Asexual life history by quadriflagellate swarmers of Ulva spinulosa (Ulvales, Ulvophyceae)

This is the first report of a Ulva species reproducing asexually solely by quadriflagellate swarmers. Ulva spinulosa Okamura et Segawa specimens were collected from Ukibuchi on the Pacific coast of Kochi Prefecture, southern Japan. Quadriflagellate swarmers were released from these specimens. The swarmers showed negative phototaxis before settlement. Thalli cultured from these swarmers also released quadriflagellate swarmers in culture. Microspectrophotometric studies demonstrated equivalent DNA in nuclei of vegetative cells in thalli of U. spinulosa and in sporo‐phytes of the other Ulva species with sexual life history (U. fasciata Delile). Furthermore, the quadriflagellate swarmers of U. spinulosa had the same DNA value, demonstrating that the quadriflagellate swarmers are produced without meiosis.     

A new green-tide-forming alga, Ulva ohnoi Hiraoka et Shimada sp. nov. (Ulvales, Ulvophyceae) from Japan

Ulva ohnoi Hiraoka et Shimada sp. nov. (Ulvales, Ulvophyceae) is described from southern and western Japan and is characterized by the following combination of features: (i) the large, fragile, easily torn thalli, which are 30–55 μm thick in the upper and middle regions and often have microscopic marginal teeth; (ii) the production of zoids in the upper marginal region; (iii) a regular alternation of dioecious gametophytes and a sporophyte; (iv) the production of free‐floating thalli from torn‐off attached thalli, which reproduce vegetatively by fragmentation and form green tides in summer to autumn; (v) disorderly arranged cells that are polygonal or quadrangular in the upper and middle regions; and (vi) the chloroplast covering the outer face of cell, with 1–3 pyrenoids. Ulva ohnoi differs from U. armohcana Dion et al., U. fasciata Delile, U. reticulata Forsskal, U. scandinavica Eliding and U. spiulosaOkamura et Segawa, which all possess microscopic marginal serrations, in thallus shape, cell shape or life history pattern. It is also distinguished from morphologically similar species by sequences of the nuclear encoded internal transcribed spacers and the 5.8S ribosomal RNA gene and the plastid encoded large subunit of ribulose‐l,5‐bisphosphate carboxylase/oxgenase gene. Furthermore, crossing tests demonstrate that there is a reproductive boundary between U. ohnoi and the most closely related species, U. fasciata and U. reticulata.     

Regeneration of plantlets fromEnteromorpha (Ulvales,Chlorophyta) protoplasts in axenic culture

High yields of protoplasts have been obtained from vegetative thalli of three species ofEnteromorpha by enzymatic degradation of the cell wall. Several commercial and crude enzymes prepared from the digestive system and hepatopancrease of abalone and top-shell were tested at different concentrations and combinations to evaluate the yield. Commercial enzymes in combination with either abalone or top-shell crude enzymes, consistently produced a high yield of protoplasts from all three species. High regeneration rate (85–95%) occurred in the protoplasts cultured at a density greater than 1.72 × 10³ cells cm⁻² at 20 and 25°C. Light intensities tested in the present study did not affect protoplast wall formation and regeneration. Protoplasts, after regenerating the cell wall, followed different types of developmental patterns under identical culture conditions. In one type some cells underwent repeated cell divisions and formed a round and oval shaped hollow thallus with a single layer of cells. In the second type many cells underwent one or two cell divisions (occasionally no division) and soon matured and discharged many motile spores, which on germination grew into normal plantlets. In the third type some cells divided irregularly to form a mass of callus-like cells (exceptE. prolifera). Culture medium supplemented with either mannitol, sorbitol, dextrose, saccharose or NaCl at higher concentrations (> 0.4 M) inhibited cell division and further differentiation in all species. author for correspondence     

Induction of fast-growing and morphologically different strains through intergeneric protoplast fusions of Ulva and Enteromorpha (Ulvales,Chlorophyta)

