European Journal of Phycology

Published by Taylor & Francis
Online ISSN: 1469-4433
Publications
Article
A recent reclassification of diatoms based on phylogenies recovered using the nuclear-encoded SSU rRNA gene contains three major classes, Coscinodiscophyceae, Mediophyceae and the Bacillariophyceae (the CMB hypothesis). We evaluated this with a sequence alignment of 1336 protist and heterokont algae SSU rRNAs, which includes 673 diatoms. Sequences were aligned to maintain structural elements conserved within this dataset. Parsimony analysis rejected the CMB hypothesis, albeit weakly. Morphological data are also incongruent with this recent CMB hypothesis of three diatom clades. We also reanalyzed a recently published dataset which purports to support the CMB hypothesis. Our reanalysis found that the original analysis had not converged on the true bipartition posterior probability distribution, and rejected the CMB hypothesis. Thus we conclude that a reclassification of the evolutionary relationships of the diatoms according to the CMB hypothesis is premature.
 
Article
The sequence of events taking place during infection of the algal host Chondrus crispus by the algal endophyte Acrochaete operculata was characterised at the ultrastructural level. Infection required settlement of quadriflagellate zoospores on the outer cell wall of the host. The host outer cell wall did not represent a barrier against penetration by the endophyte, which digested its way through. The extent of further spread of A. operculata in C. crispus was determined by the life-history phase of the host. In sporophytic fronds, colonisation of inner tissues occurred by digestion of the intercellular matrix, and infection spread throughout the frond. In gametophytic thalli, infection remained localised, forming a characteristic papule. In spite of these differences, degenerative changes at the cellular level were similar in the two phases of the host. Damage was caused by compression and by penetration of host cells. Final breakdown of the host resulted from the combined effects of A. operculata and bacteria. Bacteria alone did not cause damage to healthy C. crispus , but they accelerated cell wall degradation in the presence of A. operculata . Thus, A. operculata should be considered a primary pathogen of C. crispus , causing direct cellular damage and facilitating secondary infections.
 
Article
The cyanelles of glaucocystophyte (=glaucophyte) algae and the thecate amoeba Paulinella were previously thought to be endosymbiotic cyanobacteria or intermediates in the evolution of plastids from endosymbiotic cyanobacteria. While the more recent determinations of the number and nature of genes in the cyanelle genome show that cyanelles are genetically plastids, they have retained cyanobacterial features which are never (peptidoglycan walls) or only very rarely (carboxysomes) found in other plastids. The hypothesis presented here relates the peptidoglycan wall and the function of carboxysomes to the involvement of active water efflux using contractile vacuoles or their functional equivalent in volume regulation of non-cyanelle compartments of the cell in at least some parts of the life cycle. The peptidoglycan wall could permit a relatively high intracyanelle osmolarity which is presumably needed for cyanelle growth. A high intracyanelle osmolarity permits the high inorganic carbon concentration in the cyanelle during photosynthesis which is required for a carboxysome-based inorganic carbon concentration mechanism. The occurrence of a contractile-vacuole-based mechanism of volume regulation in at least part of the life cycle argues on energetic grounds for a low osmolarity of the cytosol, with a consequent lack of potential for a high inorganic carbon concentration in the cell as a whole. These predictions are susceptible to experimental test.
 
Article
A phylogenetic analysis of the genus Alexandrium , including both the most common and rare species from coastal areas of the Mediterranean Sea was carried out. Nucleotide sequences of 5.8 S gene and Internal Transcribed Spacer regions of the rRNA operon were examined and analysed together with isolates of Alexandrium spp. from elsewhere in the world. These rDNA ribosomal markers were useful in delineating the phylogenetic position of species in the genus, as well as in determining relationships among isolates within each species collected from different localities. Results of phylogeographical analyses within the ' Alexandrium tamarense ' species complex identified three lineages in the Mediterranean Sea: the Mediterranean (ME), Western European (WE) and Temperate Asian (TA) clades. The phylogenetic grouping of the isolates is consistent with the ribotype clades, but not with the morpho-species that constitute the complex. Additional non-toxic isolates were included in the ME clade. The NA (North Atlantic) clade is the fourth group within the ' Alexandrium tamarense ' species complex identified by phylogenetic analyses. Based on its higher genetic diversity and phylogeographical relationships, it can be hypothesized that the NA clade represents the ancestral group of the ' Alexandrium tamarense ' species complex. Alexandrium minutum isolates of the NW Mediterranean clustered with strains from Brittany and Australia. Alexandrium minutum constituted a sister clade of A. tamutum , which is another species strongly associated with the Mediterranean area. Another typical Mediterranean species, A. taylori , was placed as a sister clade of A. pseudogoniaulax by the phylogenetic analysis. Finally, the phylogenetic relationships of some Alexandrium morpho-species that were infrequently observed in the Mediterranean Sea have been resolved.
 
Article
The marine red alga Stictosiphonia hookeri was isolated from three locations in cold-temperate Chile and Argentina, and the comparative ecophysiology of the isolates was studied. The growth response in relation to various salinity, light and temperature conditions was investigated and photosynthesis-irradiance curves were determined. The effect of increasing salinity on the intracellular concentration of the sugar alcohol sorbitol, and the accumulation/degradation kinetics of this compound, were studied. All isolates are characterized as euryhaline, eurythermal and low-light-adapted organisms because of the growth and photosynthesis response patterns. With increasing salinities the macroalgae accumulated high concentrations of sorbitol, which acts as an organic osmolyte and compatible solute. However, this process requires more than 24 h, and hence is too slow to contribute to osmotic acclimation within a tidal cycle. The geographic distribution and the growth versus temperature data indicate infraspecific differences which are interpreted as a stage in the development of different (physiological) ecotypes.
 
Break force (F break ) vs. planform area (A) for 118 populations of macroalgae. Log 10 (F break ) ¼ 1.85 þ 0.36 Â Log 10 (A) ( p<0.0001, r 2 ¼ 0.38). The superimposed letters refer to the taxonomic order of each population. C ¼ Caulerpales, F ¼ Fucales, G ¼ Gigartinales, L ¼ Laminariales, P ¼ Palmariales and U ¼ Ulvales.
Article
The forces required to break or dislodge benthic macroalgae can be linked to many important ecological processes such as survival, transport and dispersal, and it is, therefore, important to understand what factors affect these forces. We review existing studies, using simple vote-counting, to establish what ecological factors have been found to influence macroalgal break forces. We also used the compiled information to test for the existence of a general allometric relationship between thallus area and break force. We found 30 studies reporting break forces for 27 species of macroalgae from 118 populations. Species within the orders Laminariales, Fucales and Gigartinales were well studied whereas other orders had been neglected. Two-thirds of all studies were from rocky intertidal habitats and two-thirds were from North American coastlines. The most frequently tested ecological factor was wave exposure (14 studies with 7 significant effects). Other factors that were found to affect break force significantly in at least one study included time of sampling, thallus size, substratum and species. Generally, most factors had been addressed only a few times, emphasizing a need for further study. We found a highly significant positive relationship between thallus size and break force across a wide range of species and ecological conditions, providing evidence that size alone is an important determinant of macroalgal break force. To strengthen inferences and ensure that meta-analyses can be conducted, we suggest that future studies (i) use nested replication in space and time, (ii) include taxa and regions that have not been studied in detail, and (iii) report, as a minimum, corresponding information on break forces, thallus sizes, substratum type, wave exposure and time of sampling.
 
Article
Analysis of biogeographic affinities is a key tool to establish and improve the resolution of hierarchical biogeographic systems. We describe patterns of species richness of the marine macroalgal flora across Lusitanian Macaronesia (Azores, Madeira, the Salvage Islands and the Canary Islands), and test (i) whether such differences are related to differences in proximity to the nearest continental shore and size among islands. We also explore biogeographic affinities in the composition of macroalgal assemblages (= presence/absence of each taxon in multivariate datasets) to determine (ii) whether each archipelago is a biogeographic unit within this ecoregion and (iii) whether patterns in assemblage composition are related to proximity (i.e. distances) among islands. Presence/absence matrices were created to test and visualize multivariate affinities among archipelagos. A total of 872 taxa were compiled. Species richness peaked at the Canary Islands and decreased towards the Azores; the pattern matched a progressive increase in distance from the nearest continental shores, matching the classical island biogeography theory. Intra-archipelago differences in species richness were largely related to variations in island size. Biogeographic similarities among archipelagos were hierarchically structured. Madeira and the Salvage Islands constituted one biogeographic unit. Floras from the Azores, Madeira and the Salvage Islands were barely separable from each other, but were different from those at the Canary Islands. Such biogeographic similarities among islands were negatively correlated with the geographical separation (i.e. distances) among them. Proximity to nearby continental shores, in conjunction with large- and meso-scale oceanographic patterns, seems to interact to create patterns in richness and composition of algal assemblages across Lusitanian Macaronesia.
 
