Changes in amino acid content of an algal feed species (Navicula sp.) and their effect on growth and survival of juvenile abalone (Haliotis rubra)

The growth and survival of juvenile Haliotis rubra, when fed with the diatom Navicula sp. cultured in f/2 medium containing combined nitrogen at 24.71 mg NO₃-N L^–1 (high), 12.35 mg NO₃-N L^–1 (standard) or 2.47 mg NO₃-N L^–1 (low), were compared in a 33-day trial. The alga in the low nitrogen medium contained 37% less total amino acid than that in the high and standard nitrogen media. There was a slightly greater reduction in essential amino acids (40%) compared to non-essential amino acids (35%). Juvenile abalone feeding on Navicula grown in medium with low nitrate and lower total amino acid content grew more slowly than when fed on the same species grown in standard or higher nitrogen medium with a higher amino acid content. The growth rate of juveniles was highest (43 m d^–1) in the high nitrate treatment followed (40 m d^–1) by the standard nitrate treatment and lowest (31 m d^–1) in the low nitrate treatment. The survival of the juveniles was also effected by the diet. Survival was better in the high and standard nitrogen media (88%) than the low nitrogen medium (75%). The results suggest that in order to achieve uniformity in nutritional quality of diatoms and good growth of abalone juveniles in commercial abalone nurseries, the nitrogen concentration in tanks should be monitored and additional nitrate added to provide an optimum concentration of between 2 and 12 mg NO₃-N L^–1.     

Growth ofEnteromorpha linza in sewage effluent and sewage effluent-seawater mixtures

Enteromorpha linza (L.) J. Ag. was grown in full strength sewage effluent, various combinations of sewage effluent and seawater, and in natural seawater. It was found that full strength sewage effluent with a salinity of 14 supported best growth of the alga. After a 12 day cultivation period, growth ofE. linza in full strength sewage effluent and 75% sewage effluent- seawater mixture showed 3.5-fold and 2-fold increase in fresh weight over that grown in natural seawater; respectively. Uptake of PO _inf4 ^sup3– -P, NH₃-N and NO _inf3 ^sup– -N by cells ofE. linza was extremely efficient in all tested media. Data obtained from the experiments indicated that inorganic nitrogen rather than phosphorus was the limiting nutrient factor for growth ofE. linza in full strength sewage effluent and in other sewage effluent- seawater mixtures. NH₃-N at concentrations above 4.5 ppm was found to inhibit uptake of NO _inf3 ^sup– -N in the same culture medium by the algal cells. The fact that sewage grownE. linza contained comparatively much higher protein content (30.2% dry weight) than that grown in natural seawater (12.5% dry weight) leads to the conclusion that sewage grownE. linza could serve as an economically feasible feed for livestock in Hong Kong where the sewage is characterized by having a salinity of approximately 14. It is proposed that this multicellular green alga is a suitable algal species to serve the dual function of wastewater purification through the production of algal protein from sewage effluent having high salinities.     

Attempts to produce solasodine in callus and suspension cultures of Solanum mauritianum Scop.

Callus cultures of Solanum mauritianum Scop. were initiated from green berry explants on a hormone-free Murashige and Skoog (1962) medium excluding glycine, and containing 0.1 g L^–1 myo-inositol and 3% sucrose. Such cultures contained 10.08±0.59 g g^–1 DW of solasodine, which is equivalent to that in the leaves of mature S. mauritianum plants, but far less than that extracted from the green berries (185 g g^–1 DW). In vitro solasodine productivity could be increased by reducing the strength of the medium by half, substituting 3% glucose for 3% sucrose as carbon source, or by the addition of certain combinations of BA and NAA. Phosphate limitation and alterations in the carbon: nitrogen ratio were not able to increase solasodine productivity. Suspension cultures of S. mauritianum were initiated and maintained in a Murashige and Skoog (1962) medium with the RT vitamins of Khanna and Staba (1968), 0.1 g L^–1 myo-inositol, 3% sucrose and 1 mg L^–1 2,4-D. No solasodine was detectable in these cultures, or slight modifications thereof.Abbreviations BA benzyladenine - NAA naphthaleneacetic acid - 2,4-D 2,4-dichlorophenoxyacetic acid - MS Murashige and Skoog's (1962) medium     

