of the generality of mtDNA recombination in Mytilus by reporting recombinant mtDNA molecules in a related mussel species, Mytilus trossulus, from the Baltic. The mtDNA region studied begins in the 16S rRNA gene and terminates in the cytochrome b gene and includes a major noncoding region that may be analogous to the D-loop region observed in other animals. Many bivalve species, including some Mytilus species, are unusual in that they have two mtDNA genomes, one of which is inherited maternally (F genome) the other inherited paternally (M genome). Two recombinant variants reported in the present study have population frequencies of 5% and 36% and appear to be mosaic for F-like and M-like sequences. However, both variants have the noncoding region from the M genome, and both are transmitted to sperm like the M genome. We speculate that acquisition of the noncoding region by the recombinant molecules has conferred a paternal role on mtDNA genomes that otherwise resemble the F genome in sequence.     

Complete sequences of the highly rearranged molluscan mitochondrial genomes of the Scaphopod Graptacme eborea and the bivalve Mytilus edulis

We have determined the complete sequence of the mitochondrial genome of the scaphopod mollusk Graptacme eborea (14,492 nts) and completed the sequence of the mitochondrial genome of the bivalve mollusk Mytilus edulis (16,740 nts). (The name Graptacme eborea is a revision of the species formerly known as Dentalium eboreum.) G. eborea mtDNA contains the 37 genes that are typically found and has the genes divided about evenly between the two strands, but M. edulis contains an extra trnM and is missing atp8, and it has all genes on the same strand. Each has a highly rearranged gene order relative to each other and to all other studied mtDNAs. G. eborea mtDNA has almost no strand skew, but the coding strand of M. edulis mtDNA is very rich in G and T. This is reflected in differential codon usage patterns and even in amino acid compositions. G. eborea mtDNA has fewer noncoding nucleotides than any other mtDNA studied to date, with the largest noncoding region only 24 nt long. Phylogenetic analysis using 2,420 aligned amino acid positions of concatenated proteins weakly supports an association of the scaphopod with gastropods to the exclusion of Bivalvia, Cephalopoda, and Polyplacophora, but it is generally unable to convincingly resolve the relationships among major groups of the Lophotrochozoa, in contrast to the good resolution seen for several other major metazoan groups.     

Interspecies transfer of female mitochondrial DNA is coupled with role-reversals and departure from neutrality in the mussel Mytilus trossulus.

Mussels of the genus Mytilus have distinct and highly diverged male and female mitochondrial DNA (mtDNA) genomes with separate routes of inheritance. Previous studies of European populations of Mytilus trossulus demonstrated that 33% of males are heteroplasmic for a second mtDNA genome of increased length and that hybridization with Mytilus edulis does not block mtDNA introgression, in contrast to reports for American populations. Here, we demonstrate that the female mtDNA type of M. edulis has replaced the resident female mtDNA type of European M. trossulus. This is supported by COIII sequence data indicating that the female mtDNA of European M. trossulus is very similar to that of M. edulis and that in phylogenetic trees, the mtDNAs of these two species cluster together but separately from American M. trossulus sequences, the latter not being disturbed by introgressive hybridization. We also provide evidence that the mtDNA genome of increased length found in heteroplasmic males of European M. trossulus derives from a recent partition of an introgressed M. edulis female type into the male route of transmission. Neutrality tests reveal that European populations of M. trossulus display an excess of replacement polymorphism within the female mtDNA type with respect to conspecific American populations, as well as a significant excess of rare variants, of a similar magnitude to those previously reported for the invading European M. edulis mtDNA. Results are consistent with a nearly neutral model of molecular evolution and suggest that selection acting on European M. trossulus mtDNA is largely independent of the nuclear genetic background.     

Polyploidy in a natural population of mussel, Mytilus trossulus.

We have analyzed natural polyploidy in a population of Mytilus trossulus from Vancouver Island (British Columbia, Canada) by means of cytogenetic techniques. Results obtained are the first reporting on this type of numerical chromosome aberrations in mussels.     

