Fecal discharge of zooxanthellae in the giant clam Tridacna derasa, with reference to their in situ growth rate

The population dynamics of zooxanthellae living in the mantle of a giant clam, Tridacna derasa, was studied. The giant clams with shell lengths of 5 to 6 cm which had been reared in the Palau Mariculture Demonstration Center, in the Republic of Palau, were transferred to aquaria on deck of the R.V. “Sohgen-maru” and kept in running sea water at 29 to 30 °C. Two clams were removed from the aquaria, and zooxanthellae in the mantle were isolated every 2 h for 24 h. Numbers of the zooxanthellae in or not in the cell division stage were counted for calculations of the zooxanthellae population in the mantle and their mitotic index (MI). The MI increased after sunset and reached the maximum values of 6.1 to 11.5% at 03:00 to 05:00 hrs. The specific growth rate, μ, estimated from the MI was 0.083 to 0.14 d⁻¹. Five clams were kept in each of 2 Plexiglas containers in the aquarium for collection of the discharged feces every 3 to 4 h. The discharged zooxanthellae in the feces were counted. The zooxanthellae discharged in 24 h were 0.38 to 1.46% of the total zooxanthella population in the mantle, and 2.7 to 16.9% of the newly formed zooxanthella population in a day. Increase of zooxanthella population in the mantle was estimated from clam shell growth rate and from the correlation between zooxanthella population and clam shell size. Daily increase of zooxanthella population in the mantle was estimated to be approximately 7.6 to 19% of the newly formed zooxanthella population. Therefore, the sum of zooxanthellae populations accounting for daily increase in the mantle and discharge in the feces was 11 to 36% of the newly formed population. About 64 to 89% of the newly formed cells were missing; some of these may have been digested by the clam. Received: 14 July 1996 / Accepted: 19 August 1996     

Interplay of host morphology and symbiont microhabitat in coral aggregations

The Belizean reef coral Agaricia tenuifolia Dana forms aggregations in which rows of thin, upright blades line up behind each other. On average, the spacing between blades increases with depth and hence with decreasing ambient irradiance. We designed and built a small, inexpensive light meter and used it to quantify the effect of branch spacing on light levels within colonies at varying distances from branch tips. Concurrently, we measured photosynthetic pigment concentrations and population densities of symbiotic dinoflagellates (zooxanthellae) extracted from coral branches of colonies with tight (≤3 cm) vs wide (≥6 cm) branch spacing, collected at 15 to 17 m and from colonies with tight branch spacing collected at 1 to 2 m. Light levels decreased significantly with tighter branch spacing and with distance from the branch tips. Total cellular pigment concentrations (chlorophylls a, c ₂ and peridinin) as well as chlorophyll a:c ₂ and chlorophyll a: peridinin ratios all increased significantly with distance from the branch tip, indicating very localized differences in photoacclimation within individual branches. Zooxanthellae from colonies with widely-spaced branches displayed significantly lower chlorophyll a:c ₂ and chlorophyll a:peridinin ratios, and were present at significantly higher population densities than those from colonies with tightly-spaced branches collected at the same depth (15 m). Tightly-spaced colonies collected from shallow environments (1 to 2 m) displayed pigment ratios similar to those from widely-spaced colonies from deeper water (15 m), but maintained zooxanthellae populations at levels similar to those in tightly-branched colonies from deeper water. Thus, variation in colony morphology (branch spacing and distance from branch tip) can affect symbiont physiology in a manner comparable to an increase of over 15 m of water depth. These results show that a host's morphology can strongly determine the microhabitat of its symbionts over very small spatial scales, and that zooxanthellae can in turn display steep gradients in concordance with these altered physical conditions. Received: 12 June 1997 / Accepted: 24 June 1997     

