Taxonomic study of Scytosiphon (Phaeophyceae) from temperate coasts of Argentina.

Scytosiphon is a common intertidal genus widely distributed on temperate coasts worldwide. Recently, eight species have been delimited with molecular tools. Although S. lomentaria is the only species that predominates in the macroalgal literature of the Southwestern Atlantic Ocean (SwAO), unpublished molecular data obtained for a population study of S. lomentaria revealed hidden species diversity of Scytosiphon among the individuals collected from four localities at the SwAO. The aim of this study was to revise the identity and phylogenetic relationships of Scytosiphon from temperate coasts of the SwAO using DNA data. Thalli were collected from the Argentinean coast between 39° S and 43° S, from which cox1 and rbcL gene sequences were obtained. Phylogenies and haplotype networks were inferred and morphology of gametophytes was studied. Four species were recognized, S. lomentaria, S. promiscuus, S. shibazakiorum, and one species that belongs to a complex of species known as "Scytosiphon Atlantic complex." This complex was known to occur only in the North Atlantic, however, the results found in this study revealed that it has an extended distribution range that includes the southern hemisphere, where its populations have high genetic diversity and unique haplotypes. The morphological differences among the four species were subtle; denoting that previous Scytosiphon records from the SwAO attributed to the renowned S. lomentaria could represent different species. In addition, sex ratio and genome-wide single nucleotide polymorphisms (SNPs) analyses were done for populations of S. promiscuus presumably introduced to the SwAO, and the results indicated that they included female-dominant parthenogenetic populations, which were probably introduced from Japan.

Resolving some of the earliest names for Corallina species (Corallinales, Rhodophyta) in the North Pacific by sequencing type specimens and describing the cryptic C. hakodatensis sp. nov. and C. parva sp. nov.

Partial rbcL sequences from type specimens of three of the earliest described Corallina species showed that C. arbuscula (type locality: Unalaska Island, Alaska, USA) and C. pilulifera (type locality: Okhotsk Sea, Russia) are synonymous, with C. pilulifera as the taxonomically accepted name and that C. vancouveriensis (type locality: Botanical Beach, Vancouver Island, Canada) is a distinct species. To identify molecular species limits and clarify descriptions and distributions of C. pilulifera and C. vancouveriensis, we sequenced and analyzed portions of one mitochondrial and two plastid genes from historical and recent collections. The single-gene phylogenetic reconstructions support the recognition of both species as distinct, as well as two additional species, C. hakodatensis sp. nov. and C. parva sp. nov., which are sister to, and often morphologically indistinguishable from C. pilulifera and C. vancouveriensis, respectively. DNA sequence data currently illustrate that C. pilulifera is found in the cold northern Pacific waters from the Okhotsk Sea of Russia to Hokkaido, Japan, eastward across the Aleutian Islands to Knoll Head, Alaska, and as far south as Nanaimo, British Columbia. Corallina vancouveriensis is distributed as far west as Attu Island in the Aleutian Islands to Sitka, Alaska, and southeasterly at numerous sites from British Columbia to the north of Point Conception, California, USA. The cryptic species C. hakodatensis and C. parva occur sympatrically with their sister species but with narrower ranges. The complex phylogenetic relationships shown by the single gene trees recommend Corallina as a model genus to explore coralline algal biogeography, evolution, and patterns of speciation.

Geographical parthenogenesis in the brown alga Scytosiphon lomentaria (Scytosiphonaceae): Sexuals in warm waters and parthenogens in cold waters.

Geographical parthenogenesis, a phenomenon where parthenogens and their close sexual relatives inhabit distinct geographical areas, has been considered an interesting topic in evolutionary biology. Reports of geographical parthenogenesis from land and freshwater are numerous, but this occurrence has been rarely reported from the sea. Brown algae are mostly marine and are thought to include numerous obligate parthenogens; still, little is known about the distribution, origin and evolution of parthenogens in this group. Here we report a novel pattern of geographical parthenogenesis in the isogamous brown alga Scytosiphon lomentaria. Sex ratio investigation demonstrated that, in Japan, sexual populations grew in the coast along warm ocean currents, whereas female-dominant parthenogenetic populations grew mainly in the coast along a cold ocean current. In the two localities where sexual and parthenogenetic populations were parapatric, parthenogens grew in more wave-exposed areas than sexuals. Population genetic and phylogenetic analyses, including those based on genome-wide single nucleotide polymorphism data, indicated that parthenogens have initially evolved at least twice and subsequent hybridizations between the parthenogens and sexuals have generated multiple new parthenogenetic lineages. The origin of the initial parthenogens is not clear, except that it would not be interspecies hybridization. Interestingly, we found that the production of sex pheromones, which attract male gametes, has been independently lost in the initial two parthenogenetic lineages. This parallel loss of the sexual trait may represent the direct origin of parthenogens, or the regressive evolution of a useless trait under asexuality.

