Ammonium, but not nitrate, stimulates an increase in glutamine concentration in the haemolymph of Tridacna gigas

The concentration of glutamine in Tridacna gigas haemolymph increased >35-fold following exposure to sea water supplemented with ammonium (20 μM), but no increase was observed with nitrate (20 μM). Lack of a diel cycle, no decrease in haemolymph glucose levels, the expression patterns of glutamine synthetase in zooxanthellae and host, and the lack of glutamine release in response to nitrate supplementation all support the proposition that the increase in haemolymph glutamine is a product of the host and not the zooxanthellae. Unlike ammonium, nitrate accumulates rapidly in the haemolymph. It has no effect on the concentration of glutamine in the haemolymph, but there is an increase in arginine, histidine and lysine in the haemolymph, suggesting the release of these essential amino acids from zooxanthellae. Glutamine synthetase (GS) activity decreased markedly in the gill and less so in the mantle over a period of 6 d exposure to elevated ammonium (20 μM). In contrast, GS activity in zooxan- thellae doubled. The response of zooxanthellae in situ was confirmed by incubating freshly isolated zooxanthellae for 4 d in ammonium, which resulted in a ten-fold increase in GS activity. Comparison of the in situ response of zooxanthellae with that obtained in vitro indicates that the symbionts are likely to be exposed to ammonium concentrations lower than that found in the haemolymph. Received: 14 November 1997 / Accepted: 28 April 1998     

Microsensor studies of photosynthesis and respiration in the symbiotic foraminifer Orbulina universa

Oxygen and pH microelectrodes were used to investigate the microenvironment of the planktonic foraminifer Orbulina universa and its dinoflagellate endosymbionts. A diffusive boundary layer surrounds the foraminiferal shell and limits the O₂ and proton transport from the shell to the ambient seawater and vice versa. Due to symbiont photosynthesis, high O₂ concentrations of up to 206% air saturation and a pH of up to 8.8, i.e. 0.5 pH units above ambient seawater, were measured at the shell surface of the foraminifer at saturating irradiances. The respiration of the host–symbiont system in darkness decreased the O₂ concentration at the shell surface to <70% of the oxygen content in the surrounding air-saturated water. The pH at the shell surface dropped to 7.9 in darkness. We measured a mean gross photosynthetic rate of 8.5 ± 4.0 nmol O₂ h⁻¹ foraminifer⁻¹. The net photosynthesis averaged 5.3 ± 2.7 nmol O₂ h⁻¹. In the light, the calculated respiration rates reached 3.9 ± 1.9 nmol O₂h⁻¹, whereas the dark respiration rates were significantly lower (1.7 ± 0.7 nmol O₂ h⁻¹). Experimental light–dark cycles demonstrated a very dynamic response of the symbionts to changing light conditions. Gross photosynthesis versus scalar irradiance curves (P vs E _o curves) showed light saturation irradiances (E _k) of 75 and 137 μmol photons m⁻² s⁻¹ in two O. universa specimens, respectively. No inhibition of photosynthesis was observed at irradiance levels up to 700 μmol photons m⁻² s⁻¹. The light compensation point of the symbiotic association was 50 μmol photons m⁻² s⁻¹. Radial profile measurements of scalar irradiance (E _o) inside the foraminifera showed a slight increase at the shell surface up to 105% of the incident irradiance (E _d). Received: 26 January 1998 / Accepted: 11 April 1998     