Amino acid synthesis in the symbiotic sea anemone Aiptasia pulchella

Symbiotic Aiptasia pulchella and freshly isolated zooxanthellae were incubated in NaH¹⁴CO₃ and NH₄Cl for 1 to 240 min, and samples were analysed by reverse-phase high-performance liquid chromatography (HPLC) and an online radiochemical detector. NH₄ ⁺ was first assimilated into ¹⁴C-glutamate and ¹⁴C-glutamine in the zooxanthellae residing in A. pulchella. The specific activities (dpm nmol⁻¹) of ¹⁴C-glutamate and ¹⁴C-glutamine in vivo, were far greater in the zooxanthellae than in the host tissue, indicating that NH₄ ⁺ was principally incorporated into the glutamate and glutamine pools of the zooxanthellae. ¹⁴C-α-ketoglutarate was taken up from the medium by intact A. pulchella and assimilated into a small amount of ¹⁴C-glutamate in the host tissue, but no ¹⁴C-glutamine was detected in the host fraction. The ¹⁴C-glutamate that was synthesized was most likely produced from transamination reactions as opposed to the direct assimilation of NH₄ ⁺. The free amino acid composition of the host tissue and zooxanthellae of A. pulchella was also measured. The results presented here demonstrate that NH₄ ⁺ was initially assimilated by the zooxanthellae of A. pulchella. Received: 3 February 1997 / Accepted: 24 October 1997     

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     

Photoadaptation of zooxanthellae in the sponge Cliona vastifica from the Red Sea, as measured in situ

In the Red Sea, the zooxanthellate sponge Cliona vastifica (Hancock) is mainly present at >15 m depth or in shaded areas. To test whether its scarcity in unshaded areas of shallower waters is linked to the functional inefficiency of its photosymbionts at high irradiances, sponges were transferred from 30 m to a six times higher light regime at 12 m depth, and then returned to their original location. During this time, photosynthetic responses to irradiance were measured as rapid light curves (RLCs) in situ by pulse amplitude modulated (PAM) fluorometry using a portable underwater device, and samples were taken for microscopic determinations of zooxanthellar abundance. The zooxanthellae harboured by this sponge adapted to the higher irradiance at 12 m by increasing both their light saturation points and relative photosynthetic electron transport rates (ETRs). The ETRs at light saturation increased almost fourfold within 15–20 days of transfer to the shallower water, and decreased back to almost their original values after the sponges were returned to 30 m depth. This, as well as the fact that the photosynthetic light responses within an individual sponge were in accordance with the irradiance incident to specific surfaces, shows that these photosymbionts are highly adaptable to various irradiances. There was no significant change in the number of zooxanthellae per sponge area throughout these experiments, and the different photosynthetic responses were likely due to adaptations of the photosynthetic apparatus within each zooxanthella. In conclusion, it seems that parameters other than the hypothesised inability of the photosymbionts to adapt adequately to high light conditions are the cause of C. vastifica's rareness in unshaded shallow areas of the Red Sea. Received: 25 April 2000 / Accepted: 13 October 2000     

UV-absorbing substances in zooxanthellate and azooxanthellate clams

The effects of UV-A and UV-B radiation on photosynthesis of zooxanthellae within the siphonal mantle of the giant clam, Tridacna crocea, and in isolation were studied. While UV-B irradiation (2.4 W m⁻², 20 min) completely suppressed photosynthesis of the isolated zooxanthellae, it had little effect on their photosynthetic ability if they were irradiated while within the siphonal mantle of the host tissue. Chemical analysis of the siphonal mantle of T. crocea showed the presence of significant amounts of mycosporine-like amino acids (MAAs), which absorb UV-A and -B light. However, no MAA was detected in the isolated zooxanthellae. MAAs were concentrated in the siphonal mantle and kidney tissues in comparison with other tissues. In the siphonal mantle, MAA concentrations were the highest in the outermost surface layer where most of the zooxanthella cells resided. This indicates that the zooxanthellae are protected from UV radiation by a screen of concentrated MAAs in the host clam. Aside from T. crocea, significant amounts of MAAs were found not only in other zooxanthellate clams, such as T. derasa, Hippopus hippopus, Colculum cardissa and Fragum unedo, but also in a closely related azooxanthellate clam, Vasticardium subrugosum. On the other hand, no MAA was detected in any of the zooxanthellae from these zooxanthellate clams. No MAA was detected in the tissues of a deep-sea bivalve, Calyptogena soyoae. Although MAAs seem to block strong UV radiation in the shallow-water clam, they are probably not essential for the clam's life in the dark. MAAs in shallow-water clams may be derived from food and accumulated in their tissues, especially in the siphonal mantle and kidney. Received: 29 November 1996 / Accepted: 13 January 1997     

