Sexual selection in seaweed? Testing Bateman's principles in the red alga Gracilaria gracilis.

In anisogamous species, sexual selection is expected to be stronger in males. Bateman's principles state that the variance in (i) reproductive and (ii) mating success is greater for males, and (iii) the relationship between reproductive success and mating success (the Bateman gradient) is also stronger for males than for females. Sexual selection, based on Bateman's principles, has been demonstrated in animals and some angiosperms, but never in a seaweed. Here we focus on the oogamous haploid-diploid rhodophyte Gracilaria gracilis in which previous studies have shown evidence for non-random mating, suggesting the existence of male-male competition and female choice. We estimated mating and reproductive success using paternity analyses in a natural population where up to 92% of fertilizations occurred between partners of that population. The results show that the variance in mating success is significantly greater in males than in females and that the Bateman gradient is positive only in males. Distance to female partners also explains a minor part of the variance in male mating success. Although there is no evidence for sexual dimorphism, our study supports the hypothesis that sexual selection occurs in G. gracilis, probably on male traits, even if we cannot observe, characterize or quantify them yet.

Pollinators of the sea: A discovery of animal-mediated fertilization in seaweed.

The long-held belief that animal-mediated pollination is absent in the sea has recently been contradicted in seagrasses, motivating investigations of other marine phyla. This is particularly relevant in red algae, in which female gametes are not liberated and male gametes are not flagellated. Using experiments with the isopod Idotea balthica and the red alga Gracilaria gracilis, we demonstrate that biotic interactions dramatically increase the fertilization success of the alga through animal transport of spermatia on their body. This discovery suggests that animal-mediated fertilization could have evolved independently in terrestrial and marine environments and raises the possibility of its emergence in the sea before plants moved ashore.

O father where art thou? Paternity analyses in a natural population of the haploid-diploid seaweed Chondrus crispus.

The link between life history traits and mating systems in diploid organisms has been extensively addressed in the literature, whereas the degree of selfing and/or inbreeding in natural populations of haploid-diploid organisms, in which haploid gametophytes alternate with diploid sporophytes, has been rarely measured. Dioecy has often been used as a proxy for the mating system in these organisms. Yet, dioecy does not prevent the fusion of gametes from male and female gametophytes originating from the same sporophyte. This is likely a common occurrence when spores from the same parent are dispersed in clumps and recruit together. This pattern of clumped spore dispersal has been hypothesized to explain significant heterozygote deficiency in the dioecious haploid-diploid seaweed Chondrus crispus. Fronds and cystocarps (structures in which zygotes are mitotically amplified) were sampled in two 25 m(2) plots located within a high and a low intertidal zone and genotyped at 5 polymorphic microsatellite loci in order to explore the mating system directly using paternity analyses. Multiple males sired cystocarps on each female, but only one of the 423 paternal genotypes corresponded to a field-sampled gametophyte. Nevertheless, larger kinship coefficients were detected between males siring cystocarps on the same female in comparison with males in the entire population, confirming restricted spermatial and clumped spore dispersal. Such dispersal mechanisms may be a mode of reproductive assurance due to nonmotile gametes associated with putatively reduced effects of inbreeding depression because of the free-living haploid stage in C. crispus.

Spatiotemporal changes in the genetic diversity of harmful algal blooms caused by the toxic dinoflagellate Alexandrium minutum.

Organisms with sexual and asexual reproductive systems benefit from both types of reproduction. Sexual recombination generates new combinations of alleles, whereas clonality favours the spread of the fittest genotype through the entire population. Therefore, the rate of sexual vs. clonal reproduction has a major influence on the demography and genetic structure of natural populations. We addressed the effect of reproductive system on populations of the dinoflagellate Alexandrium minutum. More specifically, we monitored the spatiotemporal genetic diversity during and between bloom events in two estuaries separated by 150 km for two consecutive years. An analysis of population genetic patterns using microsatellite markers revealed surprisingly high genotypic and genetic diversity. Moreover, there was significant spatial and temporal genetic differentiation during and between bloom events. Our results demonstrate that (i) interannual genetic differentiation can be very high, (ii) estuaries are partially isolated during bloom events and (iii) genetic diversity can change rapidly during a bloom event. This rapid genetic change may reflect selective effects that are nevertheless not strong enough to reduce allelic diversity. Thus, sexual reproduction and/or migration may regularly erase any genetic structure produced within estuaries during a bloom event.

Mating system and gene flow in the red seaweed Gracilaria gracilis: effect of haploid-diploid life history and intertidal rocky shore landscape on fine-scale genetic structure.

The impact of haploid-diploidy and the intertidal landscape on a fine-scale genetic structure was explored in a red seaweed Gracilaria gracilis. The pattern of genetic structure was compared in haploid and diploid stages at a microgeographic scale (< 5 km): a total of 280 haploid and 296 diploid individuals located in six discrete, scattered rock pools were genotyped using seven microsatellite loci. Contrary to the theoretical expectation of predominantly endogamous mating systems in haploid-diploid organisms, G. gracilis showed a clearly allogamous mating system. Although within-population allele frequencies were similar between haploids and diploids, genetic differentiation among haploids was more than twice that of diploids, suggesting that there may be a lag between migration and (local) breeding due to the long generation times in G. gracilis. Weak, but significant, population differentiation was detected in both haploids and diploids and varied with landscape features, and not with geographic distance. Using an assignment test, we establish that effective migration rates varied according to height on the shore. In this intertidal species, biased spore dispersal may occur during the transport of spores and gametes at low tide when small streams flow from high- to lower-shore pools. The longevity of both haploid and diploid free-living stages and the long generation times typical of G. gracilis populations may promote the observed pattern of high genetic diversity within populations relative to that among populations.