Article
Alexandrium tamarense is a cyst-forming dinoflagellate that can cause toxicity in shellfish. Belfast Lough, in northeast Ireland, has experienced toxicity events due to the presence of A. tamarense, which are monitored because of shellfish farms in the Lough. Since 1992 anthropogenic influences on the Lough have changed with the introduction of a ‘fast cat’ ferry service and an increase in mussel farming. In 2002 Belfast Lough was surveyed for A. tamarense cyst distribution in the sediments and the results are compared to a previous cyst survey carried out in the Lough in 1992. Cyst numbers were generally lower in 2002 than in 1992 (t-test, P −1 dry sediment compared with 3330 in 1992. Although sediment disturbance increased in the period between the surveys and plays a role in cyst distribution, A. tamarense cysts are still present in the seabed of the Lough. These cysts remain an important repository of inoculating cells for A. tamarense blooms that have not been removed by the recent anthropogenic activity in the Lough and therefore the requirement for monitoring remains. Comparisons between cyst counts per volume of wet sediment and per weight of dry sediment were carried out, and although the first is needed for ecological studies allowing a deeper analysis, it is also recommended that cyst counts per dry weight of sediment are always reported for wider comparative purposes.
 
Article
Given the problems of species delimitation in algae using morphology or sexual compatibility, molecular data are becoming the standard for delimiting species and testing their traditional boundaries. The idea that species are separately evolving metapopulation lineages, along with theoretical progress in phylogenetic and population genetic analyses, has led to the development of new methods of species delimitation. We review these recent developments in DNA-based species delimitation methods, and discuss how they have changed and continue to change our understanding of algal species boundaries. Although single-locus approaches have proven effective for a first rapid and large-scale assessment of species diversity, species delimitation based on single gene trees falls short due to gene tree–species tree incongruence, caused by confounding processes like incomplete lineage sorting, trans-species polymorphism, hybridization and introgression. Data from unlinked loci and multi-species coalescent methods, which combine principles from phylogenetics and population genetics, may now be able to account for these complicating factors. Several of these methods also provide statistical support regarding species boundaries, which is important because speciation is a process and therefore uncertainty about precise species boundaries is inevitable in recently diverged lineages.
 
Article
Batch cultures of Chlamydomonas reinhardtii were used to study carbon allocation in relation to growth phase and phosphorus availability. Cultures were grown at initial phosphorus (PO4-P) concentrations of 500 µg l (high-P) and 50 µg l (low-P). Cellular carbon allocation was monitored using Fourier transform infrared (FTIR) microspectroscopy with the ratio of the band intensities at 1736 cm (lipid) and the 1180–950 cm region (carbohydrate) to 1652 cm (amide I) used as an index of changing carbon balance. Cellular phosphorus concentrations (P quota) were measured by energy dispersive X-ray microanalysis (EDXRMA). Both treatments entered stationary phase on day 18. Increased cell counts in the high-P treatment (max. 3.0 × 10 cells ml at stationary phase) led to a rapid decrease in external P availability to
 
Article
This study presents the first record of a living dinoflagellate cyst with a hypocystal, antapical archeopyle. It is also the first detailed account of the archeopyle of a living freshwater cyst from the genus Peridinium. The cysts were isolated from sediment traps deployed in Lake Nero di Cornisello, a low-alkalinity high mountain lake of the Adamello mountain range (2233 m above sea level, South Eastern Alps, Italy). The archeopyle is large, clearly hypocystal, polygonal, and slightly peanut-shaped. The species producing this cyst belongs to the Peridinium umbonatum group and is described based on scanning electron microscopy and light microscopy. Partial sequences of SSU rDNA were obtained and compared with previously published sequences from the P. umbonatum group. The taxonomic position of the species is discussed.
 
In vivo absorption spectra of Chaetoceros brevis at the end of batch growth experiments under various conditions. (A) Absorption normalized to cell number; (B) Absorption normalized to chlorophyll a concentration. For abbreviations in key see Table 1.  
Photosynthesis-irradiance response curves of Chaetoceros brevis, measured by PAM fluorometry 0 (A) 3 (B) and 5 (C) days after the start of the experiment. For abbreviations in key see Table 1.  
Time course of the cellular carbon (A) and nitrogen content (B) of Chaetoceros brevis grown under various conditions. Symbols and lines as in Fig. 2.  
Diel variations in water-extractable polysaccharide content of Chaetoceros brevis cells under various conditions after 0 (A), 3 (B) and 5 (C) days from the start of the experiment. Symbols and lines as in Fig. 2.  
Relationship between cellular water-extractable polysaccharide C content and total carbon content. Data for all treatments were pooled. The solid line is the principal axis (y = 1.7x + 9.2). Note break in y-axis.  
Article
Iron, one of the structural elements of organic components that play an essential role in photosynthesis and nitrogen assimilation of plants, is available at extremely low concentrations in large parts of the Southern Ocean’s surface waters. We tested the hypothesis that photosynthesis is the primary target of iron stress in phytoplankton living in this specific environment, resulting in a reduced carbohydrate production. Cultures of a small Antarctic diatom, Chaetoceros brevis, were exposed to two different photon irradiances under iron-rich and iron-poor conditions. Under both light regimes growth rate was reduced only slightly by iron starvation, as expected because the iron requirement of a small-celled species such as C. brevis is low. Even so, iron-starved cells differed markedly from iron-replete cells: for low and high irradiance, respectively, they had a 20 and 27% lower content of light-harvesting pigments (chlorophyll a and c2 and fucoxanthin), a 8 and 15% decrease in light absorption and a 15 and 17% decrease in quantum yield of photosystem II. The diurnal production of water-extractable carbohydrates was reduced by 28 and 31%, which resulted in a low supply of energy and carbon skeletons from these storage products. This may well have influenced protein synthesis. The nocturnal consumption of carbohydrates was also reduced, which, together with the almost proportional decrease in cellular C and N content, suggests that the C and N metabolism were tightly tuned in iron-stressed cultures. The decrease in C and N content correlated with a decrease in cell volume. Our results indicate that iron limitation is likely to affect the ability of phytoplankton to maintain high rates of protein synthesis within the deep wind-mixed layer of the Southern Ocean. In addition, growth at the surface could be inhibited by too much light: iron-poor cultures of C. brevis grown at low irradiance showed enhanced sensitivity to photoinhibition.
 
Water temperature, salinity and pH in four ponds studied on Rangiroa Atoll.
Article
Cyanobacterial populations, the primary producers and builders of 'kopara' microbial mats were studied in four selected ponds along the rim of the Rangiroa Atoll, French Polynesia using a polyphasic approach. Seven isolates were maintained in uni-cyanobacterial cultures, characterized morphotypically and phylogenetically by evaluating sequences of the 16S rRNA gene of about 620 base pairs in length. Cyanobacteria in natural populations were analyzed microscopically, characterized morphotypically, and compared with cultured strains. Three of the isolates were identified in the field samples: Lyngbya aestuarii, Johannesbaptistia pellucida and Chroococcus submarinus were present in the mats only as minor components, whereas the species of Schizothrix that dominated the mat community could not be cultured. The sequence of Johannesbaptistia pellucida is published for the first time. The phylogenetic and taxonomic relations are discussed on the basis of a reconstructed phylogenetic tree in relation to morphotypic characters. Sequences of Kopara isolates plot separately from those cultured from the lagoon of the neighbouring atoll Tikehau, indicating a narrow niche differentiation of benthic cyanobacterial taxa. The results support the application of a polyphasic approach to characterization, ecology and diversity of cyanobacteria.
 
Article
The Indo-Pacific Ocean is a biodiversity hotspot for marine organisms. In this area, most of the research has focused on marine animals, such as reef fish, molluscs and other associated coral fauna, but very little has been done on macroalgae. The Thai-Malay Peninsula is an important north–south barrier in this area, which faces two different oceans – the Indian Ocean and the Pacific Ocean. This study aims to investigate genetic distribution patterns of Padina boryana Thivy around the Thai-Malay Peninsula, where it is common. Three DNA marker regions, the mitochondrion-encoded cytochrome c oxidase subunit 3 gene (cox3); the plastid rbcL, and the nuclear internal transcribed spacer 2 (ITS2) were used to evaluate genetic diversity and the relationships within and between populations. Samples were collected from both the Andaman Sea and Gulf of Thailand sides of the peninsula. Parsimony networks and maximum likelihood and Bayesian analyses showed clearly that there are two separated P. boryana lineages, one restricted to the Gulf of Thailand and the other to the Andaman Sea and other areas of the Indo-Pacific. The effect of different ocean currents along the Andaman Sea and Gulf of Thailand may have shaped these populations of P. boryana. This phylogeographic separation, based on persistent currents in the area, may affect other marine organisms along the Thai peninsula.
 
