Accumulation of paralytic shellfish poisoning toxins in planktonic copepods during a bloom of the toxic dinoflagellate<Emphasis Type="Italic"> Alexandrium tamarense</Emphasis> in Hiroshima Bay,western Japan

Concentrations of paralytic shellfish poisoning (PSP) toxins in toxic dinoflagellate cells and in marine planktonic copepods were monitored during the bloom of Alexandrium tamarense in Hiroshima Bay, western Japan. Concentration of the toxins retained by copepods was a function of the ambient toxin concentration, i.e. the product of A. tamarense cell density and cellular toxicity. The toxin concentration in copepods increased with the increase of toxicants in the seawater then leveled off, but decreased significantly at higher concentrations. In the field, the maximum toxin concentration was 1.2 pmol ind^-1, whereas in the laboratory, the copepod Acartia omorii accumulated a much higher concentration of PSP toxins (24 pmol ind^-1). Feeding avoidance against Alexandrium tamarense and a shift to alternative food sources such as diatoms in the field might keep their toxin levels lower than their potentially maximum level. The copepod toxin levels in the field were not so high as to cause an instantaneous lethal effect on their predator fishes but may reach possibly lethal levels after a few days' continuous feeding. Overall toxin retention by copepods after 12 h feeding and 2 h starvation was only 2.5% of total ingested toxins, which suggested that a significant amount of toxins was released into the seawater. Measurements of toxin reduction and gut evacuation suggested that the toxins were removed through both fecal evacuation and metabolism (e.g. excretion, decomposition and transformation). The results, as a whole, imply that copepods can be a link for PSP toxin flux in both pelagic and benthic food webs and can also be a sink for toxins by metabolizing and removing them from the environment.Communicated by T. Ikeda, Hakodate     

Feeding and absorption of the toxic dinoflagellate Alexandrium tamarense by two marine bivalves from the South China Sea

Paralytic shellfish poisoning (PSP) toxins can be accumulated by bivalves through the feeding process; therefore, knowledge on feeding and the assimilation of PSP-toxin-containing algae is critical to understand the kinetics of PSP toxins in these bivalves. In the South China Sea, it has been documented that the scallop Chlamys nobilis has a much higher PSP toxin burden than the clam Ruditapes philippinarum. Experiments were therefore carried out to assess whether the difference in toxin burden between these two species of bivalves was due to differences in feeding and absorption. In a mixed diet of Alexandrium tamarense (a PSP-toxin-producing dinoflagellate) and Thalassiosira pseudonana (a non-toxic diatom), the maximum clearance and filtration rates were about two times higher in the scallop C. nobilis than in the clam R. philippinarum. Furthermore, the clams produced pseudofeces at a lower cell density than the scallops. However, we found that the clams were unable to selectively exclude the toxic dinoflagellates by pseudofeces production. The scallop C. nobilis also possessed a greater ability to assimilate A. tamarense with a comparable carbon absorption efficiency to the diatom T. pseudonana. In contrast, the carbon absorption in the clam R. philippinarum was lower when feeding on A. tamarense than on the diatom. In general, the absorption efficiency decreased with increasing concentration of A. tamarense. Thus, it is likely that the higher PSP toxin levels in the scallops compared with clams can be partly explained by differences in their feeding and absorption behavior. Other processes, especially the biotransformation and biokinetics of PSP toxins, may also play a significant role in defining the inter-species differences in PSP body burden in marine bivalves.     

