ASSESSMENT OF TEMPERATURE AND NUTRIENT LIMITATION ON SEASONAL DYNAMICS AMONG SPECIES OF SARGASSUM FROM A CORAL REEF IN SOUTHERN TAIWAN1

The effects of temperature and nutrient availability on seasonal abundance were compared among Sargassum berberifolium J. Ag., S. polycystum C. Ag., S. siliquosum J. Ag., and S. sandei from a reef in southern Taiwan. Growth temperature limits and optimum were variable between species and between developmental stages. Growth temperature ranges agree with Sargassum periodicity except S. sandei. Sargassum siliquosum can tolerate high temperature as indicated by higher optimum temperature and tolerance limits. Temporal changes in tissue nutrient content and alkaline phosphatase activity and the results from nutrient enrichment bioassay show that S. sandei, S. berberifolium, and S. polycystum are P limited in the early growth period and then N limited, whereas S. siliquosum showed a contrasting trend, reflecting the type of nutrient limitation is variable by time and between species. A smaller nutrient threshold for maximum growth in S. berberifolium, S. polycystum, and S. siliquosum than in S. sandei and higher growth rate in S. berberifolium and S. polycystum indicate the higher growth vigor and nutrient utilization efficiency, the higher abundance. High N reserves in S. berberifolium and S. polycystum exposed to elevated seawater N in the early growth period supported the subsequent growth. The multiple regression analysis indicates that percent cover of S. siliquosum increased with increasing water temperature and dissolved inorganic nitrogen concentrations, whereas that of S. polycystum decreased with increasing water temperature. In conclusion, seasonal dynamics of Sargassum spp. from southern Taiwan are attributable to species and temporal variations in temperature limitation and nutrient utilization strategy.     

Use of RAPD in differentiation of selected species of <Emphasis Type="Italic">Sargassum</Emphasis> (Sargassaceae,Phaeophyta)

Sargassum C. Agardh is one of the most common but little understood genera of Phaeophyta in Malaysia. The difficulty in species delineation is due to morphological plasticity. A combination of morphology and random amplified polymorphic DNA (RAPD) studies of selected Sargassum species was carried out to have a better understanding of the taxonomy. Primer OPA13 was found to be good for discriminating between Sargassum species. Sargassum binderi was shown to be different from S. oligocystum (S_D>0.5 = 14.11%), indicating the importance of the vesicle and receptacle in species differentiation. S. baccularia was clearly separated out from S. polycystum and S. stolonifolium using primer OPA13. RAPD analysis showed that the presence of the stolon is an important character for separating S. baccularia (no stolon) from S. polycystum (stolon) and S. stolonifolium (stolon). Presented at the 6th Meeting of the Asian Pacific Society of Applied Phycology, Manila, Philippines.     

MOLECULAR PHYLOGENY OF GRACILARIA SPECIES INFERRED FROM MOLECULAR MARKERS BELONGING TO THREE DIFFERENT GENOMES1

The nucleotide sequence data of molecular markers 18S rRNA, RUBISCO spacer, and cox2‐3 intergenic spacer were integrated to infer the phylogeny of Gracilaria species, collected from the western coast of India, reducing the possibility of misidentification and providing greater phylogenetic resolution. A phylogenetic tree was constructed using cox2‐3 and RUBISCO spacer sequences, exhibiting the same clustering but differing slightly from that of the rRNA‐based phylogenetic tree. The phylogeny inferred from the combined data set confers an analogous pattern of clustering, compared with those of trees constructed from individual data sets. The combined data set resulted in a phylogeny with better resolution, which supported the clade with higher consistency index, retention index, and bootstrap values. It was observed that Gracilaria foliifera (Forssk.) Børgesen is closer to G. corticata (J. Agardh) J. Agardh varieties, while G. salicornia (C. Agardh) E. Y. Dawson and G. fergusonii J. Agardh both originated from the same clade. The position of G. textorii (Suringar) De Toni faltered and toppled between G. salicornia and G. dura (C. Agardh) J. Agardh; however, G. gracilis (Stackh.) M. Steentoft, L. M. Irvine et W. F. Farnham was evidently distant from the rest of the species.     