Article
Arthrospira platensis is a cyanobacterium known for its nutritional value and secondary metabolites. Extracellular polymeric substances (EPS) are an important trait of most cyanobacteria, including A. platensis. Here, we extracted and analysed different fractions of EPS from a locally isolated strain of A. platensis. Three different fractions of EPS were distinguished. These were EPS released into the medium (REPS), EPS loosely bound to the organism (LEPS) and EPS tightly bound to the organism (TEPS), which were extracted by different procedures. The LEPS fraction was smaller than the other two fractions. The EPS of A. platensis exhibited high diversity. Total protein and carbohydrate content was determined in each of these fractions. The largest amount of total carbohydrates and total proteins was in the TEPS fraction. Eight sugar moieties were detected and analysed in all EPS fractions using HPAE-PAD. Fructose, mannose and ribose were rare sugar residues in all fractions of EPS. With the exception of fructose, all sugars tested for were detected in TEPS. The amount of sugars detected was significantly higher in TEPS compared with the two other fractions, especially for galactose, xylose and glucose. The EPS were localized by confocal laser scanning microscopy (CLSM) after staining with different fluorescent dyes and it was found that A. platensis possessed a thick and smooth layer of EPS around the spiral trichomes.
 
The frequency of occurrence (A) and extent (B) of gaps in the canopy cover of Fucus serratus at four locations, separated by 3-5 km, in the vicinity of Svaneke (Bornholm, southern Baltic Sea). Data for each location are the sum of gaps measured along three 60-m transects perpendicular to the coastline. Boelshavn: n ¼ 27 gaps; Vige Havn: n ¼ 41 gaps; Maageflejl: n ¼ 21 gaps; and Aarsdale: n ¼ 23 gaps. 
Results of ANOVA testing for effects of location and clearing size on the abundance of major taxa in the centres of clearings 
Abundance of dominant assemblage-formers, determined within a 20 Â 20 cm quadrat, at the centres of experimental clearings and at the edges of large clearings 21 days after impact. Clearings were set up in a subtidal Fucus serratus habitat at two locations, 3 km apart, in the vicinity of Svaneke (Bornholm, southern Baltic Sea). Data were pooled across locations as there were no effects of location. Horizontal lines above histograms indicate results from planned contrasts where the p-value shows the significance level of the contrast. Error bars are SE (n ¼ 6). 
nMDS plot of similarity of post-disturbance assemblage structure in control plots (intact canopy), small clearings (0.6 m diameter), and centres and edges of large clearings (1.2-4.8 m diameter), 21 days after implementing clearings in the Fucus serratus canopy cover near Svaneke (Bornholm, southern Baltic Sea). 
Article
Disturbances occasionally create gaps of varying sizes in the canopy cover of macroalgae on temperate reefs. This study quantified the size distribution of naturally occurring gaps in a subtidal Fucus serratus canopy, and experimentally tested the hypotheses that (i) early post-disturbance abundance of dominant assemblage-formers in the canopy-dominated habitat will depend on the spatial extent of the impact, and (ii) the effects of disturbance on the abundance of dominant assemblage-formers at the edges of large impacted areas will be similar to the effects at small impacted areas. Gaps of up to 6 m diameter accounted for 17% of the habitat and, while small gaps were numerically dominant, all gap size-classes accounted for a similar proportion of habitat. Experimental disturbances (canopy clearing) ranging from 0.6 to 4.8 m diameter caused consistent scale-dependent divergence of assemblage structure after 21 days. Of particular interest was a dense cover (∼75%) of filamentous algae in the centres of large (≥1.2 m) clearings. Filamentous algae are widely known to have negative effects on Fucus recruitment and growth, and their rapid increase in abundance suggests that future recovery of the Fucus canopy could be adversely affected. The abundance of most taxa at the edges of large clearings was either indistinguishable from abundances in small clearings and intact canopy, or intermediate between abundances in these treatments and in the centres of large clearings. Juvenile Fucus were, however, more abundant around edges than anywhere else. These experimental outcomes support the model that scale-dependent divergence of assemblage structure is conveyed by an escape from processes associated with edges of the original habitat. I propose that, in the subtidal Fucus beds of Bornholm, recovery from large disturbances is restricted to encroachment from the edges because the adjacent canopy suppresses the growth of filamentous algae and thus facilitates regeneration and recruitment of Fucus.
 
Morphological features of Cabra reticulata sp. nov. compared with C. matta Species Cabra reticulata a, b Cabra matta c
Cabra reticulata Chomérat et Nézan sp. nov.: line drawings. Fig. 1. Right lateral view showing the outline shape, the location of the cingulum (C), the sulcus (S) and the nucleus (n). Figs 2-5. Drawings showing the thecal plates arrangement. Fig. 2. Right lateral view. Fig. 3. Left lateral view. Fig. 4. Apical view. Fig. 5. Antapical view. Scale bars represent: Figs 1-3, 5, 10 µm; Fig. 4, 5 µm.
Article
A new sand-dwelling dinoflagellate species, Cabra reticulata sp. nov., is described from sandy habitats in the south of Brittany (northwestern France). This new species possesses the characteristics of the 'unusual' genus, Cabra Murray et Patterson. The thecal plate formula is Po Pt 3' 1a 4'' 'x' 3c ?s 5''' 1'''' (or alternatively interpreted as Po Pt 4' 0a 4'' 'x' 3c ?s 5''' 1''''). Because of the strong lateral compression, cells of C. reticulata are frequently seen laterally and have a polygonal shape, with three prominent antapical pointed flanges and a dorsal spine in the anterior part of the hypotheca. The cingulum is ascending and incomplete. The thecal surface is ornamented with strong reticulations forming polygonal areolae, which differs from C. matta Murray et Patterson, the type species of the genus. In addition to their different size and shape, a detailed study of C. reticulata by scanning electron microscopy also revealed several different plate features. Plates of the cingulum, c1 and c3, have no reticulations and are ornamented with shallow round areolae, while c2 is only faintly reticulated. Plate 1'''' is also characteristic because dorsally it forms a prominent pointed flange and ventrally bears a peculiar small area of densely arranged pores or tiny areolae. A similar finding has previously been described in Roscoffia capitata Balech. In the description of the genus, it was suggested that Cabra is closely related to species of the genus Roscoffia, and our observations strengthen this hypothesis. Prior to this study, C. reticulata was probably observed in the Virgin Islands, Caribbean Sea, but was tentatively identified as Thecadinium sp. and not fully described. Thus, C. reticulata appears to be present on both sides of the Atlantic Ocean and to occupy benthic habitats in temperate and tropical areas.
 
The different Pseudo-nitzschia species examined in this study for verification of frustule morphology, reactivity in both immunolabelling and nucleotide probing, and toxicity
Article
Biological monitoring in the Dutch Wadden Sea between November 1993 and July 1994 revealed Pseudo-nitzschia-like pennate chain-forming diatom species with cell numbers ranging from 10^2 to 10^5 per litre. Cultured isolates and field samples were examined by electron microscopy, which revealed the majority to be Pseudo-nitzschia pungens. This species dominated over other diatoms in the phytoplankton population during November 1993 and at the end of June 1994. At the beginning of June 1994, P. fraldulenta was also present; occasionally, P. delicatissima was observed. One isolate showed the characteristic morphology of P. multiseries. Species-specific polyclonal antibodies and large-subunit (LSU) rRNA-targeted oligonucleotides for North American strains of P. multiseries and P. pungens applied to the European isolates, confirmed species designations based on electron microscopy. The isolate of P. multiseries from the Dutch Wadden Sea produced domoic acid; after 55 days of growth about 19 pg per cell was measured. This is the first report of a domoic acid-producing P. multiseries isolated from European coastal waters.
 