Outdoor pond cultivation of the subtropical marine red algaGracilaria tenuistipitata in brackish water in Sweden. Growth,nutrient uptake,co-cultivation with rainbow trout and epiphyte control

Pond cultivation of the subtropical, euryhaline macroscopic red algaGracilaria tenuisipitata var.liui Zhanget Xia was carried out in brackish seawater (6–7) in the Gryt archipelago on the east coast of Sweden, using four outdoor tanks of 30–40 m³. Growth rate and nutrient uptake in batch culture were measured with the aim of estimating the water purification capacity ofG. tenuisipitata in outdoor conditions. Its ability to withstand epiphytic infections was also studied. An average growth rate of 4 biomass increase per day was recorded during two seasons with a maximum growth rate of 9 d^–1. The initial biomass was usually 1 kgFW m^–3 (FW, fresh weight). The nutrient uptake capacity was on average ca. 1 g N_i kgFW^–1 d^–1 and 0.08 g P_i kgFW^–1 d^–1 and the uptake rates for NH₄ ⁺-N were higher than those for NO₃ ^–-N. Both the growth rate and the nutrient uptake rate were highest at the highest water temperature. Co-cultivation with rainbow trout (Oncorhynchus mykiss) was tested: with trout fodder as the only nutrient inputG. tenuistipitata could grow and maintain low levels of N_i and P_i with optimum efficiency at a trout: alga ratio of 1:1 (w:w). Epiphytic growth of filamentous green and brown algae was limited, probably as a result of the high pH values caused by inorganic carbon uptake byG. tenuistipitata. The growth ofEnteromorpha intestinalis, the only significant epiphyte, was completely inhibited and the majority of plants died by a few days treatment with 100 µg 1^–1 Cu²⁺, a concentration that did not severely affectG. tenuistipitata. We conclude thatG. tenuistipitata can be cultivated in outdoor ponds in southern Sweden during 5–6 months of the year using aerated or unaerated batch cultures and that wastewater from trout cultivation may be used as a nutrient source, resulting in purification with respect to N and P.     

Influence of temperature and nitrogen concentration on photosynthesis ofDunaliella viridis Teodoresco

The photosynthetic behaviour ofDunaliella viridis has been studied under a combination of three variables: irradiance (0–900 mol m^–2 s^–1), temperature (15, 23, 31, 38, 42 °C) and nitrogen concentration (0.05, 0.5, 1.5, 5, 10 mM NO ₃ ^- ) at a salinity of 2 M NaCl.The highest rates of photosynthesis have been found at 31 °C and a nitrate concentration of 10 mM. There exists a synergistic effect between temperature and nitrogen availability on the photosynthesis ofD. viridis; under nitrogen deficiency oxygen evolution is low, even null at high temperature. The interaction between these two variables of control occurs in a multiplicative way. There is also a general increase in photosynthetic pigments following the increase in nitrogen concentration in the culture medium. The normalization of net photosynthesis data in relation to chlorophylla shows that nitrogen concentration makes an indirect control of the photosynthetic rate ofD. viridis through the variation of pigment concentration.     