Growth of zooxanthellae in culture with two nitrogen sources

Physiological characteristics of zooxanthellae were examined under nutrient-saturated conditions created by mixing ammonium (¹⁵NH₄) with nitrate (¹⁵NO₃) to give 0.88 mM total nitrogen. Growth rate varied with the form of nitrogen provided. Ammonium alone resulted in the lowest C:N and C:chl-a ratios. Although zooxanthellae took up nitrate in the absence of ammonium, ammonium assimilation was 1.3 times higher than nitrate assimilation. Ammonium strongly inhibited nitrate assimilation. While high-ammonium treatments resulted in the highest ¹⁴C incorporation into intermediate compounds, high nitrate levels resulted in the highest ¹⁴C incorporation into protein, suggesting that the intermediate compounds are produced prior to the subsequent production of protein when ammonium is the dominant N source. The enhanced production of intermediate compounds at the expense of carbon directed to protein synthesis in the presence of ammonium might be analogous to the “host factor” observed in zooxanthellae–host symbioses, since growth rate is depressed due to low production of protein. Received: 16 March 2000 / Accepted: 26 August 2000     

Dark survival strategies in marine phytoplankton assessed by cytometric measurement of metabolic activity with fluorescein diacetate

Cytometric quantification of cellular fluorescence upon cleavage of fluorescein diacetate (FDA) is presented as a sensitive and rapid technique to assess phytoplankton metabolic activity during exposure to prolonged darkness of 10 to 12 d. Two distinct types of metabolic response to darkness are distinguished: Type I cells (Brachiomonas submarina, Pavlova lutheri, Chrysochromulina hirta) adapt to prolonged darkness by reducing their metabolism to a lower level of activity (∼10% of initial in P. lutheri, C. hirta, ∼0.5% in B. submarina) within few days, whereas Type II cells (Prymnesium parvum, Bacteriastrum sp., unidentified pennate diatom) continue with unchanged activity. Type I cells were able to maintain their initial cell abundance and commenced rapid cell growth upon re-illumination after 12 d of darkness. Among Type II cells, diatoms were able to maintain cell abundance and growth capacity as well, whereas P. parvum was not. Type I cells are expected to exhibit competitive advantages in environments with frequent or long dark periods. Bacterivory further supports dark survival in C. hirta. Received: 10 May 1999 / Accepted: 20 September 1999     

Symbiotic anemones can grow when starved: nitrogen budget for Anemonia viridis in ammonium-supplemented seawater

The ability of endosymbioses between anthozoans and dinoflagellate algae (zooxanthellae) to retain excretory nitrogen and take up ammonium from seawater has been well documented. However, the quantitative importance of these processes to the nitrogen budget of such symbioses is poorly understood. When starved symbiotic Anemonia viridis were incubated in a flow-through system in seawater supplemented with 20 μM ammonium for 91 d under a light regime of 12 h light at 150 μmol photons m⁻² s⁻¹ and 12 h darkness, they showed a mean net growth of 0.197% of their initial weight per day. Control anemones in unsupplemented seawater with an ammonium concentration of <1 μM lost weight by a mean of 0.263% of their initial weight per day. Attempts to construct a nitrogen budget showed that, over a 14 d period, ≃40% of the ammonium taken up could be accounted for by growth of zooxanthellae. It was assumed that the remainder was translocated from zooxanthellae to host. However, since the budget does not balance, only 60% of the growth of host tissue was accounted for by this translocation. The value for host excretory nitrogen which was recycled to the symbionts equalled that taken in by ammonium uptake from the supplemented seawater, indicating the importance of nitrogen retention to the symbiotic association. Received: 23 December 1997 / Accepted: 12 September 1998     

Amino acid uptake by the toxic dinoflagellate Alexandrium fundyense

A toxic axenic strain of Alexandrium fundyense is shown to be capable of removing dissolved free amino acids (DFAAs) until concentrations are similar (low nM) to those found in natural waters. Uptake is greatest during exponential growth, rather than during C and/or N-stress as is usual in diatoms and other flagellates. A wide range of amino acids can be taken up, their concentration being decreased within a few hours to the levels observed prior to DFAA addition. The maximum rate of DFAA-N uptake, during early exponential phase, was 0.8 pmol-N cell⁻¹ h⁻¹, equivalent to ≃20% of the total N requirement. More typically, the contribution of DFAA-N was only ≃5%. However, these uptake rates are not sustainable. It is apparent that this organism cannot use amino-N to support significant growth, even though it can take up DFAAs. This, and the fact that the composition of the internal amino acid pool differed from that externally, is further evidence that the N-physiology of this genus is abnormal (differences to other dinoflagellates include an abnormally high concentration of glutamine and arginine, an effective absence of amine X, and release of nitrite during the␣concurrent assimilation of nitrate and ammonium in␣darkness). There is no evidence that the use of DFAAs enhance toxin content, except when cells are supplied with very high (unnatural) concentrations of arginine. Received: 8 May 1989 / Accepted: 14 September 1998     