Chlorophyll content analyses showing the concentration of different pigments (ng cell −1 ), namely chl a, b and c, in temporary cysts and motile cells of Pyrodinium bahamense var. compressum grown in f/2 and total low nutrient conditions.
Matrix of figures showing the trends for motile cell, temporary cysts and total cell counts (◊ motile cells, □ temporary cysts, Δ total).  
Light and confocal laser scanning microscopy (CLSM) of Pyrodinium bahamense var. compressum showing intracellular changes during pellicle formation. Figs 1, 2. Cells were first observed to exhibit aggregation of the red autofluorescent (RAF) organelles (plastids) during pellicle formation, followed by formation of the opening in the membrane (arrows) and in between the thecal plates (Figs 3, 4) and extrusion of the RAF materials via the opening in the wall as indicated by the arrows (Figs 5, 6). Figs 7, 8. Extruded materials might possibly include bacteria-like forms (BLFs), plastids and other cytoplasmic materials (BLFs pseudocoloured yellow stained with Sybr Green, theca pseudocoloured green stained with Calcofluor White, RAF-bodies/plastids pseudocoloured red). Scale bars are 20 µm.  
Light and pseudocoloured CLSM images of Pyrodinium temporary cysts. Fig. 9. Midsection showing the more compact nuclear region (NR), the RAF-cytoplasm (Cyt) and three visible transitional red bodies (TRB). Extracellular bacteria (green dot-like/ cocci particles) are also seen in the periphery of the cell. Fig. 10. A DIC image showing the space that forms between the pellicular body and the old theca as a result of reduction in size. Fig. 11. The cyst still encapsulated in its old theca (Th) with some visible structures such as the autofluorescent cytoplasmic matrix (Cyt) and TRBs. Fig. 12. A 3D reconstructed image of the RAF-pellicle body showing the outer surface with attached bacteria (green cocci particles) and the old theca (blue). Fig. 13. A DIC image showing the clear outline of the outermost covering that is not stainable by Calcofluor White as compared with the theca (blue). BLFs (green dots, stained with Sybr Green) are also seen attached to outside wall of the cyst. The pellicle body is also about to be dislodged off from the theca. Fig. 14. Regeneration of the chloroplast (Chl) and the expansion of the nuclear region (NR) during reversal from temporary cyst to motile form. The other visible components in the image are the cytoplasm (Cyt) and the outermost covering (OC). Scale bars are 20 µm.  
Article
Pellicle or temporary cysts of Pyrodinium bahamense var. compressum (Böhm) Steidinger, Tester & F.J.R. Taylor and their role in bloom dynamics have not yet been adequately characterized and understood. We investigated the role of temperature- and nutrient-mediated stress as factors that could induce pellicle formation in batch cultures. Cellular features and their implications for temporary cyst viability were examined using confocal laser scanning microscopy (CLSM). Our data suggest that temperature change is one of the key factors influencing pellicle formation, preserving viability at low temperature (i.e. 13°C). Hypnocysts (resting cysts) were not observed. During pellicle formation, motile cells generally undergo ecdysis, extrusion of cytoplasmic materials and bacteria, compaction of the nucleus and non-motility. The outermost covering of the temporary cysts shows red autofluorescence and it contains lower concentrations of chlorophyll (chl) a and no detectable chl c. The nuclear region is surrounded by transitional red bodies and other unidentified cellular structures. Temporary cysts can immediately revert back to the motile state upon exposure to optimum conditions. This is accompanied by the expansion of the nuclear region, regeneration of the chloroplasts and enlargement of the cell. Developmental changes during reversal of temporary cysts to motile forms were also observed to cause breaks in the cell covering that could serve as sites for bacterial entry. Though observed in vitro, such behaviour may also be occurring in nature especially as a response to drastic short-lived environmental changes. This is the first detailed report on the characteristics of temporary cysts of P. bahamense var. compressum.
 
Article
The Mediterranean red algal flora is diverse but current knowledge of its diversity is at best fragmentary. Here, a new species of Kallymenia from Croatia is described based on morphological and molecular data. Members of the genus Kallymenia share similar morphology making their generic identification relatively easy, whereas species level identification is notoriously difficult. In this paper an integrative systematics study using three gene markers, cox1 (COI), rbcL and nuclear LSU, allowed us to (i) confirm the identity of four existing members of this genus inhabiting the Mediterranean Sea: K. feldmannii, K. lacerata, K. patens and K. requienii; (ii) detect the presence of K. reniformis only in the Atlantic, and (iii) reveal the presence of a new member of Kallymenia from the Mediterranean Sea, K. ercegovicii Vergés & Le Gall, sp. nov., which is described based on distinctive morphological and molecular characters. Kallymenia ercegovicii is distinguished, using three molecular markers, from all Kallymenia species for which these sequences are available. Morphologically, the new species can be distinguished from other polycarpogonial Kallymenia by a set of characters including a deeply lobed thallus, large inner cortical cells, stellate but non-glanglionic medullary cells and non-ostiolate cystocarps surrounded by a filamentous net composed of elongated cells forming fascicles. An initial phylogeny of the genus Kallymenia was inferred from cox1 (COI), rbcL and nuclear LSU sequences. Kallymenia ercegovicii was resolved with full support within the same lineage as K. reniformis (the generitype), K. feldmannii and K. patens, whereas K. lacerata and K. requienii were only distantly related.
 
Article
A new benthic phototrophic dinoflagellate is described from sediments of a tropical marine cove at Martinique Island and its micromorphology is studied by means of light and electron microscopy. The cell contains small golden-brown chloroplasts and the oval nucleus is posterior. It is laterally compressed, almost circular in shape when viewed laterally. It consists of a small epitheca tilted toward the right lateral side and a larger hypotheca. In the left view, the cingulum is more anterior and the epitheca is reduced. The cingulum is displaced and left-handed. This organism is peculiar in having no apical pore and its thecal plate arrangement is 2′ 1a 7′′ 5c 3s 5′′′ 1′′′′. The plates are smooth with small groups of pores scattered on their surface. An area with 60–80 densely arranged pores is found near the centre of the 2′′′ plate, on the left lateral side. Morphologically, these features are different from all other laterally compressed benthic genera. In addition, molecular genetic sequences of SSU and partial LSU form a distinct and well-supported clade among dinoflagellates and support the erection of a new genus. However, molecular phylogenies inferred from ribosomal genes failed to confirm any clear relationship with other benthic taxa and affinity with other laterally compressed dinoflagellates has not been demonstrated. Hence, the taxonomic affinity of Madanidinium loirii with a defined order and family is unclear at the moment.
 
Article
Based on physiological characteristics, we hypothesized that different strains of Gracilaria birdiae from two distinct geographical areas of the Brazilian coast (2500 km apart) would have different responses to long-term exposure to UV-B radiation (UV-B). The locations differ in their environmental conditions: one is a warmer area, Ceará State (CE), closer to the equator; the other is a colder area, Espirito Santo State (ES), closer to the Tropic of Capricorn. To test the hypothesis that the CE population is more resistant to UV-B than the ES population, apical segments of the red (RDCE, RDES), green (GRCE) and greenish-brown (GBCE) strains were cultivated in the laboratory under two treatments: control (PAR) and artificial UV-B (PAR + UV-B). Algal performance was evaluated by considering growth rates, pigment content and ultrastructural analysis. Compared with the control, all strains showed a decrease in growth rates after exposure to UV-B. Of all strains, RDES showed the greatest sensitivity to UV-B. However, a decrease in growth rate and morphological changes were observed to a lesser extent in the RDCE strain. Moreover, exposure to UV-B resulted in a decrease in the concentrations of phycobiliproteins in the RDCE strain. The GBCE strain showed an increase in phycoerythrin (PE)/allophycocyanin (APC) and phycocyanin (PC)/allophycocyanin (APC) ratios after exposure to UV-B, suggesting this strain had a higher tolerance to the radiation. No differences in the chlorophyll a and carotenoid content were found between the control and UV-B treated samples for all strains. Ultrastructural changes, such as damage to chloroplasts and mitochondria, were present in all strains after exposure to UV-B. In summary, our findings support the hypothesis that the population from Ceará State has adapted to the higher irradiation and is thus more resistant to increased UV-B. Additionally, of the strains tested, the GBCE and RDCE strains appear to be more resistant to this radiation.
 
Schematic illustration showing the allocation of Fucus vesiculosus pieces (small rectangles) for a single replicate at one of 12 points in time. Induction aquaria contained 48 pieces (only 4 shown) at the beginning of the experiment from which 4 were allocated to feeding arenas (circles) or gene expression analysis at each of the 12 points in time. Stippled and solid lines indicate containers with and without grazers, respectively. Letters and numbers indicate ungrazed and grazer-exposed pieces of F. vesiculosus, respectively. Reconstituted food items (pentagons) used for feeding assays (subscript a) and corresponding autogenic controls (subscript b) were derived from the same previously grazed or ungrazed seaweed piece. 
Article
This study aimed to reveal the dynamics of inducible anti-herbivory traits in the brown seaweed Fucus vesiculosus, in response to grazing by the isopod Idotea baltica. As the molecular basis of defence induction in seaweeds is poorly understood, a transcriptomic approach was used to gain insight into cellular processes underlying defence induction and thus promote the mechanistic understanding of anti-herbivory responses in seaweeds. In a 27 day feeding-assayed induction experiment, temporal patterns of the induced anti-herbivory resistance of F. vesiculosus were assessed under laboratory conditions. Feeding assays were performed at three day intervals, using fresh and reconstituted food. Microarray hybridizations investigating the expression of genes 3 days before, as well as at the same time as, feeding assays revealed a decreased palatability of previously grazed F. vesiculosus pieces compared with non-grazed control pieces. Despite permanent exposure to grazers, F. vesiculosus palatability varied over time. Non-grazed F. vesiculosus pieces were significantly preferred to grazed pieces after 18 and again after 27 days of previous grazing, while their relative palatability for isopods was comparable at all other times during the experiment. Relative to controls, 562/402 genes were ≥ 1.5-fold up-/down-regulated in seaweed pieces that were grazed for 18 days, i.e. when induction of defences was detected in feeding assays. Massive reprogramming of the regulatory expression orchestra (translation, transcription) as well as up-regulation of genes involved in lipid and carbohydrate metabolism, intracellular trafficking, defence and stress response was found. At the same time, down-regulation of photosynthesis was observed in grazed seaweed pieces. This study suggests a high level of temporal variability in induced anti-herbivory traits of F. vesiculosus and reveals increased expression of genes with putative defensive functions in conjunction with the reallocation of resources from primary to secondary metabolism.
 