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     

Movements of tiger sharks (Galeocerdo cuvier) in coastal Hawaiian waters

The giant clam Tridacna crocea harbors in the mantle tissue symbiotic microalgae commonly called zooxanthellae. Isolated zooxanthellae release glycerol into the medium in the presence of mantle tissue homogenate (MH), but it is not clear whether the cells do so in situ. In order to determine the photosynthetic products released by zooxanthellae in the mantle of the giant clam we traced photosynthetic fixation products from ¹³C- and ¹⁴C-bicarbonate both in the clam and in isolated zooxanthellae (IZ) in the presence or absence of MH. After 15 min incubation in the absence of MH the IZ released less than 0.6% of the fixed labeled carbon, mainly as glucose. The major intracellular photosynthates were neutral lipids, which constituted 20 to 40% of the total extractable ¹⁴C. In the presence of MH, the IZ released up to 5.6% of the total fixed ¹⁴C, mostly as glycerol, and the major intracellular photosynthate was glucose. In an intact clam incubated in sea water containing ¹⁴C-bicarbonate, 46 to 80% of the fixed ¹⁴C was translocated from the zooxanthellae to the host tissues. Most of the ¹⁴C in the hemolymph, in the isolated zooxanthellae and in intact mantle tissue (containing zooxanthellae) was recovered as glucose. No ¹⁴C-glycerol was detected in the mantle after 1 to 30 min incubation, and, even after 60 min, far less ¹⁴C-glycerol was synthesized than by IZ in the presence of MH. The possibility that in clam tissue glycerol is converted to glucose was examined by tracing the labeled carbon from ¹⁴C-glycerol injected into the adductor muscle. After 5 min incubation, no labeled glucose was found in the hemolymph, but after 60 min, some 20% was found as glucose. Thin slices containing zooxanthellae, cut from the surface of the mantle, fixed inorganic carbon supplied as NaH¹⁴CO₃ in the medium and mainly released ¹⁴C-glucose. The addition of MH to the surrounding medium did not affect the release rate or form of release product. When the slices were cut into smaller pieces, however, the ratio of glycerol to glucose in the release product increased. These results indicate that in the presence of MH the metabolism of isolated zooxan- thellae was different from that of zooxanthellae in the mantle. In the presence of MH, isolated zooxanthellae release mostly glycerol, whereas in the mantle they release glucose. Received: 18 February 1998 / Accepted: 4 December 1998     

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     

The oxygen microenvironment adjacent to the tissue of the scleractinian Dichocoenia stokesii and its effects on symbiont metabolism

The biology of symbiotic scleractinians is profoundly influenced by their intracellular zooxanthellae, and many studies have focused on the mechanistic basis of this influence. This has usually been accomplished by examining the metabolism of zooxanthellae under physical conditions measured in the open reef and assumed to be similar to conditions in hospite. Recent advances in the measurement of conditions near and within coral tissue suggests that this assumption may result in substantial errors. To address this possibility, the role of water flow in determining oxygen saturation adjacent to the tissue of Dichocoenia stokesii was investigated, and the effect of these measured oxygen saturations on the respiration and photosynthesis of zooxanthellae isolated from the same species was quantified. Using a microelectrode (700 μm diam), we measured oxygen saturations above (≤4 mm) the tissue in two flow speeds over 24 h periods in a flume receiving sunlight at in situ levels. The results were used as a proxy for ecologically relevant intracellular oxygen saturations, which were applied to zooxanthellae in vitro to assess their effect on symbiont metabolism. Microenvironment oxygen saturations (% air saturation) ranged from 74–159% in slow flow (2.7 cm s⁻¹) to 88–110% in faster flow (7.5 cm s⁻¹) over day–night cycles. Therefore, the metabolic rates of zooxanthellae were measured at 50 to 54% (hypoxia), 98 to 102% (normoxia) and 146 to 150% (hyperoxia) oxygen saturation. Oxygen saturation significantly affected the metabolism of zooxanthellae, with gross photosynthesis increasing 1.2-fold and dark respiration increasing 2-fold under hyperoxia compared to hypoxia. These results suggest that the metabolism of zooxanthellae in hospite is affected markedly by their microenvironment which, in turn, is influenced by flow-mediated mass transfer. Received: 13 July 1998 / Accepted: 30 April 1999     

