Superoxide production by marine microalgae

A survey was conducted for production of the reactive oxygen species superoxide by 37 species (65 strains) of microalgae including dinoflagellates, raphidophytes, chlorophytes, prasinophytes, eustigmatophytes and prymnesiophytes. Ichthyotoxic raphidophyte species of Chattonella were found to produce the highest environmental levels of superoxide (177×10⁴ total chemiluminescence units). However, ichthyotoxic dinoflagellates (Karenia, Alexandrium) and the prymnesiophyte Prymnesium were also found to produce significant levels of superoxide (4×10⁴, 3×10⁴ and 5×10⁴ chemiluminescence units, respectively), equivalent to that of other raphidophyte species of Heterosigma and Fibrocapsa (6×10⁴ and 2×10⁴, respectively). A direct relationship between cell size and superoxide production was observed (r²=0.94), with larger cells producing more superoxide per cell. Chattonella produced the most superoxide per cell (expressed as cellular chemiluminescence units), followed by the dinoflagellate species Karenia, Alexandrium, Takayama and Gymnodinium. Small cells, such as the raphidophyte Heterosigma and the prymnesiophyte Prymnesium produced very little superoxide per cell (cellular chemiluminescence units), but potentially could still produce high total levels of superoxide if present at high biomass levels. Species commonly used as aquaculture bivalve feeds such as Dunaliella, Tetraselmis, Nannochloropsis and Pavlova produced negligible levels of superoxide, even at high biomass. We speculate that superoxide, while not the sole ichthyotoxic principle, may play a wider role in algal toxicity than previously considered, and propose a broad classification of microalgae based upon superoxide production.Communicated by M.S. Johnson, Crawley     

Superoxide production by marine microalgae

This study investigated the possible roles of superoxide produced by raphidophyte and prymnesiophyte microalgae as an ichthyotoxic agent to damselfish and an allelopathic agent to bacteria. We found that the rate of superoxide production varied with algal cell density, with cell densities of the raphidophyte Chattonella marina >10,000 cells ml^–1 producing less environmental levels of superoxide per cell (94±14 chemiluminescence units) than cell densities <10,000 cells=">^–1 (390±54 units per cell). Microalgal cells have the capacity to change their superoxide production rate over a period of 1 h, dependent on cell density and metabolic activity. We also examined the effect of superoxide on suppression of bioluminescence of the marine bacterium Vibrio fischeri as a model for bacterial alleopathy and found that both superoxide and free fatty acids such as eicosapentaenoic acid (EPA; 20:53) present in raphidophyte microalgal cells cause suppression of bacterial bioluminescence. The combination of superoxide in the presence of EPA further enhanced bioluminescence suppression. Superoxide was also found to enhance the toxicity of free fatty acid EPA to damselfish (Acanthochromis polycanthus) at concentrations as low as 0.2 mg l^–1. In conclusion, consideration should be given to density dependent and/or metabolic variations of toxicity when publishing minimum alert levels for superoxide producing ichthyotoxic microalgal species. A secondary role of superoxide production may be to enhance the toxicity of algal exudates or serve as an allelopathic agent against bacterial fouling.     

Plasmid mediated antibiotic resistance in marine bacteria

This research work was conducted in Uppanar estuary to ascertain the role of plasmids in the antibiotic resistance of bacteria. Water and sediment samples were collected for a period of three months. When tested against 20 antibiotics 22 MAR strains were isolated from the samples, which were found resistant to 5-13 antibiotics. They belong to 7 genera and 10 species. Gram-negative bacteria namely Neisseria mucosa, N. sicca, Branhamella catarrhalis, Klebsiella ozaenae, Citrobacterintermedius, Pseudomonas fluorescens and Enterobacter aerogenes were isolated. Gram-positive bacteria were of Bacillus subtilis, B. megaterium and Micrococcus luteus. When plasmid curing was done using acredine orange, the resistance against penicillin-G, ampicillin, tetracycline, amoxycillin, kanamycin, and chloramphenicol were totally lost in all strains, which confirmed the role of plasmid in these strains against antibiotics. Ten strains belong to different species were selected for the plasmid isolation and electrophoresis was done. Presence of plasmids in all strains was confirmed and the molecular weight was in the range of 2850 to 3170 bp. The study revealed that MAR strains are common in Uppanar estuary and they are plasmid mediated. This environment is seemed to be deteriorating at an alarming rate.     

