LOW MOLECULAR WEIGHT CARBOHYDRATES OF THE BANGIOPHYCIDAE (RHODOPHYTA)1

In the order Porphyridiales there are three clades based on molecular evidence. These show parallels with the low molecular weight carbohydrate (LMWCs) in different genera. Clade Porphyridiales 1 includes Dixoniella, Glaucosphaera, Rhodella, and one undescribed genus (3987) that all contain mannitol. Clade Porphyridiales 2 comprises taxa of the Stylonematales Rhodosorus and Stylonema species and contains digeneaside and sorbitol, whereas Chroodactylon has only sorbitol. In clade Porphyridiales 3 Flintiella, Porphyridium, and the undescribed genus (3797) all possess only floridoside. In the Erythropeltidales Rhodochaete contains floridoside and digeneaside, Erythrotrichia species contain only floridoside, Sahlingia subintegra has floridoside and traces of D‐floridoside, and Smithora has L‐isofloridoside plus floridoside. In the Compsopogonales Boldia and Compsopogon have only floridoside. Within these genera as presently circumscribed, the LMWCs appear to be a reliable character to supplement the usual cytological characters.     

DISTRIBUTION AND SYSTEMATICS OF THE FRESHWATER GENUS SIRODOTIA(BATRACHOSPERMALES,RHODOPHYTA) IN NORTH AMERICA1

Multivariate morphometrics and image analysis were used to determine the number of well-delineated infrageneric taxa of Sirodotia in North America. Three groupings were distinguished from 25 populations examined from Newfoundland and Quebec in the north to central Mexico in the south. These groupings were statistically related to 10 type specimens, and the following species were recognized: Sirodotia huillensis (Welwitsch ex W. et G. S. West) Skuja (syn. S. ateleia Skuja), S. suecica Kylin (syn. S. acuminata Skuja ex Flint and S. fennica Skuja), and S. tenuissima (Collins) Skuja ex Flint. These species are differentiated on the basis of whorl shape and degree of separation at maturity (S. suecica, rounded and appressed; S. huillensis and S. tenuissima, truncated apex and separated), the density of spermatangia (S. huillensis, dense clusters; S. suecica and S. tenuissima, sparsely aggregated), and the mode of germination of the gonimoblast initial (S. suecica and S. tenuissima, from the nonprotuberant side of the fertilized carpogonium; S. huillensis from the protuberant side). Sirodotia huillensis was found only in the desert-chaparral, whereas S. suecica and S. tenuissima occurred from south-temperate to boreal regions in cool (temperature 8–18° C), low ion (specific conductance 10–99 μS · cm^?1), and mildly acidic to neutral (pH 5.7–7.3) waters.     

A CELLULOSE SYNTHASE (CESA) GENE FROM THE RED ALGA PORPHYRA YEZOENSIS (RHODOPHYTA)1

The cell walls of Porphyra species, like those of land plants, contain cellulose microfibrils that are synthesized by clusters of cellulose synthase enzymes (“terminal complexes”), which move in the plasma membrane. However, the morphologies of the Porphyra terminal complexes and the cellulose microfibrils they produce differ from those of land plants. To characterize the genetic basis for these differences, we have identified, cloned, and sequenced a cellulose synthase (CESA) gene from Porphyra yezoensis Ueda strain TU‐1. A partial cDNA sequence was identified in the P. yezoensis expressed sequence tag (EST) index using a land plant CESA sequence as a query. High‐efficiency thermal asymmetric interlaced PCR was used to amplify sequences upstream of the cDNA sequence from P. yezoensis genomic DNA. Using the resulting genomic sequences as queries, we identified additional EST sequences and a full‐length cDNA clone, which we named PyCESA1. The conceptual translation of PyCESA1 includes the four catalytic domains and the N‐ and C‐terminal transmembrane domains that characterize CESA proteins. Genomic PCR demonstrated that PyCESA1 contains no introns. Southern blot analysis indicated that P. yezoensis has at least three genomic sequences with high similarity to the cloned gene; two of these are pseudogenes based on analysis of amplified genomic sequences. The P. yezoensis CESA peptide sequence is most similar to cellulose synthase sequences from the oomycete Phytophthora infestans and from cyanobacteria. Comparing the CESA genes of P. yezoensis and land plants may facilitate identification of sequences that control terminal complex and cellulose microfibril morphology.     

