UDPGLUCOSE PYROPHOSPHORYLASE ACTIVITY IN THE DIATOM CYCLOTELLA CRYPTICA. PATHWAY OF CHRYSOLAMINARIN BIOSYNTHESIS 1

UDPglucose pyrophosphorylase activity was detected in cell-free extracts of the diatom Cyclotella cryptica TI3L Reimann, Lewin and Guillard. When assayed in the direction of UDPglucose formation, the enzyme had maximal activity at pH 7.8 and was stimulated by Mg²⁺and Mn²⁺ions. 3-Phosphoglycerate and inorganic phosphate had little effect on enzymatic activity, and the enzyme was relatively insensitive to feedback inhibition from UDPglucose (K, > I millimolar). A glucan was formed from UDP-[¹⁴C]glucose in cell-free extracts of C. cryptica. This glucan had a median molecular weight of 4600 (as determined by gel filtration chromatograbhy) and could be hydrolyzed by laminarinase. Partial acid hydrolysis of the glucan resulted in the formation of glucose and laminaribiose. but not cellobiose. These results suggest that the synthesis of chrysolaminarin (the major storage carbohydrate of diatoms) occurs via the activity of UDPglucose pyrophosphorylase. followed by glucosyl transfer from UDPglucose to the growing β-(1–3)-linked glucan.     

GROWTH RESPONSES OF THE DIATOM,CYCLOTELLA CRYPTICA (BACILLARIOPHYCEAE), TO GIBBERELLIC ACID1

The plant hormone, gibberellic acid (GA), stimulated growth of a marine diatom, Cyclotella cryptica Reimann, Lewin and Guillard. Four concentrations of GA (5 × 20 × 25 × and 35 × 10^?6 g/mL) were added to axenic cultures of C. cryptica. Changes in cell densities, measured by cell counts and turbidimetric readings, confirmed that GA at 20 × 10^?6 g/mL produced maximum stimulation. There was an increase in the total number of cells produced and a shorter lag phase of growth at this concentration. Coulter counter measurements of cell size, as well as ocular micrometer measurements, indicated there was no significant variation in cell volumes of GA grown cells over that of the controls.     

EFFECTS OF SILICON DEFICIENCY ON LIPID COMPOSITION AND METABOLISM IN THE DIATOM CYCLOTELLA CRYPTICA1

The effects of silicon deficiency on the metabolism and composition of lipids in Cyclotella cryptica T13L Reimann, Lewin, and Guillard were examined. Silicon-deficient cells had higher levels of neutral lipids (primarily triacylglycerols) and higher proportions of saturated and monounsaturated fatty acids than silicon-replete cells. After 4 h of silicon deficiency, the percentage of newly assimilated NaH¹⁴CO₃ partitioned into lipids increased from 27.6% to 54.1%, whereas the percentage partitioned into chrysolaminarin decreased from 21.6% to 10.6%. In addition, pulse-chase experiments with NaH¹⁴CO₃ indicated that the amount of ¹⁴C in the total cellular lipid fraction increased by 32% after 12 h of silicon deficiency despite the absence of additional photoassimilable ¹⁴C. Therefore, the accumulation of lipids in response to silicon deficiency appears to be due to two distinct processes: (a) an increase in the proportion of newly assimilated carbon partioned into lipids, and (2) a slow conversion of previously assimilated carbon from non-lipid compounds into lipids     

MONITORING ULTRAVIOLET-B-INDUCED DNA DAMAGE IN INDIVIDUAL DIATOM CELLS BY IMMUNOFLUORESCENT THYMINE DIMER DETECTION1

We developed a method to investigate the effect of ultraviolet-B radiation (UVBR) on the formation of thy-mine dimers in microalgal DNA that can be used for both laboratory and in situ research. Antibody labeling of dimers was followed by a secondary antibody (fluorescein isothiocyanate) staining to allow visualization of DNA damage with flow cytometry or fluorescence microscopy. Thymine dimer-specific fluorescence in nuclear DNA of the marine diatom Cyclotella sp. was linearly related to the UVBR dose. Simultaneous measurements of cellular DNA content showed that the vulnerability of G2 cells to DNA damage did not differ significantly from the vulnerability of G1 cells. The formation and removal of thymine dimers in Cyclotella sp. cells was monitored for 3 consecutive days at two realistic UVBR irradiance levels. Thy-mine dimers were removed within 24 h when exposed to a saturating photosynthetically active radiation intensity following the UVBR treatment. This new method allows the study of UVBR-induced DNA damage on a cell-to-cell basis. It is also feasible for field studies because cells remain intact and can be recognized readily after antibody treatment.     