Examination of hydroxamate-siderophore production by neritic eukaryotic marine phytoplankton

Species of neritic eukaryotic marine phytoplankton were investigated during 1982 for hydroxamate-type siderophore production under iron-sufficient and iron-deficient culture conditions. Three of the 5 Prorocentrum species examined produced siderophores. Prorocentrin, the extracellular hydroxamate-type siderophore isolated from P. minimum, was also produced by P. mariae-lebouriae and P. gracile. P. maximum and P. micans grew poorly in iron-deficient medium and did not produce intracellular or extracellular hydroxamate-type siderophores. Thalassiosira pseudonana and Dunaliella tertiolecta produced extracellular siderophores under iron-deficient conditions, but siderophore production was not detected in the other two species, Skeletonema costatum and Olisthodiscus luteus. Each species which produced extracellular Csaky-positive hydroxamate showed a similar pattern of production. Under iron-sufficient conditions there was no measurable siderophore found either intracellularly or extracellularly. Under iron-deficient culture conditions hydroxamate-type siderophore was produced 1 to 2 d after the cessation of growth in the stationary phase. Production was over a short period of time (1 to 2 d) and the siderophore did not remain in the medium. The rate of siderophore disappearance from the medium was similar to the rate of production. Each species which produced siderophores showed an increase in in vivo fluorescence coincidental with the disappearance of the extracellular siderophore from the culture medium. There was no corresponding increase in in vivo fluorescence in iron-sufficient cultures. It is suggested that in vivo fluorescence may be used as a screening procedure for determining hydroxamate-type siderophore production in eukaryotic phytoplankton. An hypothesis on the iron uptake mechanism is proposed.     

A flow-through system for field measurements of production by marine macroalgae

A continuous flow-through chamber for the in situ measurement of primary production in macrophytic algae (as exemplified by Ulva lactuca) is described. Rapid mixing occurred within the chamber independent of flow rate or amount of algal tissue, so that the complete-mix reactor model of sanitary engineering was closely approximated. Field experiments with the apparatus revealed a rapid response to short-term variations in production rate as well as a close correlation between production and changing light levels. The flowthrough apparatus was compared to a closed system (the traditional bell jar approach); when the water in the latter was mixed, the two methods gave similar results for incubations not longer than several hours. However, during longer incubations, the productivity in the static system was sometimes depressed, possibly due to nutrient depletion or to abnormally high levels of oxygen. Thus, the flow-through system permits reliable measurements of macroalgal production for periods of 12 h or longer.     

Performance of an ANAMMOX reactor treating wastewater generated by antibiotic and starch production processes

A pilot-scale anaerobic ammonia oxidation (ANAMMOX) reactor was used to treat mixed wastewater resulting from a chlortetracycline and starch production process. The results, collected over the course of 272 days, show that the ratio of influent ammonium to nitrite, pH, and temperature can all affect the efficiency of nitrogen removal. The ratio of influent ammonium to nitrite was maintained at about 1:1 at a concentration below 200 mg·L^-1 for both influent ammonium and nitrite. The total nitrogen (TN) loading rate was 0.15–0.30 kgN·m^-3·d^-1, pH remained at 7.8–8.5, and temperature was recorded at 33±1°C. The rate of removal of ammonia, nitrite, and TN were over 90%, 90%, and 80%, and the effluent ammonium, nitrite and TN concentrations were below 50, 30, and 100 mg·L^-1.     

Extent and nature of extracellular organic production by the marine coccolithophorid Hymenomonas carterae

Production of extracellular dissolved organic substances by cultures of the marine coccolithophorid Hymenomonas carterae SMBA 254 was investigated by the ¹⁴C-tracer technique. In 4 h incubations, extracellular ¹⁴C production represented 20 to 64% of ¹⁴C incorporation into cell material for cultures nearing, or during stationary growth; for rapidly growing cultures with relatively low cell densities the ¹⁴C-production was only 4 to 10% of incorporation. Intra-and extra-cellular material was fractionated by ion-exchange membrane electrodialysis. Ionic compounds accounted for 40 to 60% of extracellular material; in contrast only 16 to 27% of intracelular metabolites were in this category. Of a range of methods which were investigated, separation of compounds by gel-filtration in conjunction with two-dimensional thin-layer chromatography proved most effective. Up to 80% of extracellular material was of less than 1 800 mol. wt and included carbohydrates and amino acids, with glutamic acid predominating. The intracellular material was more heterogeneous, with a substantially greater proportion of high molecular weight material. The quite distinct compositions show clearly that extracellular production observed in these experiments arose through excretion and was not attributable to cell lysis.     