Fate of domoic acid ingested by the copepod <Emphasis Type="Italic">Acartia clausi</Emphasis>

Two important issues in the studies of harmful algae include ecological role of the toxic compounds and their fate through the food web. The aims of this study were to determine whether the production of domoic acid is a strategy evolved to avoid predation and the role of copepods in the fate of this toxic compound through the food web. The copepod Acartia clausi was fed with single and mixed cultures of the toxic diatom Pseudo-nitzschia multiseries and the non-toxic diatom Pseudo-nitzschia delicatissima. Ingestion rate as a function of diatom abundance was the same for the toxic and non-toxic Pseudo-nitzschia species, indicating no selective feeding behaviour against P. multiseries. The toxins ingested by the copepods did not affect mortality, feeding behaviour, egg production and egg hatching of the copepods. Copepods assimilated the 4.8% of the total domoic acid ingested. Although the amount of toxins daily detoxificated by the copepods was 63.6%, the copepods accumulated domoic acid in their tissues. We conclude that domoic acid is not toxic for copepods and, probably for this reason, this toxin does not act as feeding deterrent for copepods. However, even though the production of domoic acid has apparently not evolved to deter predation, copepods may play an important role on the fate of this toxic compound through the marine food web.     

Toxic dinoflagellates (Alexandrium fundyense and A. catenella) have minimal apparent effects on oyster hemocytes

The possible effect of Alexandrium spp. containing paralytic shellfish poisoning (PSP) toxins on the hemocytes of oysters was tested experimentally. In one trial, eastern oysters, Crassostrea virginica Gmelin, were exposed to bloom concentrations of the sympatric dinoflagellate, Alexandrium fundyense Balech, alone and in a mixture with a non-toxic diatom, Thalassiosira weissflogii (Grun) Fryxell et Hasle. Subsequently, another experiment exposed Pacific oysters, Crassostrea gigas Thunberg, to a mixed suspension of the sympatric, toxic species Alexandrium catenella (Whedon et Kofoid) Balech, with T. weissflogii. Measurements of numbers of oyster hemocytes, percentages of different cell types, and functions (phagocytosis, reactive oxygen species (ROS) production, and mortality) were made using flow-cytometry. During and after exposure, almost no significant effects of Alexandrium spp. upon hemocyte numbers, morphology, or functions were detected, despite observations of adductor-muscle paralysis in C. virginica and measured toxin accumulation in C. gigas. The only significant correlation found was between toxin accumulation at one temperature and higher numbers of circulating live and dead hemocytes in C. gigas. The PSP toxins are known to interfere specifically with sodium-channel function; therefore, the finding that the toxins had no effect on measured hemocyte functions suggests that sodium-channel physiology is not important in these hemocyte functions. Finally, because oysters were exposed to the living algae, not purified toxins, there was no evidence of bioactive compounds other than PSP toxins affecting hemocytes in the two species of Alexandrium studied.     

An integrative response by Mytilus chilensis to the toxic dinoflagellate Alexandrium catenella

Physiological responses of Mytilus chilensis exposed to the toxic dinoflagellate Alexandrium catenella were measured over 21 days in the laboratory and were compared with control mussels not exposed to the dinoflagellate. Mussels were collected from culturing ropes at Yaldad Bay, southern Chile (43o08′S 73o44′W), in August 2004 and acclimated to laboratory conditions for one week prior to the experiment. After 8 days, the paralytic shellfish poisoning (PSP) toxins (i.e. saxitoxin) in the tissues of exposed mussels exceeded safe levels for human consumption. Clearance rates, ingestion of organic matter, and absorption efficiency of exposed mussels were significantly lower than those of controls on day 0, but this was followed by an increase on day 3. The exposed mussels also increased their excretion rate over time, and this increase was significantly correlated with the accumulation of PSP toxins in their tissues. Oxygen consumption was not affected by the PSP toxins. The scope for growth (SFG) on day 0 was negative in exposed mussels, but it increased during the experiment. Although feeding activity and absorption efficiency were adversely affected during the first few days of exposure to PSP toxins from A. catenella in the laboratory, the M. chilensis cultured in Yaldad Bay may have evolved mechanisms that allow them to exploit the toxic dinoflagellate as a food source.     

