Eleftherios Zouros

MULTIPLE ORIGINS OF GENDER-ASSOCIATED MITOCHONDRIAL DNA LINEAGES IN BIVALVES (MOLLUSCA: BIVALVIA)

Previous studies have shown that marine mussels (genus Mytilus) and a freshwater mussel (Pyganodon grandis) contain two distinct gender‐associated mitotypes, which is a characteristic feature of the phenomenon of doubly uniparental inheritance (DUI) of mitochondrial DNA (mtDNA). Here we present evidence for the presence of distinct male (M) and female (F) mitotypes in three other bivalve species, the mytilid Geukensia demissa, and the unionid species P. fragilis and Fusconaia flava. Nucleotide sequences of a segment of the COI gene from the M and F mitotypes from each of the three mytilid species (M. edulis, M. trossulus, G. demissa) and three unionid species (P. grandis, P. fragilis, F. flava) were used for phylogenetic analysis. The analysis suggests three independent origins of M and F mitotypes for the six species examined; one for the three unionid species, one for the two Mytilus species, and one for Geukensia. The first of these F/M divergence events, while of uncertain age, predates the divergence of the two unionid genera and is likely older than either of the two F/M divergence events in the mytilid taxa. The most parsimonious explanation of multiple F/M divergence events is that they represent independent origins of DUI. Another possibility is that, in a given taxon, an F or M mitotype assumes the role of the opposite mitotype (by virtue of a mechanism that remains to be clarified) and subsequently was fixed within its new gender. The fixation of a mtDNA lineage derived from a mitotype of switched function would reset the divergence of the gender‐associated lineages to zero, thereby mimicking a de novo split of F and M lineages from a preexisting mtDNA genome that was not gender specific. Further broad‐scale taxonomic studies of the occurrence of distinct M and F mitotypes may allow for the evaluation of the latter hypothesis.

Genetic differentiation in relation to marine landscape in a broadcast-spawning bivalve mollusc (Placopecten magellanicus)

Marine bivalves are sessile or sedentary as adults but have planktonic larvae which can potentially disperse over large distances. Consequently larval transport is expected to play a prominent role in facilitating gene flow and determining population structure. The sea scallop (Placopecten magellanicus) is a dioecious species with high fecundity, broadcast spawning and a c. 30‐day planktonic larval stage, yet it forms discrete populations or ‘beds’ which have significantly different dynamics and characteristics. We analysed variation at six microsatellite loci in 12 locations throughout the geographic range of the species from Newfoundland, Canada, to New Jersey, USA. Significant differentiation was present and the maximum pairwise θ value, between one of the Newfoundland samples in the north and a sample from the southern portion of the range, was high at 0.061. Other proximate pairs of samples had no detectable genetic differentiation. Mantel tests indicated a significant isolation by distance, but only when one of the populations was excluded. A landscape genetic approach was used to detect areas of low gene flow using a joint analysis of spatial and genetic information. The two major putative barriers inferred by Monmoniers algorithm were then used to define regions for an analysis of molecular variance (amova). That analysis showed a significant but low percentage (1.2%) of the variation to be partitioned among regions, negligible variation among populations within regions, and the majority of the variance distributed between individuals within populations. Prominent currents were concordant with the demarcation of the regions, while a novel approach of using particle tracking software to mimic scallop larval dispersal was employed to interpret within‐region genetic patterns.

Larval viability and heterozygote deficiency in populations of marine bivalves: evidence from pair matings of mussels

Juveniles of the blue mussel Mytilus edulis that had resulted from a large number of pair matings were examined at six polymorphic enzyme loci. In four loci, the overall number of heterozygotes was significantly less than expected from the parental genotypes. The degree of deviation from expectation varied considerably among families, and families which shared the same male parent tended to show similar degrees of deviation. Inbreeding and population mixture, the two most cited explanations of heterozygote deficiency (a phenomenon commonly observed in populations of marine bivalves), do not apply in the case of pair-matings, and the null allele hypothesis is inconsistent with the data. These observations suggest that genotype-dependent larval mortality constitutes the most probable cause of this phenomenon.