UV-radiation can affect depth-zonation of Antarctic macroalgae

Due to depletion of stratospheric ozone over polar regions of the Northern and Southern Hemispheres UV-B-radiation has increased at the surface of the earth. Measurements of variable chlorophyll fluorescence were conducted to document UV-induced photoinhibition of photosystem II in cultivated macroalgae with different depth distributions in Antarctica. The reactions during artificial UV-exposure were observed on a short time scale (hours) and in light–dark cycles over several days. The nine species of investigated macroalgae show great differences in UV-tolerance of the photosynthetic process. Photosynthesis of the studied green algae was inhibited to a minor degree, while the brown algae showed an intermediate inhibition of photosynthesis. The response of the studied red algae varied with species. The differences in the degree of inhibition and recovery of photosynthetic efficiency and capacity indicate that UV-radiation is one important factor affecting the vertical distribution of macroalgae in nature. Received: 11 December 1997 / Accepted: 27 March 1998     

Resting spore formation and biochemical composition of the marine planktonic diatom Chaetoceros pseudocurvisetus in culture: ecological significance of decreased nucleotide content and activation of the xanthophyll cycle by resting spore formation

The biochemical composition of vegetative cells and resting spores of Chaetoceros pseudocurvisetus Mangin was compared in cultures under various nutrient and light conditions. The cellular content of major nucleotides such as AMP, ADP, ATP and UTP decreased in the order: vegetative cells, nutrient-starved (vegetative) cells and resting spores, indicating that the general metabolism of resting spores is relatively inactive. ADP-glucose was only abundant in nutrient-starved vegetative cells, suggesting metabolic imbalance in these cells. The chl a–specific fluorescence yield of vegetative cells grown under all culture conditions was low, but very high in resting spores. The ratios of the cellular contents of diadinoxanthin to chl a and of diatoxanthin to chl a were higher in resting spores and nutrient-starved vegetative cells than in nutrient-replete vegetative cells. The diadinoxanthin–diatoxanthin xanthophyll cycle was active in resting spores; the xanthophyll cycle was synchronized with a 14 h light:10 h dark photoperiod. Also, the ratios of cellular content of diadinoxanthin and diatoxanthin to cellular content of chl a in resting spores were relatively high in high irradiance, and decreased gradually in conditions of darkness over long culture periods. Under conditions of strong light and high temperature, most resting spores survived more than 40 d while nutrient-starved vegetative cells died within 33 d. These results suggest that resting spore formation is a strategy for enhancing protection and lowering metabolic rate for survival. These physiological changes accompanying spore formation enable resting spores not only to overwinter but also to “oversummer” in the coastal euphotic layer. Received: 23 March 1999 / Accepted: 11 August 1999     

