Response of muscle-based biochemical condition indices to short-term variations in food availability in post-flexion reared sea bass Dicentrarchus labrax (L.) larvae

Six condition indices based on RNA, total soluble protein and two metabolic enzymes [lactate dehydrogenase (LDH) and citrate synthase (CS)] were analysed in muscle tissue of individual larvae, post-flexion reared sea bass Dicentrarchus labrax using DNA and total soluble protein as standards for size. In addition, the effect of 2 days of food deprivation on the cell proliferation rates was assessed. The RNA:DNA best reflected short-term changes in feeding conditions. If standardized by DNA content, LDH activity was a better indicator of condition than any other index but RNA:DNA. Further, the analysis of cell proliferation rates in muscle from 26 day-old larvae proved useful in distinguishing continuously fed larvae from individuals subjected to 2 days of fast.     

Distribution of epiplanktonic Chaetognatha along a transect in the Indian Ocean

Chaetognaths were identified and counted in 23 samples collectedin the west Indian Ocean along a transect from the Gulf of Adento the Cape of Good Hope, in February–March 1967. Althoughthe hydrographic front located at 5°S does not seem to representa barrier for the distribution of the majority of the species,some of them are in a preferential area south or north of thefront. The principal component analysis indicates the locationswhere the more intense overlapping of species takes place. Itis important to mention the occurrence of S.pseudoserratodentatain the proximity of Madagascar Island.     

Variations in biochemical parameters of Heterocapsa sp. and Olisthodiscus luteus grown in 12:12 light:dark cycles II. Changes in pigment composition

Photosynthetic pigment composition was studied in batch cultures of Heterocapsa sp. and Olisthodiscus luteus growing exponentially in a 12:12 light:dark cycle. Both species divided in the dark. The synthesis of pigments was continuous for both species. However for chlorophyll c and peridinin, in Heterocapsa sp., and chlorophyll c and fucoxanthin, in O. luteus, (pigments belonging to light harvesting complexes) the synthesis was significantly higher during the light period. Concentrations per total cell volume (TCV) of chlorophyll a, chlorophyll c, peridinin and diadinoxanthin in Heterocapsa sp., and chlorophyll a, chlorophyll c, fucoxanthin and violaxanthin in O. luteus, showed a maximum at the onset of light and decreased during the light period. The values of the chlorophyll a:chlorophyll c, chlorophyll a:peridinin and chlorophyll a:fucoxanthin ratios are compared with data reported in the literature.     

DETERMINATION OF RNA AND DNA CONCENTRATIONS IN NATURAL PLANKTON SAMPLES USING THIAZOLE ORANGE IN COMBINATION WITH DNASE AND RNASE DIGESTIONS1

A fluorometric technique, based on the combination of RNase and DNase incubation with the use of thiazole orange (RNase/DNase method), was investigated to determine DNA and RNA concentrations in marine plankton. Tests were performed to optimize both RNase and DNase assay conditions. The RNase assay should be conducted at 37° C for 20 min with 0.5 μg·mL^?1 of DNase-free RNase. An incubation at 25° C for 20 min with 10 units ·mL^-1 of RNase-free DNase were the optimal conditions required for DNA digestion by DNase. The detection limits in terms of minimum biomass for reliable measurements of DNA and RNA were 7.5 and 10 μg of protein · (mL assay)^?1, respectively. RNA and DNA concentration were estimated in oligotrophic water samples using the RNase/DNase and other available methods (e.g. a double fluorochrome method). The different techniques provided similar DNA estimations. However, the RNase/DNase method provided the highest sensitivity and a low variability for the estimation of RNA.     