Marked mitochondrial DNA differences between Mediterranean and Atlantic populations of the swordfish, Xiphias gladius

Restriction analysis of mitochondrial DNA (mtDNA) from 204 individuals of swordfish (Xiphias gladius) revealed no differentiation among samples from three sites in the Mediterranean Sea (Greece, Italy, Spain), but a high degree of differentiation between Mediterranean samples and a sample from the Gulf of Guinea. A fifth sample from the Atlantic side of the Straits of Gibraltar (Tarifa) consisted mostly of mitotypes that are common in the Mediterranean, but contained several of mtDNA types of the Guinea sample not found in the Mediterranean. We conclude that, in spite of free migration of swordfish across the Straits of Gibraltar, little genetic exchange occurs between the populations inhabiting the Mediterranean Sea and the tropical Atlantic ocean. This is the first evidence of genetic differentiation among geographic populations of this highly mobile species that supports a world‐wide fishery.

Allozyme and physiological variation in the scallop placopecten magellanicus and a general model for the effects of heterozygosity on fitness in marine molluscs

Heterozygosity and growth rate have been correlated in many molluscs, although the phenomenon is not universal. Enhanced growth of heterozygous molluscs has been attributed to lower energetic requirements for basal metabolism. We have investigated heterozygote deficiency, and the correlation between single-locus and multiple-locus heterozygosity and phenotype in juveniles of the scallop Placopecten magellanicus (Gmelin). Six samples were collected between 22 November 1984 and 11 December 1985 at Passamaquoddy Bay, New Brunswick, Canada. On average, heterozygote deficiency was small, although somewhat higher at the octopine dehydrogenase (Odh) locus, and decreased with age. No correlation was observed between genotype and growth rate. This result and published records indicate that allozyme heterozygosity of pectinids does not influence growth to the same degree as in other bivalves. However, we have observed in P. magellanicus a correlation between allozyme heterozygosity and octopine accumulation, a trait that relates to functional anaerobiosis, and may be related to the scallops scope for movement. We combine these results and results from the literature into a model that relates the hypothesis of “associative overdominance” (at the population genetics level) with the hypothesis of “energetic efficiency” (at the physiological level). The model assumes that energy savings attributed to heterozygosity are used to maximize fitness. In freely moving molluscs this results in enhanced activity (such as searching for prey or swimming away from a predator), and in sessile molluscs either in enhanced somatic growth in juveniles or gonadal growth in adults.

Genetics of growth in blue mussels: family and enzyme-heterozygosity effects

Shell length and electrophoretic heterozygosity at six enzyme loci were scored in juveniles of 30 hatchery-grown families and in adults of 10 field-grown families of the blue mussel Mytilus edulis. A total of 4 809 offspring were scored. We found no consistent pattern in the correlations between shell length and enzyme heterozygosity among sibs within families. However, a significant family effect on shell length was observed, suggesting an influence of background genotype on this character. A family effect on viability was also observed. When the environment and family effects were accounted for, a small positive correlation between heterozygosity and shell length at the juvenile stage remained. This correlation was significant in one of the three experiments. No such correlation was evident at the adult stage. Our interpretation of these results is that electrophoretic alleles appear to have no direct of independent effect of their own on growth. We suggest that the negative correlation of homozygosity with these characters, seen in natural populations, results from homozygosity for hidden recessive deleterious genes with which the electromorphs are in a steady state of quasilinkage disequilibrium.