Levels and distribution patterns of mitochondrial cox3 gene variation in brown seaweed,Sargassum polycystum C. Agardh (Fucales,Phaeophyceae) from Southeast Asia

Sargassum polycystum C. Agardh is one of the most abundant marine brown algae and is distributed widely in warm and temperate waters, particularly in the Indo-west Pacific region and Australia. Although its commercial potential and ecological and evolutionary importance are recognized, many pivotal aspects of its biology remain unexplored. Current knowledge of the historical biogeographical affinities and patterns is limited, but some data are available about its genetics, the genetic variation among populations, and spatial patterns. This study aimed to analyze the genetic population structure and distribution patterns of S. polycystum populations in 13 different locations from Indonesia to Japan using the mitochondrial gene cox3. The seven haplotypes of cox3 identified in this study indicated a low level of genetic diversity. Homogeneity of this haplotype was observed particularly in the Gulf of Thailand, Cambodia, and Japan, whereas higher haplotype diversity was found in Phuket (Thailand), Bali (Indonesia), and Singapore. Those data suggest that S. polycystum is likely to have expanded from the south of Indonesia and the west of the Malay Peninsula towards the northeast of the region. Geological studies showed that Sundaland, now corresponding to the Gulf of Thailand, was submerged due to sea level rises after the last glacial period. Therefore, the decrease in the genetic diversity of S. polycystum populations is interpreted here as a population expansion after the rise in sea levels.     

The yield,physical and chemical properties of agar gel from Gracilaria species (Gracilariales,Rhodophyta) of the Kenya coast

Six species of Gracilaria, G. corticata J. Agardh, G. crassa Harvey, G. millardetii J. Agardh, G. salicornia (J. Ag.) Dawson, G. verrucosa (Huds.) Papenfuss and Gracilaria sp, collected from different stations along the Kenya coast were studied. The yield of hot water native agar extract ranges from 8.1–30% of dry weight, with G. verrucosa and G. salicornia having the greatest and the least yield, respectively. The gel-strength of 1.5% agar solution was highest in G. verrucosa (220 g cm^–2) and lowest in G. corticata (< 60 g cm – 2) whereas the highest gelling temperature was recorded for Gracilaria sp. (40.4 °C) and the lowest in G. verrucosa (28.9 °C). 3,6 anhydrogalactose content was the highest in G. verrucosa and the lowest in G. corticata while sulphate content was higher in the latter.     

Molecular characterisation of Sargassum polycystum and S. siliquosum (Phaeophyta) by polymerase chain reaction (PCR) using random amplified polymorphic DNA (RAPD) primers

Genomic DNA was extracted from 13 samples of Sargassum polycystum and S. siliquosum collected from various localities around Peninsular Malaysia and Singapore by using four different extraction methods. The yields and the suitability of the DNA to be used as template for the polymerase chain reaction (PCR) was compared. DNA samples were subjected to PCR analysis by using random primers. Only DNA samples that were extracted using the CTAB method were successfully amplified by random amplified polymorphic DNA (RAPD)-PCR. Five of 31 random primers (OPA02, OPA03, OPA04, OPA13 and OPM10) tested amplified sequences of DNA from the DNA samples. Reproducible, amplified products were obtained using these primers and showed some potential to be useful in discriminating individual samples within the genus, in determining relationships between species within a genus and in developing individual fingerprints for individual samples.     

Seasonal studies on the alginate and its biochemical composition I: Sargassum polycystum (Fucales), Phaeophyceae

Investigations were made on the brown seaweed Sargassum polycystum C. Agardh collected from Rameswaram Coast, Tamil Nadu. The alginates extracted from ‘leaf’, ‘stem’ and entire thallus of S. polycystum were investigated for their viscosity and chemical constituents, namely β‐D‐mannuronic acid (M‐block), α‐L‐guluronic acid (G‐block) and alternating sequences of β‐D‐mannuronic acid and α‐L‐guluronic acid (MG‐block) for six different seasons between August 1998 and November 1999. Significant seasonal variation (P< 0.05) was observed with high yield of alginate in February. The alginate extracted from the ‘leaf’ region showed a maximum yield whereas the ‘stem’ region exhibited maximum viscosity. The amount of G‐block was found to be more than M‐ and MG‐blocks in all the samples tested. The amount of G‐block was high in ‘stem’ followed by leaf and entire thallus. A positive correlation was recorded between viscosity and G‐block. Among the three alginates, the ratio of M/G was low in the ‘stem’ followed by ‘leaf’ and entire thallus.     

