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.     

Shell growth response of mussels (Mytilus edulis) exposed to toxic microalgae

Mussels (Mytilus edulis) were exposed to the algaeAlexandrium ostenfeldii, Chrysochromulina polylepis, Gyrodinium aureolum, Gymnodinium galatheanum andHeterosigma akashiwo for 24 h; significant reductions in growth rate, as compared to the control, were observed after exposure toA. ostenfeldii, C. polylepis, G. aureolum andG. galatheanum at initial concentrations of 4.5 × 10⁶, 110 × 10⁶, 9 × 10⁶ and 120 × 10⁶ cells l^–1, respectively. Exposure to high initial concentrations of the non-toxic algaeTetraselmis suecica (174 × 10⁶ cells l^–1) andIsochrysis galbana (610 × 10⁶ cells l^–1) showed no adverse effect on growth rate. When mussels with reduced growth were transferred to clean seawater, they recovered to > 90% of control growth within 2 to 4 d. Exposure to algal filtrates of the toxic algal cultures produced no reduction in growth rate.     

Nuclear ITS region of the alga Heterosigma akashiwo (Chromophyta: Raphidophyceae) is identical in isolates from Atlantic and Pacific basins

The internal transcribed spacer (ITS) region from 19 isolates of the algal genus Heterosigma (Chromophyta: Raphidophyceae) was amplified by polymerase chain-reaction (PCR) and sequenced. Isolates were obtained from both the Atlantic and Pacific basins, including Europe, eastern North America, western North America, Japan and New Zealand. This study presents evidence that all Heterosigma isolates in this study are representatives of one species (H. akashiwo). All 19 isolates, except one (LB 2005) had identical ITS sequence (98.31% similar by pairwise comparison); Isolate LB 2005 may represent a separate subspecies. Such high degree of ITS sequence identity implies that the organism has spread between oceanic regions in geologically recent times, possibly by human means. In addition to those from Heterosigma spp., the ITS regions from other marine Raphidophyceae (Chattonella antiqua, C. marina, C. subsalsa, Fibrocapsa japonica, and Olisthodiscus luteus) were amplified and sequenced using PCR. Total ITS lengths differed among the Raphidophyceae (C. antiqua, 577 base pairs (bp); C. marina, 577 bp;. C. subsalsa, 579 bp; F. japonica, 830 bp; H. akashiwo, 561 and 563 bp; O. luteus, 829 bp), but 5.8S rDNA sequences were similar in size (13 to 142 bp). The high ITS sequence identity between C. antiqua and C. marina (>99.9% by pairwise comparison) suggests the need for a taxonomic review of these species encompassing all morphological, genetic, physiological and biochemical information. Additionally, a number of cultures of Raphidophyceae were positively identified. In general, ITS comparisons among the Raphidophyceae may be most useful at the level of species determination rather than at the population level. Received: 12 July 1999 / Accepted: 16 March 2000     

Growth and herbivory by heterotrophic dinoflagellates in the Southern Ocean,studied by microcosm experiments

Growth and herbivory of heterotrophic dinoflagellates (Gymnodinium sp.) from the Weddell Sea and the Weddell/Scotia Confluence were studied in 1988 in 100-liter microcosms. The microcosms were screened through 200-µm or 20-µm mesh nets and incubated for 12 d at 1 °C under artificial light. Mean cell volume of dinoflagellates was 1 000 to 1 500µm³, and that of their phytoplankton prey 360 to 430µm³. Dinoflagellate growth rate followed a Holling type II functional response, with a maximum growth rate of 0.3 d^–1 and half-saturation food concentrations of 1.0µg chlorophylla l^–1, 50µg C l^–1, or 1 500 cells ml^–1. Carbon budgets based on¹⁴CO₂ assimilation and biomasses of phytoplankton and heterotrophic dinoflagellates suggested a balance between phytoplankton grazing loss and dinoflagellate consumption, assuming a dinoflagellate carbon conversion efficiency of 40%. Applying this to the functional response yielded estimates of maximum ingestion rate (0.8µg Cµg^–1 C d^–1, or 6 pg C dinoflagellate^–1 h^–1) and maximum clearance (0.8 to 1.2 × 10⁵ body volumes h^–1, or 80 to 120 nl ind.^–1 h^–1). The microcosm experiments suggested that heterotrophic dinoflagellates may contribute significantly to maintenance of low phytoplankton biomass in the Southern Ocean.     