Isolated protoplasts of Ulva pertusa and Enteromorpha prolifera were electrically fused. Treatment of protoplasts in 1% protease for 15–20 min prior to fusion enhanced fusion ability. Protoplasts from each fusion partner were mixed together in 1:1 ratio in low conductivity electrofusion solution at a density of 1 × 10⁵ cells ml⁻¹ before subjecting them to electrofusion. The protoplasts were aligned in AC field (1MHz, 25 V for 10–15 s) and subsequently fused by a high intensity single DC pulse of 250 V for 25 μs duration. Fusion buffer supplemented with 1 mM calcium and 1 mM magnesium yielded optimum fusion frequencies (about 18–24%). Entrapment of fusion treated cells inside agarose/agar plate facilitated marking and regeneration of fusion products. The regeneration patterns of fused protoplasts were similar to normal (unfused) protoplast development. Most of the regenerated plants from fusion products had a thallus similar to either U. pertusa type or E. prolifera type. Although some of the plants of the former were morphologically similar to U. pertusa, but most had a higher growth rate (1.9 to 1.5 times) than U. pertusa. Furthermore the thallus of some plants had a characteristic irregular and dentate margin, which was never observed in the parental type.     

Molecular and morphological diversity of Narragansett Bay (RI,USA) Ulva (Ulvales,Chlorophyta) populations

Macroalgal bloom‐forming species occur in coastal systems worldwide. However, due to overlapping morphologies in some taxa, accurate taxonomic assessment and classification of these species can be quite challenging. We investigated the molecular and morphological characteristics of 153 specimens of bloom‐forming Ulva located in and around Narragansett Bay, RI, USA. We analyzed sequences of the nuclear internal transcribed spacer 1 region (ITS1) and the chloroplast‐encoded rbcL; based on the ITS1 data, we grouped the specimens into nine operational taxonomic units (OTUs). Eight of these OTUs have been previously reported to exist, while one is novel. Of the eight OTUs, all shared sequence identity with previously published sequences or differed by less than 1.5% sequence divergence for two molecular markers. Previously, 10 species names were reported for Ulva in Rhode Island (one blade and nine tube‐forming species) based upon morphological classification alone. Of our nine OTUs, three contained blade‐forming specimens (U. lactuca, U. compressa, U. rigida), one OTU had a blade with a tubular stipe, and six contained unbranched and/or branched tubular morphologies (one of these six, U. compressa, had both a blade and a tube morphology). While the three blade‐forming OTUs in Narragansett Bay can frequently be distinguished by careful observations of morphological characteristics, and spatial/temporal distribution, it is much more difficult to distinguish among the tube‐forming specimens based upon morphology or distribution alone. Our data support the molecular species concept for Ulva, and indicate that molecular‐based classifications of Ulva species are critical for proper species identification, and subsequent ecological assessment or mitigation of Ulva blooms.     

Molecular assessment of Ulva (Ulvales,Chlorophyta) diversity in Vietnam including the new species U. vietnamensis

Species diversity of Ulva in Vietnam was investigated using three commonly used genetic markers, the nuclear encoded rDNA ITS region and the plastid encoded rbcL and tufA genes. Single locus species delimitation methods, complemented with morphological and ecological information resulted in the delimitation of 19 species. This diversity is largely incongruent with the traditional understanding of Ulva diversity in Vietnam. Only four species identified in this study, U. lactuca, U. reticulata, U. spinulosa, and U. flexuosa, have been previously reported, and seven species, U. ohnoi, U. tepida, U. chaugulii, U. kraftiorum, U. meridionalis, U. limnetica, and U. aragoënsis, are recorded for the first time from Vietnam. Seven genetic clusters could not be associated with species names with certainty. A new species, U. vietnamensis, is described from marine to brackish coastal areas from southern Vietnam based on its morphological and molecular distinctiveness from the currently known Ulva species. A comparison with recent molecular-based studies of Ulva diversity showed that species composition in Vietnam is similar to that of adjacent countries, including Japan, China, as well as Australia. Our study emphasizes the importance of molecular data in the assessment of Ulva diversity, and indicates that a lot of diversity may still remain to be discovered, especially in tropical regions.     