Some effects of plant growth regulators on tissue cultures of the marine red alga Agardhiella subulata (Gigartinales,Rhodophyta)

We examined whether auxins and cytokinins, either singly or in combination, stimulate cell division in tissue cultures of a red seaweed. Our experimental model consisted of filamentous and callus-like growths that developed from cross-sectional discs cut from young branches of Agardhiella subulata. Plant growth regulators were added to the medium to give combinations of an auxin with a cytokinin over a range of concentrations (1 µg L^–1 –10 mg L^–1). Several mixtures of auxins and cytokinins, as well as some single auxins, cytokinins and phenolics, stimulated cell division and growth in the tissue cultures beyond that of controls. The treatments that were effective included: phenylacetic acid/zeatin; phenylacetic acid/6-benzylaminopurine; -naphthaleneacetic acid/zeatin; 2,4,5-trichlorophenoxyacetic acid/6-benzylaminopurine; and indoleacetic acid/kinetin. High concentrations of cytokinins (i.e. 10 mg L^–1) inhibited the regeneration of plants in some of the cell cultures. These results provide further evidence that growth regulators can be used for the tissue culture of seaweeds and for the study of developmental phenomena in these plants.     

Somatic embryogenesis and plantlet regeneration from cell suspension cultures of palmarosa grass (Cymbopogon martinii)

A cell suspension culture was established from nodal callus ofCymbopogon martinii (Roxb.) Wats in a liquid medium containing Murashige and Skoog (1962) basal salts, vitamins, 100 mg 1^–1 myo-inositol and 20 g l^–1 of sucrose (MS) that was supplemented with 13.6 M 2,4-dichlorophenoxyacetic acid and 1.15 M kinetin. An initial inoculum density of 2 x 10⁴ cells ml^–1exhibited optimum cell growth. Calli were obtained 12–15 days after the suspension was plated onto semisolid medium of a similar composition. When calli were transferred to semisolid regeneration medium containing MS + 6.7 M N ⁶-benzyl-adenine + 1.15 M kinetin, somatic embryogenesis and plantlet regeneration occurred after 10–25 days. There was no significant decrease in the regeneration potential of the calli even when the cultures were initiated from 47-week-old cell suspensions. Chromosome counts of cells in suspensions, calli and somatic embryos derived from cultures of different ages revealed the presence of diploids, tetraploids and octaploids. However, the 33 regenerated plants tested were all diploid, indicating that only diploid cells were capable of regeneration in vitro.Abbreviations MS Murashige and Skoog (1962) basal salts with vitamins (100 mg1^–1 myo-inositol, 20 g1^–1 sucrose) - 2,4-D 2,4-dichlorophenoxyacetic acid - BA N ⁶-benzyl-adenine - Kn kinetin - MSC MS + 13.6 M 2,4-D + 1.15 M Kn - MSR MS +6.7 M BA + 1.15 M Kn     

Somatic embryogenesis in sisal (Agave sisalana Perr. ex. Engelm)

A protocol has been developed for somatic embryogenesis and plant regeneration of sisal (Agave sisalana Perr. ex. Engelm). Embryogenic callus cultures were initiated from young shoots raised in vitro from the stem portion of the bulbil on medium supplemented with 1–2 mg l^-1 kinetin (KN) and 0.2–0.5 mg l^-1 -naphthaleneacetic acid plus KN or 1–1.5 mg l^-1 benzylaminopurine (BAP) or 0.25–0.5 mg l^-1 2,4-dichlorophenoxyacetic acid plus BAP or 0.5–1.0 mg l^-1 KN. Embryos at various developmental stages (globular-, heart- or torpedo-shaped) produced mature and germinating embryos on being transferred to a new medium containing 0–0.25 mg l^-1 KN. After 28 days, a maximum of 76% germinated embryos was obtained on a medium supplemented with 0.1 mg l^-1 KN. The capacity for embryogenesis remained constant in the callus upon subculturing on the same medium for more than 48 months. Histological observations showed a distinct multicellular origin for most of the somatic embryos as they developed from epidermal, sub-epidermal and inside callus cells, while a few of them originated from a superficial callus cell. Plantlets regenerated from embryos were transferred to the field where their survival rate was 100%.Abbreviations BAP Benzylaminopurine - 2,4-D 2,4-Dichlorophenoxyacetic acid - IAA Indoleacetic acid - KN Kinetin - NAA -Naphthaleneacetic acidCommunicated by W. Barz     