Cell cycle and toxin production in the benthic dinoflagellate Prorocentrum lima

Profiles of diarrhetic shellfish poisoning (DSP) toxins produced throughout the growth cycle and the cell cycle of the toxigenic marine dinoflagellate Prorocentrum lima were studied in triplicate unialgal batch cultures. Cells were pre-conditioned at 18 ± 1 °C, under a photon flux density (PFD) of 90 ± 5 μmol m⁻² s⁻¹ on a 14 h light:10 h dark photoperiod. In exponential growth phase, cultures were synchronized in darkness for 17 d. After dark synchronization, cultures were transferred back to the original photoperiod regime. Cells were harvested for DSP toxin analysis by LC-MS (liquid chromatography with mass spectrometry), and double-stranded (nuclear) DNA was quantified by flow cytometry. The cell populations became asynchronous within approximately 3 d after transition from darkness to the 14 h light:10 h dark photoperiod. This may be due to the prolonged division cycle (5 to 7 d) that is not tightly phased by the photoperiod. Unlike other planktonic Prorocentrum spp., cytokinesis in P. lima occurred early in the dark and ceased by “midnight”. Cellular levels of the four principal DSP toxins, okadaic acid (OA), OA C8-diol-ester (OA-D8), dinophysistoxin-1 (DTX1) and dinophysistoxin-4 (DTX4), ranged from 0.37 to 6.6, 0.02 to 1.5, 0.04 to 2.6, and 1.8 to 7.8 fmol cell⁻¹, respectively. No toxin production was evident during the extended period of dark synchronization nor during the initial period when NH₄ was consumed as the major nitrogen source. Soon after the cells were returned to the 14 h light:10 h dark cycle and they began to take up NO₃, cellular levels of all four toxins gradually increased. This increase in DSP toxins usually occurred in the light, marked by a rise in DTX4 levels that preceded an increase in the cellular concentration of OA and DTX1 (delayed by 3 to 6 h). Thus, DTX4 synthesis is initiated in the G1 phase of the cell cycle and persists into S phase (“morning” of the photoperiod), whereas OA and DTX1 production occurs later during S and G2 phases (“afternoon”). No toxin production was measured during cytokinesis, which happened early in the dark. The evidence indicates that toxin synthesis is restricted to the light period and is coupled to cell cycle events. Received: 3 September 1998 / Accepted: 30 March 1999     

Effects of host-tissue homogenate of the scleractinian coral Plesiastrea versipora on glycerol metabolism in isolated symbiotic dinoflagellates

Symbiotic dinoflagellate algae (Symbiodinium sp.) isolated from the scleractinian coral Plesiastrea versipora and incubated in homogenized host tissue released 4 to 7 times as much glycerol (14 to 46 nmol glycerol/10⁶ algae) as those incubated in seawater (3 to 6 nmol glycerol/10⁶ algae) after 4 h incubation in the light. During this period, no release of triglycerides was detected. Intracellular glycerol increased 2- to 3-fold in algae incubated in host homogenate, but remained unchanged in algae incubated in seawater at a concentration of 0.82 ± 0.47 nmol glycerol/10⁶ algae. In each incubation condition, intracellular triglyceride levels increased. However, in algae incubated in host homogenate, the intracellular levels of triglycerides reached only about 75% of the amount reached in algae incubated in seawater (max. 18.55 ± 2.40 nmol glycerol/10⁶ cells). Host homogenate did not stimulate the release of glycerol from algae during dark incubation. These data show that the glycerol released by algae incubated in host-tissue homogenate was derived from increased synthesis of glycerol or from diversion of some glycerol or other photosynthetic intermediates from incorporation into algal triglyceride stores, and did not come from existing stores. Received: 20 December 1996 / Accepted: 9 January 1997     

Rhythmicity of larval release in three species of intertidal brachyuran crabs (Crustacea: Brachyura) from Inhaca Island (Mozambique)

 A study of rhythmicity of larval release in three species of intertidal brachyuran crabs, based on laboratory and field experiments, was undertaken at Inhaca Island, southern Mozambique, using Leptodius exaratus and Macrophthalmus grandidieri from December 1994 to January 1995, and Arcotheres palaensis from April to July 1995. L. exaratus and M. grandidieri showed a semi-lunar cycle in larval release. The release of larvae for L. exaratus, a species having conspicuous larvae, occurred in the first half of the night, after the post-crepuscular high tide, which suggests maximisation of protection of larvae from visual predation. The larval release activity matched the late spring and early neap tides. Results from the field were similar to those from the laboratory. M. grandidieri, having inconspicuous larvae, did not show a pattern related to the light–dark cycle and hatched during spring tides (around full and new moons) to maximise larval dispersion. A. palaensis, living inside the host mussel which inhabits the lowest section of the intertidal zone, did not show a relation with moon phase, tidal or light–dark cycles. Received: 16 February 1999 / Accepted: 8 December 1999     