Systematics, distribution, and sexual compatibility of six Scytosiphon species (Scytosiphonaceae, Phaeophyceae) from Japan and the description of four new species.

The brown alga Scytosiphon lomentaria (Scytosiphonaceae, Ectocarpales) has been reported from cold and warm waters worldwide. Although previous molecular phylogenetic studies and crossing experiments have shown that it is a complex of multiple distinct species, the taxonomic position and accurate distribution of each species have remained largely unclear. For the S. lomentaria complex from Japan, our cox1 and rbcL phylogenetic analyses detected six species. In addition to five previously detected species (species Ia-Va), one species (species VI) was newly found in the subtropical area, Okinawa Island. Species VI was recovered as a sister to species Ia-Va in rbcL. Based on the morphological and phylogenetic data, we concluded that species Ia is S. lomentaria, species IIIa is S. promiscuus, and the other four species are newly described: S. shibazakiorum for species IIa, S. tosaensis for species IVa, S. arcanus for species Va, and S. subtropicus for species VI. The cox1-based analysis also showed that S. lomentaria, S. shibazakiorum, and S. promiscuus have worldwide distributions, while the other three species were not found outside of Japan. In Japan, except for S. subtropicus, distributions of these species overlapped. Our crossing experiments showed that gametic incompatibility (pre-zygotic barrier) was less developed between the allopatric species (i.e., S. subtropicus and others) compared with the sympatric species. It may suggest that pre-zygotic barriers have evolved among the sympatric species due to reinforcement.

Cryptic Haploid Stages in the Life Cycle of Leathesia marina (Chordariaceae, Phaeophyceae) Under In Vitro Culture.

We evaluated the life cycle of Leathesia marina through molecular analyses, culture studies, morphological observations, and ploidy measurements. Macroscopic sporophytes were collected from two localities in Atlantic Patagonia and were cultured under long-day (LD) and short-day (SD) conditions. Molecular identification of the microscopic and macroscopic phases was performed through the cox3 and rbcL genes and the phylogeny was assessed on the basis of single gene and concatenated datasets. Nuclear ploidy of each phase was estimated from the DNA contents of individual nuclei through epifluorescence microscopy and flow cytometry. Molecular results confirmed the identity of the Argentinian specimens as L. marina and revealed their conspecificity with L. marina from New Zealand, Germany, and Japan. The sporophytic macrothalli (2n) released mitospores from plurilocular sporangia, which developed into globular microthalli (2n), morphologically similar to the sporophytes but not in size, constituting a generation of small diploid thalli, with a mean fluorescent nuclei cross-sectional area of 3.21 ± 0.7 µm2 . The unilocular sporangia released meiospores that developed two morphologically different types of microthalli: erect branched microthalli (n) with a nuclear area of 1.48 ± 0.07 µm2 that reproduces asexually, and prostrate branched microthalli (n) with a nuclear area of 1.24 ± 0.10 µm2 that reproduces sexually. The prostrate microthalli released gametes in LD conditions, which merged and produced macroscopic thalli with a nuclear cross-sectional area of 3.45 ± 0.09 µm2 . Flow cytometry confirmed that the erect and prostrate microthalli were haploid and that the globular microthalli and macrothalli were diploid.

Contrasting patterns of genetic structure and phylogeography in the marine agarophytes Gelidiophycus divaricatus and G. freshwateri (Gelidiales, Rhodophyta) from East Asia.