Growth and grazing rates of the tintinnid ciliate<Emphasis Type="Italic"> Favella taraikaensis</Emphasis> on the toxic dinoflagellate<Emphasis Type="Italic"> Alexandrium tamarense</Emphasis>

Growth and feeding activities of the tintinnid ciliate Favella taraikaensis fed the toxic dinoflagellate Alexandrium tamarense were examined in laboratory experiments. Both growth and ingestion rates of F. taraikaensis as a function of the A. tamarense concentration were fitted to a rectangular hyperbolic equation. The maximum growth and ingestion rates of F. taraikaensis were 1.0 day^–1 and 2.8 cells ind. h^–1 (carbon specific ingestion rates: 3.5 day^–1), respectively, which are both included in the range of previous data reported for Favella spp. feeding on other algae. The gross growth efficiency (GGE) of F. taraikaensis ranged from 0.26 to 0.49 (mean value 0.40) at the concentration of 10–800 cells ml^–1, which is within the range of previous data on Favella spp. Also, the growth and ingestion rates and GGE of F. taraikaensis on A. tamarense were not significantly different from the values on another non-toxic dinoflagellate (Heterocapsa triquetra) at two different prey concentrations. This indicates that the toxicity of A. tamarense probably did not influence the feeding and growth activities of F. taraikaensis at concentrations of less than ca. 800 cells ml^–1. To evaluate the grazing by F. taraikaensis on A. tamarense blooms in the field, the population dynamics of A. tamarense were simulated based on the growth and ingestion parameters of F. taraikaensis. As a result, the grazing impact by F. taraikaensis was considered to potentially regulate the development of A. tamarense blooms. If the toxicity of A. tamarense does not influence the growth and feeding activities of F. taraikaensis, the occurrence of such grazer plankton are considered to be important for predicting the course of A. tamarense bloom dynamics under natural conditions.Communicated by T. Ikeda, Hakodate     

Effects of the toxic dinoflagellate <Emphasis Type="Italic">Gymnodinium catenatum</Emphasis> on uptake and fate of paralytic shellfish poisons in the Pacific giant lions-paw scallop <Emphasis Type="Italic">Nodipecten subnodosus</Emphasis>

Juvenile Pacific giant lions-paw scallops Nodipecten subnodosus were fed the toxic dinoflagellate Gymnodinium catenatum, a producer of paralytic shellfish poison (PSP), supplied with Isochrysis galbana (a nontoxic microalgae). Short-term (<24 h) experiments were performed to determine clearance and ingestion rates of G. catenatum. Kinetics of PSP was examined in longer-term experiments (>2 days). At high food concentrations, juvenile scallops showed production of pseudofeces, partial shell valve closure, and reduction in feeding. According to HPLC analysis, the only toxins present in the dinoflagellate G. catenatum and in the scallops were the gonyautoxins (GTXs), except in the labial palps and digestive gland, where trace amounts of saxitoxin (STX) were present in scallops. These tissues could play an important role in toxin biotransformation. The ranking of toxin concentration in tissues was: digestive gland > labial palps > intestine > gills > mantle > adductor muscle, where the total contribution of viscera was more than 80% of the total toxin body burden. Juvenile scallops exhibited no apparent detrimental physiological responses during the long-term feeding experiment. The dinoflagellate may contribute nutrients to the scallop, in addition to the microalgae I. galbana. The dinoflagellate may enhance cell uptake and byssus production. Once PSP accumulated during the first 12 days, it was slowly eliminated. The limited capacity for accumulating toxins in the adductor muscle favors domestic marketing of scallops.     

Copepod grazing during a declining spring phytoplankton bloom in the Northern North Sea