PHYLOGEOGRAPHIC PATTERNS INDICATE TRANSATLANTIC MIGRATION FROM EUROPE TO NORTH AMERICA IN THE RED SEAWEED CHONDRUS CRISPUS (GIGARTINALES,RHODOPHYTA)1

Inferring how the Pleistocene climate oscillations have repopulated the extant population structure of Chondrus crispus Stackh. in the North Atlantic Ocean is important both for our understanding of the glacial episode promoting diversification and for the conservation and development of marine organisms. C. crispus is an ecologically and commercially important red seaweed with broad distributions in the North Atlantic. Here, we employed both partial mtDNA Cox1 and nrDNA internal transcribed spacer region 2 (ITS2) sequences to explore the genetic structure of 17 C. crispus populations from this area. Twenty‐eight and 30 haplotypes were inferred from these two markers, respectively. Analysis of molecular variance (AMOVA) and of the population statistic Φ_ST not only revealed significant genetic structure within C. crispus populations but also detected significant levels of genetic subdivision among and within populations in the North Atlantic. On the basis of high haplotype diversity and the presence of endemic haplotypes, we postulate that C. crispus had survived in Pleistocene glacial refugia in the northeast Atlantic, such as the English Channel and the northwestern Iberian Peninsula. We also hypothesize that C. crispus from the English Channel refugium repopulated most of northeastern Europe and recolonized northeastern North America in the Late Pleistocene. The observed phylogeographic pattern of C. crispus populations is in agreement with a scenario in which severe Quaternary glaciations influenced the genetic structure of North Atlantic marine organisms with contiguous population expansion and locally restricted gene flow coupled with a transatlantic dispersal in the Late Pleistocene.     

PROTEOMICS OF THE RED ALGA,GRACILARIA CHANGII (GRACILARIALES,RHODOPHYTA)1

The application of proteomics in alga research is still quite limited. The present report describes the establishment of the proteome of a red alga of economic importance, Gracilaria changii (Xia et Abbott) Abbott, Zhang et Xia. Initially, four protein extraction methods including direct precipitation by trichloroacetic acid/acetone, direct lysis using urea buffer, Tris buffer, and phenol/chloroform extraction were compared for their suitability to generate G. changii proteins for two‐dimensional gel electrophoresis (2‐DE). The phenol/chloroform protein extraction method gave the best 2‐DE resolution of the proteins. Using these 2‐DE gels and mass spectrometry, several proteins including pigment proteins, metabolic enzymes, and ion transporters were identified. These findings highlight the potential of using proteomic approaches for the investigation of G. changii protein function.     

PHYLOGENETIC IMPLICATIONS OF THE CAD COMPLEX FROM THE PRIMITIVE RED ALGA CYANIDIOSCHYZON MEROLAE (CYANIDIALES,RHODOPHYTA)1

The de novo pyrimidine biosynthetic pathway consists of six enzymes: carbamoyl‐phosphate synthetase II (CPS II), aspartate carbamoyltransferase (ACT), dihydroorotase (DHO), dihydroorotate dehydrogenase, orotate phosphoribosyltransferase, and orotidine‐5′‐monophosphate decarboxylase. The origin and organization of the first three enzymes differ markedly between Opisthokonta (Metazoa and Fungi) and the Amoebozoa and green plants. However, no information has been available regarding the characteristics of such genes in other photosynthetic eukaryotes. In this study, we examined the pyrimidine biosynthetic cluster in the primitive red alga Cyanidioschyzon merolae P. DeLuca et al. isolate 10D. Unlike the situation in green plants, the CPS II, ACT, and DHO of C. merolae were fused to form a single open reading frame (the CAD complex), as in the Opisthokonta and Amoebozoa. Phylogenetic analysis of the CPS domain sequences suggested that this red algal CAD complex did not result from a recent lateral gene transfer from Metazoa or Fungi but that the fusion of the three genes occurred before the divergence of Opisthokonta, Amoebozoa, and the red algae. These results cast doubt on the recent hypothesis that the Opisthokonta and Amoebozoa form a monophyletic group, based on the presence in both of the CAD complex.     