The evolutionary and population demographic history of marine red algae in East Asia is poorly understood. Here, we reconstructed the phylogeographies of two upper intertidal species endemic to East Asia, Gelidiophycus divaricatus and G. freshwateri. Phylogenetic and phylogeographic inferences of 393 mitochondrial cox1, 128 plastid rbcL, and 342 nuclear ITS2 sequences were complemented with ecological niche models. Gelidiophycus divaricatus, a southern species adapted to warm water, is characterized by a high genetic diversity and a strong geographical population structure, characteristic of stable population sizes and sudden reduction to recent expansion. In contrast, G. freshwateri, a northern species adapted to cold temperate conditions, is genetically relatively homogeneous with a shallow population structure resulting from steady population growth and recent equilibrium. The overlap zone of the two species roughly matches summer and winter isotherms, indicating that surface seawater temperature is a key feature influencing species range. Unidirectional genetic introgression was detected at two sites on Jeju Island where G. divaricatus was rare while G. freshwateri was common, suggesting the occurrence of asymmetric natural hybrids, a rarely reported event for rhodophytes. Our results illustrate that Quaternary climate oscillations have left strong imprints on the current day genetic structure and highlight the importance of seawater temperature and sea level change in driving speciation in upper intertidal seaweed species.

Correction to: Rapid turnover of life-cycle-related genes in the brown algae.

Following publication of the original article [1], it was noticed that the author names were published with initials instead of full names. The article [1] has been updated.

Rapid turnover of life-cycle-related genes in the brown algae.

BACKGROUND: Sexual life cycles in eukaryotes involve a cyclic alternation between haploid and diploid phases. While most animals possess a diploid life cycle, many plants and algae alternate between multicellular haploid (gametophyte) and diploid (sporophyte) generations. In many algae, gametophytes and sporophytes are independent and free-living and may present dramatic phenotypic differences. The same shared genome can therefore be subject to different, even conflicting, selection pressures during each of the life cycle generations. Here, we analyze the nature and extent of genome-wide, generation-biased gene expression in four species of brown algae with contrasting levels of dimorphism between life cycle generations. RESULTS: We show that the proportion of the transcriptome that is generation-specific is broadly associated with the level of phenotypic dimorphism between the life cycle stages. Importantly, our data reveals a remarkably high turnover rate for life-cycle-related gene sets across the brown algae and highlights the importance not only of co-option of regulatory programs from one generation to the other but also of a role for newly emerged, lineage-specific gene expression patterns in the evolution of the gametophyte and sporophyte developmental programs in this major eukaryotic group. Moreover, we show that generation-biased genes display distinct evolutionary modes, with gametophyte-biased genes evolving rapidly at the coding sequence level whereas sporophyte-biased genes tend to exhibit changes in their patterns of expression. CONCLUSION: Our analysis uncovers the characteristics, expression patterns, and evolution of generation-biased genes and underlines the selective forces that shape this previously underappreciated source of phenotypic diversity.

Parthenogenesis is rare in the reproduction of a sexual field population of the isogamous brown alga Scytosiphon (Scytosiphonaceae, Ectocarpales).

Parthenogenetic development of unfused gametes is commonly observed in laboratory cultures among various brown algal taxa. There is, however, little information on the contribution of parthenogenesis to the reproduction of field populations. In this study, we investigated whether parthenogenesis is present in a sexual population of the isogamous brown alga Scytosiphon with a 1:1 sex ratio. In culture, both female and male gametes showed higher mortality and slower development compared to zygotes. More than 90% of surviving partheno-germlings formed parthenosporophytes irrespective of the culture conditions tested. Therefore, if parthenogenesis occurs in the field, most unfused gametes are expected to form parthenosporophytes. Contrary to this expectation, parthenosporophytes were rare in the field population. We collected 126 sporophytic thalli and isolated and cultured a unilocular sporangium from each of them. We confirmed that cultures of 120 unilocular sporangia produced both female and male gametophytes by the observation of zygotes or amplification of PCR-based sex markers indicating that these sporangia originated from zygotic sporophytes. Only females were detected in cultures from two sporangia and only males from four sporangia suggesting that these sporangia originated from parthenosporophytes. In the Scytosiphon population, although parthenogenesis is observable in culture, our results demonstrate that the contribution of parthenogenesis to reproduction is small (≤4.8%) compared to sexual reproduction. Unfused gametes may not survive to form mature parthenosporophytes in significant numbers in the field partly due to their higher mortality and slower development compared from zygotes.