Growth and productivity of the high Antarctic Bryozoan Melicerita obliqua

We analysed growth of the Antarctic bryozoan Melicerita obliqua (Thornely, 1924) by x-ray photography and stable isotope analysis. M. obliqua colonies form one segment per year, thus attaining maximum length of about 200 mm within 50 years. In the Weddell and Lazarev Seas, annual production/biomass ratio of M. obliqua is 0.1 yr⁻¹, which is in the range of other Antarctic benthic invertebrate populations. Production amounts to 3.34 mg C_orgm⁻² yr⁻¹ and 90.6 mg ash m⁻² yr⁻¹ on the shelf (100 to 600 m water depth), and to 0.13 mg C_orgm⁻² yr⁻¹ and 36.8 mg ash m⁻² yr⁻¹ on the slope (600 to 1250 m water depth). Received: 27 February 1998 / Accepted: 8 May 1998     

Light and carbon balance in the seagrass Thalassia testudinum : evaluation of current production models

The production dynamics and carbon balance of Thalassia testudinum in the lower Laguna Madre, Texas, USA, were examined during the 1995 summer period based on in situ photosynthesis vs irradiance (PI) measurements and continuous measurements of underwater photon-flux density (PFD). The validity of applying the H _sat model, used to calculate production for Zostera marina as the product of the maximum rate of photosynthesis (P _max) and daily hours of saturating irradiance (H _sat) was assessed for T. testudinum by comparison with integrated production estimates derived through numerical integration. Gross integrated production values were combined with dark-respiration measurements of photosynthetic (PS) and non-photosynthetic (NPS) tissues and areal biomass to generate daily whole-plant carbon balance. Production and whole-plant carbon balance are discussed in relation to surface and underwater PFD measurements, biomass and other physical and chemical parameters collected during a 1 yr period from January to December 1995. The H _sat model significantly underestimated production during all summer months, averaging 70% of integrated production over the entire study period. Gross integrated production ranged between 11.5 mg C g⁻¹ leaf dry wt d⁻¹ in June (during a period of unseasonably low PFDs caused by a drift-alga mat covering the seagrass bed) to 26.7 mg C g⁻¹ leaf dry wt d⁻¹ in July. Modeled net carbon gain was highest in July at 454 mg C m⁻² d⁻¹ (1.4 g dry wt m⁻² d⁻¹), sufficient to account for measured rates of leaf production in the study area and representative of T. testudinum populations of low productivity. During part of the summer period, however, the population was in negative carbon balance. The relatively low productivity of this population and the periods of negative carbon balance are attributed to low net photosynthesis:dark respiration (P _net:R _d) ratios, sporadic low-light periods, the small fraction of PS tissue relative to whole-plant biomass (5 to 13%) and nutrient limitation. Production models are sensitive to both light availability and the proportion of PS tissue supporting NPS biomass as reflected in whole-plant P _net:R _d ratios. Received: 13 August 1997 / Accepted: 6 March 1998     

Interactions between NH+4 and NO−3 uptake and assimilation: comparison of diatoms and dinoflagellates at several growth temperatures