DIATOM MINERALIZATION OF SILICIC ACID. VIII. METABOLIC REQUIREMENTS AND THE TIMING OF PROTEIN SYNTHESIS1

We have examined the influence of temperature, protein synthesis, and energy metabolism on the process of silicon biomineralization in synchronized cultures of the diatom Navicula saprophilia Lange-Bertalot & Bonik (1976). Temperature effects on silicon polymerization were compared in vitro and in vivo. In vivo incorporation was very temperature dependent with a Q₁₀ of 7.53. In contrast, the Q₁₀ for in vitro polymerization was 1.42, indicating much lower temperature dependence. This difference in Q10 values suggests that in vivo polymerization involves more than autopolycondensation. Cycloheximide addition to synchronized cultures up to, but not later than one hour after the addition of silicic acid depressed total uptake, incorporation, but not pool size. Developing valves demonstrated morphological abnormalities with cycloheximide additions from 0 to 2 h following silicic acid addition. These data suggest that de nova proteins are required in biomineralization and that they are synthesized during or just after cytokinesis. Biomineralization is not coupled to energy derived directly from photosystem II or photosynthesis, since neither darkness nor DCMU had an effect on any aspect of silicification.     

DIATOM ASSEMBLAGES OF STREAMS INFLUENCED BY WILDFIRE1

The Greater Yellowstone Area ecosystem experienced major wildfires in 1988, resulting in a substantial number of catchments being burned. We studied diatom assemblage structure at 14 sites over 5 years in catchments ranging from 0 to over 90% burned. Coefficients of variation for selected physical measures provided a good assessment of the degree of disturbance experienced by the various sites and correlated highly with percentage of catchment burned. Principal components analysis quantitatively estimated the degree of temporal change in diatom assemblages at a site. The more extensively burned catchments showed the greatest amount of structural change in diatom taxa among years. Navicula permitis Hust. and Nitzschia inconspicua Grun. were predominant in the heavily disturbed systems, whereas Achnanthes minutissima Kuetz., A. lanceolata (Breb.) Grun., Rhoicosphenia curvata (Kuetz.) Grun. ex Rabh., and Diatoma hiemale var. mesodon (Ehr.) Grun. were common in less disturbed streams. The results demonstrate that recovery trajectories of lotic diatom assemblages following major landscape-scale disturbances are a function of disturbance intensity resulting from differences in catchment characteristics.     

A NEW SPECIES OF THE DIATOM GENUS EUNOTIA1

A new species of the diatom genus Eunotia is described. The species, E. catillifera Morrow, is distinguished by the presence of fin-like plates, or transapical ridges, which alternate with the striae on the valve faces. This alga appears to be intermediate between the more typical Eunotia species and Semiorbis Patr. This observation suggests that Semiorbis hemicyclus (Ehr.) Patr. in Patrick and Reimer be returned to Eunotia hemicyclus Ralfs in Pritchard and that the use of the generic name Semiorbis be discontinued.     

IMPORTANCE OF RESTING STAGES IN DIATOM SEASONAL SUCCESSION1

The role of environmental factors in seasonal succession of six clones of marine diatoms was examined by testing effects of various combinations of temperature (5°, 10°, 15°, 20°C) and photoperiod (10, 12, 14, 16 h) on time to germination of resting stages and subsequent vegetative cell division rates. Resting stages were induced in batch cultures using a low nutrient medium and a cold, dark period. A three-factor ANOVA showed that species, temperature, daylength, and their interactions signficantly (P =0.0001) injuenced germination time and division rate. Resting stages of Chaetoceros similis Cleve and Odontella aurita (Lyngbye) Agardh were most successful in producing populations of cells on warm, long days (20°C/16 h). Chaetoceros didymus Ehrenberg and C. cinctus Gran were most successful under more moderate conditions (15°C/14 h). Resting stages of Ditylum bright-wellii (West) Grunow germinated and cells divided rapidly under all conditions. No consistent pattern was evident with Thalassiosira rotula Meunier. We compared results of the laboratory study with environmental conditions prevailing during the presence of these species in a British Columbia fjord. Favorable growth conditions determined for C. didymus, D. brightwellii, and T. rotula were similar to conditions present when these species were commonly found in the fjord.     