Availability of two forms of dissolved nitrogen to the coral Pocillopora damicornis and its symbiotic zooxanthellae

The relative contribution of dissolved nitrogen (ammonium and dissolved free amino acids DFAAs) to the nitrogen budget of the reef-building coral Pocillopora damicornis was assessed for colonies growing on control and ammonium-enriched reefs at One Tree Island (southern Great Barrier Reef) during the ENCORE (Enrichment of Nutrient on Coral Reef; 1993 to 1996) project. P. damicornis acquired ammonium at rates of between 5.1 and 91.8 nmol N cm⁻² h⁻¹ which were not affected by nutrient treatment except in the case of one morph. In this case, uptake rates decreased from 80.5 to 42.8 nmol cm⁻² h⁻¹ (P < 0.05) on exposure to elevated ammonium over 12 mo. The presence or absence of light during measurement did not influence the uptake of ammonium ions. Nitrogen budgets revealed that the uptake of ammonium from concentrations of 0.11 to 0.13 μM could completely satisfy the demand of growing P. damicornis for new nitrogen. P. damicornis also took up DFAAs at rates ranging from 4.9 to 9.8 nmol N cm⁻² h⁻¹. These rates were higher in the dark than in the light (9.0 vs 5.1 nmol m⁻² h⁻¹, P < 0.001). Uptake rates were highest for the amino acids serine, arginine and alanine, and lowest for tyrosine. DFAA concentrations within the ENCORE microatolls that received ammonium were undetectable, whereas they ranged up to 100 nM within the control microatolls. The contribution of DFAAs to the nitrogen budget of P. damicornis constituted only a small fraction of the nitrogen potentially contributed by ammonium under field conditions. Even at the highest field concentrations measured during this study, DFAAs could contribute only ≃11.3% of the nitrogen demand of P.␣damicornis. This contribution, however, may be an important source of nitrogen when other sources such as ammonium are scarce or during periods when high concentrations of DFAAs become sporadically available (e.g. cell breakage during fish-grazing). Received: 22 April 1998 / Accepted: 3 November 1998     

Microsensor studies of photosynthesis and respiration in larger symbiotic foraminifera. I The physico-chemical microenvironment of Marginopora vertebralis, Amphistegina lobifera and Amphisorus hemprichii

 The physico-chemical microenvironment of larger benthic foraminifera was studied with microsensors for O₂, CO₂, pH, Ca²⁺ and scalar irradiance. Under saturating light conditions, the photosynthetic activity of the endosymbiotic algae increased the O₂ up to 183% air saturation and a pH of up to 8.6 was measured at the foraminiferal shell surface. The photosynthetic CO₂ fixation decreased the CO₂ at the shell down to 4.7 μM. In the dark, the respiration of host and symbionts decreased the O₂ level to 91% air saturation and the CO₂ concentration reached up to 12 μM. pH was lowered relative to the ambient seawater pH of 8.2. The endosymbionts responded immediately to changing light conditions, resulting in dynamic changes of O₂, CO₂ and pH at the foraminiferal shell surface during experimentally imposed light–dark cycles. The dynamic concentration changes demonstrated for the first time a fast exchange of metabolic gases through the perforate, hyaline shell of Amphistegina lobifera. A diffusive boundary layer (DBL) limited the solute exchange between the foraminifera and the surrounding water. The DBL reached a thickness of 400–700 μm in stagnant water and was reduced to 100–300 μm under flow conditions. Gross photosynthesis rates were significantly higher under flow conditions (4.7 nmol O₂ cm⁻³ s⁻¹) than in stagnant water (1.6 nmol O₂ cm ⁻³ s⁻¹), whereas net photosynthesis rates were unaffected by flow conditions. The Ca²⁺ microprofiles demonstrated a spatial variation in sites of calcium uptake over the foraminiferal shells. Ca²⁺ gradients at the shell surface showed total Ca²⁺ uptake rates of 0.6 to 4.2 nmol cm⁻² h⁻¹ in A. lobifera and 1.7 to 3.6 nmol cm⁻² h⁻¹ in Marginopora vertebralis. The scattering and reflection of the foraminiferal calcite shell increased the scalar irradiance at the surface up to 205% of the incident irradiance. Transmittance measurements across the calcite shell suggest that the symbionts are shielded from higher light levels, receiving approximately 30% of the incident light for photosynthesis. Received: 6 July 1999 / Accepted: 28 April 2000     