Nickel hyperaccumulation in shoot cultures of Alyssum markgrafii

Shoot cultures of Alyssum markgrafii O.E. Shulz, endemic nickel hyperaccumulating species of central Balkan, were established and maintained on Murashige and Skoog medium supplemented with 0.2 mg dm^–3 benzyladenine (BA). Nickel in form of NiCl₂ . 6 H₂O was supplemented at 22 different concentrations ranging from 0.0001 to 15 mM but none of them was lethal to cultures. High Ni²⁺ concentrations (10 mM or more) arrested shoot growth which, upon transfer to Ni-free medium, commenced via axillary bud proliferation. Shoots that developed from axillary buds through the subculture manifested increased tolerance to Ni²⁺ expressed as shoot elongation. Shoot multiplication and dry biomass production decreased with increase of Ni²⁺ in medium. Only the accumulation of Ni²⁺ in tissues increased with Ni²⁺ content of the medium. Apart from shoot cultures, high Ni²⁺ accumulation was registered in undifferentiated callus cultured on medium with 0.5 mg dm^–3 BA and 0.5 mg dm^–3 naphthylacetic acid. Highest content of accumulated Ni was 2.37 g g^–1 (d.m.) in shoots and 2.65 g g^–1 (d.m.) in callus, both measured on medium with 15 mM Ni²⁺.     

Uptake and release of63Ni2+ byXenopus embryos during early cleavage stages

Summary Uptake and release of⁶³Ni was studied in dejelliedXenopus laevis embryos exposed to⁶³Ni²⁺ (0.3–30 mol/1) for 0.5-h intervals during the period 1–4.5 h post-fertilization (i.e. from first cleavage to early blastula stage). At first cleavage, the mean uptake of⁶³Ni by embryos was 12-17 times that by non-fertilized eggs, suggesting that conversion of the vitelline envelope to the fertilization envelope enhanced integumental permeability to⁶³Ni²⁺. 63 Ni uptake by embryos at the 1-2-cell stage averaged 1.8–2.5 times that at the early blastula stage. An average of 5% of total⁶³Ni in washed embryos was recovered in isolated fertilization envelopes, indicating that⁶³Ni²⁺ passed through the envelope into internal compartments. Progressive increases of⁶³Ni uptake were seen with increasing exposure levels; after exposure during 1–1.5 h post-fertilization to the highest concentration of⁶³Ni²⁺ (30 mol/1),⁶³Ni uptake averaged 11.4 (SD±5.1) pmol/embryo. Rapid efflux of⁶³Ni was noted after⁶³Ni²⁺-exposed embryos were transferred to nickel-free medium; mean⁶³Ni contents at 0.25 h and 2 h post-exposure diminished to 50% and 15% of the initial values, regardless of the exposure level. The finding thatXenopus embryos are permeable to⁶³Ni²⁺ during early cleavage stages provides a convenient experimental system to investigate the embryotoxicity and teratogenicity of nickel.     

The spring-time abundance and feeding ofEurytemora affinis (Poppe) in Volkerak-Zoommeer,a newly-created freshwater lake system in the Rhine delta (The Netherlands)

Eurytemora affinis, a calanoid copepod, has been encountered in Volkerak-Zoommeer (Rhine delta region, S.W. Netherlands) both before this lake system was isolated in 1987 from the estuarine influence, and after. It was the main particle-feeding crustacean at all the 3 sampling stations in March–April 1990 when it reached densities of up to 215 ind.l^–1. Its decline from mid April onwards, and low densities through summer, coincided with increase in cladocerans, especiallyDaphnia spp. (D. pulex andD. galeata), a decrease in seston (<33m) and chlorophyll concentrations and in primary production rates. The clearance rates (CR) ofEurytemora measured in the spring period varied enormously (0.6–24 ml.ind^–1.d^–1) depending mainly on size (0.44–1.06 mm), food concentration (0.8–2.2 mg C.l^–1), and the water temperature which varied only narrowly (8.0–9.0°C). Mean ingestion rates of the animals measuring 0.68±0.02 mm during the study was 6.7±3.2 gC.ind^–1.d^–1; and assimilation efficiency varied between 27 and 53% (mean: 41±9%). The weight specific CR (SCR) varied between 0.96 and 6.4 litre.mg^–1 body C.d^–1. Pooled regression of SCR on the animal's body weight at the 3 study stations revealed a significant inverse relationship. Also daily ration and specific assimilation ofE. affinis varied greatly and inversely with the body weight. This calanoid contributed from about 50 to 100% to the zooplankton community grazing rates and assimilation rates, the former often exceeding the phytoplankton primary production.     