IDENTIFICATION AND CHARACTERIZATION OF THE CATALASE GENE PyCAT FROM THE RED ALGA PYROPIA YEZOENSIS (BANGIALES,RHODOPHYTA)1

Catalase is an antioxidant enzyme that plays a significant role in protection against oxidative stress by reducing hydrogen peroxide. The full‐length catalase cDNA sequence as isolated from expressed sequence tags (ESTs) of Pyropia yezoensis (Ueda) M. S. Hwang et H. G. Choi (PyCAT) through rapid amplification of cDNA ends (RACE) was identified and characterized. It encoded a polypeptide of 529 amino acids, which shared 36%–44% similarity with other known catalase proteins. Phylogenetic analysis revealed that PyCAT was closer to the catalases from plants than from other organisms. The PyCAT mRNA expression was investigated using real‐time PCR to determine life‐cycle‐specific expression and the expression pattern during desiccation. The mRNA expression level in gametophytes was significantly higher than in sporophytes, and the mRNA expression level of PyCAT was significantly up‐regulated during the desiccation process. The recombinant PyCAT protein was purified and analyzed biochemically. The recombinant PyCAT protein exhibited high enzymatic activity (28,000 U·mg^?1) with high thermal stability and a broad pH range. All these results indicate that the PyCAT is a typical member of the plant and algal catalase family and may play a significant role in minimizing the effect of oxidative damage in P. yezoensis during desiccation.     

PHYLOGEOGRAPHY OF BATRACHOSPERMUM MACROSPORUM (BATRACHOSPERMALES,RHODOPHYTA) FROM NORTH AND SOUTH AMERICA1

Phylogeographic trends in Batrachospermum macrosporum Mont. were investigated using the mitochondrial intergenic spacer between the cytochrome oxidase subunit 2 and 3 genes (cox2‐3). A total of 11 stream segments were sampled with seven in the coastal plain of North America and four in tropical areas of South America. Fifteen thalli were sampled from seven streams, 14 thalli from two streams, and eight thalli from two streams. There were 16 haplotypes detected using 149 individuals. Of the eight haplotypes from locations in North America, all were 334 base pairs (bp) in length, and of those from South America, five were 344 bp, and three were 348 bp. Two individual networks were produced: one for the haplotypes from North America and another for those from South America, and these could not be joined due to the large number of base pair differences. This split between haplotypes from North and South America was confirmed with sequence data of the rbcL gene. There was very little genetic variation among the haplotypes from the North American locations, leading us to hypothesize that these are fairly recent colonization events along the coastal plain. In contrast, there was high variation among haplotypes from South America, and it would appear that the Amazon serves as a center of diversity. We detected considerable variation in haplotypes among streams, but frequently, a single haplotype in an individual stream segment, which is consistent with data from previous studies of other batrachospermalean taxa, may suggest a single colonization event per stream.     

INCORPORATION OF FRESHWATER RHODOPHYTA INTO THE CASES OF CHIRONOMID LARVAE (CHIRONOMIDAE,DIPTERA) FROM NORTH AMERICA1

Chironomid larvae incorporate pieces of freshwater red algae into their cases from a wide geographic range in North America, extending from southern Canada to central Mexico. The Rhodophyta used in this process represent two orders (Acrochaetiales and Batrachospermales), five genera (Audouinella, Batrachospermum, Lemanea, Paralemanea, and Sirodotia), and 14 species from 21 locations. Three genera from the chironomid subfamily Orthocladiinae make these cases: Cardiocladius, Eukiefferiella, and Orthocladius. The Eukiefferiella claripennis group was the most frequently observed infrageneric taxon using red algae in its cases. The cases were tubular in shape with longitudinally oriented strips of algae held together by silken threads. Some of the cases constructed with Batrachospermum and Sirodotia also had several lateral branches of the alga radiating from the tube.     