Genetic Variation Underlying Protein Expression in Eggs of the Marine Mussel Mytilus edulis

Study of the genetic basis of gene expression variation is central to attempts to understand the causes of evolutionary change. Although there are many transcriptomics studies estimating genetic variance and heritability in model organisms such as humans there is a lack of equivalent proteomics studies. In the present study, the heritability underlying egg protein expression was estimated in the marine mussel Mytilus. We believe this to be the first such measurement of genetic variation for gene expression in eggs of any organism. The study of eggs is important in evolutionary theory and life history analysis because maternal effects might have profound effects on the rate of evolution of offspring traits. Evidence is presented that the egg proteome varies significantly between individual females and that heritability of protein expression in mussel eggs is moderate to high suggesting abundant genetic variation on which natural selection might act. The study of the mussel egg proteome is also important because of the unusual system of mitochondrial DNA inheritance in mussels whereby different mitochondrial genomes are transmitted independently through female and male lineages (doubly uniparental inheritance). It is likely that the mechanism underlying this system involves the interaction of specific egg factors with sperm mitochondria following fertilization, and its elucidation might be advanced by study of the proteome in females having different progeny sex ratios. Putative identifications are presented here for egg proteins using MS/MS in Mytilus lines differing in sex ratio. Ontology terms relating to stress response and protein folding occur more frequently for proteins showing large expression differences between the lines. The distribution of ontology terms in mussel eggs was compared with those for previous mussel proteomics studies (using other tissues) and with mammal eggs. Significant differences were observed between mussel eggs and mussel tissues but not between the two types of eggs.

The Control Region of Maternally and Paternally Inherited Mitochondrial Genomes of Three Species of the Sea Mussel Genus Mytilus

Species of the mussel genus Mytilus possess maternally and paternally transmitted mitochondrial genomes. In the interbreeding taxa Mytilus edulis and M. galloprovincialis, several genomes of both types have been fully sequenced. The genome consists of the coding part (which, in addition to protein and RNA genes, contains several small noncoding sequences) and the main control region (CR), which in turn consists of three distinct parts: the first variable (VD1), the conserved (CD), and the second variable (VD2) domain. The maternal and paternal genomes are very similar in gene content and organization, even though they differ by >20% in primary sequence. They differ even more at VD1 and VD2, yet they are remarkably similar at CD. The complete sequence of a genome from the closely related species M. trossulus was previously reported and found to consist of a maternal-like coding part and a paternal-like and a maternal-like CR. From this and from the fact that it was extracted from a male individual, it was inferred that this is a genome that switched from maternal to paternal transmission. Here we provide clear evidence that this genome is the maternal genome of M. trossulus. We have found that in this genome the tRNAGln in the coding region is apparently defective and that an intact copy of this tRNA occurs in the CR, that one of the two conserved domains is missing essential motifs, and that one of the two first variable domains has a high rate of divergence. These features may explain the large size and mosaic structure of the CR of the maternal genome of M. trossulus. We have also obtained CR sequences of the maternal and paternal genomes of M. californianus, a more distantly related species. We compare the control regions from all three species, focusing on the divergence among genomes of different species origin and among genomes of different transmission routes.

Dispersed discrete length polymorphism of mitochondrial DNA in the scallop Placopecten magellanicus (Gmelin)

Three separate regions exhibiting incremental length polymorphism have been found in the mitochondrial genome of the scallop Placopecten magellanicus. Each locus has a discrete and different unit of variation, measuring 1450 bp at locus I, 250 bp at locus II, and less than 100 bp at locus III. At least six size classes were observed at each locus, and individual variation can account for the intraspecific mtDNA size range of 31–42 kb, but not for the unusually large base size of the genome. Intramolecular hybridization patterns with clones of two of the variable regions indicate that there is a dispersed sequence similarity with the 1,450-bp locus-I repeat and its flanks, and with some part of locus II.

No evidence for absence of paternal mtDNA in male progeny from pair matings of the mussel Mytilus galloprovincialis

The claim that a Mytilus galloprovincialis male failed to transmit mtDNA to its sons in controlled crosses is shown to be false. At present there is no evidence for mussel males lacking a paternal mtDNA. This makes unlikely the hypothesis that maternal genomes may become paternally transmitted by invading the germ line of males that lack a paternal genome.