Parthenogenetic female populations in the brown alga Scytosiphon lomentaria (Scytosiphonaceae, Ectocarpales): decay of a sexual trait and acquisition of asexual traits.

In isogamous brown algae, the sexuality of populations needs to be tested by laboratory crossing experiments, as the sexes of gametophytes are morphologically indistinguishable. In some cases, gamete fusion is not observed and the precise reproductive mode of the populations is unknown. In the isogamous brown alga Scytosiphon lomentaria in Japan, both asexual (gamete fusion is unobservable) and sexual populations (gamete fusion is observable) have been reported. In order to elucidate the reproductive mode of asexual populations in this species, we used PCR-based sex markers to investigate the sex ratio of three asexual and two sexual field populations. The markers indicated that the asexual populations consisted only of female individuals, whereas sexual populations are composed of both males and females. In culture, female gametes of most strains from asexual populations were able to fuse with male gametes; however, they had little to no detectable sexual pheromones, significantly larger cell sizes, and more rapid parthenogenetic development compared to female/male gametes from sexual populations. Investigations of sporophytic stages in the field indicated that alternation of gametophytic and parthenosporophytic stages occur in an asexual population. These results indicate that the S. lomentaria asexual populations are female populations that lack sexual reproduction and reproduce parthenogenetically. It is likely that females in the asexual populations have reduced a sexual trait (pheromone production) and have acquired asexual traits (larger gamete sizes and rapid parthenogenetic development).

Next-Generation Sequencing of an 88-Year-Old Specimen of the Poorly Known Species Liagora japonica (Nemaliales, Rhodophyta) Supports the Recognition of Otohimella gen. nov.

Liagora japonica is a red algal species distributed in temperate regions of Japan. This species has not been collected from its type locality on the Pacific coast of Japan since 1927 and seems to have become extinct in this area. For molecular characterization of L. japonica, we extracted DNA from the topotype material of L. japonica collected in 1927, analyzed seven genes using Illumina next-generation sequencing, and compared these data with sequences from modern samples of similar red algae collected from the Japan Sea coast of Japan. Both morphological and molecular data from modern samples and historical specimens (including the lectotype and topotype) suggest that the specimens from the Pacific and Japan Sea coasts of Japan should be treated as a single species, and that L. japonica is phylogenetically separated from the genus Liagora. Based on the phylogenetic results and examination of reproductive structures, we propose Otohimella japonica gen. et comb. nov., characterized morphologically by diffuse carposporophytes, undivided carposporangia, and involucral filaments initiated only from the cortical cell on the supporting cell.

Evolution of Green Plants Accompanied Changes in Light-Harvesting Systems.

Photosynthetic organisms have various pigments enabling them to adapt to various light environments. Green plants are divided into two groups: streptophytes and chlorophytes. Streptophytes include some freshwater green algae and land plants, while chlorophytes comprise the other freshwater green algae and seawater green algae. The environmental conditions driving the divergence of green plants into these two groups and the changes in photosynthetic properties accompanying their evolution remain unknown. Here, we separated the core antennae of PSI and the peripheral antennae [light-harvesting complexes (LHCs)] in green plants by green-native gel electrophoresis and determined their pigment compositions. Freshwater green algae and land plants have high Chl a/b ratios, with most Chl b existing in LHCs. In contrast, seawater green algae have low Chl a/b ratios. In addition, Chl b exists not only in LHCs but also in PSI core antennae in these organisms, a situation beneficial for survival in deep seawater, where blue-green light is the dominant light source. Finally, low-energy Chl (red Chl) of PSI was detected in freshwater green algae and land plants, but not in seawater green algae. We thus conclude that the different level of Chl b accumulation in core antennae and differences in PSI red Chl between freshwater and seawater green algae are evolutionary adaptations of these algae to their habitats, especially to high- or low-light environments.

Trigonelline and other betaines in species of Laminariales.