Article
Morphological and molecular evidence is provided to further document the status of the enigmatic taxon known as Caulerpa floridana W.R. Taylor from White Shoal, Dry Tortugas, Florida. DNA sequencing of three historical herbarium specimens (WRT329, WRT345 and WRT349) housed at the University of Michigan Herbarium (MICH) demonstrated the molecular separation of this species based on the reconstruction of 931 nucleotides of the chloroplast gene tufA. Caulerpa floridana is sister to the western Atlantic endemic C. ashmeadii Harvey and an unknown Caulerpa taxon from the Florida Middle Grounds. Caulerpa floridana most reliably differs from C. ashmeadii by the presence of a sharp, unequivocal apiculus at the tip of each pinnule. A morphological review of southwestern Atlantic records of C. floridana from Brazil excludes these reports as representative of the species.
 
Comparisons of genetic parameters among three Gracilaria cervicornis populations from Gran Canaria
Article
Genetic variability of gametophytic populations of Gracilaria cervicornis from three localities sampled in the Canary Islands (Spain) was examined by isozyme electrophoresis. Seventeen putative alleles corresponding to 16 gene loci were compared. The genetic variability of populations (percentage of polymorphic loci, mean number of alleles per locus and average gene diversity) was low, with a high number of fixed alleles. These results suggest that the genetic structure of the populations of G. cervicornis from the Canary Islands may be due to a predominance of vegetative propagation and low sporeling recruitment. Some heterogeneity was observed at two polymorphic loci which could be due to random drift. The values obtained in the present study are consistent with the genetic variation described for other red seaweeds with a predominance of asexual reproduction.
 
Article
Fourier-transform infrared (FTIR) spectroscopy was carried out on single colonies of Pediastrum duplex present in air-dried preparations of mixed phytoplankton samples isolated from a eutrophic freshwater lake. FTIR absorption spectra had 12 distinct bands over the wavenumber range 3300–900 cm which were tentatively assigned to a range of chemical groups, including –OH (residual water, wavenumber 3299 cm), –CH2 (lipid, 2924), –C=O (cellulose, 1739), amide (protein, 1650 and 1542), >P=O (nucleic acid, 1077) and –C–O (starch, 1151 and 1077). Measurement of band areas identified residual water, protein and starch as the major detectable constituents. Areas of single bands and combined bands of –CH2, –C–O and >P=O species normalized to protein (to correct for differences in specimen hydration and thickness) showed wide variation between colonies, indicating environmental heterogeneity. Correlation analysis demonstrated close statistical associations between different molecular species. Particularly high levels of correlation between bands 3/4 (CH2), 6/7 (amide) and 8/9 (–CH3) was consistent with their joint origin from the same molecular species. The isolation of bands 11 and 12 in the correlation pattern was confirmed by factor analysis, suggesting that variation in the level of starch is statistically unrelated to other macromolecules being monitored. The use of FTIR spectroscopy to characterize an algal micro-population within mixed phytoplankton has potential for future studies on biodiversity and environmental interactions at the species level.
 
Article
Glutamate-l-semialdehyde (GSA) aminotransferase catalyses the final step in the C5 pathway converting glutamate to the tetrapyrrole precursor δ-aminolaevulinic acid. This enzyme is sensitive to gabaculine (2,3-dihydro-3-amino benzoic acid) and to 4-amino-5-fluoropentanoic acid (AFPA), which are irreversible, mechanism-based inhibitors of pyridoxal phosphate-dependent enzymes. Spontaneous mutants of Synechococcus PCC6301 resistant to these inhibitors contain altered enzyme that displays corresponding resistance to high concentrations of the inhibitor. The enzyme from strain GR6, resistant to both inhibitors, contains a three-amino-acid deletion at positions 5–7 and a Met248[rightward arrow]Ile substitution. The enzyme from strain K40 resistant to AFPA but not to gabaculine, contains a Ser163[rightward arrow]Thr substitution. GSA aminotransferases containing either the deletion or the substitution that are characteristic of the GR6 mutant were produced in Escherichia coli using the expression vector pMalc2. These engineered mutant enzymes were characterized in terms of their catalytic parameters and sensitivities to gabaculine and AFPA. Furthermore, maltose binding protein/aminotransferase fusion proteins were characterized spectrophotometrically to monitor the interaction of bound cofactor with diamino- and dioxo-compounds related to the substrate and both inhibitors. Results were compared with those for similarly produced recombinant wild-type, K40 and GR6 GSA aminotransferases. The engineered products with either the N-terminal deletion or the Met248[rightward arrow]Ile substitution displayed catalytic efficiencies that were intermediate between the wild-type and GR6 or K40 enzymes. However, with respect to their absorption spectra, sensitivity to inhibitors and the reactivity of bound cofactor, they were essentially wild-type. These in vitro studies demonstrate that both changes in enzyme structure are necessary to obtain the distinctive properties of the GR6 aminotransferase, including resistance to high concentrations of gabaculine and AFPA.
 
Article
The Synura petersenii species complex represents a common, cosmopolitan and highly diverse taxon of autotrophic freshwater flagellates. In this paper, we describe and characterize four new species (S. borealis, S. heteropora, S. hibernica and S. laticarina) that have been identified during our extensive sampling of freshwater habitats in 15 European countries. Morphometric analyses of siliceous scales led to the significant phenotypic differentiation of all four newly described species, and their separation from other related species of the S. petersenii complex. Two of these newly described species (S. hibernica and S. borealis) can be clearly distinguished by characteristic large colonies consisting of elongated, lanceolate-shaped cells. Development of strongly elongated, narrow cells in S. hibernica could be explained by the adaptation of this species to oligotrophic conditions. Though morphologically distinct, S. borealis possesses an exceptionally high degree of genetic diversity, possibly indicating recent speciation and evolutionary diversification within this taxon. Three of the four newly described species exhibit restricted biogeographic distribution. The evolutionarily related S. borealis and S. laticarina occur only in Northern Europe, and seem to be adapted to colder areas. The most remarkable distribution pattern was observed for S. hibernica, which has a geographic distribution that is restricted to western Ireland.
 
Article
The cyanobacterium Microcystis PCC7806, grown under continuous light, fermented endogenously stored glycogen to equimolar amounts of acetate and ethanol when incubated anaerobically in the dark. In addition, H2, CO2 and some l-lactate were produced. This fermentation pattern differed from that displayed by cells which had been grown under an alternating light/dark (16/8 h) cycle. Such cells produced much more ethanol than acetate, while no lactate was formed. These differences could not be related to the levels of key enzymes of fermentation, which were identical in the two cultures. The cultures grown under continuous light contained twice as much glycogen as the light/dark-grown cells and the former metabolized it at a rate approximately 3 times as fast as the latter culture. Fermentation in the culture grown under continuous light showed low carbon recovery (59–80%) and high oxidation/reduction balance (approximately 1.5). On the basis of calculations of ATP yield it was concluded that this culture was capable of growth driven by fermentation. The increase in structural cell material would account for the missing carbon and reduction equivalents.
 
Article
Use of isozymes (including allozymes) in studies of population genetics and systematics of seaweeds has increased sufficiently in the last decade to allow some generalization. Only a single locus has been observed for about half the enzymes analysed in seaweeds, compared with 29% in vascular plants. Compared with higher plants, macroalgal species generally have low amounts of electrophoretically detectable genetic variation; the lowest levels of genetic variation found in natural populations are those reported for seaweeds. Nonetheless, seaweeds show an association between levels of genetic diversity as revealed by isozymes and species-specific attributes, such as mating system and predominance of asexual versus sexual reproduction. In systematic studies, isozymes have revealed cryptic species and identified pairs of sibling taxa. The quaternary structure of enzymes appears to be conserved at the phylum level. With the current availability of improved techniques for enzyme electrophoresis and for data interpretation, we expect future studies utilizing isozyme electrophoresis to provide further insight into population and evolutionary processes in seaweeds.
 