Changes in the lipid composition of embryonic cells of the mussel Mytilus trossulus during cryopreservation

A comparative analysis of the lipid composition of embryonic cells of the mussel Mytilus trossulus prior to, and after, cryopreservation in liquid nitrogen was carried out in the presence of two types of cryoprotectors: 1) dimethylsulfoxide and trehalose; and 2) dimethylsulfoxide, trehalose, total lipid extract from the mussel Crenomytilus grayanus, alpha-tocopherol and ascorbic acid. It was found that not only the cell viability but also the fatty acid composition of cell lipids after cryopreservation depend on the composition of cryoprotectors used. The content of saturated fatty acids, monoenic and polyenic fatty acids, the omega 3/omega 6 ratio, and the index of nonsaturation in the fatty acid composition of lipids was shown to change remarkably after cryopreservation. Possible reasons of these changes are discussed.     

Expression of several domains of twitchin and myorod in the ontogenesis of mussel Mytilus trossulus

The expression of MLCK- and PEVK-domains of twitchin, as well as the unique N-terminal domain of myorod in early development of the mussel Mytilus trossulus has been studied. The MLCK-domain of twitchin and the unique N-terminal domain of myorod appear at the early stages of development, whereas the PEVK-domain of twitchin is present only in muscles of adult mussel. The sizes of genes of the N-terminal domain of myorod, obtained at the blastula stage and from the adult animal are similar, but the proteins have significant differences in the amino acid sequences. Consequently, myorod and twitchin appear at early stages of larval mussels before the formation of "adult" muscles capable of catch contraction, and at these stages both proteins are isoforms, which differ from the isoforms of adult animals. It is possible that the MLCK-domain in the "larval" isoform of twitchin is necessary for regulating the formation of the contractile apparatus of molluscan smooth muscles, while the PEVK-domain is important for the regulation of the catch state in muscles of adult animals.     

Local adaptation and species segregation in two mussel (Mytilus edulis x Mytilus trossulus) hybrid zones

Few marine hybrid zones have been studied extensively, the major exception being the hybrid zone between the mussels Mytilus edulis and Mytilus galloprovincialis in southwestern Europe. Here, we focus on two less studied hybrid zones that also involve Mytilus spp.; Mytilus edulis and Mytilus trossulus are sympatric and hybridize on both western and eastern coasts of the Atlantic Ocean. We review the dynamics of hybridization in these two hybrid zones and evaluate the role of local adaptation for maintaining species boundaries. In Scandinavia, hybridization and gene introgression is so extensive that no individuals with pure M. trossulus genotypes have been found. However, M. trossulus alleles are maintained at high frequencies in the extremely low salinity Baltic Sea for some allozyme genes. A synthesis of reciprocal transplantation experiments between different salinity regimes shows that unlinked Gpi and Pgm alleles change frequency following transplantation, such that post-transplantation allelic composition resembles native populations found in the same salinity. These experiments provide strong evidence for salinity adaptation at Gpi and Pgm (or genes linked to them). In the Canadian Maritimes, pure M. edulis and M. trossulus individuals are abundant, and limited data suggest that M. edulis predominates in low salinity and sheltered conditions, whereas M. trossulus are more abundant on the wave-exposed open coasts. We suggest that these conflicting patterns of species segregation are, in part, caused by local adaptation of Scandinavian M. trossulus to the extremely low salinity Baltic Sea environment.     

Development of the larval muscle system in the mussel Mytilus trossulus (Mollusca,Bivalvia)

In the present study we examined muscle development throughout the complete larval cycle of the bivalve mollusc, Mytilus trossulus. An immunofluorescence technique and laser scanning confocal microscopy were used in order to study the organization of the muscle proteins (myosin, paramyosin, twitchin, and actin) and some neurotransmitters. The appearance of the muscle bundles lagged behind their nervous supply: the neuronal elements developed slightly earlier (by 2 h) than the muscle cells. The pioneer muscle cells forming a prototroch muscle ring were observed in a completed trochophore. We documented a well‐organized muscle system that consisted of the muscle ring transforming into three pairs of velar striated retractors in the early veliger. The striations were positive for all muscle proteins tested. Distribution of FMRFamide and serotonin (5‐HT) immunocytochemical staining relative to the muscle ring differed significantly: 5‐HT‐immunioreactive cells were situated in the center of the striated muscle ring, while Phe‐Met‐Arg‐Phe‐NH2 neuropeptide FMRFamid immunoreactive fibers were located in a distal part of this ring. Our data showed clearly that the muscle proteins and the neurotransmitters were co‐expressed in a coordinated fashion in a continuum during the early stages of the mussel development. Our study provides the first strong evidence that mussel larval metamorphosis is accompanied by a massive reorganization of striated muscles, followed by the development of smooth muscles capable of catch‐contraction.     