Impaired growth and survival of tropical macroalgae (Sargassum spp.) at elevated temperatures

Increasing ocean temperatures associated with ongoing climate change have resulted in regional reductions in the cover of live coral and increasing concerns that coral reefs will be overgrown by macroalgae. The likelihood of macroalgal overgrowth will, however, depend on the thermal sensitivities of the macroalgae themselves. We exposed recently settled propagules of the common canopy-forming macroalga Sargassum swartzii and adult thalli of three species of Sargassum (S. swatzii, S. cristaefolium, S. polycystum) to three experimental temperatures: ambient, + 2 °C, and + 3.5 °C, reflective of summer minimum, mean, and maximum temperatures for the region. Growth and survival of Sargassum swartzii propagules were assessed over 48 days, and the growth, physical toughness, elemental composition, and susceptibility to herbivory of adult thalli were assessed after short-term exposure (2-weeks) to experimental temperatures. Growth and survival of S. swartzii propagules were reduced by 43% and 84%, respectively, when cultured at the elevated (+ 3.5 °C) temperature compared to ambient temperature. Similarly, elevated temperature resulted in a 17–49% decline in the growth of adult Sargassum thalli relative to controls. Susceptability of S. swartzii and S. cristaefolium to herbivory (i.e. mass removed by herbivores) was 50% less for thalli cultured at elevated (+ 3.5 °C) compared to ambient temperature, but this pattern was not related to changes in the physical or chemical properties of the thalli as a result of elevated temperature. The negative effects of elevated temperatures on the growth and survival of both Sargassum propagules and adult thalli will likely restrict the capacity of Sargassum, and potentially other macroalgae, to establish in new areas, and may also threaten the persistence of existing macroalgal meadows under future ocean temperatures. The thermal sensitivities of tropical Sargassum, together with those of corals, suggest ongoing ocean warming may lead to novel reef ecosystems that are low in both coral cover and macroalgal cover.

     

Application of polymerase chain reaction (PCR) using random amplified polymorphic DNA (RAPD) primers in the molecular identification of selected Sargassum species (Phaeophyta,Fucales)

The random amplified polymorphic DNA-polymerase chain reaction (RAPD-PCR) was used for the molecular characterisation and identification of Sargassum spp. A total of 17 samples of Sargassum (Sargassaceae, Fucales) was obtained from various localities around Peninsular Malaysia and Singapore. On the basis of morphological characteristics, the samples were tentatively grouped into five species: Sargassum baccularia, S. glaucescens, S. oligocystum, S. polycystum and S. siliquosum. By RAPD-PCR, five of 31 random primers tested generated reproducible amplification products, and polymorphic loci were detected by four of them (OPA02, OPA03, OPA04, OPA13). The RAPD-PCR profiles did not correlate with the morphological grouping into five species and extensive variation was detected between different isolates of the same species. A 450 base pair fragment generated using OPA13 was detected in 12 of 17 samples of Sargassum. This fragment was also present in profiles from Turbinaria (Sargassaceae). This study suggests that RAPD-PCR is useful in discriminating individual samples of the genus Sargassum and in developing fingerprints for them.     

Seasonal growth rate of Sargassum species at Teluk Kemang, Port Dickson, Malaysia

The seasonal growth rates of three Sargassum species were studied along two reef flats of Teluk Kemang, located at Port Dickson, Malaysia from September 2009 to September 2010. Systematic quadrat and line transects were sampled monthly. Nondestructive sampling was conducted, whereby Sargassum plants were tagged and monitored for a 13-month period. The majority of the tagged Sargassum samples belonged to lower length classes (<200 mm), especially in 0–99 mm (Sargassum polycystum, 64.20 %; Sargassum binderi, 68.29 %; Sargassum siliquosum, 56.80 %). Analysis of the monthly mean thallus length (MTL) revealed a bimodal pattern in growth rates, with two periods of high growth rates (January–February 2010 and June–July 2010) and two periods of higher degenerative rates (April 2010 and September 2010). The highest growth rates were recorded in February 2010 (4.08 mm day^?1) for S. siliquosum, and in June for S. polycystum (2.54 mm day^?1) and S. binderi (1.89 mm day^?1). Redundancy analysis (RDA) was employed to test for the overall correlation between monthly variation in MTL and the environmental parameters measured; S. binderi was correlated with ambient temperature (r = 0.5395), while S. siliquosum was correlated with seawater salinity (r = 0.5419) and ammonia (r = ?0.4603). This study reviews the seasonality of Sargassum species on two reefs of Teluk Kemang and their correlation with the selected environmental parameters.     