Algal chloroplasts in the protoplasm of three species of benthic foraminifera: taxonomic affinity,viability and persistence

The ultrastructure and pigment content of algal chloroplasts (derived from Bacillariophyceae or Chrysophyceae) are described from 3 benthic species of brackish-water foraminiferans.Elphidium williamsoni Haynes contains 4×10⁶ chloroplasts mg^-1, whereas the contents ofNonion germanicum (Ehrenberg) andE. excavatum (Terquem) are about 10% of this value. The two former contain chlorophyllsa andc and fucoxanthin, but these pigments were not detectable in the latter.E. williamsoni andN. germanicum had a net uptake of¹⁴C–HCO ₃ ^- , proportional to their content of chlorophyll and number of chloroplasts, increasing linearly up to approximately 10 Klux. At light saturation the former assimilates 2.3x10^-3 mg C mg^-1 h^-1 and the latter only about 20% of this value. Dark uptake was insignificant in all cases. Uptake could not be demonstrated inE. excavatum. The photosynthesis effected by these species is trivial in terms of the total benthic carbon fixation effected by the microflora. The chloroplasts survived longer in forminiferans kept in the dark than in light/dark adapted individuals. To keep a steady state population of chloroplasts under light/dark conditions,E. williamsoni must eat at least 65 chloroplasts individual^-1 h^-1, whereas the minimum consuption rate inN. germanicum is 20.     

Possible involvement of the glycocalyx in the ichthyotoxicity of Chattonella marina (Raphidophyceae): immunological approach using antiserum against cell surface structures of the flagellate

Although the ichthyotoxic mechanism of Chattonella marina is still unknown, several lines of evidence suggest that the reactive oxygen species (ROS), such as superoxide anion (O₂^-), hydrogen peroxide (H₂O₂)_, and hydroxyl radical (·OH), produced by C. marina are involved in the mortality of fish exposed to this flagellate. Recently, we found that the cell-free supernatant prepared from C. marina, which is considered to contain the glycocalyx, showed NADPH-dependent O₂^- generation. In this study, we prepared antiserum against the crude glycocalyx of C. marina. Using indirect immunofluorescence, it was confirmed that the antiserum specifically reacted with C. marina cells. In addition to C. marina, the antiserum also reacted with other raphidophycean flagellates such as Heterosigma akashiwo, Olisthodiscus luteus, and Fibrocapsa japonica, whereas no reactivity was observed against six other flagellate species tested. These results suggest that raphidophycean flagellates have common epitopes recognized by the antiserum. Interestingly, immunohistochemical analysis of paraformaldehyde-fixed gill lamellae from yellowtail exposed to C. marina revealed that the antiserum stained the surface of gill lamellae, while no such staining pattern was observed in control gill lamellae. These results suggest that the glycocalyx may be discharged when C. marina cells are inhaled into the fishes' mouths and then come into contact with the gill surface. Based on the present results, together with our previous findings, we propose that continuous accumulation of the discharged glycocalyx on the gill surface occurs during C. marina exposure, which may be responsible for the ROS-mediated severe gill tissue damage leading to fish death.     

Genetic variability in Gymnodiniaceae ITS regions: implications for species identification and phylogenetic analysis