Ammonium concentrations of ∼1 M are commonly cited as being the threshold for inhibition of NO₃ ⁻ uptake, but the applicability of this threshold to phytoplankton from different taxonomic classes has rarely been examined. Additionally, little is known about the influence of environmental variables (e.g. growth temperature) on the interaction between ambient NH₄ ⁺ and NO₃ ⁻ uptake. Four species of estuarine phytoplankton, two diatom [Chaetoceros sp., and Thalassiosira weissflogii (Grunow) Fryxell et Hasle] and two dinoflagellate [Prorocentrum minimum (Pavillard) Schiller, and Gyrodinium uncatenum Hulburt], were grown on NO₃ ⁻ at several different temperatures (4, 10, 15, or 20 °C), and the impact of NH₄ ⁺ additions on NO₃ ⁻ uptake/assimilation (non-TCA-extracted) and assimilation (TCA-extracted) was assessed. For all species at all temperatures, NO₃ ⁻ uptake/assimilation and assimilation rates decreased in a roughly exponential manner with increasing NH₄ ⁺ concentrations but were not completely inhibited even at elevated NH₄ ⁺ concentrations of 200 μM. Estimated half-inhibition concentrations (K _i) were significantly greater in the diatom species (mean ± SE; 2.70 ± 0.67 μM) than in the dinoflagellate species (1.26 ± 0.55 μM). Half-inhibition constants were positively related to temperature-limited relative growth rate although not significantly. The observed inhibition of NO₃ ⁻ uptake and assimilation, as a percentage of NO₃ ⁻ uptake in the absence of NH₄ ⁺, averaged about 80% and ranged from 49 to 100%. For all species, a significant (P < 0.001) positive correlation was found between percent inhibition of NO₃ ⁻ assimilation and temperature-limited relative growth rate. Two experiments on Chesapeake Bay phytoplankton during an April 1998 diatom bloom showed that in short-term (∼1 h) temperature manipulation experiments, percent inhibition of NO₃ ⁻ uptake/assimilation was also positively related (P = 0.05) to experimental temperature. The observed relationships between temperature-limited relative growth rate and percent inhibition of NO₃ ⁻ assimilation rates for the species tested suggest that at the enzyme level, the inhibitory mechanism of NO₃ ⁻ assimilation is similar among species, but at the whole cell level may be regulated by species-specific differences in the accumulation of internal metabolites. These findings add not only to our understanding of species-specific variability and the role of growth temperature, but also provide additional data with which to evaluate current models of NH₄ ⁺ and NO₃ ⁻ interactions. Received: 31 August 1998 / Accepted: 7 December 1998     

The amphipod Corophium multisetosum (Corophiidae) in Ria de Aveiro (NW Portugal). II. Abundance, biomass and production

 The abundance and biomass of Corophium multisetosum Stock, 1952 were determined from benthic corer samples collected monthly over 1 yr in the upper reaches of Canal de Mira (Ria de Aveiro, Portugal). Both density and biomass over the sampling period were negatively correlated with water temperature and positively correlated with chlorophyll a concentration in the sediment. C. multisetosum density was significantly negatively correlated with plant biomass and positively correlated with salinity. The nature of the sediment, favourable environmental conditions, high availability of food and low interspecific competition allowed the population to reach a maximal density of 200 × 10³ individuals m⁻² and a maximal biomass (ash-free dry wt, AFDW) of 62 g_AFDW m⁻². The population was highly productive, especially during the autumn/winter period. Production, estimated by two different methods (Hynes method: 251 g_AFDW m⁻² yr⁻¹; Morin–Bourassa method: 308 g_DW m⁻² yr⁻¹), was much higher than the values reported for other Corophium species. The annual P:Bˉ ratio (10) was high, but similar to values reported for Swedish populations of C. volutator and lower than the values estimated from Mediterranean populations of C. insidiosum. Received: 8 October 1999 / Accepted: 22 June 2000     

Effect of an organophosphorus insecticide, fenitrothion, on survival and osmoregulation of various developmental stages of the shrimp Penaeus japonicus (Crustacea: Decapoda)