Olpidiopsis sp., an oomycete from Madagascar that infects Bostrychia and other red algae: Host species susceptibility

Olpidiopsis sp. (Oomycota) was cultured with its original host Bostrychia moritziana (Sonder ex Kützing) J. Agardh from Madagascar. Bean‐shaped zoospores with two heterokont flagella attached to the host cell wall surface and in 2 days host cells began collapsing and one or more syncytia developed in each infected cell. Zoospores were cleaved and an exit tube with a small plug was formed. Complete development and zoospore discharge occurred in 3 days. Infection occurred in cells of polysiphonous branches, monosiphonous branches, rhizoids and reproductive stichidia. Dead cells of plants treated with microwave were not infected. Susceptibility was variable in other Bostrychia species from different countries. Bostrychia moritziana (Sonder ex Kützing) J. Agardh, and Bostrychia radicans (Montagne) Montagne from Madagascar were susceptible but one Bostrychia tenella (J. V. Lamouroux) J. Agardh isolate from Madagascar was susceptible and two were not. B. radicosa (Itono) J. A. West, G. C. Zuccarello et M. Hommersand isolates from Madagascar, Thailand, Australia and New Caledonia were susceptible but an isolate from Malaysia was not. B. radicans isolates from Mexico and Brazil were non‐susceptible as were Bostrychia flagellifera Post, Bostrychia harveyi Montagne, Bostrychia montagnei Harvey, Bostrychia simpliciuscula Harvey ex J. Agardh, Bostrychia tenuissima R. J. King et Puttock, Stictosiphonia intricata(Bory de Saint‐Vincent) P. C. Silva, Stictosiphonia kelanensis (Grunow) R. J. King et Puttock and Stictosiphonia tangatensis (Post) R. J. King et Puttock, Lophosiphonia sp., Neosiphonia sp. and Polysiphonia spp. isolates were also non‐susceptible. Many non‐susceptible strains showed initial cell‐collapse followed by rapid wound‐repair cell formation without syncytia or sporangia developing. Caloglossa leprieurii (Montagne) G. Martens from Madagascar showed cell‐collapse and wound‐repair in periaxial cells, but wing cells died and became purple without wound‐repair. Caloglossa ogasawaraensis Okamura and Caloglossa postiae M. Kamiya et R. J. King had no symptoms of infection. Dasysiphonia chejuensis I. K. Lee et J. A. West was not infected. Surprisingly, the conchocelis phase but not the blade phase of Porphyra pulchella J. A.West, G. C. Zuccarello and Porphyra suborbiculata Kjellman was infected. The conchocelis of Porphyra tenera Kjellman and Porphyra linearis Greville were infected but no blade stages were tested. Porphyra miniata (C. Agardh) C. Agardh and Porphyra dentata Kjellman conchocelis were not infected. Bangia atropurpurea (Roth) C. Agardh gametophyte filaments were not infected. Other red, brown and green algae were not infected. Time lapse videomicroscopy of development and spore release was done.     