ECOLOGICAL CONSIDERATIONS OF DIATOM CELL MOTILITY. I. CHARACTERIZATION OF MOTILITY AND ADHESION IN FOUR DIATOM SPECIES1

To better determine the ecological role of motility in pennate diatoms, we quantitatively characterized several motility and adhesion properties of four species of motile pennate diatoms (Craticula sp., Pinnularia sp., Nitzschia sp., and Stauroneis sp.) isolated from the same freshwater pond. Using computer-assisted video microscopy, we measured speed, size/shape, functional adhesion, path curvature, and light sensitivity for these species, each of which shows a distinctive set of motile behaviors. The average speeds of Stauroneis, Pinnularia, Nitzschia, and Craticula cells are 4.6, 5.3, 10.4, and 10.0 μm · s^?1, respectively. Craticula and Nitzschia cells move in a relatively straight path (<4 degrees rotation per 100 μm movement), Stauroneis exhibits minor rotation (about 7 degrees per 100 μm movement), and Pinnularia rotates considerably during movement (about 22 degrees per 100 μm moved). Functional adhesion (as measured by the release rate of attached cells from the underside of an inverted coverslip) shows a half time for cell release of approximately 50 min for Craticula, 192 min for Pinnularia, and >1 day for Nitzschia and Stauroneis. Direction reversal at light/dark boundaries, which appears to be the main contributor to diatom Phototaxis, is most responsive for Craticula, Pinnularia, and Nitzschia at wavelengths around 500 nm. Craticula and Nitzschia cells are the most sensitive in the photophobic response, with over 60% of these cells responding to a 30-1x light/dark boundary at 500 nm, whereas Pinnularia cells are only moderately responsive at this irradiance, showing a maximal response of approximately 30% of cells at 450 nm. Stauroneis cells, in contrast, had a maximal photosensitive response at 700 nm, suggesting that this cell type may use a different response mechanism than the other three cell types. In addition, Craticula and Pinnularia show a net movement out of the light spot when illuminated at 650 nm, whereas Stauroneis shows a net movement out of the light spot when illuminated at 450 nm. Such quantitative characterizations of species-specific responses to environmental stimuli should give us a firm foundation for future studies analyzing the behavior of interspecies diatom competition for limited light or nutrient resources.     

PHOTOSYNTHESIS AND COLD ACCLIMATION: MOLECULAR EVIDENCE FROM A POLAR DIATOM1

The psychrophilic diatom Fragilariopsis cylindrus (Grunow) Krieger in Helmcke & Krieger was used to investigate photosynthesis and growth under freezing temperatures. Gene expression during a temperature shift from +5° C to ?1.8° C was studied under 3 and 35 μmol photons·m^?2·s^?1 by using a macroarray. These measurements were paralleled by determination of fluorescence induction at PSII and pigment analysis. The shift to ?1.8° C at 35 μmol photons·m^?2·s^?1 caused a marginal decrease of photosynthetic quantum yield (F_v/F_m) from 0.61 to 0.52 with fast recovery after 1 day. The ratio of chl c to chl a increased from 3.1 to 5.5, and the ratio of diatoxanthin to diadinoxanthin increased from 0.7 to 5.0. Genes encoding proteins of PSII (psbA, psbC) and for carbon fixation (rbcL) were down‐regulated, whereas genes encoding chaperons (hsp70) and genes for plastid protein synthesis and turnover (elongation factor EfTs, ribosomal protein rpS4, ftsH protease) were up‐regulated. In contrast, cold exposure at 3 μmol photons·m^?2·s^?1 induced a marginal increase in F_v/F_m from 0.61 to 0.63 and a strong increase in fucoxanthin concentrations from 0.04 up to 0.12 pg·cell^?1. This was paralleled by up‐regulation of fcp genes. The ratio of chl c to chl a also increased from 3.1 to 4.2, as did the ratio of diatoxanthin to diadinoxanthin from 0.7 to 2.2. Down‐regulation of psbA, psbC, and rbcL could also be measured but not up‐regulation of hsp70, EfTs, rpS4, and the ftsH protease. The latter genes are probably necessary to avoid cold shock photoinhibition only at higher light intensities.     