The internal transcribed spacer (ITS) regions and 5.8S rRNA genes from several strains of toxic Gymnodiniaceae were sequenced and subjected to phylogenetic analysis with other Gymnodiniaceae species. Sequence comparisons showed that high sequence divergence existed in Gymnodiniaceae, especially in the genus Gymnodinium. The amplicons of the ITS regions from Amphidinium and Gyrodinium species were 438–439 and 604–605 bp, respectively, and those of the Gymnodinium species ranged from 575 to 615 bp. The mean distance value within Gymnodinium, calculated from the ITS sequence, was 0.68827 (range: 0–0.92323), 0.11342 for Amphidinium (range: 0.00467–0.17120) and 0.2005 for Karenia (range: 0.00521–0.29971). Low distance values were found within the species Gyrodinium instriatum (<0.01) and Karlodinium micrum (<0.02). Amphidinium remarkably had a shorter ITS than did other genera in Gymnodiniaceae, this implied that Amphidinium might be distant from the other Gymnodiniaceae and supported Saunders opinion that the taxonomy of Amphidinium needs to be reevaluated. Largely congruent phylogenetic trees were produced by the maximum-parsimony method (PAUP), maximum-likelihood method (PAUP) and Bayesian inference (MrBayes), whereas the three analyses showed that the genera Gymnodinium and Karenia are unresolved groups in phylogeny. Minor sequence divergence was found within the different clones of Amphidinium carterae, suggesting that the ITS regions are suitable as genus- and species-specific oligonucleotide probes to rapidly detect and identify the red tide-forming algal species.Communicated by O. Kinne, Oldendorf/Luhe     

Fluctuations of the red tide flagellates Chattonella spp. (Raphidophyceae) and the algicidal bacterium Cytophaga sp. in the Seto Inland Sea, Japan

A marine algicidal gliding bacterium Cytophaga sp. strain J18/M01 was isolated in 1990 from a station in northern Harima-Nada, the Seto Inland Sea, Japan, using the harmful red tide alga Chattonella antiqua (Hada) Ono as a susceptible organism. The bacterium can prey upon various species of microalgae. Temporal fluctuations of this bacterium and Chattonella spp. [C. antiqua and C. marina (Subrahmanyan) Hara et Chihara] were investigated weekly at the above station in the summer of 1997 and 1998, using immunofluorescence assay employing highly specific polyclonal antibodies for the bacterium. In the summer of 1997, the cell density of Chattonella spp. showed a maximum value (70 cells ml⁻¹) on 8 July, and decreased thereafter. The bacterium Cytophaga sp. J18/M01 was commonly detected around a few hundreds of cells per milliliter or less. The number of Cytophaga sp. J18/M01 increased after the peak of Chattonella spp., and the maximum cell number of the bacterium was 1350 ml⁻¹. This algicidal bacterium also followed the changes of total amounts of microalgal biomass (chlorophyll a+pheophytin) when Chattonella spp. were absent. In the summer of 1998, Chattonella spp. were relatively less abundant (maximum 21 cells ml⁻¹), and the algicidal bacterium Cytophaga sp. J18/M01 showed a close relationship with the change of total microalgal biomass. The present study suggests that the algicidal bacterium Cytophaga sp. J18/M01 preyed upon, not only harmful red tide microalgae, but also other common microalgae such as diatoms, and the bacterium presumably plays an important role in regulating microalgal biomass in natural marine environments. Received: 20 April 2000 / Accepted: 1 December 2000     

Early reproduction and development of dominant calanoid copepods in the sea ice zone of the Barents Sea—need for a change of paradigms?

Reproduction and growth of the dominant copepods Calanus finmarchicus, C. glacialis, C. hyperboreus and Pseudocalanus minutus were studied on transects across the sea ice zone of the northern Barents Sea in May and June 1997. C. glacialis and C. finmarchicus were numerically dominant and also the largest component of the biomass. C. hyperboreus was rather rare. Moderate levels of phytoplankton and eventually high concentrations of ice algae supported maximum egg production rates of 53.6 and 48.5 eggs female^–1 day^–1 of C. glacialis in May and June, respectively. Results of incubation experiments were supported by a tremendous abundance of C. glacialis eggs in the water column ranging from 7×10³ to 4.4×10⁴ m^–2 in May and from 9.8×10³ to a maximum of 9.7×10⁴ m^–2 in June. In contrast, C. finmarchicus spawned only in the vicinity of the ice edge, at a maximum rate of 30 eggs female^–1 day^–1. Egg sacs of P. minutus were often observed in the preserved samples, but contained only few eggs, which may be due to loss during sampling. The presence of considerable concentrations of young stages in May and June indicated successful recruitment of C. glacialis and P. minutus. Back calculation using published stage duration estimates indicates March/April as the begin of the reproductive and growth period for these species under the first-year ice of the Barents Sea. Hence, secondary production in the study area starts at the same time as in open water regions and polynyas in the northern North Atlantic. Although the role of ice algae in the nutrition of copepods was not clarified here, the significant relationship between phytoplankton chlorophyll and egg production of C. glacialis suggests that high reproductive activity has already been achieved at moderate food concentrations.Communicated by O. Kinne, Oldendorf/Luhe     