Tube-building by a marine meiobenthic harpacticoid copepod

Pseudostenhelia wellsi Coull and Fleeger is a meiobenthic harpacticoid copepod inhabiting muddy, estuarine sediments. All individuals observed, and all ages including nauplii, build and inhabit elongate, mucous tubes which may extend to a depth of 3.9 mm into the sediment. Many of the narrow tubes, 0.27 mm in diameter, have a tube-cap extending 0.32 mm above the sediment-water interface. The tubes are a matrix of fine silt, sand and detritus cemented with a mucopolysaccharide, as shown by the Periodic Acid Schiff stain, apparently secreted from glands in the ventrolateral margin of the cephalothorax. Upon addition to sterile, homogeneous sediment, P. wellsi quickly (within 1 to 2 h) transform the upper 0.4 cm to a cohesive conglomerate of tubes, silt-clay particles and mucous filaments.     

Measurement of photosynthetic production of a marine phytoplankton population using a stable 13C isotope

The use of stable isotope of carbon, ¹³C, for the determination of the photosynthetic rate of a marine phytoplankton population was examined. Particular concern was paid to the effects of non-phytoplanktonic organic carbon and the enrichment of inorganic carbon on the estimation of the photosynthetic rate. Photosynthetic rates determined by the ¹³C method showed a remarkable agreement with those determined by the ¹⁴C method. Insitu determinations of photosynthetic rate were made in three different water types: open ocean, coastal and neritic waters, which included oligo- and mesotrophic waters, by using the ¹³C method established.     

Influence of bacteria on growth and hemolysin production by the marine dinoflagellate Amphidinium carterae

The effect of various bacteria on the growth of and hemolysin production by Amphidinium carterae was studied. The algal culture had an indigenous bacterial flora of Moraxella and Pseudomonas and these could not be eliminated by treatment with bactericidal antibiotics like gentamicin. Various bacteria like Micrococcus, Aeromonas, Vibrio, and Moraxella-like bacteria were added to A. carterae cultures to study their effect on growth and hemolysin production. Micrococcus and Aeromonas were found to improve the growth marginally. Hemolysin titres were considerably higher in A. carterae cultures supplemented with bacteria. Received: 27 May 1997 / Accepted: 22 July 1997     

Distinguishing modes of dissolved organic carbon production in marine macrophytes by tracer kinetic analysis

Tracer kinetic analysis of radioisotope incorporation into dissolved organic compounds reveals two distinct patterns of photosynthate release by macroalgae. In experiments employing Sargassum lacerifolium, dissolved organic carbon was produced at a constant rate during light incubations. Steady state rates of production were never achieved in experiments employing either Ecklonia radiata (Turn.) J. Agardh. or Ulva lactuca L. Analysis of the time-varying radioactivity curves obtained in experiments using these algae always resulted in models consistent with dissolved organic carbon production being an autocatalytic process. Preincubation of U. lactuca in the dark resulted in a diminished (ca. 40%) rate of dissolved organic carbon production during the subsequent light incubations. In no case did the radioisotope content of the dissolved organic carbon approach a limiting value, indicating that in contrast to phytoplankton, uptake rates of photosynthate by macroalgae are always less than the rates of production.     

Economic valuation of marsh areas for marine production processes

The relationship of natural marsh-estuarine systems to the economic productivity of marine systems is not well understood, at least in any quantitative sense. An approach is developed for relating blue crab economic productivity on Florida's Gulf Coast to marsh availability in the area. Previous efforts have not always applied economic concepts appropriately in attempts at such quantification. The marginal value productivity of marsh is shown to vary with alternative levels of marsh and effort in the fishery. The interaction and subsequent interdependence is shown to be statistically significant. Data availability on marginal response to marsh changes poses a severe obstacle to further progress.     