ACTIVE GLIDING MOTILITY IN AN ARAPHID MARINE DIATOM,ARDISSONEA (FORMERLY SYNEDRA) CRYSTALLINA1

Active gliding movement over long distances was observed and filmed in the marine pennate diatom Ardissonea (Synedra) crystallina (Agardh) Kütz. Typical speeds measured ca. 1–2 μm-s^?1. Motion wax often smooth and steady; however, discontinuous jerky motions and rolling movements were common. Motion, was associated with secretion of twin or, less commonly, single straight trails of mucilage from one end of the cell. In a few instances, reversal in direction was related to cessation of mucilage secretion at one end and commencement at the other. Temporary cessation of movement due to an obstruction was accompanied by a build-up of mucilage at one end of the cell. Mucilage was apparently secreted at two specific sites at each end of the cell and was stained by alcian blue. Persistent trails were visible under scanning electron microscopy (SEM). SEM confirmed that cells had no raphes or labiate processes. The apparent site of secretion was a deep groove formed at the junction of the valve and valvocopula (first girdle band) at each end of the cell. Transmission electron microscopy confirmed the presence of mucilage vesicles in the cytoplasm, but these were not in any manner obviously related to secretion nor was any morphological structure associated with secretion. Cells often become epiphytic through secretion of a terminal stipe. Both stipe secretion and movement may involve the same structural differentiation of the frustule. These results demonstrate a previously unrecorded type of diatom motility. The mechanism, involves mucilage secretion and appears similar to that seen, for example, in some other algae such as the desmids (green algae).     

OBSERVATIONS OF A TUBE-DWELLING DIATOM: NAVICULA HAMULIFERA (BACILLARIOPHYCEAE)1

Colonies of the tube-dwelling diatom Navicula hamulifera Grunow living on mangrove prop roots in Indian River. Florida and at La Parguera, Puerto Rico, were studied using light and electron microscopy. Observations of the tube morphology and cell structure of this diatom from fresh samples and cultures are described, as well as the ultrastructural morphology of its frustule. The formation of tubes by this diatom is reported for the first time. Comparisons are made with the closest species; Navicula delognei V.H. and Navicula pseudocomoides Hendey.     

EFFECTS OF HARVEST STAGE AND LIGHT ON THE BIOCHEMICAL COMPOSITION OF THE DIATOM THALASSIOSIRA PSEUDONANA1

The marine diatom Thalassiosira pseudonana (Hustedt, clone 3H) Hasle and Heimdal was cultured under three different light regimes: 100 μmol photon · m^?2· s^?1 on 12:12 h light : dark (L:D) cycles; 50 μmol photon · m^?2· s^?2 on 24:0 h L:D; and 100 μmol photon · m^?2· s^?1 on 24:0 h L:D. It was harvested during logarithmic and stationary phases for analysis of biochemical composition. Across the different light regimes, protein (as % of organic weight) was highest in cells during logarithmic phase, whereas carbohydrate and lipid were highest during stationary phase. Carbohydrate concentrations were most affected by the different light regimes; cells grown under 12:12 h L:D contained 37–44% of the carbohydrate of cells grown under 24:0 h L:D. Cells in logarithmic phase had high proportions of polar lipids (79 to 89% of total lipid) and low triacylglycerol (≤10% of total lipid). Cells in stationary phase contained less polar lipid (48 to 57% of total lipid) and more triacylglycerol (22 to 45% of total lipid). The fatty acid composition of logarithmic phase cells grown under 24:0 h L:D were similar, but the 100 μmol photon · m^?2· s^?1 (12:12 h L:D) cells at the same stage contained a higher proportion of polyunsaturated fatty acids (PUFAs) and a lower proportion of saturated and monounsaturated fatty acids due to different levels of 16:0, 16:1(n-7), 16:4(n-1), 18:4(n-3), and 20:5(n-3). With the onset of stationary phase, cells grown at 100 μmol photon · m^?2· s^?1 (both 12:12 and 24:0 h L:D) increased in proportions of saturated and monounsaturated fatty adds and decreased in PUFAs. Concentrations (% organic or dry weight) of 14:0, 16:0, 16:1(n-7), 20:5(n-3), and 22:6(n-3) increased in cells of all cultures during stationary phase. The amino acid compositions of cells were similar irrespective of harvest stage and light regime. For mariculture, the recommended light regime for culturing T. pseudonana will depend on the nutritional requirements of the animal to which the alga is fed. For rapidly growing bivalve mollusc larvae, stationary-phase cultures grown under a 24:0 h L:D regime may provide more energy by virtue of their higher percentage of carbohydrate and high proportions and concentrations of energy-rich saturated fatty acids.     