Endogenous control of timing of metamorphosis in megalopae of the shore crab Carcinus maenas

Batches of hundreds of freshly collected megalopae of the shore crab Carcinus maenas (L.) showed persistent circatidal rhythms of moulting to the juvenile crab stage when maintained in constant laboratory conditions. Peaks of moulting occurred around expected times of high tides, with few megalopae moulting at other times. In larvae collected offshore, the highest tidally-timed peak of metamorphosis occurred during the second to fifth expected times of high tide, and metamorphosis of 50% of each batch took about 22 h or longer. In contrast, in larvae collected at the water's edge, 70% metamorphosed during the first expected episode of high tide, within 6 to 8 h after collection. However, although inshore megalopae moulted before offshore ones, the tidal timing of moulting remained unaltered whether megalopae were collected at neaps or springs, from the water's edge or farther offshore, in the presence or absence of natural substratum, and under various light–dark and salinity regimes. Metamorphosis of C. maenas megalopae around the times of high tides may enhance settlement into the upper intertidal zone. Early juveniles of the crab apparently prefer that zone as they are most abundant there and, unlike adults, do not undertake up-and-down-shore migration with tides. The present finding demonstrates, for the first time, endogenous physiological timing of circatidal periodicity in the metamorphic moult of crab megalopae, suggesting that endogenous factors, as well as exogenous ones should be taken into account in considering the process of settlement by crab megalopae. Received: 21 February 1996 / Accepted: 27 November 1996     

Effect of nutrient enrichment in the field on the biomass, growth and calcification of the giant clam Tridacna maxima

Nutrients were added separately and combined to an initial concentration of 10 μM (ammonium) and/or 2 μM (phosphate) in a series of experiments carried out with the giant clam Tridacna maxima at 12 microatolls in One Tree Island lagoon, Great Barrier Reef, Australia (ENCORE Project). These nutrient concentrations remained for 2 to 3 h before returning to natural levels. The additions were made every low tide (twice per day) over 13 and 12 mo periods for the first and second phase of the experiment, respectively. The nutrients did not change the wet tissue weight of the clams, host C:N ratio, protein content of the mantle, calcification rates or growth rates. However, ammonium (N) enrichment alone significantly increased the total population density of the algal symbiont (Symbiodinium sp.: C = 3.6 · 10⁸ cell clam⁻¹, N = 6.6 · 10⁸ cell clam⁻¹, P = 5.7 · 10⁸ cell clam⁻¹, N + P = 5.7 · 10⁸ cell clam⁻¹; and C = 4.1 · 10⁸ cell clam⁻¹, N = 5.1 · 10⁸ cell clam⁻¹, P = 4.7 · 10⁸ cell clam⁻¹, N + P = 4.5 · 10⁸ cell clam⁻¹, at the end of the first and second phases of the experiment, respectively), although no differences in the mitotic index of these populations were detected. The total chlorophyll a (chl a) content per clam but not chlorophyll a per cell also increased with ammonium addition (C = 7.0 mg chl a clam⁻¹, N = 13.1 mg chl a clam⁻¹, P = 12.9 mg chl a clam⁻¹, N + P = 11.8 mg chl a clam⁻¹; and C = 8.8 mg chl a clam⁻¹, N = 12.8 mg chl a clam⁻¹; P = 11.2 mg chl a clam⁻¹, N + P = 11.3 mg chl a clam⁻¹, at the end of the first and second phases of the experiment, respectively). The response of clams to nutrient enrichment was quantitatively small, but indicated that small changes in inorganic nutrient levels affect the clam–zooxanthellae association. Received: 2 June 1997 / Accepted: 9 June 1997     