Light acclimation and photoinhibition in threeSpirulina platensis (cyanobacteria) isolates

Three isolates ofSpirulina platensis (Norst) Geitler marked BP, P4P and Z19/2 were compared with respect to their response and acclimation capability to high photon flux densities (HPFD). Cultures exposed to HPFD (1500–3500 mol photon m^–2 s^–1) exhibited a marked decrease in light-dependent O₂ evolution rate. P4P was more sensitive to HPFD than the two other isolates. All three isolates recovered from photoinhibition when placed under low PFD. The BP isolate was able to recover also in the dark but to a lower extent and at a lower rate, while no recovery was observed in the other two isolates under dark conditions. No recovery was observed when protein synthesis was inhibited using chloramphenicol. Cultures grown at 200 mol photon m^–2 s^–1 differed from cultures grown at 120 mol photon m 2 s-1 by their lower maximal photosynthetic rate (P _max ) and higher light saturation (I _k ) value, while being more resistant to HPFD stress. The ability ofSpirulina isolates to acclimate and withstand HPFD may provide useful information for the selection of strains useful for outdoor mass cultivation.Author for correspondence     

Growth of Albugo candida (race unidentified) on Brassica juncea callus cultures

An obligate fungus Albugo candida (Pers. ex Lév.) Ktze. (race unidentified) was successfully grown on host callus tissues of Brassica juncea cv. Varuna. Of the various type of diseased explants used, young (green) hypertrophied inflorescence axis bearing non-erumpent zoosporangial blisters allowed the fungus to multiply asexually over the host calli on modified MS-medium (Murashige and Skoog, 1962). The dual cultures were maintained up to 6–8 subcultures without loss of viability of zoosporangia on MS-medium supplemented with 10.0 mg L^–1 IBA, 0.05 mg L^–1 kinetin, 25.0 mg L^–1 AA, 1.0 mg L^–1 biotin, 1.0 mg L^–1 thiamine-HCl and 1.0 g L^–1 casein hydrolysate. The fungus grew only on the callus cells and not axenically on the medium. Pathogenicity test and histopathology of cultures proved the existence of the viable fungus in vitro.Abbreviations AA ascorbic acid - BAP 6-benzyl aminopurine - CH casein hydrolysate acid hydrolysed - 2,4-D-2,4 dichlorophenoxy acetic acid - FAA formaldehyde acetic acid - IAA indole-3-acetic acid - IBA indole-3-butyric acid - HgCl₂ mercuric chloride - Kinetin 6-furfuryl aminopurine - MS Murashige and Skoog (1962) - NAA alpha naphthalene acetic acid - rh relative humidity - sdw sterile distilled water - wt. weights     

Growth inhibition by high light intensities in algae from lakes undergoing acidification