Taxonomic study of the genus Spongomorpha (Acrosiphoniales,Chlorophyta) in Japan. I. Spongomorpha spiralis*

The morphology, life history, and the geographical distribution of Spongomorpha spiralis Sakai in Hokkaido, Japan, was studied. The thallus is characterized by its digitate tufts (bundles) of slender filaments with hooked branchlets. The diameter of the filaments is usually 20-50 μm in the basal portion, 50-130 (70-110) μm in the middle portion of main axes, and 30-150 μm in the upper portion. The diameter of the middle portion of the main axes varies from locality to locality. At Muroran, for example, the range is 70-130 μm (average approximately 100 μm), while at other localities it is 50–100 μm (average approximately 70 μm). The diameter of filaments in the upper portion decreases toward the end of the growing season. The cells are multinucleate with four chromosomes in each nucleus of the haploid plant. The manner of cell division is identical to the process reported previously for the Spongomorpha-Acrosiphonia complex. Sexual reproduction is isogamous with anteriorly biflagellate gametes. Plants are unisexual. The life history involves an alternation of heteromorphic generations: the gametophytic phase is a macroscopic, filamentous thallus and the sporophytic phase is a microscopic, ellipsoidal or club-shaped cell. Optimal culture condition for growth and reproduction of both stages was 5°C and long daylength. The gametophyte developed abnormally at 15°C. The sporophyte developed normally at 15°C, but did not produce zoospores.     

Taxonomic and ecological profile of `green tide' species of Ulva (Ulvales,Chlorophyta) in central Philippines

Hydrobiologia - Ulva spp. are common in the intertidal zones of the Philippines, but, at certain times, could over-proliferate producing blooms or `green tide' in some protected bays. In Mactan...     

Cryptic diversity of Ulva (Ulvales,Chlorophyta) in the Great Bay Estuarine System (Atlantic USA): introduced and indigenous distromatic species

Distromatic foliose blades of the algal genus Ulva are notoriously difficult to identify due to their simple morphologies and few diagnostic characteristics that often exhibit intraspecific variation and interspecific overlap. Hence, species differentiation is difficult and diversity estimates are often inaccurate. Two major goals of this study were to assess the diversity of distromatic Ulva spp. in the Great Bay Estuarine System (GBES) of New Hampshire and Maine, USA, and to compare historical and present day records of these species. Molecular analysis (using ITS sequences) of field-collected specimens revealed four distinct taxa: Ulva lactuca, U. rigida, U. compressa, and U. pertusa. Prior to molecular screening, Ulva lactuca was the only distromatic Ulva species reported for the GBES. Ulva pertusa and the foliose form of U. compressa are newly recorded for the Northwest Atlantic, and the range of U. rigida has been extended. Molecular analysis of historical herbarium voucher specimens indicates that U. rigida, U. pertusa, and the foliose form of U. compressa have been present in the GBES since at least 1966, 1967, and 1972, respectively. The distromatic morphotype of U. compressa is found only in low salinity areas, which suggests that salinity may influence its morphological development. Molecular and morphological evaluations are critical if we are to distinguish between cryptic taxa, accurately assess biodiversity, and effectively monitor the spread of non-indigenous macroalgae.     

Vertical distribution of chitons (Mollusca: Polyplacophora) in the rocky intertidal zone of central Chile

The patterns of vertical distribution of chitons in various exposed rocky shore habitats of central Chile are described. The density and size of the individuals were determined at different levels on intertidal rock walls, in pools and on boulders. A group of four species was found on rock walls while another group of four species was found in tidal pools and on boulders. The species showed vertical separation on rock walls and segregation according to a range of increasing frequency of water movement in pools and on boulders. Only the two species with largest adult individuals (Acanthopleura echinata (Barnes) and Chitonlatus Sowerby) showed vertical segregation in size. Small-sized species and small individuals of large-sized species were found higher on rock walls or in zones where water movement is less; large chitons occur in zones of greatest water impact. This suggests that water movement may be a significant factor in setting the lower limits of distribution of these herbivores. Some of the species examined were extremely abundant (600·m^-2) with densities of one to two orders of magnitude greater than for other chiton species elsewhere.     