The toxicity of fenitrothion was determined in larvae (nauplii, Zoeae 1 to 3, Mysis 1 to 3), postlarvae (PL stages) and juvenile shrimp (Penaeus japonicus Bate), in two media, seawater (SW) and diluted seawater (DSW) (1100 and 550 mosM kg⁻¹, ≃ 37 and 19‰ S). The effects of fenitrothion on the osmoregulatory capacities (OC) of juveniles were recorded. A gill and epipodite histopathological study was also conducted. For larvae in seawater, 24 and 48 h LC₅₀s ranged from 32.9 μg l⁻¹ (Zoeae 2) to 10.7 μg l⁻¹ (Mysis 3), and from 3.9 μg l⁻¹ (Zoeae 3) to 2.0 μg l⁻¹ (Mysis 3), respectively; 48 and 96 h  LC₅₀s in postlarvae (PL) at the same salinity ranged from 1.8 μg l⁻¹ (PL1) to 0.6 μg l⁻¹ (PL5), and from 0.3 μg l⁻¹ (PL7) to 0.4 μg l⁻¹ (PL15). In juveniles, 96 h LC₅₀s were 0.8 μg l⁻¹ in seawater and 1.5 μg l⁻¹ in diluted seawater. From hatching to juvenile stages, the overall trend was a rapid decrease (from nauplii to PL5–PL7) followed by a slight increase (from PL7 to PL15 and juveniles) in the shrimp's ability to tolerate the insecticide. In juveniles kept in seawater and in diluted seawater, fenitrothion decreased the osmoregulatory capacity (OC = difference between the hemolymph osmotic pressure and the osmotic pressure of the medium) at both lethal and sublethal concentrations. This effect was time- and dose-dependent. In SW, the decrease in hypo-OC was ˜ 25% at sublethal concentrations and ˜ 35% at the 96 h LC₅₀. In DSW, the decrease in hyper-OC was ˜ 10 to 15% at sublethal concentrations. In SW, shrimp were able to recover their OC in less than 48 h when transferred to water free of pesticide. In DSW, recovery at 48 h was only possible after exposure to the lowest tested sublethal concentration. Haemocytic congestions (thrombosis) of the gills, lamellae necrosis and other alterations of gills and epipodites (breakage of the cuticle, reduction of the hemolymph lacunae) were noted in juveniles exposed to lethal and sublethal concentrations of fenitrothion. Received: 7 October 1996 / Accepted: 13 November 1996     

Activity and inhibitor sensitivity of ATPases in the hydrothermal vent tubeworm Riftia pachyptila : a comparative approach

Phosphorylated ATPases may be involved in the effective pH regulation seen in the hydrothermal vent tubeworm Riftia pachyptila. R. pachyptila appears not only to have a large concentration of ATPases, but the main function of these ATPases seems to have shifted from other types of transport, such as Na⁺ and K⁺ movement, to the facilitation of H⁺ elimination. Plume and trophosome ATPase activity for R. pachyptila measured 646.2 ± 29.5 and 481.4 ± 32.0 μmol Pi (inorganic phosphate) g⁻¹ wet wth⁻¹, respectively. Plume tissue ATPase activity (both mass-specific and protein-specific) in R. pachyptila was higher (between 7% and 55%) than the activity measured in any tissue for 7 other shallow- and deep-living species, in this study. This supports the hypothesis that R. pachyptila regulates acid/base balance via high concentrations of H⁺-ATPases, including Na⁺/H⁺ and K⁺/H⁺ exchangers and possibly electrogenic H⁺-ATPases, as evidenced by a higher total ATPase concentration (646 μmol Pi g⁻¹ wet wt h⁻¹), lesser Na⁺/K⁺-ATPase activity (13% of the total, as compared to 20−40% found in other animals), and higher H⁺-ATPase activity (226–264 μmol Pi g⁻¹wet wt h⁻¹). Overall, R. pachyptila appears to demonstrate elevated ATPase activity, with a greater fraction of the enzymes devoted to proton elimination, in order to effectively control its extracellular pH in the face of processes acting to acidify the internal environment. Received: 9 May 2000 / Accepted: 4 October 2000     

A sensitive method for determining nitrogen-15 isotope in urea

A method is presented in which the ¹⁵N at.% of urea is determined with high precision on liquid samples containing as little as 10 nmol of urea. The method involves removal of interference from NH₄ ⁺ initially present in the sample by cation exchange. Urea in the sample is subsequently hydrolyzed to NH₄ ⁺ by the enzyme urease. Liberated NH₄ ⁺ is separated from the alkaline sample by diffusion as NH₃ through a helium gas phase where it is finally oxidized to N₂ by reaction with hypobromite iodine. The isotopically labeled N₂ thus formed is mixed with the N₂ initially present in the sample, and the ¹⁵N at.% of urea is calculated from the relative amounts of ¹⁴N¹⁵N and ¹⁵N¹⁵N by means of the isotope pairing principle. The presented method for ¹⁵N-urea analysis proved to be precise (SE < 0.4 at.%, n = 5), when applied on both marine and freshwater samples with an ¹⁵N enrichment >1 at.%, and interference was found only from volatile methyl amines. Received: 19 September 1996 / Accepted: 25 October 1996     