Anaerobic heat production of bivalves (Polymesoda caroliniana andModiolus demissus) in relation to temperature,body size,and duration of anoxia

Anaerobic heat-production rates of two co-occurring species of estuarine bivalves (a clam and a mussel) were measured with double-twin heat-flow calorimeters, one at 20°C, the other at 30°C. There is no significant difference between the two species in metabolic rates. There is evidence of initial aerobic metabolism in some individuals, as shown by high initial rates exponentially decreasing with time, while others had fluctuating but stable average metabolic activity from the beginning. During aerobic as well as anaerobic metabolism, the bivalves showed rhythmic periods of activity and quiescence. The two species differed in their rhythmic pattern of active and resting metabolism. In the case ofPolymesoda caroliniana, periods of resting metabolism tend to be longer and periods of active metabolism shorter at 30°C than at 20°C. There is a similarity between thermograms ofModiolus demissus at 20° and 30°C. Following acute temperature changes from 5° to 20° and 30°C, the bivalves showed stable metabolic rates in a matter of hours. The stabilized average rates [pooled averages for both species of 1.34×10^-4 (standard error of the mean=0.17×10^-4) W g^-1 dry weight of tissue at 20°C and 2.10×10^-4 (SE=0.20×10^-4) W g^-1 at 30°C] signify a temperature coefficient (Q₁₀) of 1.56 between 20° and 30°C, or partial temperature acclimation. Subtracting heat production as a result of physical activity, i.e., considering only resting metabolism, the corresponding means and standard errors of the means are 1.24×10^-4 and 0.14×10^-4 W g^-1 at 20°C and 1.91×10^-4 and 0.077×10^-4 W g^-1 at 30°C. Anaerobic heat production rate at 20°C is proportional to body size (r=0.84, 9 degrees of freedom, DF). ForM. demissus, measured anaerobic heat production is on the order of 7.5% of the level of aerobic respiration reported in the literature.     

Response of the microbial food web to manipulation of nutrients and grazers in the oligotrophic Gulf of Aqaba and northern Red Sea

The control mechanisms within the pelagic microbial food web of the oligotrophic Gulf of Aqaba and the northern Red Sea were investigated in the spring of 1999. Nutrient conditions and potential grazer impact were manipulated in a series of dilution experiments. Ambient nutrient concentrations and autotrophic biomass were very low (0.23–1.21 µmol NO₃ l^–1, 0.06–0.98 µmol NH₄ l^–1, 1.08–1.17 µmol Si l^–1, 0.08–0.12 µmol P l^–1, 0.15–0.36 µg chlorophyll a l^–1). The planktonic community was characterized by low abundances [3.0–5.5×10⁵ heterotrophic bacteria ml^–1, 0.58–7.2×10³ ultraphytoplankton <8 µm ml^–1 (small eukaryotic photoautotrophs and Prochlorococcus sp., excluding Synechococcus sp.), 0.45–4.4×10⁴ Synechococcus sp. ml^–1, 0.32–1.2×10³ heterotrophic nanoflagellates ml^–1, 1.3–3.8×10³ phytoplankton >8 µm l^–1, 0.93–5.4×10² microzooplankton l^–1] and dominated by small forms (0.2–8 µm). Dinoflagellates and oligotrichous ciliates were the most common groups in initial samples among the phytoplankton >8 µm and microzooplankton, respectively. Results show that bottom-up and top-down control mechanisms operated simultaneously. Small organisms were vulnerable to grazing, with maximum grazing rates of 1.1 day^–1 on heterotrophic bacteria and 1.3 day^–1 on ultraphytoplankton. In contrast, algae >8 µm showed stronger signs of nutrient limitation, especially when the final assemblages were dominated by diatoms. Synechococcus sp. were not grazed and only showed moderate to no response to nutrient additions. The high spatial and temporal variation of our results indicates that the composition of the planktonic community determines the prevailing control mechanisms. It further implies that, at this transitional time of the year (onset of summer stratification), the populations fluctuate about an equilibrium between growth and grazing.Communicated by O. Kinne, Oldendorf/Luhe     