Nephtys hombergii, a free-living predator in marine sediments: energy production under environmental stress

Nephtys hombergii is a free-living, burrowing predator in marine sediments. The worm is, therefore, exposed to various environmental conditions which tube-dwelling polychaetes of the same habitat most likely do not encounter. The worms have to survive periods of severe hypoxia and sulphide exposure, while at the same time, they have to maintain agility in order to feed on other invertebrates. N. hombergii is adapted to these conditions by utilising several strategies. The species has a remarkably high content of phosphagen (phosphoglycocyamine), which is the primary energy source during periods of environmental stress. With increasing hypoxia, energy is also provided via anaerobic glycolysis (pO₂<7 kPa), with strombine as the main end-product. Energy production via the succinate pathway becomes important only under severe hypoxia (<2 kPa), suggesting a biphasic response to low oxygen conditions which probably is related to the worm's mode of life. The presence of sulphide resulted in a higher anaerobic energy flux and a more pronounced energy production via glycolysis than in anoxia alone. Nevertheless, after sulphide exposure under anaerobic conditions of <24 h, N. hombergii is able to recover completely. Although N. hombergii appears to be well adapted to a habitat with short-term fluctuations in oxygen and appearance of hydrogen sulphide, its high energy demand as a predator renders it likely to limit its survival in an environment with longer lasting anoxia and concomitant sulphide exposure. Received: 28 May 1997 / Accepted: 21 June 1997     

Enhanced primary production and nutrient regeneration within aggregated marine diatoms

Blooms of chain-forming diatoms often terminate with the mass flocculation and subsequent settlement of cells from the nutrient-depleted euphotic zone. While mass diatom aggregation has been suggested as an adaptive mechanism for placing resting spores in the deep sea, we hypothesized that aggregation may confer an immediate adaptive advantage to the associated diatoms as well. We tested this hypothesis by comparing the photosynthetic activity, pigment composition and nutrient-uptake rates of aggregated and suspended diatoms over time. Diatom aggregates were collected by SCUBA divers in the Santa Barbara Channel (34°23N; 119°50W) on 4 March 1987 and monitored for 9 d in the laboratory. Diatom aggregates sustained chlorophyll a-specific primary production rates two to nine times higher than those of freely suspended diatoms from the surrounding seawater. The timing of maximum productivity was strongly correlated with the appearance of remineralized ammonia within the aggregates. Chlorophyll a-specific nitrate-uptake rates were routinely three to nine times lower in diatom aggregates than in the surrounding seawater. Primary production and pigment concentrations of diatom aggregates aged in situ displayed changes similar to those observed in the laboratory. These results suggest that diatoms associated with aggregates maintain higher photosynthetic rates than freely suspended diatoms by efficiently exploiting remineralized ammonia within the aggregate microenvironment, in preference to external nitrate sources. The enhanced nutrient environment within aggregates may be important for understanding the adaptive significance of the mass flocculation of diatom blooms.     

Products of denitrification by a marine bacterium as revealed by gas chromatography

Gas chromatography was used to analyse the gases released by growing cultures of the denitrifying marine bacterium Pseudomonas perjectomarinus. When nitrate was provided as final oxidant, CO₂ was the first detectable gas released. Following the peak rate of release of CO₂ at approximately 16 h, N₂ liheration began and peaked at 30 h. When N₂O was provided as terminal oxidant, CO₂ and N₂ release began within a few hours and the rate of liberation of both increased for 45 to 50 h before leveling. Cell-free extracts of bacteria from denitrifying cultures of P. perfectomarinus were incubated anaerobically with nitrate, malic acid and electron transfer cofactors provided, and the gases in the atmosphere above the reaction mixtures were analysed. NO was found to be present transiently, and N₂O was discerned after prolonged incubation. The applicability of gas chromatography to analyses of marine ecosystems and the need for better means of sampling the atmosphere above enzymatic reaction mixtures were disoussed.     

Effects of hydraulic retention time on net present value and performance in a membrane bioreactor treating antibiotic production wastewater