CELL CYCLE AND CELL MORTALITY OF ALEXANDRIUM MINUTUM (DINOPHYCEAE) UNDER SMALL‐SCALE TURBULENCE CONDITIONS1

Decreased net population growth rates and cellular abundances have been observed in dinoflagellate species exposed to small‐scale turbulence. Here, we investigated whether these effects were caused by alterations in the cell cycle and/or by cell mortality and, in turn, whether these two mechanisms depended on the duration of exposure to turbulence. The study was conducted on the toxic dinoflagellate Alexandrium minutum Halim, with the same experimental design and setup used in previous studies to allow direct comparison among results. A combination of microscopy and Coulter Counter measurements allowed us to detect cell mortality, based on the biovolume of broken cells and thecae. The turbulence applied during the exponential growth phase caused an immediate transitory arrest in the G2/M phase, but significant mortality did not occur. This finding suggests that high intensities of small‐scale turbulence can alter the cell division, likely affecting the correct chromosome segregation during the dinomitosis. When shaking persisted for >4 d, mortality signals and presence of anomalously swollen cells appeared, hinting at the activation of mechanisms that induce programmed cell death. Our study suggests that the sensitivity of dinoflagellates to turbulence may drive these organisms to find the most favorable (calm) conditions to complete their division cycle.     

Particulate DNA and protein relative to microorganism biomass and detritus in the Catalano-Balearic Sea (NW Mediterranean) during stratification

Using microscopic and biochemical approaches, the relative contribution of the main groups of pelagic microorganisms (bacteria, heterotrophic nanoflagellates, phytoplankton and ciliates) and detritus (<150 m) to total particulate protein and DNA was investigated at two stations of the Catalano-Balearic Sea (NW Mediterranean) during the stratified period. The two stations, one located in the shelf break front (S) and the other in the open sea, above the central divergence zone (D), were sampled twice in early summer 1993. Both of them showed a well-developed deep chlorophyll maximum (DCM). Maximum DNA concentrations were observed close to the DCM, while protein concentrations were fairly homogeneous from the surface to 60 m depth in all samplings. In general, the microorganism distribution showed maximum concentrations at or near the DCM depths. At both stations, bacteria were the most important contributors to living particulate DNA (22.5-32.6%), while phytoplankton and heterotrophic nanoflagellates were the main contributors to living particulate protein (3.8-24.4 and 2.9-29.1%, respectively). In addition, an important amount of detrital DNA and protein was estimated to occur at both stations. Detrital DNA accounted for 23.9-42.9% of the particulate DNA, while detrital protein represented from 63.5 to 84.7% of the particulate protein. Because both protein and DNA contain nitrogen and DNA is also a phosphorus source, these results indicate that heterotrophic organisms and detrital particles play an important role in the nitrogen and phosphorus cycles in the open sea waters of the NW Mediterranean.      

DETECTION OF THE TOXIC DINOFLAGELLATE ALEXANDRIUM FUNDYENSE (DINOPHYCEAE) WITH OLIGONUCLEOTIDE AND ANTIBODY PROBES: VARIABILITY IN LABELING INTENSITY WITH PHYSIOLOGICAL CONDITION

The toxic dinoflagellate Alexandrium fundyense Balech was grown under temperature- and nutrient-limited conditions, and changes in labeling intensity on intact cells were determined for two probe types: an oligonucleotide probe targeting rRNA and a monoclonal antibody (MAb) targeting a cell surface protein. In nutrient-replete batch culture, labeling with the rRNA probe was up to 400% brighter during exponential phase than during stationary phase, whereas MAb labeling did not change significantly with growth stage at the optimal growth temperature. In cultures grown at suboptimal, low temperatures, there was a significant difference between labeling intensity in stationary versus exponential phase for both probe types, with exponential cells labeling brighter with the rRNA probe and slightly weaker with the MAb. The decrease in rRNA probe labeling with increasing culture age was likely due to lower abundance of the target nucleic acid, as extracted RNA varied in a similar manner. With the MAb and the rRNA probes, slower growing cultures at low, nonoptimal temperature labeled 35% and 50% brighter than cells growing faster at warmer temperatures. Some differences in labeling intensity per cell disappeared when the data were normalized to surface area or volume, which indicated that the number of target antigens or rRNA molecules was relatively constant per unit area or volume, respectively. Slow growth accompanying phosphorus and nitrogen limitation resulted in up to a 400% decrease in labeling intensity with the rRNA probe compared to nutrient-replete levels, whereas the MAb labeling intensity increased by a maximum of 60%. With both probes, labeling was more intense under phosphorus limitation than under nitrogen limitation, and for all conditions tested, labeling intensity was from 600% to 3600% brighter with the MAb than with the rRNA probe. Thus, it is clear that significant levels of variability in labeling intensity can be expected with both probe types because of the influence of environmental conditions and growth stage on cellular biochemistry, cell size,rRNA levels, and the number or accessibility of cell surface proteins. Of the two probes tested, the rRNA probe was the most variable, suggesting that in automated, whole-cell assays, it can be used only in a semiquantitative manner. For manual counts, the human eye will likely accommodate the labeling differences. The MAb probe was less variable, and thus should be amenable to both manual and automated counts.     