Grazing rates of larger (Calanus finmarchicus) and smaller (Acartia clausii Pseudocalanus elongatus etc.) copepods on naturally occurring phytoplankton populations were measured during a declining spring phytoplankton bloom. During the initial period, dominated by Chaetoceros spp. diatoms, constant ingestion rates were observed in Calanus finmarchicus at suspended particulate concentrations above 300 g carbon l^-1. Average daily intake during this time amounted to 35 to 40% of body carbon and reached a maximum of 50%. The feeding response of the smaller copepods was not so well defined, although a maximum daily intake of 56% body carbon was recorded. In both groups, feeding thresholds were at particulate concentrations around 50 g C l^-1. The feeding response of C. finmarchicus was correlated with both a change in their own population and in the food cell type. Linear regressions describing the concentration-dependent feeding response were: ingestion rate (IR)=1.16 total particulate volume (TPV)-36.15 during the initial part of the period compared with IR=0.41 TPV-12.18 for the latter period. C. finmarchicus filtered out slightly larger (x 1.2 diameter) particles than the small copepods and, in both groups, some filtering adjustment was made to accomodate to modal changes in the phytoplankton population from 20–30 m to 10 m diameter cells. Particle production during feeding was frequently evident in the smallest size ranges of particles and the ratio of particle production to ingestion rate was greater at low feeding rates.     

Alongshore transport of a toxic phytoplankton bloom in a buoyancy current: Alexandrium tamarense in the Gulf of Maine

We examined the mechanisms controlling blooms of the toxic dinoflagellate Alexandrium tamarense Lebour and the concomintant patterns of shellfish toxicity in the southwestern Gulf of Maine, USA. During a series of cruises from 1987 to 1989, hydrographic parameters were measured to elucidate the physical factors affecting the distribution and abundance of dinoflagellates along this coast. In 1988 and 1989 when toxicity was detected in the southern part of this region, A. tamarense cells were apparently transported into the area between Portsmouth, New Hampshire, and Cape Ann, Massachusetts, in a coastally trapped buoyant plume. This plume appears to have been formed by the outflow from the Androscoggin and Kennebec Rivers. Flow rates of these rivers, hydrographic sections, and satellite images led us to conclude that the plume persisted for about a month, and extended alongshore for several hundred kilometers. The distribution of cells followed the position of the plume as it was influenced by wind and topography. When winds were downwelling-favorable (to the southwest), cells were moved alongshore to the south, and were held to the coast; when winds were upwelling-favorable (to the northeast),the plume sometimes separated from the coast, advecting the cells offshore. In 1987 when no plume was present, A. tamarense cells were scarce, and no toxicity was recorded at the southern stations. The alongshore advection of toxic cells within a coastally trapped buoyant plume can explain the details of the temporal and spatial patterns of shellfish toxicity along the coast. We hypothesize that (1) the source of the A. tamarense populations is in the north, possibly associated with the Androscoggin and Kennebec estuaries, that (2) toxicity patterns follow a predictable relationship with river flow volume and timing of flow peaks and that (3) wind stresses directly influence the distribution of low salinity water and the dinoflagellate cells. Local, in situ growth of dinoflagellates can be an important factor initiating toxic dinoflagellate blooms. However, these data demonstrate the significant role of alongshore transport of established populations of A. tamarense in controlling the location and timing of paralytic shellfish poisoning (PSP) outbreaks in May and June along the southwestern coast of the Gulf of Maine.     

Short-term effects of sucralose on Calanus finmarchicus and Calanus glacialis in Disko Bay,Greenland

The potential effects of sucralose on the Arctic copepods Calanus finmarchicus and Calanus glacialis were studied in Disko Bay, Greenland. Sucralose is a non-calorie sweetener and chlorine derivate of sucrose containing three chlorine atoms. Scandinavian screening studies of sucralose in 2007, revealed sucralose in all effluent samples. To investigate whether sucralose is harmful to the Arctic aquatic ecosystems, possible short-term effects were investigated on egg production, hatching rate, food intake and mortality of two species of Arctic copepods. The copepods were exposed to six different concentrations (0–50,000 ng · L^?1) of sucralose, which spans the range of concentrations found in the screening studies. Exposure led to no mortality among the copepods. Food intake by C. glacialis increased with increasing concentrations of sucralose. In C. finmarchicus, food intake did not differ with increasing concentrations. No effect of sucralose was observed on egg production of C. finmarchicus. Despite increased food intake with increasing concentrations of sucralose, C. glacialis did not increase its egg production. The results show that both species responded weakly to sucralose, but with C. glacialis being possibly slightly more sensitive to sucralose than C. finmarchicus.     