FUNGAL CONTROL OF POPULATION CHANGES OF THE PLANKTONIC DIATOM ASTERIONELLA FORMOSA IN A SHALLOW EUTROPHIC LAKE1

Parasitic fungi infected an Asterionella formosa Hass. population for 10 months of the year, during which time the population density of the alga was over 100 cells-L^?1. High injection rates of more than 20% of the cells were observed between February and July except on a few occasions such as late April and late June. Mortality of infected diatom cells was temperature dependent in dialysis tube culture experiments; the loss rate of field populations due to fungal injection also was affected by temperature. Loss rates of diatom cells were negligibly small m February even though infection percentages were more than 20%, but increased toward summer with an increase m temperature. High infection percentages and correspondingly high loss rates (e.g. 0.3 d^?1) occurred periodically during the summer. The results suggest that this type of host-parasite interaction is an important control of algal population size in natural waters.     

VOLATILE ORGANIC SULFIDES FROM FRESHWATER ALGAE1

Dimethyl sulfide (DMS) was found to be the principle biogenic sulfur compound in a freshwater environment. Endemic and non-endemic algae were cultured under axenic and nonaxenic conditions in defined media. Sulfur gas analysis of culture fluids indicated that members of the Cyanophyta were probable sources of DMS whereas representatives of the divisions Chlorophyta, Xanthophyta and Bacillariophyta apparently did not produce this compound. Comparison of gaseous contents of young and old nonaxenic cultures of filamentous algae in the divisions Chlorophyta and Xanthophyta showed DMS occurred only in aged cultures and was probably produced by bacteria utilizing substances from senescent algal cells. Data suggest that the composition of the algal community determines whether DMS is algal and/or bacterial in origin.     

POPULATION DYNAMICS OF THE SPORE-FORMING DIATOM LEPTOCYLINDRUS DANICUS IN NARRAGANSETT BAY. RHODE ISLAND1

A field study to assess resting spore importance in long-and short-term population dynamics of Leptocylindrus danicus ( Ehr.) Gran was carried out, based on the observation that germinating spores produce only vegelative cells of maximum diameter. From May through November 1978 . L. danicus was present in lower Narragansett Bay, but resting spores were not found. May and June bioassays suggest that Bay water could not support sexuality (hence spore formation) in L. danicus during this period. Mean valve diameter decreased abruptly in early June. as did cell number. When mean valve diameter increased abruptly in August, with no resting spores seen, 70 additional clonal isolates showed that one clone had the normal sexual and spore-forming life cycle while 69 clones had an alternate life cycle lacking sperm and spore formation. Valvar fine structure of the two types differed with respect to the presence or absence of a subcentral valvar pore, and a new subspecific taxon is here distinguished: variety apora. Examination of the early June data indicates the replacement of one population by another. Populations of L. danicus may be sporadically introduced into Narragansett Bay from Gulf Stream rings .     