The natural history, thermal physiology, and ecological impacts of intertidal mesopredators, Oedoparena spp. (Diptera: Dryomyzidae)

Abstract. The ecological roles of small (1–1000 mg) predators in benthic marine systems are poorly understood. We investigated the natural history and predatory impact of one group of such mesopredators—larvae of dipteran flies in the genus Oedoparena —which prey on intertidal barnacles. We 1) quantified patterns of larval Oedoparena distribution and abundance in the Northwest Straits of Washington State, USA, 2) determined larval physiological tolerance limits in the laboratory, and 3) conducted a manipulative field experiment to assess the role of microhabitat temperature on predation rates in Oedoparena . Members of Oedoparena in Washington are univoltine, with peak larval abundance in late spring and early summer. Infestation frequencies in the barnacles Balanus glandula and Chthamalus dalli were as high as 22% and 35%, respectively. In laboratory studies, larvae of O . glauca were able to tolerate temperatures up to 37°C; however, this temperature is often exceeded in high intertidal habitats. In a field manipulation using experimental shades, we demonstrate that the alleviation of physiological stress greatly increased the abundance of larvae of Oedoparena spp. As a result of increased larval densities under shades, adult B. glandula mortality increased from 5% to nearly 30%, and C. dalli mortality increased from less than 20% to over 60%. Because high intertidal barnacles serve as food and habitat for a diverse array of species, Oedoparena spp. have the potential to play a major role in structuring high intertidal communities, particularly in cooler microhabitats.     

cDNA cloning of a lectin-like gene preferentially expressed in freshwater from the macroalga Ulva limnetica (Ulvales, Chlorophyta)

The macroalgal species Ulva limnetica Ichihara et Shimada was investigated to understand the molecular mechanism of its tolerance or adaptation to freshwater. We detected a 19 kDa protein by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, which accumulated in greater amounts in freshwater conditions compared with marine conditions. The band was excised and the partial amino acid sequence was determined by Edoman degradation. Based on the sequences, we isolated the corresponding cDNA by the rapid amplification of cDNA ends (RACE) technique. The constructed, full-length cDNA was 1272 bp in length, consisting of 198 bp 5'-non-coding region, an open reading frame of 840 bp (279 amino acids), 233 bp 3'-non-coding region and poly (A) tail. The protein encoded by the cDNA showed 30% identity and 45% similarity to lectin isolated from Ulva pertusa Kjellman, and we named this gene ULL (encoding Ulva limnetica lectin-like protein). Northern blot analysis demonstrated that the expression level of the ULL in the freshwater-cultured sample was higher than in the seawater-cultured sample.     

Morphological features and chromosome numbers in cultures of five Cephaleuros species (Trentepohliaceae,Chlorophyta) from Japan

The morphological features and chromosome numbers were examined in cultures of five species of Cephaleuros; C. aucubae, C. biolophus, C. japonicus, C. microcellularis, and C. virescens, collected from Japan. On agar media of Bold's basal medium and CA medium, radial growth of the algal colonies was vigorous in C. virescens, moderate in C. aucubae, and poor in C. microcellularis, but varied from poor to moderate in isolates of C. biolophus and C. japonicus. Filaments of C. virescens branched at an angle less than 40° and just below the cross walls, while those of the other species branched at an angle greater than 40° without relation to the cross walls. The length of filamentous cells was significantly smaller in C. microcellularis than in the other species. The chromosome number differed with the species: n = 22 in C. aucubae, n = 34 in C. biolophus, n = 18 in C. japonicus, n = 12 in C. microcellularis, and n = 24 in C. virescens. The chromosome number did not vary between isolates from filaments and gametes in C. aucubae and C. biolophus and among those from filaments, gametes, and zoospores in C. japonicus and C. virescens.     