Use of high-performance liquid chromatography to investigate the production of paralytic shellfish toxins by Protogonyaulax spp. in culture

Procedures have been developed for the extraction and high-performance liquid chromatography (HPLC) analysis of paralytic shellfish poisoning (PSP) toxins from Protogonyaulax spp. grown in batch culture. Using these procedures, the toxin content of two isolates of P. tamarensis (NEPCC 183 and 255) and one isolate of P. catenella (NEPCC 355) were examined. Total toxin and individual toxin concentrations were measured for each isolate during the exponential and stationary phases of growth in batch culture. The total toxicity of each isolate as measured by HPLC analysis was found to agree with toxicity as determined by the standard mouse bioassay. Two of the isolates (255 and 355) were found to be toxic and the third (183) was non-toxic. The toxic isolates (255 and 355) both showed higher average total PSP toxin content during the exponential phase (35 and 23 fmol toxin cell^-1, respectively) than during the stationary phase (21 and 8 fmol toxin cell^-1, respectively). These cultures differed dramatically in their toxin composition. P. tamarensis (255) contained a large proportion of the N(21) sulfo toxins (B₁, B₂, C₁, C₂) while P. catenella (355) contained primarily Gonyautoxins 1 through 4. The percent composition of individual toxins was found to be constant throughout the growth cycle for both toxic isolates, even though the total toxin concentration varied. Our results suggest that PSP toxin profiles might be useful as chemotaxonomic indicators.     

Effects of exposure to toxic and non-toxic dinoflagellates on oxygen consumption and locomotion in stage 1 larvae of the crabs Cancer oregonensis and C. magister

Sublethal effects on larval crabs upon exposure to toxic dinoflagellates were examined in the laboratory in early 1999. Specifically, oxygen consumption rates and geotaxis responses were determined for stage 1 larvae of the crabs Cancer oregonensis (Dana) and C. magister Dana that were exposed to non-toxic (Alexandrium tamarense, strain 115) or toxic (A. fundyense, strain 1719) dinoflagellates or to freshly hatched nauplii of the brine shrimp Artemia sp. In C. oregonensis, larvae exposed to the toxic dinoflagellate showed reduced rates of oxygen consumption compared to those exposed to non-toxic dinoflagellates or brine shrimp nauplii. Larvae exposed to a filtrate of the non-toxic dinoflagellate showed no change in oxygen consumption, but a reduced rate when exposed to filtrate from the toxic alga at densities >5᎒² cells ml^-1. In C. magister, larvae exposed to the non-toxic A. tamarense or the toxic A. fundyense had reduced oxygen consumption rates. Larvae exposed to filtrates of non-toxic and toxic dinoflagellates had no change in oxygen consumption. In geotaxis tests, C. oregonensis larvae exposed for 1 day to the toxic A. fundyense reduced their level of locomotion compared to those exposed to non-toxic A. tamarense or to brine shrimp nauplii. C. magister larvae showed no change in activity after a 1-day exposure to the toxic A. fundyense. After a 4-day exposure to A. fundyense, C. magister larvae had much reduced locomotion. Reduced locomotory activity in larvae exposed to toxic algae is consistent with the changes in oxygen consumption rates. Responding to exposure to toxic algae by reducing locomotion may affect vertical migration in these negatively buoyant crab larvae, resulting in sinking below a toxic alga bloom, at least temporarily.     