SYSTEMATICS OF THE GRACILARIACEAE (GRACILARIALES,RHODOPHYTA): A CRITICAL ASSESSMENT BASED ON RBCL SEQUENCE ANALYSES1

Generic concepts in the economically important agarophyte red algal family Gracilariaceae were evaluated based on maximum parsimony, Bayesian likelihood, and minimum evolution analyses of the chloroplast‐encoded rbc L gene from 67 specimens worldwide. The results confirm the monophyly of the family and identify three large clades, one of which corresponds to the ancestral antiboreal genera Curdiea and Melanthalia, one to Gracilariopsis, and one to Gracilaria sensu lato, which contains nine distinct independent evolutionary lineages, including Hydropuntia. The species currently attributed to Hydropuntia comprise a single well‐supported clade composed of two distinct lineages. The two most basal clades within Gracilaria sensu lato deserve generic rank: a new genus centered around G. chilensis Bird, McLachlan et Oliveira and G. aff. tenuistipitata Chang et Xia and a resurrected Hydropuntia encompassing primarily Indo‐Pacific (G. urvillei [Montagne] Abbott, G. edulis [S. Gmelin] P. Silva, G. eucheumatoides Harvey, G. preissiana [Sonder] Womersley, and G. rangiferina [Kützing] Piccone) and western Atlantic species (G. cornea J. Agardh, G. crassissima P. et H. Crouan in Mazé et Schramm, G. usneoides [C. Agardh] J. Agardh, G. caudata J. Agardh, and G. secunda P. et H. Crouan in Mazé et Schramm). Cystocarpic features within the Gracilaria sensu lato clades appear to be more phylogenetically informative than male characters. The textorii‐type spermatangial configuration is represented in two distinct clusters of Gracilaria. The rbc L genetic divergence among the Gracilariaceae genera ranged between 8.46% and 16.41%, providing at least 2.5 times more genetic variation than does the 18S nuclear rDNA. rbc L also resolves intrageneric relationships, especially within Gracilaria sensu lato. The current number of gracilariacean species is underestimated in the western Atlantic because of convergence in habit and apparent homoplasy in vegetative and reproductive anatomy.     

Sargassum boreale sp. nov. (Fucales, Phaeophyceae) from Hokkaido, Japan

A new species, Sargassum boreale Yoshida et Horiguchi is described. It belongs to the subgenus Bactrophycus section Teretia, with cylindrical receptacles and is distinct from Sargassum confusum C. Agardh, S. pallidum (Turner) C. Agardh and Sargassum microceratium (Turner) C. Agardh in having a rather elongated stem with smooth surface and distantly issuing main branches, with narrow leaves. The distinction between S. boreale and these species is also revealed by a difference in internal transcribed spacer 2 (ITS‐2) sequences. In addition to the base substitutions, the existence of a large gap in S. boreale distinguishes this species from others. Sargassum boreale is distributed around Hokkaido and Saghalien to 50°N latitude. A key to the species of section Teretia is provided.     

TWO TYPES OF AUXILIARY CELL AMPULLAE IN GRATELOUPIA (HALYMENIACEAE,RHODOPHYTA), INCLUDING G. TAIWANENSIS SP. NOV. AND G. ORIENTALIS SP. NOV. FROM TAIWAN BASED ON rbcL GENE SEQUENCE ANALYSIS AND CYSTOCARP DEVELOPMENT1

Few species in the genus Grateloupia have been investigated in detail with respect to the development of the auxiliary cell ampullae before or after diploidization. In this study, we document the vegetative and reproductive structures of two new species of Grateloupia, G. taiwanensis S.‐M. Lin et H.‐Y. Liang sp. nov. and G. orientalis S.‐M. Lin et H.‐Y. Liang sp. nov., plus a third species, G. ramosissima Okamura, from Taiwan. Two distinct patterns are reported for the development of the auxiliary cell ampullae: (1) ampullae consisting of three orders of unbranched filaments that branch after diploidization of the auxiliary cell and form a pericarp together with the surrounding secondary medullary filaments (G. taiwanensis type), and (2) ampullae composed of only two orders of unbranched filaments in which only a few cells are incorporated into a basal fusion cell after diploization of the auxiliary cell and the pericarp consists almost entirely of secondary medullary filaments (G. orientalis type). G. orientalis is positioned in a large clade based on rbcL gene sequence analysis that includes the type species of Grateloupia C. Agardh 1822 , G. filicina. G. taiwanensis clusters with a clade that includes the generitype of Phyllymenia J. Agardh 1848 , Ph. belangeri from South Africa; that of Prionitis J. Agardh 1851 , Pr. lanceolata from Pacific North America; and that of Pachymeniopsis Y. Yamada ex Kawab. 1954, Pa. lanceolata from Japan. A reexamination of the type species of the genera Grateloupia, Phyllymenia, Prionitis, and Pachymeniopsis is required to clarify the generic and interspecific relationships among the species presently placed in Grateloupia.     