Comparative study of microbial communities from cultured and natural population of the mussel Mytilus trossulus in Peter the Great bay

The 525 strains of heterotrophic bacteria isolated from natural and cultured populations of the mussel Mytilus trossulus and the surrounding seawater were identified to a genus level on the basis of phenotypic analysis and the fatty acid composition of cell lipids. Gram-negative isolates were dominated by six genera of the family Enterobacteriaceae and by the genera Pseudoalteromonas, Vibrio, Photobacterium, Cytophaga/Flavobacterium/Bacteroides, Pseudomonas, and Moraxella, Gram-positive isolates were mainly represented by the genus Streptomyces. The taxonomic compositions of natural and cultured populations of the mussel M. trossulus in Peter the Great Bay were similar.     

A connective tissue tumor in the mussel Mytilus trossulus from a polluted region of Nakhodka Bay, the Sea of Japan

A tumor was found for the first time in a mussel Mytilus trossulus from a heavily polluted area of Nakhodka Bay, Sea of Japan. Tumor cells were found in the connective tissue of different organs and also in gill vessels and hemal sinuses of the visceral mass. They were both attached and diffuse. The tumor was at an advanced stage, replacing the normal connective tissue cells, and formed nodes. The tumor cells were polymorphic, with a high nucleocytoplasmic ratio, and had a prominent nucleolus. The size of their nuclei was three to five times that of the nuclei of agranular hemocytes. The mitotic activity of the tumor cells was more than an order of magnitude higher than in the normal cells: the mean mitotic index was 1.4 +/- 0.5%, ranging from 0.97 to 2.3% in different organs. The mitotic indices in the connective tissue cells of three normal mussels were 0, 0, and 0.12%. A significant proportion (up to 78%) of the mitotic cells were at metaphase. The frequency of abnormal mitoses was 17%. Metaphases with displaced (often multiple) chromosomes constituted 71% of abnormal mitoses; anaphases, 8%; and tri- and tetrapolar mitoses, 11%. The tumor described is similar to diffuse sarcomatoid diseases of mussels from other geographical regions.     

Heteroplasmy suggests paternal co-transmission of multiple genomes and pervasive reversion of maternally into paternally transmitted genomes of mussel (Mytilus) mitochondrial DNA

Marine mussels of the genus Mytilus have two types of mitochondrial DNA with separate paternal and maternal inheritance. Females are homoplasmic for an F genome that is transmitted to all offspring, whereas males are heteroplasmic for this F genome and for a highly diverged (> 20%) M genome that is transmitted only to sons. Here we provide phylogenetic evidence based on lrRNA sequence data that most of the paternal genomes in European M. trossulus have an introgressive female M. edulis origin and are nearly indistinguishable in sequence from F types of M. trossulus. This observation is best explained by the hypothesis that introgressed F type molecules have recently invaded the paternal route and have assumed the role of M molecules, then resetting to zero the time of sequence divergence between M and F lineages. European M. trossulus shows a high prevalence of males heteroplasmic for three different mitochondrial DNA types all having the same two paternal types and the same maternal type, consistent with paternal co-transmission of multiple genomes. Co-transmission of the same genomes must apparently operate uninterruptedly for several generations in spite of the very different evolutionary origin of the specific molecules that are transmitted paternally and maternally in European M. trossulus.     

Expression of thick filament proteins during ontogenesis of the mussel Mytilus trossulus (Mollusca: Bivalvia)

The appearance of thick filament proteins organized into supramolecular complexes was studied by SDS-PAGE and Western-blot analysis at different developmental stages of the mussel Mytilus trossulus. Paramyosin appeared at the egg stage, while twitchin and myorod appeared at the blastula stage (12 h after fertilization). In addition, RT-PCR analysis showed that the twitchin genes were expressed starting from the blastula stage. Thus, the proteins forming thick filaments of the contractile apparatus of mussel muscles are expressed long before the formation of the first well-organized muscle system of the veliger larvae (55 h). Further, the ratios actin/myosin heavy chain (MHC) and paramyosin/MHC at the veliger stage (96 h) distinctly differed from those in the adult mussel.     