Polymerase chain reaction-single strand conformation polymorphism analyses of nuclear and chloroplast DNA provide evidence for recombination, multiple introductions and nascent speciation in the Caulerpa taxifolia complex.

Independent lines of evidence support an Australian origin for the Mediterranean populations of the tropical alga Caulerpa taxifolia. To complement previous biogeographical studies based on nuclear rDNA internal transcribed spacer (ITS), a new chloroplast marker was developed--the cp 16S rDNA intron-2. Sequence variability for both nuclear and chloroplast markers were assessed in 110 individuals using single strand conformation polymorphism. Comparison of intrapopulation genetic diversity between invasive Mediterranean and 'native' Australian populations revealed the occurrence of two divergent and widespread clades. The first clade grouped nontropical invasive populations with inshore-mainland populations from Australia, while the second clustered all offshore-island populations studied so far. Despite our finding of nine distinct nuclear and five distinct chloroplast profiles, a single nucleocytoplasmic combination was characteristic of the invasive populations and sexual reproduction was found to be very rare. C. taxifolia is clearly a complex of genetically and ecologically differentiated sibling species or subspecies.

Phylogenetic analyses of Caulerpa taxifolia (Chlorophyta) and of its associated bacterial microflora provide clues to the origin of the Mediterranean introduction.

The accidental introduction of Caulerpa taxifolia into the Mediterranean is no longer under dispute. What has eluded researchers until now, is definitive evidence for the original, biogeographical source population. Here we present two independent lines of evidence that support an Australian origin for the Mediterranean populations of C. taxifolia. First, we reanalysed algal rDNA-internal transcribed spacer (rDNA-ITS) sequences, combining previously published sequences from different studies with 22 new sequences. The ITS sequence comparison showed that the Australian sample is the sister group of the Mediterranean-aquarium clade. Second, cloned bacterial 16S rDNA gene sequences were analysed from the associated microflora of C. taxifolia collected from Australia, Tahiti, the Philippines and the Mediterranean. Five bacterial lineages were identified, of which three were dominant. Alpha Proteobacteria were the most abundant and were found in all samples. In contrast, members of the beta Proteobacterial line and Cytophaga-Flexibacter-Bacteroides line (CFB) were mainly associated with Mediterranean and Australian samples. Frequency distributions of the five bacterial lineages were significantly different among biogeographical locations. Phylogenetic analyses of the 54 bacterial sequences derived from the four C. taxifolia individuals resulted in a well-resolved tree with high bootstrap support. The topologies of the beta Proteobacteria and CFB mirror the geographical sources of their algal hosts. Bacterial-algal associations provide an identification tool that may have wide application for the detection of marine invasions.

Identification of random amplified polymorphic DNA (RAPD) markers highly linked to sex determination in the red alga Gracilaria gracilis.

The random amplified polymorphic DNA (RAPD) technique was employed in the haplo-diploid dioecious species Gracilaria gracilis to identify sex-linked PCR markers. Sixty-nine decamer oligonucleotide primers were tested on two bulks of DNA, one from five haploid males and the other from five haploid females. One of these primers (OPD13) generated a 430-bp fragment specific to males and a 620-bp fragment specific to females. The diploid individuals (tetrasporophytes) showed the co-occurrence of these two fragments. In order to verify the linkage between the sexual phenotypes and these markers, a progeny array of 59 haploid individuals (male and female) born on a diploid individual was analysed, in all of which the two markers produced by the OPD13 primer segregated perfectly with sex.

Evolution of alternation of haploid and diploid phases in life cycles.

Eukaryotic sex leads to an alternation of haploid and diploid nuclear phases. Because all multicellular animals are diploid, diploidy is often considered a 'biological success' and many arguments have been advanced to explain the evolution of a prolonged diploid phase. Nevertheless, among eukaryotes three basic situations are encountered, where the vegetative individuals are diploid or haploid or both. These three basic life cycles are widely distributed among kingdoms and in some taxa the occurrence of different life cycles within the same species has been reported. This article briefly summarizes the different hypotheses on the evolution of reproductive life cycles and underlines how possibilities of variation for this trait may open new perspectives for research.

Rubisco spacer sequence divergence in the rhodophyte alga Gracilaria verrucosa and closely related species.

In the red alga Gracilaria verrucosa, the genes encoding the large and the small subunit of ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) are separated by a short spacer of less than 131 bp. Sequencing of PCR-amplified Rubisco spacers from a number of populations of G. verrucosa was performed to assess the feasibility of using this sequence for discriminating among closely related species or populations. Intrapopulation comparisons of the nucleotide sequences of these spacers from five isolates of G. verrucosa, and similar species, demonstrated four main groups. The first group included isolates from Europe and Argentina while the other groups are correlated with the geographical location of their origin.