INCORPORATION OF FRESHWATER RHODOPHYTA INTO THE CASES OF CADDISFLIES (TRICHOPTERA) FROM NORTH AMERICA

Cases of larvae and pupae from six caddisfly (Insecta, Trichoptera) genera in three families from North America were observed to contain pieces of freshwater rhodophyte thalli. Seven genera of Rhodophyta, representing 13 species and 35 populations, were observed in this association. Four of the 25 species of Batrachospermum were incorporated into cases of Ochrotrichia ( Hydroptilidae) and Agrypnia ( Phryganeidae). Two of the three freshwater species of Bostrychia were used by Ochrotrichia larvae. Both Compsopogon coeruleus ( Balbis) Mont. and C. prolificus Yadava et Kumano were present in the cases of Hydroptial ( Hydroptilidae) and Ochrotrichia; Compsopogonopsis leptocladus ( Mont.) Krishnamur-thy was observed in the cases of Hydroptila. The more cartilaginous thalli of Lemanea fluviatilis ( L.) C. Ag. and Tuomeya americana ( Kütz.) Papenfuss were used by the brachycentrid larvae of Brachycentrus and Mi-crasema. Lemanea and Paralemanea spp. were also in the cases of Dibusa angata Ross (Hydroptilidae). The architecture of each caddisfly case was studied with light and scanning electron microscopy. Strips of algae were fit together in a transverse, concentric, or spiralled fashion. Based on transmission electron micrographs, cortical cells of Lemanea and Tuomeya in the cases of Brachycentrus and Micrasema appeared to be healthy with intact chloroplasts and typical batrachospermalean pit plugs. Geographic distributions of each rhodophyte-caddisfly association are given .     

NEW RECORD OF THE MARINE RED ALGA CUBICULOSPORUM (GIGARTINALES) FROM THE SOUTHERN GREAT BARRIER REEF1

Cubiculosporum koronicarpis Kraft (Cubiculosporaceae, Gigartinales), known previously only from the type locality (southeastern Luzon, Philippines), has been collected at North West Island on the southern Great Barrier Reef. The habitat, distribution and taxonomic status of the species are discussed, and habit features of the new specimens are illustrated.     

STUDIES ON DEVELOPING AND RELEASED SPERMATIA IN THE RED ALGA,TIFFANIELLA SNYDERAE (RHODOPHYTA)1

Developing and released spermatia of the red alga, Tiffaniella snyderae (Farl.) Abb. were studied. Spermatia were observed under hydrodynamically defined conditions and found to be released from the exposed spermatangial heads in a spermatium-plus-strand unit that remained connected to the spermatangial head. Interactions of single-spermatial strands resulted in the formation of multi-spermatial strands as long as 600 μm with as many as 47 spermatia along their length; however, most were 100–200 μm with 8–21 spermatia. Strand length and number of spermatia were correlated. Spermatial strands contracted or extended and rotated as the water velocity past the plant was changed, and in still water the strands retracted into a clump on the spermatangial head surface. Each strand type exhibited a characteristic threshold water velocity at which it reached maximum length, and above which it broke and was carried away. Fluorescence microscopy showed that the strands did not contain nucleic acid (DNA) and could thus be differentiated from filamentous blue-green algal and bacterial epiphytes. Histochemical staining indicated that the strands and spermatial vesicles contained an acidic, sulfated polysaccharide. Chelation of Ca²⁺ with EGTA resulted in strand breakdown suggesting that this divalent cation may be involved in strand integrity. Scanning electron microscopy revealed that release from the spermatangia occurred through tears in the cuticle covering the spermatangial head if it was still present, or from exposed spermatangia. Individual spermatia were attached tangentially to a well-defined strand 0.64 μm in diameter in the contracted state to 0.2 μm in the extended state. Transmission electron microscopy of spermatangial heads showed that immature spermatangia were characterized by a centrally positioned nucleus and abundant ER cisternae filled with a moderately electron dense granular material. Later in development the spermatangia acquire two spermatial vesicles containing highly convoluted fibrillar contents. The cell becomes polarized with the nucleus displaced apically and the spermatial vesicles occupying the basal half of the spermatangium. At maturity one of the vesicles is released basally. Liberated spermatia contain a membrane-bound nucleus and mitochondria and are associated with an oblong accumulation of fibrous material similar in size and position to the strand observed with the SEM. These strands are discussed in relation to red algal fertilization and other phases of the red algal life-history.     