The contribution of haploids, diploids and clones to fine-scale population structure in the seaweed Cladophoropsis membranacea (Chlorophyta)

Local populations of Cladophoropsis membranacea exist as mats of coalesced thalli composed of free-living haploid and diploid plants including clonally reproduced plants of either phase. None of the phases are morphologically distinguishable. We used eight microsatellite loci to explore clonality and fine-scale patch structure in C. membranacea at six sites on the Canary Islands. Mats were always composites of many individuals; not single, large clones. Haploids outnumbered diploids at all sites (from 2:1 to 10:1). In both haploid and diploid plants, genetic diversity was high and there was no significant difference in allele frequencies. Significant heterozygote deficiencies were found in the diploid plants at five out of six sites and linkage disequilibrium was associated with the haploid phase at all sites. Short dispersal distances of gametes/spores and small effective population sizes associated with clonality probably contribute to inbreeding. Spatial autocorrelation analysis revealed that most clones were found within a radius of approximately 60 cm and rarely further than 5 m. Dominance of the haploid phase may reflect seasonal shifts in the relative frequencies of haploids and diploids, but may alternatively reflect superiority of locally adapted and competitively dominant, haploid clones; a strategy that is theoretically favoured in disturbed environments. Although sexual reproduction may be infrequent in C. membranacea, it is sufficient to maintain both life history phases and supports theoretical modelling studies that show that haploid-diploid life histories are an evolutionarily stable strategy.     

Life history of Chnoospora implexa (Chnoosporaceae, Phaeophyceae) in culture

The life history of the brown alga Chnoospora implexa J. Agardh (Chnoosporaceae, Scytosiphonales) from Japan was studied in laboratory cultures. This species showed a heteromorphic and diphasic life history, alternating between erect gametophytes and discoid sporophytes. The gametophytes were dioecious and produced isogametes. The zygotes developed into sporophytes at 20°C under long‐day conditions, which formed plurilocular zoidangia. Zoids released from the plurilocular zoidangia developed again into sporophytes that always formed plurilocular zoidangia at 20°C and 25°C in long‐day conditions, and mainly unilocular zoidangia at 25°C in short‐day conditions. Zoids released from unilocular zoidangia developed into dioecious gametophytes. At 15°C zygotic erect thalli were formed and were revealed to be diploid by microspectrofluorometric measurements of nuclear DNA contents. The development and reproduction of unfused gametes were similar to those of zygotes. Some strains showed a direct‐type life history; gametophytic thalli were produced, but not via a sporophytic phase.     

A life table study on three genetic strains of Ophryotrocha diadema (Polychaeta,Dorvilleidae)

A life-table study was performed on three strains of Ophryotrocha diadema. The wild type strain, YY (yellow eggs), was compared with a recessive mutant strain, ww (white eggs), and the F₁, hybrid, Yw.

The viability of the YY strain was greater than that of the ww strain. The hybrids were intermediate. Expectation of life. e., of zygotes were: YY 27.9, Yw 19.1, and ww 15.1 wk, respectively. The total outcome of reproduction followed the same pattern; the net reproductive rates, R ₀, were related as 1:0.62:0.47 for YY: Yw: ww.

Reproduction began in the fourth week. A maximum reproductive output during the next 4 wk was followed by a fast decline. The intrinsic rate of natural increase, r, was 0.880 per week for the YY strain, 0.903 for the Yw strain and 0.872 for the ww strain. For all strains, the first 7 wk of reproduction contributed more than 99% of the Lotka equation σ e ^?rx l _x m _x = l, i.e., l _x m _x values at older ages would not substantially affect population growth. In this 7–wk period (and the previous nonreproductive weeks), mortality differences were small among strains. During the period of maximum reproduction the average size of egg masses was approximately the same in all strains. The heterozygote superiority was due to shorter intervals between successive spawnings.