Grazing rates of the copepods Calanus glacialis and C. finmarchicus in arctic waters of the Barents Sea

Natural feeding rates of Copepodite Stages IV and V, and adult female Calanus glacialis (Jaschnov) and Copepodite Stage V and adult female C. finmarchicus (Gunnerus) were estimated using fluorescence analysis of gut contents. Measurements were made on copepods sampled from arctic waters east of Svalbard (Barents Sea) during the spring phytoplankton increase, in the period from 27 May to 13 June, 1983. Observations on Copepodite Stages IV and V and adult female C. glacialis suggest that the gastric evacuation rate is independent of developmental stage, whereas C. finmarchicus Copepodite Stage V showed a lower gastric evacuation rate than adult females. Gut fullness displayed a low correlation with the ambient chlorophyll concentrations. Ingestion rates calculated for C. glacialis were 0.3, 2.3, and 11.0 g C h^-1 for Copepodite Stages IV and V and adult females, respectively. Copepodite Stage V and adult female C. finmarchicus ingested 0.9 and 1.1 g C h^-1 at a temperature of ca.-1.0°C. The maximum ingestion rate in terms of percent body carbon d^-1 was higher for adult female C. glacialis and C. finmarchicus than for the respective Copepodite Stage V's. The results are discussed both in relation to the physiological state of the species and to the environmental conditions.     

Biosynthesis of trimethylamine oxide in calanoid copepods. seasonal changes in trimethylamine monooxygenase activity

Live copepods, Calanus finmarchicus (Gunnerus) and C. hyperboreus (Krøyer), exposed to dissolved ¹⁴C-labeled trimethylamine (TMA) in seawater, oxidized TMA to trimethylamine oxide (TMAO), which accumulated in the organisms. The amount of TMAO synthesized was dependent on the time of exposure to TMA and the concentration of TMA in the seawater. It was inferred that copepods can produce TMAO by oxidation of TMA found in their plant food. Choline and methionine did not appear to be of importance as precursors of TMAO. There were large seasonal changes in TMA monooxygenase activity of both copepods. The activity was high in spring, and decreased through summer and autumn to a winter low in October to March. The changes in TMAO content of C. finmarchicus were, in comparison, small (this aspect was not tested for C. hyperboreus).     

Comparisons among species ofAlexandrium (Dinophyceae) grown in nitrogen- or phosphorus-limiting batch culture

Three species of the dinoflagellate genusAlexandrium (Halim)-two strains of toxic.A. minutum, one each of nontoxicA. tamarense andA. affini-were grown in batch culture in either a low-nitrogen or a low-phosphate medium. Maximum carbon-specific growth rates forA. tamarense were lower (at <0.25 d^-1) than for the other strains, which all exceeded 0.38 d^-1. C-quotas (C content per cell) during exponential growth were similar for all strains (2.5 ng C cell^-1), with cells becoming smaller during the N-limiting stationary phase, but enlarging during prolonged P-deprivation. Values of ¹³C during the exponential phase were low (-25to-30), with most cells during the light phase swimming at the surface when nutrient-replete and migrating to the bottom of the flasks when nutrient-deplete with ¹³C rising to around-15. Biomass could not be estimated reliably from pigmentation, but could be estimated from biovolume (r>0.95), although this was complicated in cultures ofA. minutum by the presence of particles comprized of thecal plates of a similar size to intact cells. Alkaline phosphatase activity was not a reliable indicator a P-status. The most toxic strain tested (A. minutum AL1V) contained the highest concentrations of free amino acids, of arginine (a precursor of paralytic shellfish toxins) and of proline, and also had the lowest C:N mass ratio (at 4.3).A. affini contained the lowest concentrations of arginine, andA. tamarense the highest exponential phase C:N (7.8). For all strains, the mole ratio of intracellular glutamine: glutamate (Gln: Glu, which was abnormally high compared to other algae) could only be used to indicate the presence or absence of N-stress rather than the degree of stress. Additions of ammonium and phosphate resulted in increases in Gln: Glu within 20 min in N-stressed cells and also enhanced toxin content inA. minutum (mainly gonyautoxin) 4 over a 24 h period.     