Deleterious effects of a nonPST bioactive compound(s) from <Emphasis Type="Italic">Alexandrium tamarense</Emphasis> on bivalve hemocytes

The known negative effects of shellfish toxin-producing dinoflagellates on feeding, burrowing and survival of some bivalve mollusks has prompted questions concerning whether they might also impair the internal defense system of affected bivalves and make them more susceptible to disease agents. The primary components of the cellular defense system are hemocytes. Many toxic dinoflagellates are too large to be ingested whole by hemocytes and would most likely be exposed to intracellular toxins only after the algae are consumed, broken down, and the water-soluble toxins, released. Therefore, we conducted a series of experiments in which hemocytes from two suspension-feeding bivalves—the Manila clam, Ruditapes philippinarum, and the softshell clam, Mya arenaria—were exposed in vitro to filtered extracts of one highly toxic paralytic shellfish toxin (PST)-producing and one nonPST-producing strain of Alexandrium tamarense (isolates PR18b, 76 ± 6 STXeq cell⁻¹ and CCMP115, with undetectable PST, respectively). We measured adherence and phagocytosis, two hemocyte attributes known to be inhibited by bacterial pathogens and other stressors. We found no measurable effect of a cell-free extract from a highly concentrated suspension of the PST-producing strain on hemocytes of either bivalve species. Instead, extract from the nonPST-producing strain had a consistent negative effect on both clams, resulting in significantly lower adherence and phagocytosis compared to strain PR18b and filtered seawater controls. The bioactive compound produced by strain CCMP115, which has yet to be characterized, may be similar to the PST-independent allelopathic compounds described for Alexandrium spp., which act on other plankters. These compounds and those produced by other harmful algae are known to cause immobilization, cellular deformation and lysis of co-occurring target organisms. Thus, nonPST producing Alexandrium spp., which do not cause paralysis and burrowing incapacitation of clams, may still produce a compound(s) that has negative effects not only on hemocytes, but on other molluscan cell types and their functions, as well.     

Naturally occurring radionuclides in drinking water: An exercise in risk benefit analysis

The scientific background information describing the occurrence, measurement, health effects, treatment technology, risk assessment and economic consequences of the presence of naturally occurring radionuclides in drinking water are described for 60,000 public drinking water supplies. The relevant data for the occurrence of radium, uranium and radon in drinking water supplies are discussed and analysed. Radon is of importance because it is released in the process of taking showers and baths and in washing dishes and clothes. Its progeny is then inhaled, leading to the risk of lung cancer. Radium and uranium can both cause bone cancer. The range of average occurrence of natural radioactivity in drinking water is as follows:²²⁶Ra, 0.3 to 0.8 pCi L^–1;²²⁸Ra, 0.4 to 1.0 pCi L^–1; uranium, 0.3 to 2.0 pCi L^–1 and²²²Rn, 500 to 600 pCi L^–1. The estimated lifetime risks due to the mean groundwater concentrations of naturally occurring radionuclides are:²²⁶Ra and^228Ra, 1.0 10^–5; uranium, 2.0 × 10^–6 and radon, 4.0 × 10^–4. The cost to reduce total radium levels to 5.0 pCi L^–1 is about $9 million. An equivalent expenditure would be required to reduce radon levels to about 4,000 pCi L^–1, or uranium levels to about 100 pCi L^–1. The problem of maximizing the total mortality and the reduction per unit dollar outlay per unit dollar cost for the uranium/radon case is examined.The thoughts and ideas expressed in this paper are those of the authors and are not necessarily those of the US Environmental Protection Agency.This paper is published as a contribution to discussion on this problem and not as a paper providing new research data.     