Pigment profiles in surface sediments from the Barrow (a–c), Eden (d–f ) and Tagus (g–i) estuaries measured at different times over 1 day. The first sample was always taken immediately after emersion of each study site. a, d, g: Chlorophyll a; b, e, h: xanthophyll pigments in relation to Chl a; c, f, i: de-epoxidation state of the xanthophyll pigments. Zero-values indicate dt below detection limit. Abbreviations: dd ¼ diadinoxanthin; dt ¼ diatoxanthin.  
Characteristics of sites where short-term experiments were conducted. All experiments were performed in May 2003
Relationship between de-epoxidation state, dt/(dt þ dd), and chlorophyll a biomass in three European estuaries. Exponential relationships between the deepoxidation state (y) and chlorophyll a(x), y ¼ y o þ ae Àbx , were significant for the Eden and Tagus estuaries. Samples were taken from 0–500 mm; closed symbols represent surface samples. Abbreviations: dd ¼ diadinoxanthin; dt ¼ diatoxanthin.  
Parameters of the exponential relation y ¼ y o þ ae Àbx between the de-epoxidation state, y, and the chlorophyll a biomass, x, as determined in the Barrow, Eden and Tagus estuaries
Article
Estuarine microphytobenthos are frequently exposed to excessively high irradiances. Photoinhibition in microalgae is prevented by various photophysiological responses. We describe here the role of the xanthophyll pigments in photoacclimation. The pigment composition of the microphytobenthos was studied in three European estuaries (Barrow, Ireland; Eden, UK; Tagus, Portugal). Using HPLC-analyses, microscale changes in biomass and pigment composition were monitored over short (hourly) and long (seasonal) time scales. In the Barrow estuary, the biomass of microphytobenthos (measured as chlorophyll a) increased significantly in the top 400–500 mm of the sediment surface within 1 h of emersion; simultaneously, the xanthophyll pool size (diadinoxanthin plus diatoxanthin, ddþdt) almost doubled. A more gradual conversion of dd into dt was observed, with the dt:dd ratio increasing from <0.1 at the start of emersion to >0.3 after 3 h emersion. Similar trends in the dt:dd ratio were observed in the surface sediments of the Eden and the Tagus estuaries. Higher ratios were recorded in the Tagus estuary, which may be explained by higher incident irradiance. In addition, seasonal studies carried out in the Eden and Tagus estuaries showed that the xanthophyll pool size increased by 10% in the summer months. The pool size was highest in the Tagus estuary. Concurrently, high values for the de-epoxidation state were recorded, with values for dt/(dtþdd)>0.35 recorded in the summer. At the Eden, the ratio never exceeded 0.3. The de-epoxidation state was higher in winter than in summer, which was ascribed to the low winter temperatures. During a vertical migration study, a negative relationship between chlorophyll a and the de-epoxidation state was observed. It is suggested that this relationship originates from ‘micro-migration’ within the biofilm. Migration within the euphotic zone may provide an alternative means for cells to escape photodamage. In this paper, we propose that both xanthophyll cycling and ‘micro’-migration play an important role in photoacclimation and it appears that these processes operate in parallel to regulate the photosynthetic response.
 
Article
Stephanodiscus triporus was described in 1978, based on a study from the phytoplankton of Volgograd Reservoir that used transmission electron microscopy. This species is small, diameter 3.7–10.6 µm, with 14–30 striae and 30–60 areolae in 10 µm, and differs from other Stephanodiscus species by the presence of three satellite pores in the single central fultoportula. Later a new species similar in morphology to S. triporus, was described from Iowa, USA, namely S. vestibulis. A large population of S. vestibulis was found in Lake Balaton and the species also occurs in different Hungarian and French waters. Detailed comparison of S. triporus and S. vestibulis, based on our results and literature, showed they are very similar. Therefore we reinvestigated the type material of both and compared them with the Hungarian and French specimens. Conventional and geometric morphometric analyses were carried out, also including comparisons with the morphologically closest taxon, S. minutulus. There is a continuum of variation between S. triporus and S. vestibulis (in diameter, the number and morphology of the striae, the position of the valve face fultoportula with three satellite pores, the presence of a vestibulum having more or less the same shape). Hence we suggest that they are conspecific, with S. vestibulis a later synonym of S. triporus.
 
Article
Reaching up to 50% of the total biomass in oligotrophic waters and armed with a set of ecological and biological properties related to their small size, picophytoplankton (Minutocellus sp., strain RCC967), and another from oceanic waters in the Indian Ocean (Minutocellus sp., strain RCC703) were used to test hypotheses on the functional relation between ecological niche adaptation and photosynthetic regulation capacity and efficiency. Cultures were subjected to five sine light climates, each one set to peak at a different photon flux density, respectively 10, 50, 100, 250 and 500 µmol photons m−2 s−1. Growth rate, photosynthesis, non-photochemical fluorescence quenching, pigment composition, and particulate organic carbon and nitrogen content were followed daily for 5 days. Growth rate and physiological response curves were different in the two species, in agreement with their distinct habitats of origin. Such differences could be related to the diverse photoacclimative strategies displayed by the two species, revealing a clear adaptive divergence despite their close taxonomic relationship. Photoacclimative strategies of the two picoplanktonic diatoms are discussed in the light of functional diversity and ecosystem adaptation.
 
PCA plots for UPLC-MS data for all injections from the northern (N) and southern (S) group from the 2.5, 8 and 13°C experiments. The three injections of the strain AMB-80 are marked in the figure for 13°C.
Similarity (as % of maximum obtainable similarity) in m/z ratios when comparing sample by sample between only northern, between southern and northern, or between only southern strains of C. socialis, all cultivated at the same temperatures. Acronyms on the x-axis are N (northern) and S (southern), numbers are temperatures in°C and error bars show standard errors (SE).
Article
Morphology and molecular phylogeny constitute the structural elements of diatom taxonomy. These approaches do not, however, give information on the functioning of taxa. Additional methods to serve a more integrated and wide-ranging taxonomy have therefore been called for. Metabolic fingerprinting is one approach used within the field of metabolomics, often applied in classification of samples. Here we apply metabolic fingerprinting in a taxonomic study of a cryptic diatom species. Strains of the cosmopolitan diatom Chaetoceros socialis from two geographical areas; the north-east Atlantic and Arctic and the Gulf of Naples, were cultivated at three different temperatures; 2.5, 8 and 13°C. The strains from the two different geographical areas exhibited different growth rates as well as different photosynthetic efficiencies. Algal extracts, collected at the end of the growth experiments, were analysed by Ultra-Performance Liquid Chromatography High Resolution Mass Spectrometry. The two groups of strains were separated by principal component analysis of their metabolic fingerprints. Analysis of the data revealed both qualitative and quantitative differences in metabolite markers. These phenotypic differences reinforce differences also found for morphology, phylogenetic markers and growth rates, and point at different adaptive characteristics in organisms living under different temperature regimes.
 