RELATIONSHIPS OF SEDIMENTED DIATOM SPECIES (BACILLARIOPHYCEAE) TO ENVIRONMENTAL GRADIENTS IN DILUTE NORTHERN NEW ENGLAND LAKES1

Limnological gradients of small, oligotrophic, and low conductance lakes in northern New England were defined by principal components analysis; relationships of sedimented diatom species to the gradients were investigated by correlation analysis. Diatom distributions were most strongly related to the gradient of pH and alkalinity and the covarying variables, conductance, Mg, Ca, total Al, and exchangeable Al. Weaker relationships to lake morphology, dissolved organic carbon and water color, altitude and marine aerosol inputs, and the distinctive water chemistry of some New Hampshire lakes were also present. Results for 16 taxa of importance in our studies of lake acidity are given in detail and are compared to results from other regions of eastern North America. Planktonic taxa were absent below pH 5.5, with the exception of the long form of Asterionella ralfsii var. americana Korn. The two forms of this taxon differed ecologically: the long form (>45μm) had an abundance weighted mean (AWM) pH 4.90 and occurred mostly in lakes that were deep relative to transparency; the short form (<45μm)had an AWM pH and occurred on lakes that were shallow relative to transparency. The ecological advantage of a “splitter” approach to diatom taxonomy was demonstrated by examination of other taxa as well, including Tabellaria flocculosa (Roth) Kütz. These results have important implications for paleolimnological interpretations.     

PATTERNS OF EPIPELIC ALGAL DISTRIBUTION IN AN ACIDIC ADIRONDACK LAKE1

The biovolume and species composition of epipelic algae along sediment depth gradients were sampled seasonally in an acidic oligotrophic lake in the Adirondack Park in New York State. The epipelic algal community of Woods Lake (Herkimer Co., NY) was dominated by diatoms and cyanobacteria. Distinct depth zonation patterns of community composition were evident. Total algal biovolume increased with depth due to a dense cyanobacterial mat on the sediments in deeper water (5–8 m). This mat was dominated by a single species of cyanobacteria, Hapalosiphon pumilus (Kütz.) Kirchner, which accounted for the late summer maximum in total biovolume at 7 m. The shallower (1–4 m) epipelic communities were dominated by diatoms, which showed a spring maximum in total biovolume and were dominated by Fragilaria acidobiontica Charles, Navicula tenuicephala Hust. and N. subtilissima Cl.     

REMOVAL OF ORGANIC AND INORGANIC MATTER FROM ANTARCTIC LAKES BY AERIAL ESCAPE OF BLUEGREEN ALGAL MATS1

Field studies on three perennially ice-covered Antarctic oasis lakes with little or no outflow disclosed a unique biological phenomenon. Benthic algal mats dominated by the prokaryotic Phormidium frigidum Fritsch and several pennate diatoms growing in shallower, more brightly illuminated areas beneath 4 to 5.5 m of ice accumulate and entrap bubbles of photosynthetically produced oxygen. Clumps of this gas-filled mat tear loose from the gravelly substrate, lift off and float to the bottom of the lake ice. Some of these floating mat pieces become frozen into newly forming ice with the onset of winter. Through the combination of ablation of ice from the upper surface and the formation of new ice from below, algal mat pieces reach the upper lake surface in 5–10 years. Here, they are lyophilized by polar winds and dispersed in at least a partially viable state. The process of mat lift-off and escape is important in removing nutrients and salts from these lakes and helps to perpetuate their oligotrophic state. Neutron activation and X-ray dispersive analyses of elements in the algal mats along with other analyses, field observations, and calculations suggest that significant quantities of organic matter, select minerals and salts are lost from the lakes annually through this process whose magnitude has not been recognized previously.     

PROTOCOLS FOR THE REMOVAL OF BACTERIA FROM FRESHWATER BENTHIC DIATOM CULTURES1

In this study, we describe different combinations of physical separation and antibiotic treatment to remove associated bacteria from freshwater diatoms. Diatoms were purified either from natural epilithic biofilms or from unialgal cultures. We determined that for most strains, different purification procedures have to be combined individually. In a new approach, we show that for some diatom strains, the substitution of associated aquatic bacteria by an antibiotic‐sensitive Escherichia coli strain and subsequent treatment with antibiotics may be a successful strategy to obtain axenic diatom cultures. Axenic diatom cultures are essential to study the physiology and biochemistry of individual strains as well as their responses to environmental changes without interference of accompanying bacteria.