Circatidal rhythm of an intertidal crab, Hemigrapsus sanguineus : synchrony with unequal tide height and involvement of a light-response mechanism

The circatidal rhythm of intertidal animals may reflect the inequality of the tides. In addition, a light-sensitive mechanism may be involved in their internal timing systems. To test these hypotheses, the larval release activity of the intertidal crab Hemigrapsus sanguineus was monitored under different light conditions in the laboratory. Under a 24-h light–dark (LD) cycle with the phase similar to the field, the activity coincided with the times of high tide in the field and showed a tidal rhythm. This rhythm free-ran in constant, dim-light conditions, suggesting that the timing is controlled by an endogenous clock. When the population was exposed to a 24-h LD cycle with the phase changed from that in the field, the tidal rhythm was phase-shifted; while the light cycle advanced in phase from the field caused a phase-advance of the rhythm, that delayed in phase induced a phase-delay of the rhythm. Thus, a light-response mechanism is definitely involved in circatidal timing systems. But the population rhythm showed a large variability among individuals, associated with the phase-shift, and the magnitude of the phase-shift did not accurately correspond to that of the light cycle. These results suggest that the light-response system can control the phase of the rhythm less stronger than that in estuarine crabs. Most releases occurred at higher high tides, but the release of some females obviously occurred at lower high tides. The larval release pattern thus could not be accounted for by a simple synchrony with higher high tides. Hatching of H. sanguineus occurred after a “hatching program” of 49.5 to 52.5 h. This program is initiated by some factor (hatching-program inducing stumuli: HPIS) transmitted from the female to the embryos. We speculated that this factor is effectively transmitted to the embryos when the habitat is exposed to air, i.e., at lower low-tide periods, and that once each embryo is stimulated, hatching occurs synchronously 2 d later during high tide. The release of HPIS is probably controlled by the circatidal clock of the female, and the 24-h LD cycle may participate in shifting this timing to the opposite low tide. Received: 14 January 1997 / Accepted: 18 February 1997     

Effect of diel photoperiod on nitrogen metabolism of cultured and symbiotic zooxanthellae

Ammonium uptake and assimilation by zooxanthellae (Symbiodinium sp.) cultured with an excess of nitrate was enhanced in light. Uptake was decreased by the same amount when zooxanthellae were incubated in darkness either after 6 h pretreatment in light, or at the end of the dark period of a 12 h light: 12 h dark cycle. This suggested that short-term incubations of zooxanthellae were valid tests for light enhancement of dissolved inorganic nitrogen (DIN) uptake. Assimilation of ammonium into glutamine (Gln) and glutamate (Glu) was also decreased in darkness. During a 12 h light: 12 h dark cycle, free pools of both Gln and Glu fell quickly at the start of the light period, followed by steady increases until the beginning of the next dark period. Of the four other major components of free amino acid pools tested, only the nonprotein amino acid taurine showed diel fluctuations. Gln and Glu pools in zooxanthellae freshly isolated from reef-forming corals also showed differences between day and night, suggesting changes in patterns of DIN assimilation over the diurnal cycle.     