ITS1 sequences of type specimens of Gigartina and Sarcothalia and their significance for the classification of South African Gigartinaceae (Gigartinales,Rhodophyta)

Uncertainties about the identity of type specimens of red algae have frequently led to taxonomic and nomenclatural confusion. A procedure for extracting PCR-amplifiable DNA from formalin-fixed material and herbarium specimens was used to investigate the taxonomic status of several South African Gigartinaceae. We compared nucleotide sequences in the internal transcribed spacer 1 (ITS1) region in type specimens and other historically important collections presently referred to Gigartina bracteata, G. radula and G. stiriata. The following opinions are supported: (1) Fucus bracteatus S.G. Gmelin, Chondrodictyon capense Kützing and Iridaea clathrata Decaisne represent a single species of South African Gigartina (G. bracteata (S.G. Gmelin) Setchell & Gardner) in which the disintegration of tetrasporangial sori results in a reticulate thallus. (2) Mastocarpus polycarpus Kützing, M. incrassatus Kützing and Iridaea lapathifolia Kützing represent a single species of South African Gigartina (G. polycarpa (Kützing) Setchell & Gardner) that has often, but erroneously, been called G. radula (Esper) J. Agardh. (3) Mastocarpus verrucosus Kützing is a later heterotypic synonym of Iridaea papillosa Bory (Sarcothalia papillosa (Bory) Leister) and was based on material that probably came from southern South America rather than from South Africa, the provenance given by Kützing. (4) Fucus stiriatus Turner and Sphaerococcus burmannii C. Agardh represent a single species of South African Sarcothalia (S. stiriata (Turner) Leister).     

PLASTID DNA RESTRICTION ANALYSIS LINKS THE HETEROMORPHIC PHASES OF AN APOMICTIC RED ALGAL LIFE HISTORY1

Gymnogongrus sp. (Phyllophoraceae) from Nova Scotia, Canada, identified tentatively as G. devoniensis (Greville) Schotter, grows in association with an Erythrodermis-like that forms chains of tetrasporangia or bisporangia. The crust resembles tetrasporophytic phases of other Gymnogongrus species, but in culture both it and the G. devoniensis gametophytes cycle independently by apomictic reproduction. A method was developed for extracting organelle DNA from this carrageenophyte genus involving purification of nucleic acids by binding to hydroxylapatite. Plastid DNA from G. devoniensis and bisporangial Erythrodermis-like crusts was compared with that of G. devoniensis and G. crenulatus (Turner) J. Agardh from France and of G. furcellatus (C. Agardh) J. Agardh from Chile. Plastid genomes of all Gymnogongrus species and the Erythrodermis-like crust were approximately 175 kb long. A single 3.5-kb plasmid DNA species was found in G. devoniensis and the Erythrodermis-like bisporophyte but not in other samples. Digestion of plasted DNA with several restriction endonucleases produced identical patterns in G. devoniensis and the Erythrodermis-like bisporophyte from the same location, indicating clearly that these entities represent two phases of an uncoupled life history. These results were confirmed with heteologous probes. A restriction fragment length polymorphism was identified between two Nova Scotian G. devoniensis populations. There was no similarity in restriction patterns between G. devoniensis from Nova Scotia, G. devoniensis from France. G. crenulatus or G. furcellatus, suggesting that molecular taxonomic methods could be important in delineating members of this morphologically variable genus. Further study is necessary to determine whether either Nova Scotian G. devoniensis or French G. devoniensis corresponds to type populations of G. devoniensis from Devon, England.     

Morphological and molecular characterisation and differentiation of <Emphasis Type="Italic">Sargassum baccularia</Emphasis> and <Emphasis Type="Italic">S. polycystum</Emphasis> (Phaeophyta)

Two Sargassum species (S. baccularia and S. polycystum) collected from Teluk Kemang and Cape Rachado, Port Dickson, Negeri Sembilan, Malaysia, which are alike in morphology except for the rhizoidal system and vesicles, were characterised using random amplified polymorphism DNA (RAPD). The genomic DNA of both species was isolated from the leaves using a modified CTAB method. Four random primers, that is, OPA2, OPA3, OPA4 and OPA13, successfully amplified the DNA. The polymorphisms generated by these four primers were analysed using the Dice Coefficient of Similarity and cluster analysis was carried out using GelCompar II Version 2.0 (Applied Maths, Kortrijk, Belgium) based on UPGMA. DNA analysis showed that three primers were able to differentiate the two species. Morphological analysis using Principal Components Analysis (PCA) and discriminant function analysis supported the molecular data. Both species are characterised by heavily muricate main branches, oblong-lanceolate leaves with dentate margins and discoid holdfasts and spherical vesicles; both are dioecious. The only difference is that S. polycystum has secondary holdfasts transformed into stolons. This last characteristic is therefore a very important criterion and may contribute to the difference shown by DNA analysis.