It is suggested that in the opportunistic species O. diadema deviations from an r–selected reproductive pattern are due to small adult size and to its ancestry from a mainly K–selected, very old polychaete group.

O. diadema has proven to be a useful test animal for marine pollution research. Observed effects on survival and reproduction have been summarized in a model to demonstrate population consequences of sublethal pollutant levels.     

Effects of Chiton granosus (Frembly, 1827) and other molluscan grazers on algal succession in wave exposed mid-intertidal rocky shores of central Chile

Molluscan grazers can have important effects on the abundance, colonization rates, and successional pathways of algal assemblages and the entire intertidal community. In general, early successional algae are more readily consumed than corticated algae and kelps, which usually get established later in the community succession. To generalize, however, the effect of different grazers on algal assemblages must be examined on different coasts and under different scenarios. This information could help us understand the mechanisms of ecosystem processes and situations in which general models do not apply. Along the coast of Chile, humans harvest large keyhole limpets, which seem to be the only invertebrate grazers capable of controlling the dominant corticated alga Mazzaella laminarioides, a canopy-forming species that can cover extensive areas of the mid intertidal zone. In this scenario, where large limpets are harvested, the overall effects of the diverse molluscan assemblage of limpets, chitons and snails on algal succession and on corticated algae in particular are not clear. We conducted a 26-month-long experiment to evaluate the effects of molluscan grazers on mid-intertidal algal succession and to isolate the effects of Chiton granosus, the most conspicuous member of the assemblage at these tidal elevations. At sites heavily impacted by humans the molluscan grazer assemblage had strong negative effects on colonization and abundance of green algae such as ulvoids and Blidingia minima. In doing so, the grazer assemblage had a strong negative indirect effect on the establishments of chironomid fly larvae, which were only observed on green algal mats and rarely on bare rock. No significant effects were detected on epilithic microalgae, and effects on sessile invertebrates were highly variable over space and time. C. granosus also had significant negative effects on green algae but did not account for the total grazing pressure exerted by the guild. Limited foraging excursions (ca. 35 cm) from refuges and moderate site (crevice) fidelity in this species may contribute to the patchiness in green algal distribution observed in the field. Nearly 13 months after rock surface were experimentally cleared, M. laminarioides appeared in all experimental plots, but increased over three times faster in enclosures containing C. granosus than in exclosures plots or controls, suggesting that moderate levels of herbivory could actually facilitate the establishment of this alga in the succession and that the green algal cover found in the absence of grazers may delay its establishment.     

Morphology and life history of Scytosiphon canaliculatus comb. nov. (Scytosiphonales,Phaeophyceae) from Japan*

Morphology of field material and life history in culture were studied in Scytosiphon canaliculatus (Setchell et Gardner) comb. nov. from northern Japan. Erect gametophytes of S. canaliculatus are cylindrical, tubular, up to 7 mm wide and 40 cm long, and without regular constrictions. S. canaliculatus has pronounced anisogamy and ascocysts accompanied with plurilocular gametangia. The life history of S. canaliculatus showed an alternation between erect gametophytes and crustose prostrate sporophytes bearing unilocular sporangia. Since field sporophytes of S. canaliculatus were found to be identical with Hapterophycus canaliculatus Setchell et Gardner (Ralfsiaceae, Phaeophyceae), it is proposed to transfer H. canaliculatus to the genus Scytosiphon. In the field, gametophytes with plurilocular gametangia appeared in spring and disappeared in summer. Sporophytes with unilocular sporangia were collected in late autumn and winter. Unilocular sporangia were produced at 15°C in short-day culture conditions and unispores developed into erect gametophytes at 5–15°C. It is suggested that the seasonal Occurrence of gametophytes in the field is due to the seasonal formation of unilocular sporangia, which is regulated by temperature and photoperiod.