Planktonic bioluminescence in the pack ice and the marginal ice zone of the Beaufort Sea

An icebreaker cruise into the Beaufort Sea in the fall of 1986 provided a unique opportunity for studying planktonic bioluminescence in ice fields and in the marginal ice zone. Bathyphotometer casts (bioluminescence intensity, seawater temperature, beam attenuation coefficient, and salinity) and biological collections were made to a depth of 100 m. A light budget, which describes the planktonic species responsible for the measured bioluminescence, and a dinoflagellate species budget were constructed from the mean light output from luminescent plankton and plankton counts. The vertical distribution of bioluminescence among the ice stations was similar. The maximum intensities were 2 to 8×10⁶ photons s^-1 cm^-3 in the upper 50 m of the sea-ice interface. The marginal ice zone station (MIZ) exhibited a maximum intensity of 2 to 3×10⁸ photons s^-1 cm^-3 between 5 and 30 m depth. At Ice Station 2, Metridia longa and their nauplii contributed approximately 80% of stimulable bioluminescence in the upper 10 m but, overall, Protoperidinium spp. dinoflagellates contributed most of the light to a depth of 100 m. In the MIZ, Protoperidinium spp. dinoflagellates contributed 90% of the light within the upper 10 m, decreasing to 43% of the contributed light at a depth of 40 m. Below 40 m, dinoflagellate bioluminescence decreased to a few percent of the total to a depth of 90 m. Metridia spp. copepods contributed more than 50% of the light at depths from 40 to 90 m. Ostracods, larvaceans, and euphausiid furcilia contributed <1% of all bioluminescence at all depths sampled. Correlation analyses between measured bioluminescence (photons s^-1 cm^-3), the number of bioluminescent dinoflagellates and the light budget for the MIZ indicated highly significant associations: r=0.919, p=0.001, and r=0.912, p<0.001, respectively (Student's two-tailed t-tests). Bioluminescence was negatively correlated with seawater salinity at all stations (p=0.001). Maximum bioluminescence was measured in the less saline surface waters at all stations.     

Lectin-induced enhancement of superoxide anion production by red tide phytoplankton

Chattonella marina, a raphidophycean flagellate, is a highly toxic red tide phytoplankton which causes severe damage to fish farming. Recent studies demonstrated that Chattonella spp. continuously release superoxide anions (O₂ ⁻) while they are living. Heterosigma akashiwo, another raphidophycean flagellate, also produces O₂ ⁻. In the present study, we found that lectins such as concanavalin A (Con A), wheat germ agglutinin (WGA), and castor bean hemagglutinin (CBH) stimulated  C. marina and H. akashiwo to generate enhanced amounts of O₂ ⁻ in a concentration-dependent manner. The lectin-specific sugars potently inhibited the lectin-induced increase of O₂ ⁻ production, suggesting that the effects of lectins are mediated mainly through the interaction of these lectins with carbohydrate moiety present on the flagellate cell surface. In contrast to the potent ability of native Con A (tetravalent), succinylated Con A (divalent) showed only a slight stimulative effect on these flagellates. O₂ ⁻ production was totally inhibited by treatment with proteinase K for 30 min, without affecting the viabilities of flagellates. These results suggest that cell-surface redox enzymes may be involved in O₂ ⁻ production, and such enzymes are responsible for the lectin-stimulation. Received: 21 August 1997 / Accepted: 8 January 1998     

Effects of the dinoflagellates Karlodinium veneficum and Prorocentrum minimum on early life history stages of the eastern oyster (Crassostrea virginica)

The bloom-forming dinoflagellates Prorocentrum minimum and Karlodinium veneficum can have detrimental effects on some marine life, including shellfish, but little is known about their effects on early life history stages of bivalves. In the Chesapeake Bay region, blooms of these dinoflagellates overlap with the spawning season of the eastern oyster, Crassostrea virginica. In laboratory experiments, we compared the effects of P. minimum and K. veneficum on the survival and development of embryos and larvae of the eastern oyster. At 10⁴ cells ml⁻¹, P. minimum did not have a negative effect on embryos and larvae in 2-day exposures. The yield of D-hinge larvae was equal to or greater than in control treatments. At 2 × 10⁴ cells ml⁻¹ (approximately equal biomass to the P. minimum treatment) K. veneficum caused significant mortality to oyster embryos within 1 day and almost no embryos developed into D-hinge larvae. This effect was not alleviated by the provision of an alternate food source (Isochrysis sp.). Significant mortality was observed when larvae were exposed to K. veneficum at concentrations of 10⁴ cells ml⁻¹ (approximately 5 ng ml⁻¹ of karlotoxin). The K. veneficum cultures used in these experiments were relatively low in toxin content, more toxic strains could be expected to cause mortality at lower cell concentrations. Survival and maturation of embryos and larvae may be reduced when spawns of the eastern oyster coincide with high bloom densities of K. veneficum.     