Chattonella subsalsa from the Adriatic Sea, TEM. 14. Whole cell of strain CRIM-A in transverse section with clearly separated endoplasm (en) and ectoplasm (ec) and cell organelles. 15. Section in the flagellar area of a cell of strain CRIM-B. Note the mastigonemes (ma) attached to the anterior flagellum (af) and the rhizoplast (rh). 16. Detail of cell of strain CRIM-C with osmiophilic granules (arrowheads) at the surface. 17. Transverse section across the anterior flagellum (af) of a cell of strain CRIM-B, showing several rows of mastigonemes attached to the flagellar membrane. 18. Section of a cell of strain CRIM-A showing the Golgi body (g) adjacent to the nucleus (nu) in the flagellar area and other cellular organelles, such as pyrenoids (py) within the chloroplasts (ch), and mitochondria (m). Scale bars = 5 µm (Figs 14, 18), 1 µm (Fig. 15), 2 µm (Fig. 16) and 0.2 µm (Fig. 17).  
Micrographs of Chattonella subsalsa from the Gulf of Mexico (strain CCMP217) obtained by LM (Figs 19–21), SEM (Figs 22–24) and TEM (Fig. 25). 19. Living cell showing the anterior flagellum (af) beating and the posterior flagellum (pf) pointing backwards. 20. Living cell with a posterior tail (arrow). 21. Living cell showing peanut-shaped chloroplasts. 22. Whole cell with irregular and peanut-shaped chloroplasts (ch). 23. Detail of the flagellar area, showing the mesh-like structure at the base of the anterior flagellum (af). 24. Detail of cell surface showing small granules. 25. Section showing chloroplasts (ch) with pyrenoids (py), and the nucleus (nu). Scale bars = 10 µm (Figs 19–21), 5 µm (Fig. 22) and 2 µm (Figs 23–25).  
Neighbour joining (NJ) phylogenetic trees of selected sequences (all haplotypes) of five genetic markers with accompanying base pair differences and bootstrap values (in parentheses). Branch lengths indicate genetic distances (scale bars are shown in individual trees). See also ML trees in Figs S1-S5. Raphidophytes other than Chattonella were used as outgroups (Heterosigma akashiwo or Fibrocapsa cf. japonica). 1. NJ tree of eight selected nuclear SSU sequences of Chattonella spp. 2. NJ tree of six selected nuclear LSU sequences of Chattonella spp. 3. NJ tree of six selected ITS sequences of Chattonella spp. 4. NJ tree of five selected psaA sequences of Chattonella spp. 5. NJ tree of six selected rbcL sequences of Chattonella spp.
Chattonella subsalsa from the Adriatic Sea, LM (Figs 6-9) and SEM (Figs 10-13). 6. Living cell of strain CRIM-B with the undulated flagellum directed anteriorly (af), the posterior flagellum directed backwards (pf) and the chloroplasts (ch). 7. Slightly swollen living cell of strain CRIM-B showing chloroplasts (ch), and larger button-like (arrows) and smaller rounded (arrowheads) surface bodies. 8. Cells of strain CRIM-A stained with Neutral Red showing a massive discharge of oboe-shaped mucocysts (mu). 9. Detailed micrograph of mucocysts discharged following Neutral Red staining of a cell of strain CRIM-A. 10. Cell surface of a cell of strain CRIM-A showing rounded bodies arranged in circles around a central depression (arrows). 11. Detail of the cell surface of strain CRIM-A with a ring of small rounded bodies surrounding an aperture (arrow). 12. Whole cell of strain CRIM-A with stretched flagella disappearing in filter holes. 13. Cell of strain CRIM-A surrounded by numerous mucocysts (mu). Scale bars = 10 µm (Figs 6-9, 12, 13), 1 µm (Fig. 10) and 0.5 µm (Fig. 11).
List of selected strains and isolates for the presented phylogenetic trees, including information on origin, applied genetic marker and GenBank accession numbers. GenBank entries in bold indicate sequences obtained for this study.
Article
We analysed the molecular and morphological features of strains of Chattonella subsalsa isolated from the western Adriatic coast (Mediterranean Sea), with the aim of confirming their classification and elucidating their phylogenetic positions within the Raphidophyceae. We sequenced parts of the ribosomal operon, including the small subunit (SSU), the internal transcribed spacer region (ITS) and the large subunit (LSU) of the rDNA. Additionally, we analysed sequences of the chloroplast-encoded subunit psaA of Photosystem I (PSI) and rbcL, encoding the large subunit of the Rubisco gene. For three phylogenetic markers (LSU, ITS, rbcL), the sequences of the strains from the Adriatic Sea were identical and for two markers (SSU, psaA) only minor differences occurred. All strains were sister to, but well separated from, sequences from isolates in culture collections and from GenBank, thus far classified as belonging to C. subsalsa. Light and electron microscopy provided evidence for morphological differences between a strain of C. subsalsa (CCMP217) from the Gulf of Mexico and the isolates from the Adriatic Sea. Differences concerned the shape and arrangement of chloroplasts and the presence of mucocysts and other surface microstructures, which were only observed in isolates from the Adriatic Sea. This is the first evidence for two different taxa classified as C. subsalsa, which are clearly separated on the basis of several genetic markers and also show morphological differences. As compared with strains assigned to C. subsalsa from the NCMA (formerly CCMP) culture collection, the Adriatic strains more closely match the original species description. This would imply that strain CCMP217 and other genetically similar strains should be described under a new species name. Nevertheless, given the high morphological plasticity of Chattonella species, the definition of the true C. subsalsa must be decided based on detailed morphological and molecular analysis of more strains from other geographical areas.
 
Article
Cyanobacteria synthesize many secondary metabolites of various chemical structures and biological activity and release them into the aquatic environment. Two new oligopeptides belonging to anabaenopeptins and cyanopeptolins were isolated from an enrichment culture dominated by Woronichinia naegeliana (Unger) Elenkin. The application of mass spectrometry and nuclear magnetic resonance spectroscopy allowed determination of their structure and identification of these compounds as cyanopeptolin 1081 and anabaenopeptin 899 with a molecular formula C52H75N9O16 and C48H65N7O10, respectively. The preliminary evaluation of their toxicity was conducted via tests based on crustacean Thamnocephalus platyurus. Although these peptides did not exert acute toxic effects at the concentration up to 40 μg ml–1, deterioration of the physiological state of the studied crustaceans, indicated by reduced mobility, was observed. Considering the limited knowledge of cyanopeptide presence in the aquatic environment, it is important to identify and characterize these compounds.
 
Article
Previous studies have shown that dinoflagellates with different plastid ancestries have distinct differences in the fatty acid compositions and regiochemistries of their chloroplast-associated galactolipids, mono- and digalactosyldiacylglycerol (MGDG and DGDG, respectively), thus reflecting plastid origin as a major factor in plastid membrane composition. Specifically, dinoflagellates with aberrant plastids (e.g. Karenia brevis, Kryptoperidinium foliaceum and Lepidodinium chlorophorum) possess certain MGDG- and DGDG-associated fatty acids which are not found in peridinin-containing dinoflagellates (the largest group of photosynthetic dinoflagellates with a red algal plastid ancestry which is thought to be an evolutionary precursor to aberrant plastids), but which are common to other algal groups. For example, hexadecatetraenoic acid (16:4(n-3)) is common to green algae and is found in the MGDG and DGDG of L. chlorophorum, which agrees with its green algal plastid ancestry, while hexadecatrienoic acid (16:3) and hexadecadienoic acid (16:2) are found in the MGDG and DGDG of K. foliaceum, which agrees with its diatom plastid ancestry. Notably, 16:4 has been found by others in the total fatty acids and galactolipids of Karenia mikimotoi, but in no other examined members of the Kareniaceae (all of which have plastids of haptophyte origin). However, these findings lack information as to the regiochemistry of 16:4. We have utilized positive-ion electrospray ionization/mass spectrometry (ESI/MS) and ESI/MS/MS to demonstrate that 16:4, which aside from L. chlorophorum is not found conclusively in the MGDG and DGDG of any other dinoflagellates examined to date irrespective of plastid ancestry, is found in K. mikimotoi as 18:5/16:4 (sn-1/sn-2 regiochemistry) MGDG and DGDG, and that its presence is not modulated (i.e. does not become more saturated) with an increase in growth temperature. Considering an aberrant pigment composition as described by others, we present a perspective where galactolipid-associated 16:4 in K. mikimotoi indicates a plastid ancestry more convoluted than for other members of the Kareniaceae.
 
Article
Abstract Diatoms represent an important class of aquatic phototrophs. They are not only one of the major contributors to global carbon fixation, but they also play a key role in the biogeochemical cycling of silica. Molecular identification methods based on conserved DNA sequences, such as internal transcribed spacer (ITS) have revolutionized our knowledge and understanding of conventional taxonomy. In this study, we aimed to compare the conventional identification methods with molecular identification methods. To do so, we isolated 4 diatom samples from the coast at Urla and characterized them Using light microscopy (LM) and scanning electron microscopy (SEM) according to morphological features. Then we amplified ITS regions using a conventional polymerase chain reaction (PCR), sequenced the PCR products, and analyzed the sequences using bioinformatic tools. Bioinformatic analysis indicated that the isolated species had high sequence similarity to Pseudo-nitzschia delicatissima, Achnanthes taeniata, Amphora coffeaeformis, and Cylindrotheca closterium. We think that molecular identification methods enable rapid and more reliable identification of diatom species and are crucial for monitoring harmful algal blooms.
 
Article
The only eukaryotes found in highly acidic environments (pH 0.5–3) with elevated temperatures (up to 56 °C) are three species of unicellular red algae: Cyanidioschyzon merolae, Cyanidium caldarium and Galdieria sulphuraria. These habitats are scattered all over the world and are usually very small. Because all three species are strictly acidophilic and will not tolerate desiccation, distribution by wind or water seems very unlikely. The populations in the various habitats might have been isolated for very long times, providing a unique opportunity to observe a significant degree of independent evolution under strong selective pressure. We investigated the biogeography of 18 isolates of these red algae by comparison of partial sequences of the 18S rRNA. A gene tree based on 18S rRNA assigns 15 strains to one branch with high bootstrap values. These isolates share the feature of facultative heterotrophy, whether or not they were originally placed in the genus Galdieria. The remaining three strains — Galdieria maxima, C. merolae and C. caldarium — form a sister clade to this group. The exact position of these two groups in relation to other red algae remains unresolved. The evolutionary distance between individual Galdieria strains is high, indicating that within this genus several races have developed significantly altered 18S sequences. Our comparison of 18S sequences from thermo-acidophilic red algae indicates that even within a seemingly homogeneous group of eukaryotic organisms the limits of phylogenetic analysis may be reached.
 