Effects of nitrogen and phosphorus starvation on nucleic acid and protein content of Heterocapsa sp.

Changes in the protein, RNA and DNA content related to nitrogen(N) and phosphorus (P) starvation were studied in the marinedinoflagellate Heterocapsa sp. grown in batch cultures. In bothcases of nutrient starvation, metabolic adaptations affectedprotein and RNA pools, while the DNA content per cell remainedapproximately constant. N starvation led to a parallel decreasein protein and RNA concentration which caused the protein/RNAratios to remain constant. A dramatic decrease in the RNA contentcharacterized the P-starved cultures, although protein synthesiscontinued. The ribosomal RNA content was lower than expectedgiven the continuation of protein synthesis. It is suggestedthat protein/RNA ratios could be used as an indicator of P starvation,while protein/chlorophyll ratios would characterize N starvation ¹Present address: University of Hawaii at Manoa Soest, BiologicalOceanography, 1000 Pope Road, Honolulu, HI 96822, USA     

Variations in biochemical parameters of Heterocapsa sp. and Olisthodiscus luteus grown in 12:12 light:dark cycles I. Cell cycle and nucleic acid composition

The division cycle of two phytoplankton species, Olisthodiscus luteus and Heterocapsa sp. was studied in relation to a 12:12 light:dark cycle. Batch cultures in exponential phase were sampled every three hours during 48 hours. Cell number, cellular volume and DNA and RNA concentrations were measured. Microscopic observations of the nuclei of Heterocapsa sp. were also performed. In both species, cell division took place in the dark. In Heterocapsa sp., DNA and RNA showed a similar diel variability pattern, with synthesis starting at the end of the light period, previously to mitosis and cytokinesis. In O. luteus. Major RNA synthesis occurred during darkness, and DNA was produced almost continuously. Both species presented different values and diel rhythmicity on the RNA/DNA ratios.     

Species‐specific physiological response of dinoflagellates to quantified small‐scale turbulence1

Turbulence has been shown to alter different aspects of the physiology of some dinoflagellates. The response appears to be species‐specific and dependent on the experimental design and setup used to generate small‐scale turbulence. We examined the variability of the response of three dinoflagellate species to the turbulence, following the same experimental design used by Berdalet (1992) on Akashiwo sanguinea (Hirasaka) Ge. Hansen et Moestrup (=Gymnodinium nelsonii G. W. Martin). In all experiments, turbulence was generated by an orbital shaker at 100 rpm, which corresponded on bulk average, to dissipation rates (ε, quantified using an acoustic Doppler velocimeter) of ≈2 cm² · s^?3. Turbulence did not appreciably affect Gymnodinium sp., a small dinoflagellate. However, Alexandrium minutum Halim and Prorocentrum triestinum J. Schiller exhibited a reduced net growth rate (33% and 28%, respectively) when shaken during the exponential growth phase. Compared to the still cultures, the shaken treatments of A. minutum and P. triestinum increased the mean cell volume (up to 1.4‐ and 2.5‐fold, respectively) and the mean DNA content (up to 1.8‐ and 5.3‐fold, respectively). Cultures affected by turbulence recovered their normal cell properties when returned to still conditions. The swimming speed of the cells exposed to agitation was half that of the unshaken ones. Overall, the response of A. minutum and P. triestinum was similar, but with lower intensity, to that observed previously on A. sanguinea. We found no clear trends related to taxonomy or morphology.