Blooms ofChrysochromulina breviturrita Nich. (Prymnesiophyceae) have been found to be restricted to lakes above pH 5.5 even though the alga is able to tolerate pH 4.0 in laboratory culture. A possible explanation is the increased transparency in acidifying lakes and a sensitivity ofC. breviturrita to high light intensities. A comparison was made withMougeotia sp., a filamentous green alga which co-occurs in moderately acidic lakes and has a similar pH tolerance range. This alga forms dense, floating mats or amorphous clouds in the upper littoral zone, where it would be exposed to full sunlight irradiances. In cultures ofC. breviturrita, prolonged exposures to 1600 μE · m⁻² · s⁻¹ (I₀′) resulted in reductions in cell yield which were dependent age at the onset of exposure to high light intensity. Only cultures exposed to high light intensities during late stationary phase were able to recover to control levels and no recovery occurred if these cultures were nitrogen deficient.Mougeotia was more tolerant of both high light intensity and nitrogen limitation during the recovery period. The inability ofC. breviturrita to recover from the effects of high light intensity during nitrogen deprivation may be particularly important in small, stratified lakes which are undergoing acidification. The slow rate of vertical circulation, and increasing transparency, would prolong exposure of the alga to the high irradiance levels of nutrient-deficient epilimnetic waters. This suggests that the geographic distribution ofC. breviturrita may be explained in part by the increasing light intensities in lakes undergoing acidification.     

Growth responses ofPterocladiella capillacea(Rhodophyta) in laboratory and outdoor cultivation

The growth and photosynthetic responses ofPterocladiella capillaceato NH₄, PO₄, CO₂-enrichment, pH, irradiance and temperature were evaluated for winter or summer plants cultivated under laboratory and outdoor settings. In the laboratory, using a gradient table, optimal growth temperature and irradiance for winter plants occurred at 10–20 °C and 100 mol photon m^–2s^–1, averaging 24.3% per week. The optimal growth conditions found for summer plants were 10–20 °C and 20 mol photon m^–2s^–1, averaging 29.0% per week. In a pH-stat cultivation system photosynthetic rates and growth rates were largely unaffected by pH in the range 6.5–8.5, however, they both decreased significantly above 8.5. In outdoor settings, using 40 L tanks,P. capillaceawas more responsive to the frequency the algae were fed with NH₄and PO₄rather than the relative concentrations of these nutrients. The average growth rates during winter were 28.3% and 12.5% per week when NH₄and PO₄were included once and twice a week for 24-h periods, respectively, while summer plants grew 15.0% and 25.3% per week at these nutrient regimes. Algae grown in seawater (containing 13.8 ± 1.8 M CO₂) or CO₂-enriched seawater (averaging 33.7 ± 13.2 M CO₂) had similar growth rates or even reduced productivity under CO₂-enrichment during winter. Concentrations of chlorophyllawere in average significantly higher in winter as compared to summer especially when nutrients were included twice a week. Phycoerythrin levels were also higher for plants fed with nutrients twice a week particularly during summer time. Although agar yields were limited and not seasonally dependent, this study shows high growth capacity forP. capillaceaas compared to previous investigations. Future mariculture prospective using current tank cultivation techniques for this species will likely depend on market demands for high quality agar.     

Phosphite accelerates programmed cell death in phosphate-starved oilseed rape (<Emphasis Type="Italic">Brassica napus</Emphasis>) suspension cell cultures

Phosphite (H₂PO₃^–, Phi) prevents the acclimation of plants and yeast to orthophosphate (Pi, HPO₄^2–) deprivation by specifically obstructing the derepression of genes encoding proteins characteristic of their Pi-starvation response. In this study, we report that prolonged (i.e., 3–4 weeks) culture of Brassica napus L. suspension cells in Pi-deficient (–Pi) media leads to programmed cell death (PCD). However, when the B. napus cells were subcultured into –Pi media containing 2 mM Phi, they initiated PCD within 5 days, with 95% cell death observed by day 9. Dying cells exhibited several morphological and biochemical features characteristic of PCD, including protoplast shrinkage, chromatin condensation, and fragmentation of nuclear DNA. Immunoblotting indicated that B. napus cells undergoing PCD upregulated a 30-kDa cysteine endoprotease that is induced during PCD in the inner integument cells of developing B. napus seeds. It is concluded that PCD in B. napus suspension cells is triggered by extended Pi starvation, and that Phi treatment greatly accelerates this process. Our results also infer that the adaptive value of acclimating at the molecular level to Pi-stress is to extend the viability of –Pi B. napus cell cultures by about 3 weeks.Abbreviations APase acid phosphatase (EC 3.1.3.2) - BnCysP B. napus cysteine proteinase - DAPI 4,6-diamidino-2-phenylindole - FDA fluorescein diacetate - PCD programmed cell death - Phi phosphite - +Pi and –Pi Pi-sufficient and -deficient, respectively - PI propidium iodide - PSI Pi-starvation inducible     