• The membrane bioreactor cost decreased by 38.2% by decreasing HRT from 72 h to 36 h. • Capital and operation costs contributed 62.1% and 37.9% to decreased costs. • The membrane bioreactor is 32.6% cheaper than the oxidation ditch for treatment. • The effluent COD also improved from 709.93±62.75 mg/L to 280±17.32 mg/L. • Further treatment also benefited from lower pretreatment investment. A cost sensitivity analysis was performed for an industrial membrane bioreactor to quantify the effects of hydraulic retention times and related operational parameters on cost. Different hydraulic retention times (72–24 h) were subjected to a flat-sheet membrane bioreactor updated from an existing 72 h oxidation ditch treating antibiotic production wastewater. Field experimental data from the membrane bioreactor, both full-scale (500 m³/d) and pilot (1.0 m³/d), were used to calculate the net present value (NPV), incorporating both capital expenditure (CAPEX) and operating expenditure. The results showed that the tank cost was estimated above membrane cost in the membrane bioreactor. The decreased hydraulic retention time from 72 to 36 h reduced the NPV by 38.2%, where capital expenditure contributed 24.2% more than operational expenditure. Tank construction cost was decisive in determining the net present value contributed 62.1% to the capital expenditure. The membrane bioreactor has the advantage of a longer lifespan flat-sheet membrane, while flux decline was tolerable. The antibiotics decreased to 1.87±0.33 mg/L in the MBR effluent. The upgrade to the membrane bioreactor also benefited further treatments by 10.1%–44.7% lower direct investment.     

Catabolism of trimethylamine by a marine bacterium,Pseudomonas NCMB 1154

Macerated suspensions of Pseudomonas NCMB 1154, grown on trimethylamine as sole nitrogen source, cause the stepwise demethylation of trimethylamine, with the production of ammonia and the occurrence of dimethylamine and methylamine as transient intermediates. The initial demethylation involves a glucose activated, non-oxidative rupture of the methyl-amino linkage, is relatively specific for methyl groups, and is partially inhibited by ATP. The first methyl group is oventually oxidised, but only by whole cells, after removal from the amine molecule. The second and third demethylations involve oxidative rupture of the methyl-amino linkages, with the production of formaldehyde, are less specific with respect to the alkyl groups, are glucose activated, but are not inhibited by ATP. The third and final demethylation reaction is partially inhibited by semi-carbazide.     

Changes in pH as a measure of photosynthesis by marine macroalgae

An automatically operated method for high precision measurements of steady-state photosynthesis by macroalgae was developed. Changes in pH and oxygen content of seawater passing the algae in a flowthrough system, could be measured with extremely high accuracy over very long periods of time. The method is especially suitable for measurements on flowthrough systems with high rates of water exchanges (i.e. short retention time), and can be used to study exchange processes for marine plants, animals and small ecosystems. Since the same measuring unit is used for several flowthrough chambers, the method is very suitable for comparisons between different species, or between differently pretreated specimens of the same species (e.g. in toxicological studies). The method was used to study the ratio: [oxygen production] to [CO₂+H⁺ uptake] at different light intensities for several macroalgae belonging to different systematic groups and from different habitats. At lower photosynthetic rates this ratio was similar for all of the algae studied (1.17±0.02). For brown algae of the fucacean family, the ratio increased by 0.08 units at higher photosynthetic rates. This increase was thought to be related to the crassulacean acid metabolism (CAM)-like strategies connected to these algae. For all other algae studied, the ratio remained constant or decreased slightly (at most by 0.04 units) at higher photosynthetic rates. The relations between the abovementioned ratio and the photosynthetic quotient are discussed on a theoretical basis.     

Thymidine incorporation as a measure of heterotrophic bacterioplankton production in marine surface waters: Evaluation and field results

To assess bacterioplankton production in the sea, we have developed a procedure for measuring growth based on incorporation of tritiated thymidine into DNA; the accuracy of this procedure was tested under a variety of laboratory and field conditions. By autoradiography, we have found that for all practical purposes our technique is specific for the nonphotosynthetic bacteria and that virtually all of the active bacteria (one-third or more of the total countable bacteria) take up thymidine. We also measured (1) the intracellular isotope dilution of thymidine assessed by parallel experiments with labeled phosphorus, and (2) DNA content of natural marine bacteria (0.2 to 0.6 m size fraction); a conversion factor derived from these data permitted estimation of production from thymidine incorporation results. A very similar conversion factor was independently derived from the empirical relationship between thymidine incorporation and growth of natural bacterioplankton under controlled conditions. Combined results show that this technique, which can be performed rapidly and easily at sea, provides good estimates of production. Data from Southern California Bight waters, which contain oligotrophic as well as moderately eutrophic regions, show that average bacterioplankton doubling times, like those of the phytoplankton, are on the order of a few days, with fastest growth at depths just below those of greatest phytoplankton abundance. Offshore bacterial production is roughly 5 to 25% of the primary production; thus, at a 50% assimilation efficiency, the bacterioplankton would consume 10 to 50% of the total fixed carbon.