A collection of Laminariales species was made with examples in each of the presently recognized families of the order. Extracts of each species were examined for betaines, using primarily 'H NMR spectroscopy for their identification. Glycinebetaine was detected in all species tested and would appear to be a consistent feature of the Laminariales. Gamma-Aminobutyric acid betaine was found in all species of Laminaria examined and in three of the five Saccharina species (family Laminariaceae), but was not detected in species of either other genera of the family or in those of other Laminariales families. Trigonelline was found in some Laminaria and Saccharina species, as well as in the north Pacific species Postelsia palmaeformis (Laminariaceae), Pseudochorda nagaii (Pseudochordaceae) and Akkesiphycus lubricus (Akkesiphycaceae).

Structural Confirmation of a Unique Carotenoid Lactoside, P457, in Symbiodinium sp. Strain nbrc 104787 Isolated from a Sea Anemone and its Distribution in Dinoflagellates and Various Marine Organisms.

The molecular structure of the carotenoid lactoside P457, (3S,5R,6R,3'S,5'R,6'S)-13'-cis-5,6-epoxy-3',5'-dihydroxy-3-(ß-d-galactosyl-(1→4)-ß-d-glucosyl)oxy-6',7'-didehydro-5,6,7,8,5',6'-hexahydro-ß,ß-caroten-20-al, was confirmed by spectroscopic methods using Symbiodinium sp. strain NBRC 104787 cells isolated from a sea anemone. Among various algae, cyanobacteria, land plants, and marine invertebrates, the distribution of this unique diglycosyl carotenoid was restricted to free-living peridinin-containing dinoflagellates and marine invertebrates that harbor peridinin-containing zooxanthellae. Neoxanthin appeared to be a common precursor for biosynthesis of peridinin and P457, although neoxanthin was not found in peridinin-containing dinoflagellates. Fucoxanthin-containing dinoflagellates did not possess peridinin or P457; green dinoflagellates, which contain chlorophyll a and b, did not contain peridinin, fucoxanthin, or P457; and no unicellular algae containing both peridinin and P457, other than peridinin-containing dinoflagellates, have been observed. Therefore, the biosynthetic pathways for peridinin and P457 may have been coestablished during the evolution of dinoflagellates after the host heterotrophic eukaryotic microorganism formed a symbiotic association with red alga that does not contain peridinin or P457.

NEW EVIDENCE FOR MORPHOLOGICAL AND GENETIC VARIATION IN THE COSMOPOLITAN COCCOLITHOPHORE EMILIANIA HUXLEYI (PRYMNESIOPHYCEAE) FROM THE COX1b-ATP4 GENES(1).

Emiliania huxleyi (Lohmann) W. W. Hay et H. Mohler is a cosmopolitan coccolithophore occurring from tropical to subpolar waters and exhibiting variations in morphology of coccoliths possibly related to environmental conditions. We examined morphological characters of coccoliths and partial mitochondrial sequences of the cytochrome oxidase 1b (cox1b) through adenosine triphosphate synthase 4 (atp4) genes of 39 clonal E. huxleyi strains from the Atlantic and Pacific Oceans, Mediterranean Sea, and their adjacent seas. Based on the morphological study of culture strains by SEM, Type O, a new morphotype characterized by coccoliths with an open central area, was separated from existing morphotypes A, B, B/C, C, R, and var. corona, characterized by coccoliths with central area elements. Molecular phylogenetic studies revealed that E. huxleyi consists of at least two mitochondrial sequence groups with different temperature preferences/tolerances: a cool-water group occurring in subarctic North Atlantic and Pacific and a warm-water group occurring in the subtropical Atlantic and Pacific and in the Mediterranean Sea.

Isolation and characterization of a novel thraustochytrid-like microorganism that efficiently produces docosahexaenoic acid.

A thraustochytrid-like microorganism (strain 12B) was isolated from the mangrove area of Okinawa, Japan. On the basis of its ectoplasmic net structure and biflagellate zoospores we determined strain 12B to be a novel member of the phylum Labyrinthulomycota in the kingdom Protoctista. When grown on glucose/seawater at 28 degrees C, it had a lipid content of 58% with docosahexaenoic acid (DHA; 22:6 n-3) at 43% of the total fatty acids. It had a growth rate of 0.38 h(-1). The DHA production rate of 2.8 +/- 0.7 g l(-1) day(-1) is the highest value reported for any microorganism.