Free amino acids and energy metabolism in eggs and larvae of seabass, Lates calcarifer

Contents of free amino acids (FAA), protein and ammonium ions together with rates of ammonia excretion and oxygen consumption were measured in order to study the role of FAA as an energy substrate in developing eggs and larvae of seabass (Lates calcarifer) maintained in seawater (30 ppt) at 28 °C without feeding. Initially eggs contained 25.3 nmol ind⁻¹ of FAA of which 21.5 nmol was rapidly utilised by the developing eggs and larvae during the period up to 40 h post spawning (PS) when nearly all the yolk had been resorbed. During the same period, a net increase in protein content of 1.7 μg ind⁻¹ was observed, indicating that the major part of the amino acids lost from the free pool had been polymerised into body proteins. Assuming that the balance of the FAA after protein synthesis was used entirely for energy metabolism, FAA appeared to be an important energy substrate during the embryonic stages (2 to 16 h PS); after hatching, the contribution of FAA to energy metabolism was less significant. From 50 h PS until the end of the study period at 100 h PS, amino acids derived from somatic protein were used for energy metabolism. For the overall period from just after spawning up to 100 h PS, the data indicate that ca. 14% of the total aerobic energy metabolism was derived from amino acid catabolism. Received: 26 September 1997 / Accepted: 1 April 1998     

DMSP-lyase activity in five marine phytoplankton species: its potential importance in DMS production

Activity of DMSP-lyase, which cleaves dissolved DMSP (henceforth DMSP_d-lyase), was examined in five axenically cultured phytoplankton species, including both DMSP-producing and non-DMSP-producing species. High DMSP_d-lyase activity was found in two DMSP producers, Heterocapsa triquetra strain NIES-7 and Scrippsiella trochoidea strain NIES-369 (Dinophyceae). The DMS production rates at 100 nM DMSP_d were 0.5 fmol cell⁻¹ min⁻¹ for H. triquetra and 0.3 fmol cell⁻¹ min⁻¹ for S. trochoidea. In a non-DMSP producer, Heterosigma akashiwo strain NIES-6 (Raphidophyceae), the DMSP_d-lyase activity was not found. Two DMSP-producing Prymnesiophyceae species, Isochrysis galbana strain CCMP-1323 and Gephyrocapsa oceanica strain NIES-353, did not show any obvious activity either, in contrary to other authors' findings on Phaeocystis sp., another DMSP-producing Prymnesiophyceae species. The comparison of the DMSP_d-lyase activity of the two Dinophyceae species with bacterial DMSP consumption and DMS production activity in Tokyo Bay showed that the DMSP_d-lyase activity of H. triquetra and S. trochoidea could be an important mechanism for DMS production during their blooms. Received: 9 April 1999 / Accepted: 10 December 1999     

Hydrodynamic and chemical conditions during onset of a red-tide assemblage in an estuarine upwelling ecosystem

The hydrodynamics and nitrogen/silicon biogeochemistry accompanying the development of a red-tide assemblage were examined in the Ría de Vigo (northwest Spain), a coastal embayment affected by upwelling, during an in situ diel experiment in September 1991. Despite a low N:Si molar ratio (0.5) of nutrients entering the surface layer, which was favourable for diatom growth, the diatom population began to decline. Limited N-nutrient input, arising from moderate coastal upwelling in a stratified water column, restricted net community production (NCP = 630 mg C m⁻² d⁻¹). In addition, light-limitation of gross primary production (GPP = 1525 mg C m⁻² d⁻¹) was observed. The relatively high f-ratio (= NCP:GPP) recorded (0.41, characteristic of intense upwelling conditions) would have been as low as 0.15 had not GPP been limited by light intensity. Temporal separation of carbohydrate synthesis during the photoperiod from protein synthesis in the dark could be inferred from the time-course of the C:N ratio of particulate organic matter. Severe light-limitation would lead to diatom collapse were the diatoms not able to meet all their energy requirements during the hours of darkness. Under the hydrodynamic, nutrient and light conditions of the experiment, an assemblage of red-tide-forming species began to develop, aided by their ability to migrate vertically and to synthesize carbohydrates during the light in surface waters and protein during the dark at the 4 m-deep pycnocline. Thermal stratification, reduced turbulence, intense nutrient mineralization, and the limited nitrogen input through moderate upwelling were all favourable to the onset of a red-tide assemblage. Received: 15 February 1997 / Accepted: 26 September 1997