PHYLOGENETIC RELATIONSHIPS OF THE FRESHWATER ALGA BOLDIA ERYTHROSIPHON (COMPSOPOGONALES, RHODOPHYTA) BASED ON 18S rRNA GENE SEQUENCES

The nuclear small-subunit ribosomal DNA sequence from the freshwater red alga Boldia erythrosiphon Herndon emend Howard et Parker was determined. Phylogenetic analysis confirms the positioning of this species within the bangiophycidean order of the Compsopogonales. The results strongly suggest that occupation of freshwater habitats by marine red algae has happened at least twice during red algal evolution.     

PHYCOBILIPROTEIN COMPOSITION AND CHLOROPLAST STRUCTURE IN THE FRESHWATER RED ALGA COMPSOPOGON COERULEUS (RHODOPHYTA)1

Serial sections of uncorticated axial cells of Compsopogon coeruleus revealed a single interconnected parietal chloroplast. Phycobilisomes in such chloroplasts were hemidiscoidal in shape with a broad-face diameter of ca. 25–30 nm. The molar ratio of phycobiliproteins in whole cell extracts was IPE:3PC:1APC, similar to isolated phycobilisomes. Two spectrally distinct C-phycocyanin forms (A618 nm, F648 nm and A630 nm, F652 nm) were resolved in dissociated phycobilisomes along with B-phycoerythrin and allophycocyanin.     

CLATHROMORPHUM NEREOSTRATUM (CORALLINALES,RHODOPHYTA): THE OLDEST ALGA?1

The longevity of organisms is intrinsically interesting and can provide useful information on their population structure and dynamics and the dynamics of associated communities. With the exception of perennial Laminariales that have rings in the stipe, the life spans of most perennial macroalgae are unknown or based on anecdotal observations. Using morphological analyses combined with the location and time of the rise in ¹⁴C from atmospheric nuclear testing within the thallus, we determined that the growth rate of a specimen of Clathromorphum nereostratum Lebednik from Adak Island was 0.30 mm·yr^?1, the 3⁰ bands within the thallus were annual, and the specimen sampled was 61–75 years old. Living crusts of this species from the same geographic region are reported to be up to 20 cm thick. Assuming our growth rate is typical, C. nereostratum can be approximately 700 years old, the oldest known living alga. This longevity and consistent banding within the thallus suggest that smaller scale sampling and additional chemical analyses of this alga could provide a detailed long‐term record of environmental variation at high latitudes in the North Pacific.     

VAUCHERIA SYNANDRA (XANTHOPHYCEAE,VAUCHERIALES): FIRST RECORD FOR NORTH AMERICA1

The brackish water species, Vaucheria synandra Woronin, common to Europe, is reported for the first time in North America from a coastal swamp area in Louisiana. The material from Louisiana is described and pictured, and is compared to other Vaucheria species with androphores.     