Abundances,growth rates,and production of tropical neritic copepods off Kingston,Jamaica

Weekly samples were collected near Kingston, Jamaica in 27 m vertical hauls, using 200 and 64µm mesh plankton nets, from July 1985 to January 1987. Thirtytwo copepod species were identified; nauplii and all copepodite stages were enumerated. Total copepod abundance ranged from 2.56 to 87.3 × 10⁴ m^–2. The annual abundance cycle was bimodal with peaks in October–November and May–June corresponding to the rainy seasons. Mean annual copepodite biomass was 0.15 g AFDW m^–2 ranging from 0.03 to 0.41 g AFDW m^–2. Mean generation time (from egg to adult) at 28°C was 19.5 d for the common speciesCentropages velificatus, Paracalanus aculeatus, andTemora turbinata. Isochronal development was demonstrated for copepodites ofP. aculeatus andT. turbinata, but not forC. velificatus. Mean daily specific growth rates (G) were 0.63, 0.63, and 0.48 d^–1 forC. velificatus, P. aculeatus, andT. turbinata, respectively. In general, daily specific growth rates decreased in the later copepodite stages. Thus, it is postulated that growth of later stages and egg production may be food limited. Annual copepodite production was estimated as 419 kJ m^–2 yr^–1, while annual exuvial production and naupliar production were 35 and 50 kJ m^–2 yr^–1, respectively. Egg production was estimated as 44% (184 kJ m^–2 yr^–1) of the total copepodite production. Thus, mean total annual copepod production was 688 kJ m^–2 yr^–1. This estimate is within the range of copepod production estimates in coastal temperate regions.     

Food limitation stimulates metamorphosis of competent larvae and alters postmetamorphic growth rate in the marine prosobranch gastropodCrepidula fornicata

The effects of food limitation on growth rates and survival of marine invertebrate larvae have been studied for many years. Far less is known about how food limitation during the larval stage influences length of larval life or postmetamorphic performance. This paper documents the effects of food limitation during larval development (1) on how long the larvae ofCrepidula fornicata (L.) can delay metamorphosis in the laboratory after they have become competent to metamorphose and (2) on postmetamorphic growth rate. To assess the magnitude of nutritional stress imposed by different food concentrations, we measured growth rates (as changes in shell length and ash-free dry weight) for larvae reared in either 0.45-m filtered seawater or at phytoplankton concentrations (Isoehrysis galbana, clone T-ISO) of 1 × l0³, 1 × 10⁴, or 1.8 × 10⁵ cells ml^–1. Larvae increased both shell length and biomass at 1 × 10⁴ cells ml^–1, although significantly more slowly than at the highest food concentration. Larvae did not significantly increase (p > 0.10) mean shell length in filtered seawater or at a phytoplankton concentration of only 1 × 10³ cells ml^–1, and in fact lost weight under these conditions. To assess the influence of food limitation on the ability of competent individuals to postpone metamorphosis, larvae were first reared to metamorphic competence on a high food concentration ofI. galbana (1.8 × 10⁵ cells ml^–1). When at least 80% of subsampled larvae were competent to metamorphose, as assessed by the numbers of indlviduals metamorphosing in response to elevated K⁺ concentration in seawater, remaining larvae were transferred either to 0.45-m filtered seawater or to suspensions of reduced phytoplankton concentration (1 × 10³, 1 × 10⁴, or 5 × 10⁴ cells ml^–1), or were maintained at 1.8 × 10⁵ cells ml^–1. All larvae were monitored daily for metamorphosis. Individuals that metamorphosed in each food treatment were transferred to high ration conditions (1.8 × 10⁵ tells ml^–1) for four additional days to monitor postmetamorphic growth. Competent larvae responded to all food-limiting conditions by metamorphosing precociously, typically 1 wk or more before larvae metamorphosed when maintained at the highest food ration. Surprisingly, juveniles reared at full ration grew more slowly if they had spent 2 or 3 d under food-limiting conditions as competent larvae. The data show that a rapid decline in phytoplankton concentration during the larval development ofC. fornicata stimulates metamorphosis, foreshortening the larval dispersal period, and may also reduce the ability of postmetamorphic individuals to grow rapidly even when food concentrations increase.     