Complete DNA sequences of the mitochondrial genomes of the pathogenic yeasts Candida orthopsilosis and Candida metapsilosis: insight into the evolution of linear DNA genomes from mitochondrial telomere mutants

We determined complete mitochondrial DNA sequences of the two yeast species, Candida orthopsilosis and Candida metapsilosis, and compared them with the linear mitochondrial genome of their close relative, C.parapsilosis. Mitochondria of all the three species harbor compact genomes encoding the same set of genes arranged in the identical order. Differences in the length of these genomes result mainly from the presence/absence of introns. Multiple alterations were identified also in the sequences of the ribosomal and transfer RNAs, and proteins. However, the most striking feature of C.orthopsilosis and C.metapsilosis is the existence of strains differing in the molecular form of the mitochondrial genome (circular-mapping versus linear). Their analysis opens a unique window for understanding the role of mitochondrial telomeres in the stability and evolution of molecular architecture of the genome. Our results indicate that the circular-mapping mitochondrial genome derived from the linear form by intramolecular end-to-end fusions. Moreover, we suggest that the linear mitochondrial genome evolved from a circular-mapping form present in a common ancestor of the three species and, at the same time, the emergence of mitochondrial telomeres enabled the formation of linear monomeric DNA forms. In addition, comparison of isogenic C.metapsilosis strains differing in the form of the organellar genome suggests a possibility that, under some circumstances, the linearity and/or the presence of telomeres provide a competitive advantage over a circular-mapping mitochondrial genome.     

Development of the muscle system and contractile activity in the mussel Mytilus trossulus (Mollusca, Bivalvia)

The development of contractile apparatus was subjected to comparative analysis during ontogenesis of the mussel Mytilus trossulus. Indirect immunofluorescence with the polyclonal antibody against mussel twitchin, a protein of thick filaments, and florescent phalloidin as a marker of filamentous cell actin were used to monitor changes in the developing muscle system at different larval stages. The first definitive muscle structures were found at the late trochophore stage (36 h after fertilization) and starting from the midveliger stage (96 h), striated muscles, which are never present in adult mussels, were distinctly seen. The striated muscle periodicity was 1.25 microm in both mussle larvae and adult scallop. The contractile activities of veliger and adult muscles were measured using an electronic signal-processing videosystem. This work is the first complex study of morphological, biochemical, and physiological characteristics of the muscle system in the larvae and adult mollusks.     

Complete sequences of maternally inherited mitochondrial genomes in musselsUnio pictorum (Bivalvia, Unionidae)

Mitochondrial genomes are frequently used to infer phylogenetic relationships. Some taxa are, however, poorly represented. To facilitate better understanding of the potential of mitochondrial genome data in freshwater mussels, we present here, for the first time, the mitochondrial sequences of 4 complete F-type mitochondrial genomes from the European freshwater bivalveUnio pictorum (Unionidae). These genomes are very compact (15 761 bp) but have a typical gene complement for bilaterian mitochondrial genomes and a very similar organization to other unionid genomes available in databases. Very low nucleotide diversity within the species suggests a small effective population size of PolishU. pictorum, a phenomenon of potential importance for environmental management policies.     

Changes in the level of lectins in the mantle of the mussel Mytilus trossulus in response to anthropogenic contaminants

The lectin activity in an extract from the mantle of the mussel Mytilus trossulus was tested for the first time. Using the method of the hemagglutination inhibition assay, it was shown that lectins were Gal/GalNAc-specific and best agglutinated with rabbit erythrocytes. The influence of foreign compounds on the lectin level in the M. trossulus mantle was examined. Upon cadmium acetate exposure, the level of lectin activity exhibited phasic alterations and depended on the contaminant concentration or the time of exposure. During exposure of mussels to a synthetic detergent or diesel fuel, changes in lectin contents were dependent on the time of exposure.