Article
Morphometric analysis and phylogenetic analysis of sequences of the rbcL chloroplast gene (which codes for the large subunit of the ribulose-1,5-bisphosphate carboxylase/oxygenase enzyme) and the nuclear 18S ribosomal RNA (rRNA) gene were carried out on 26 specimens of marine and freshwater Hildenbrandia from North America. Nineteen marine specimens were collected from Alaska to Costa Rica on the Pacific coast and from Newfoundland to Connecticut on the Atlantic coast. Seven freshwater samples were collected from Texas, Costa Rica, St Lucia and Puerto Rico. Three groups of samples were distinguished by morphometric analysis: one containing all freshwater samples (H. angolensis Welwitsch ex West et West), one consisting of a marine sample with parallel tetrasporangial divisions (H. occidentalis Setchell ex Gardner) and one group with non-parallel tetrasporangial divisions (H. rubra (Sommerfelt) Meneghini. These groupings were partially incongruent with those obtained by analyses of the molecular data. Parsimony and distance analyses of the rbcL gene resulted in trees in which Atlantic and Pacific clades were largely resolved. However, an Alaskan sample was included in the Atlantic group, which may indicate a trans-Arctic invasion event. The freshwater samples were paraphyletic for the rbcL gene, among the marine collections, which supports the concept of multiple invasions establishing the freshwater populations in North America. The 18S rRNA gene sequence data indicate that the freshwater samples are monophyletic with the exception of the unresolved position of the H. occidentalis sample. The freshwater samples form a monophyletic clade when multiple outgroups are used. The rbcL data appear to be mutationally saturated above approximately 17% divergence, which makes interpretation of phylogenetic signal among distant groups difficult. This may be a result of the asexual reproduction of the alga.
 
Article
Twenty-one marine and freshwater specimens of Hildenbrandia from Europe were compared by analyses of morphometric data and sequences of the rbcL chloroplast gene (which codes for the large subunit of the ribulose-1,5-bisphosphate carboxylase/oxygenase enzyme) and the nuclear 18S ribosomal RNA (rRNA) gene. Eleven freshwater specimens, all corresponding to H. ribularis, were collected from the British Isles, Germany, France, Austria Spain and Italy. Six marine specimens, representing H. rubra, were obtained from Northern Ireland, Wales, France, Norway, Sweden and the Netherlands, while four marine specimens, corresponding to H. crouanii, were collected from Scotland and Germany. Morphometric analyses of vegetative and reproductive characters distinguished three groups of specimens, which correspond to the three species. Parsimony and distance analyses of the rbcL and 18S rRNA genes were performed on these samples and sequences of North American representatives were included to determine their inter-relationships. The rbcL gene sequence analyses separated the marine and freshwater specimens, with the H. rivularis and H. angolensis representatives forming a well-supported monophyletic clade. The rbcL sequence analyses did not resolve the groupings of marine specimens determined in the morphometric analysis. Analyses of the 18S rRNA gene sequences also supported separation of the marine samples from freshwater samples, although the bootstrap and decay support was not as strong as for the rbcL gene. H. rubra formed a monophyletic clade that was weakly supported. In terms of biogeographic trends, the fairly low intraspeci8e divergence of H. rivularis (0-1.9% for the rbcL gene and 0-3.6% for the 18S rRNA gene) and the positioning of these samples in the gene trees supports the concept that they are derived from marine populations.
 
Article
A distinct pattern of seasonal fluctuations in intra-specific cell size and/or colony size was observed in a large number of phytoplankton species from Lake Kinneret, Israel. The same species showed larger cell size or colony size in late winter and smaller size in late summer, with intermediate sizes in the interim periods. This phenomenon was exhibited by species of chlorophytes, dinoflagellates and cyanobacteria that were abundant enough to be sampled (fortnightly) and measured throughout the year. The annual pattern of fluctuations in size repeated itself over 8 consecutive years (2004-2012). The size fluctuations were independent of the temporal changes observed in cell abundance of each species. Rather, peak sizes coincided with lowest water temperature and highest nutrient availability, minimum sizes with highest water temperature and lowest nutrient availability. These observations fit well with current ecological theory on organism size, where larger organisms occur in colder climates and the reverse in warmer climates.
 
Article
The chloroplast genome contains information that is applicable in many scientific fields, such as plant systematics, phylogenetic reconstruction and biotechnology, because its features are highly conserved among species. To date, several complete green algal chloroplast genomes have been sequenced and assembled. In this study, the nucleotide sequence of the chloroplast genome (cpDNA) of Chlorella sorokiniana SAG 211-8k is reported and compared for the first time to the chloroplast genomes of 10 Chlorellaceae. The recently updated Chlorella sorokiniana cpDNA sequence, assembled as a circular map of 109 811 bp, encodes 113 genes. Similar to other Chlorella strains, this chloroplast genome does not show a quadripartite structure and lacks the large rRNA operon-encoding Inverted Repeat (IR). The Chlorella sorokiniana plastid encodes the tRNA(Ile)-lysidine synthetase (tilS), which is responsible for modifying the CAU anticodon of a unique tRNA. Gene ordering and clustering highlight the close relationships among Chlorella clade members and the preservation of crucial gene clusters in photosynthetic strains. The features of Chlorella sorokiniana presented here reinforce the monophyletic character of Chlorellaceae and provide important information that sheds light on chloroplast genome evolution among species of Chlorella.
 
Article
Arthrospira are multicellular cyanobacteria that typically reside in alkaline lakes of (sub)tropical regions and are mass cultivated around the globe in a variety of outdoor facilities and photobioreactors for their high nutritional, pharmaceutical and clinical value. Arthrospira sp. strain PCC 8005 was selected by the European Space Agency as an oxygen producer and nutritional end-product in a bioregenerative life support system for long-haul missions. Being highly resistant to ionizing radiation, it is also an ideal candidate for other space applications such as in situ resource utilization and terraformation. During long-term strain maintenance involving continuous subculturing we noted an irreversible morphological change in PCC 8005 subcultures i.e. from only helical to only straight trichomes. These morphotypes displayed differences in growth rate, buoyancy and resistance to gamma radiation. We also found marked differences in antioxidant capacity, pigment content and trehalose concentration, while whole-genome comparison revealed a difference of 168 SNPs, 48 indels and four large insertions affecting, in total, 41 coding regions across both genomes. Although nine of these regions encoded proteins with a known function, no conclusive genotype-phenotype associations could be determined. Nonetheless, genomic changes within the gvpC gene (encoding a gas vesicle protein) and within the regulatory region of the psbD gene (encoding the D2 protein of PSII) provided some clues for the observed differences in buoyancy and growth.
 
Article
Cyanobacteria secrete exopolysaccharide (EPS), which can be intimately associated with the cell surfaces (known as capsular polysaccharide, CPS), or released into the surrounding environment (released polysaccharide, RPS). The aim of this study was to explore the role of EPS in cell aggregation and biofilm formation in the unicellular cyanobacterium Synechocystis sp. strain PCC 6803. Three mutants (Δsll5043, Δslr1063 and Δslr1076) were obtained through disrupting three putative glycosyltransferase genes (sll5043, slr1063 and slr1076) separately. Decreased contents and altered monosaccharide composition of both RPS and CPS were observed in the three mutants compared with the wild type (WT). RPS from the WT (WT-RPS) had high emulsification activity, while RPSs from the mutants had no emulsification activity. Unlike the WT, the three mutants formed cell aggregates and biofilms. Compared with the WT, the three mutants possessed lower cell hydrophobicity and less negative cell surface charge, which could result from the altered production of CPS. 40 μg ml–1 WT-RPS significantly decreased cell hydrophobicity of the three mutants, and significantly inhibited cell aggregation and biofilm formation, while the RPS from each mutant itself had no effect on cell hydrophobicity of the mutant, and did not affect cell aggregation and biofilm formation. The inhibitory effect of WT-RPS on cell aggregation and biofilm formation was thus suggested to be related to its ability to reduce cell hydrophobicity. The difference in cell aggregation and biofilm formation between the WT and the three mutants could be related to differences in cell hydrophobicity, cell surface charge and physicochemical properties of RPS. Our results in this study indicated that both CPS and RPS affected cell aggregation and biofilm formation in Synechocystis sp. strain PCC 6803, and WT-RPS inhibited cell aggregation and biofilm formation.
 
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Richard J. Geider
  • University of Essex
Julie Laroche
  • Dalhousie University
Daniel C.O. Thornton
  • Texas A&M University
Rodney Forster
  • University of Hull
Brigitte Gavio
  • National University of Colombia