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     

Effect of temperature fluctuations and food supply on the growth and metabolism of juvenile sea scallops (Placopecten magellanicus)

On the eastern shore of Nova Scotia late summer atmospheric systems cause upwelling of shelf water; the associated temperature variations of 10 °C with a 6 to 8 d period are comparable in magnitude to the seasonal variation. A laboratory study was undertaken to assess the effects of these temperature fluctuations on sea scallop (Placopecten magellanicus) growth and metabolism. In a factorial design, scallops were subjected to constant (10 °C) or a variable (6 to 15 °C) 8 d temperature cycle, and either a low (seston in filtered seawater) or high (seston supplemented with cultured phytoplankton) food diet. During the 48 d experiment scallop mortality was low and growth positive in all treatments. Shell and total tissue growth rate did not differ between temperature treatments, but growth in the high food treatments was 40 to 50% higher than in the low food treatments. However, soft tissue (excluding adductor) growth did show a temperature treatment effect; growth rates were significantly higher in the fluctuating temperature treatment, due in part to greater gonad development. Weight-standardized rates of scallop oxygen consumption (V _sO₂ , μmol O₂ g⁻¹ h⁻¹) were 20 to 25% higher in high food than in low food treatments, consistent with the expected increase in respiration due to the higher growth rates. Scallop metabolism did not acclimate to the fluctuating temperature cycle; V _sO₂ and ammonium excretion (V _sNH⁺ ₄, μmol O₂ g⁻¹ h⁻¹) remained dependent on ambient temperature throughout the experiment. V _sNH⁺ ₄ Q₁₀ (2.77) was higher than V _sO₂ Q₁₀ (2.01) which was reflected in a decrease in the O:N ratio at 15 °C, indicating a shift toward increased protein catabolism and a stressed state. At 10 °C, V _sO₂ and V _sNH⁺ ₄ in the variable temperature treatments were 15 to 18% lower than in the constant temperature treatments, a difference that was not detected in growth measurements. Results demonstrate that the metabolism of Placopecten magellanicus, unlike some bivalve species, is tightly coupled to fluctuations in ambient temperature. Although an absence of compensatory acclimation had a minimal effect on growth in this study, if high temperatures were combined with low food conditions a reduction in scallop production could result. Received: 23 June 1998 / Accepted: 8 February 1999     

Inbreeding in a lek-mating ant species, Pogonomyrmex occidentalis

In this paper we have two goals. First, we examine the effects of sample size on the statistical power to detect a given amount of inbreeding in social insect populations. The statistical power to detect a given level of inbreeding is largely a function of the number of colonies sampled. We explore two sampling schemes, one in which a single individual per colony is sampled for different sample sizes and a second sampling scheme in which constant sampling effort is maintained (the product of the number of colonies and the number of workers per colony is constant). We find that adding additional workers to a sample from a colony makes it easier to detect inbreeding in samples from given number of colonies; however, adding more colonies rather than more workers per colony always gives greater power to detect inbreeding. Because even relatively large amounts of sib-mating generate relatively small inbreeding coefficients, detection of even substantial deviations from random mating will require very large samples. Second, we look at the amount of inbreeding in a large population of the western harvest ant, Pogonomyrmex occidentalis. We find deviations from Hardy-Weinberg equilibrium equivalent to approximately 27% sib-mating in our population ( f = 0.09). Review of past studies on the population structure of other Pogonomyrmex species suggests that inbreeding may be a regular feature of the mating system of these ants. Although P. occidentalisis a swarm-mating species, there are a number of features of its population biology which suggest that the effective population size may be small. These include topographical variation that potentially breaks the population into demes, variation in the reproductive output of colonies, and variation in the size of reproductives produced by colonies. Received: 6 May 1996 / Accepted after revision: 6 October 1996