Production of DMSP and DMS during a mesocosm study of an Emiliania huxleyi bloom: influence of bacteria and Calanus finmarchicus grazing

We investigated the influence of bacteria and metazooplankton on the production of dimethylsulfoniopropionate (DMSP) and dimethylsulfide (DMS) during blooms of Emiliania huxleyi (Lohmann) Hay and Mohler in seawater mesocosms. The phytoplankton succession was marked by the rapid collapse of an initial Skeletonema costatum (Greville) Cleve bloom followed by a small E. huxleyi bloom. The collapse of the diatom bloom was accompanied by an increase in concentrations of dissolved DMSP (DMSP_d) and bacterial abundance and activity (as determined by the thymidine incorporation technique). The increase in bacterial activity was followed by a rapid decrease in DMSP_d concentrations which remained low for the rest of the experiment, even during the subsequent collapse of the E. huxleyi blooms. The absence of DMSP_d and DMS peaks during the declining phase of the E. huxleyi blooms was attributed to the high bacterial activity prevailing at that time. The influence of metazooplankton grazing on DMSP and DMS production was investigated by adding moderate (24 mg dry weight m^-3) and high (520 mg dry weight m^-3) concentrations of Copepodite Stage V and adults of Calanus finmarchicus to two of four filtered (200 m mesh net) enclosures during the E. huxleyi blooms. The addition of C. finmarchicus, even in high concentrations, had no apparent effect on the dynamics of E. huxleyi, suggesting that the copepods were not grazing significantly on nanophytoplankton. The addition of copepods in high concentrations favored an accumulation of chlorophyll a and particulate DMSP. These results suggest that copepods were preying on the herbivorous microzooplankton which, in turn, was controlling the biomass of nanophytoplankton. DMS production was also enhanced in the enclosure with maximum metazooplankton biomass, suggesting that the grazing of C. finmarchicus on microzooplankton containing DMSP may contribute to DMS production. These results provide strong support to the emerging idea that bacteria and metazooplankton grazing play a dominant role in determining the timing and magnitude of DMS pulses following phytoplankton blooms.     

Quantification of copepod gut content by differential length amplification quantitative PCR (dla-qPCR)

Quantification of feeding rates and selectivity of zooplankton is vital for understanding the mechanisms structuring marine ecosystems. However, methodological limitations have made many of these studies difficult. Recently, molecular based methods have demonstrated that DNA from prey species can be used to identify zooplankton gut contents, and further, quantitative gut content estimates by quantitative PCR (qPCR) assays targeted to the 18S rRNA gene have been used to estimate feeding rates in appendicularians and copepods. However, while standard single primer based qPCR assays were quantitative for the filter feeding appendicularian Oikopleura dioica, feeding rates were consistently underestimated in the copepod Calanus finmarchicus. In this study, we test the hypothesis that prey DNA is rapidly digested after ingestion by copepods and describe a qPCR-based assay, differential length amplification qPCR (dla-qPCR), to account for DNA digestion. The assay utilizes multiple primer sets that amplify different sized fragments of the prey 18S rRNA gene and, based on the differential amplification of these fragments, the degree of digestion is estimated and corrected for. Application of this approach to C. finmarchicus fed Rhodomonas marina significantly improved quantitative feeding estimates compared to standard qPCR. The development of dla-qPCR represents a significant advancement towards a quantitative method for assessing in situ copepod feeding rates without involving cultivation-based manipulation.     

Winter studies on zooplankton in Arctic seas: the Storfjord (Svalbard) and adjacent ice-covered Barents Sea

Zooplankton was sampled in the Storfjord and ice-covered Barents Sea during March 2003. Environmental conditions represented a typical winter situation with low air temperatures, close pack ice, and extremely low chlorophyll concentrations. Polar water dominated the hydrographic regime in the upper layers. Zooplankton distribution reflected spatial variability of hydrography. The copepods Pseudocalanus spp., Oithona similis, Microsetella norvegica together with gastropod larvae were most numerous. Biomass averaged for the entire water column varied from 3.3 to 14.3 mg dry mass m⁻³, Calanus glacialis and Parasagitta elegans contributed most, followed by C. finmarchicus, Oithona similis, and Pseudocalanus spp. Various holoplankters showed reproductive activity, especially cyclopoid copepods and chaetognaths. A few C. glacialis females laid eggs in situ, but when fed diatom cultures rapidly increased their egg production. Meroplankton including larvae of nudibranchia, bivalvia, ophiuroida, polychaeta, and bryozoa were also present. Our data demonstrate that the pelagic community of the seasonally ice-covered Barents Sea was not in a “sleeping” state at the end of the winter, but in addition to dormant stages, a portion of mainly omnivorous and several carnivorous species was reproducing.     