THE GENUS BALLIELLA ITONO & TANAKA (RHODOPHYTA: CERAMIACEAE) FROM EASTERN AUSTRALIA1

Three new species of the genus Balliella Itono & Tanaka are described from eastern Australia. Balliella amphiglanda from Lord Howe I. and Port Hacking, N.S.W., is distinctive in producing either abaxial or adaxial gland cells from the basal cells of lateral branches and in having tetrasporangia restricted to short branches. Balliella repens, from Tryon I., Heron I and Wistari Reef Qld., and Lord Howe I., N.S.W., is distinguished from the other species of the genus by its regularly developed prostrate systems and clustered tetrasporangia. Balliella grandis, from Wistari Reef, North West I. and One Tree I., Qld., is the largest recorded species of Balliella. It has correspondingly large gland cells and abaxial as well as adaxial tetrasporangia. Our work supports the placement of Balliella in the tribe Antithamnieae, a move which necessitates emending Wollaston's definition of the tribe to include species with procarps that form at successive levels along main axes rather than at only one or two points behind the apices.     

DIURNAL FLUCTUATION OF NITRATE REDUCTASE ACTIVITY IN THE MARINE RED ALGA GRACILARIA TENUISTIPITATA (RHODOPHYTA)1

The biochemical characteristics and diurnal changes in activity of the enzyme nitrate reductase (NR; EC 1.6.6.1) from the marine red alga Gracilaria tenuistipitata var. liui Zhang et Xia are described. Different assay conditions were tested to determine the stability of NR. The crude extract of G. tenuistipitata has a NR specific activity of 10.2 U.mg⁻¹, which is higher than the NR activities found for other algae, plants, and fungi. This NR is highly active at a slightly alkaline pH and is stable over a wide range of temperature, with an optimal activity at 20° C. The apical portions of the thallus contain 64.9 ± 6.6% of the total NR specific activity. The apparent Michaelis-Menten (K_m) constant found for KNO₃ was 197 μM, and it was 95 μM for NADH. The NR from G. tenuistipitata can be included in the NADH-specific group, because no activity was found when NADPH was used as an electron donor. In extracts of algae grown under either continuously dim light or a light-dark cycle, the activity of NR exhibits a daily rhythm, peaking at the middle of the light phase, when activity is 30-fold higher than during the night phase.     

DISTRIBUTION AND SYSTEMATICS OF FRESHWATER CERAMIALES (RHODOPHYTA) IN NORTH AMERICA1

Twenty-five freshwater populations of Ceramiales were collected in North America, 24 of which were from the tropical rainforest region of Central America and the Caribbean. The streams tended to be moderate in mean current velocity (X?= 23.3 cm·S^?1) and maximum width (X?= 6.3 m) but high in temperature (X?= 23.1°C), pH (X?= 7.9), and specific conductance (X?= 320 μS·cm^?1). Three Bostrychia species were restricted to the Caribbean islands: B. moritziana (Sonder ex Kütz.) J. Ag. (syn. B. cornigera Mont. and B. radicans f. moliforme Post), with ecorticate indeterminate axes, monosiphonous ultimate branches, and cladohaptera; B. radicans (Mont.) Mont. (syn. B. leprieurii Mont and B. rivularis Harv.), with ecorticate and polysiphonous axes throughout and cladophaptera; and B. tenella (Lamour.) J. Ag., with corticate indeterminate axes, monosiphonous ultimate branches, and peripherohaptera. Ballia prieurii Kütz. was found in Belize and Costa Rica and was characterized by rebranched determinate laterals, opposite branching, and long apical cells (X?= 61 μm) and axial cells (X?= 43 μm). Caloglossa leprieurii (Mont.) J. Ag. was localized in Puerto Rico while. C. ogasawaerensis Okam. was collected only in Costa Rica. The two species were separated by site of branching (midrib vs. margin) and blade width (X?= 384 vs. 861 μm). Polysiphonia subtilissima Mont. from Florida and Jamaica had four pericentral cells, no cortication, rhizoids arising from pericentral cells, and branches initiated at trichoblast scars.