Inhibitory effect of the iron chelator desferrioxamine (Desferal) on the generation of activated oxygen species by Chattonella marina

Recent studies demonstrated that the toxic red tide phytoplankton Chattonella spp. produce activated oxygen species such as superoxide anion (O ₂ ^- ), hydrogen peroxide (H₂O₂), and hydroxyl radicals (·OH), which may be responsible for the toxicity of this flagellate. However, the mechanism behind the production of these oxygen radicals and H₂O₂ by Chattonella spp. is largely unknown, and the physiological significance of activated oxygen species for Chattonella spp. is also unclear. In the present study, we investigated the involvement of iron in the generation of O ₂ ^- and H₂O₂ by C. marina. The generation of O ₂ ^- by C. marina was related to the growth phase; the highest rate of O ₂ ^- production was observed during the exponential growth phase. However, no such increase during the exponential growth phase was observed in C. marina growing in an iron-deficient medium, even though the growth of C. marina was not significantly affected by iron-deficiency during the first 4 d. In addition, the iron chelator desferrioxamine (Desferal) strongly inhibited the generation of both O ₂ ^- and H₂O₂ by C. marina in a concentration-dependent manner. The growth of C. marina was also inhibited by Desferal. Furthermore, in the presence of 500 M Desferal, C. marina-induced growth inhibition of the marine bacteria Vibrio alginolyticus was almost completely abolished. These results suggest that iron is required for the generation of activated oxygen species by C. marina, as well as for its own growth.     

Ingestion,growth and development of Penaeus indicus larvae as a function of Thalassiosira weissflogii cell concentration

Penaeus indicus larvae have been successfully reared in the laboratory using Thalassiosira weissflogii, Brachionus plicatilis and Artemia salina nauplii as food, with an average survival of 95.8% from nauplius 6 to postlarva 1. The effect of T. weissflogii cell concentration on larval ingestion, development and growth (total length) was investigated. Cell ingestion rates showed a saturation response to concentration. Both maximum ingestion rates and incipient limiting levels (the lowest concentration before ingestion rates were limited) were established for the feeding larval stages. Both were found to increase with progressive increase in larval development. Maximum ingestion rates increased from 0.25×10⁴ cells. larva^-1.h^-1 during protozoea 1 to reach a peak of 1.2×10⁴ cells. larva^-1.h^-1 during mysis 3 and then declined to 0.6×10⁴ cells. larva^-1.h^-1 at postlarva 1. Incipient limiting levels (ILLs) increased from approximately 0.6×10⁴ cells.ml^-1 during protozoea 2, to 0.65×10⁴ cells.ml^-1 during mysis 1, to 1.3×10⁴ cells. ml^-1 during mysis 3 to 1.6×10⁴ cells.ml^-1 at post-larva 1. Filter feeding efficiency was found to reach a maximum during mysis 1. Filter mechanisms are discussed. Generally, the most advanced larval development per unit time occurred at concentrations at and above the ILLs, while retarded development occurred below these levels. Growth increased asymptotically with cell concentration. Incipient growth limiting levels (IGLLs; the lowest concentration before growth was significantly limited) also increased with larval development and with the exception of mysis 3 they coincided with the ILLs. IGLLs increased from 0.55×10⁴ cells.ml^-1 during protozoea 2, to 0.66×10⁴ cells.ml^-1 during mysis 1, to 0.99×10⁴ cells.ml^-1 during mysis 3, to 1.62×10⁴ cells.ml^-1 at postlarva 1. Below the ILLs where ingestion was limited, animals were significantly smaller, with larval development and growth positively correlated to ingestion rates. When culturing penaeoid larvae, ambient cell concentrations should be kept above these known limiting levels to yield consistently good larval survival and growth.