Field depuration and biotransformation of paralytic shellfish toxins in scallop<Emphasis Type="Italic"> Chlamys nobilis</Emphasis> and green-lipped mussel<Emphasis Type="Italic"> Perna viridis</Emphasis>

Under laboratory conditions, the scallop Chlamys nobilis and the mussel Perna viridis were exposed to N-sulfocarbamoyl toxins (C2 toxin), a paralytic shellfish toxin (PST), by feeding a local toxic strain of the dinoflagellate Alexandrium tamarense (ATDP) that produced C2 toxin exclusively. The bivalves were subsequently depurated in the field, and their depuration kinetics, biotransformation and toxin distribution were quantified. Depuration was characterized by a rapid loss within the first day, followed by a secondary slower loss of toxins. In the fast depuration phase, scallops detoxified PSTs more quickly than the mussels (depuration rate constants for scallops and mussels were 1.16 day^–1 and 0.87 day^–1, respectively). In contrast, the mussels detoxified PSTs more quickly than the scallops in the slow depuration phase, and the calculated depuration rate constants (mean+SE) from day 2 to day 13 were 0.063+0.009 day^–1 and 0.040+0.019 day^–1 for mussels and scallops, respectively. The differences in the appearances of gonyautoxins, GTX2 and GTX3, and their decarbamoyl derivatives, dcGTX2, dcGTX3 and GTX5, which are all derivatives of C2 toxin, indicated active and species-specific biotransformation of the algal toxins in the two bivalves. In both species of bivalves, the non-viscera tissue contained fewer toxins and lower concentrations than the viscera-containing tissue compartment. In scallops, very little toxin was distributed in the adductor muscle. In mussels, most of the PSTs were found in the digestive gland with significant transport of toxins into the digestive gland from other tissues during the course of depuration. The toxin profiles of scallops and mussels differed from each other and from that of the toxic algae fed. A significant fraction of GTX5 was detected in the mussels but not in the scallops. Our study demonstrates a species specificity in the depuration kinetics, biotransformation and tissue distribution of PSTs among different bivalves.Communicated by T. Ikeda, Hakodate     

High reproduction of <Emphasis Type="Italic">Calanus finmarchicus</Emphasis> during a diatom-dominated spring bloom

Feeding, egg production, hatching success and early naupliar development of Calanus finmarchicus were measured in three north Norwegian fjords during a spring bloom dominated by diatoms and the haptophyte Phaeocystis pouchetii. Majority of the copepod diet consisted of diatoms, mainly Thalassiosira spp. and Chaetoceros spp., with clearance rates up to 10 ml ind⁻¹ h⁻¹ for individual algae species/groups. Egg production rates were high, ranging from ca 40 up to 90 eggs f⁻¹ d⁻¹, with a hatching success of 70–85%, and fast naupliar development through the first non-feeding stages. There was no correlation between the egg or nauplii production and diatom abundance, but the hatching success was slightly negatively correlated with diatom biomass. However, the overall high reproductive rates suggested that the main food items were not harmful for C. finmarchicus reproduction in the area, although direct chemical measurements were not conducted. The high population egg production (>1,20,000 eggs m⁻² d⁻¹) indicated that a large part of the annual reproduction took place during the investigation, which stresses the importance of diatom-dominated spring phytoplankton bloom for population recruitment of C. finmarchicus in these northern ecosystems.