A procedure for axenic isolation of the marine microalga Isochrysis galbana from heavily contaminated mass cultures

Isochrysis galbana, one of the most widely usedmarine microalgae in the rearing of finfish and shellfish larvae, is masscultured frequently in outdoor tanks. Under prolonged and repeated culture,severe contamination occurs. Axenic isolation of I.galbanafrom such cultures was best achieved by using a ternary procedure involvingpercoll-gradient centrifugation, treatment with antibiotics, and growth on agarmedium. Protozoa and other algae were removed most effectively by isolation ofI. galbana at the 30–40% density layer on apercoll-gradient. Removal of bacteria was accomplished using a mixture of 5antibiotics (250 g mL^–1 ampicillin, 50g mL^–1 gentamycin, 100 gmL^–1 kanamycin, 500 gmL^–1 neomycin, 50 gmL^–1 streptomycin). Axenic colonies were isolated fromasolid medium prepared from 1% purified agar. The ternary procedure isconsideredapplicable to the isolation of other axenic single-celled microalgae fromheavily contaminated cultures.     

In situ monitoring of chlorophyll fluorescence to assess the synergistic effect of low temperature and high irradiance stresses inSpirulina cultures grown outdoors in photobioreactors

A chlorophyll fluorescence technique was applied to anin situ study on the effects of low temperature and high light stresses onSpirulina cultures grown outdoors in controlled tubular photobioreactors at high (1.1 g L^–1) and low (0.44 g L^–1) biomass concentrations. Diurnal changes in PSII photochemistry (F _v/F _m) after 15 min of darkness, or in the light (dF/F _m), and non-photochemical (qN) quenching were measured using a portable, pulse-amplitude-modulated fluorometer. The depression of theF _v/F _m ratio ofSpirulina cultures grown outdoors at 25°C (i.e. 10°C below optimum for growth) and 0.44 g L^–1, reached 30% at the middle of the day. At the same time of the day thedF/F _m ratio showed a reduction of up to 52%. The depression of bothF _v/F _m anddF/F _m was lower in the cultures grown at 1.1 g L^–1. Photoinhibition reduced the daily productivity of the culture grown at 0.44 g L^–1 and 25°C by 33% with respect to that grown at 35°C. Changes in the growth yields of the cultures grown under different temperatures and growth rates correlate well with analogous changes in photon yield (dF/F _m). Simple measurements of photochemical yield (F _v/F _m) can be used to test the physiological status ofSpirulina cultures. The results indicate that the saturating pulse fluorescence technique, when usedin situ, is a powerful tool for assessment of the photosynthetic characteristics of outdoor cultures ofSpirulina.     

Cd-tolerant arbuscular mycorrhizal (AM) fungi from heavy-metal polluted soils

Spores of arbuscular mycorrhizal (AM) fungi were isolated from two heavy-metal polluted soils in France via trap culture with leek (Allium porrum L.). Preliminary identification showed that the predominant spore type of both cultures (P2 and Cd40) belongs to the Glomus mosseae group. Their sensitivity to cadmium was compared to a laboratory reference strain (G. mosseae) by in vitro germination tests with cadmium nitrate solutions at a range of concentrations (0 to 100 mg L^–1) as well as extracts from a metal-polluted and unpolluted soils. Both cultures of AM fungi from heavy-metal polluted soils were more tolerant to cadmium than the G. mosseae reference strain. The graphically estimated EC₅₀ was 0.8 mg L^–1 Cd (concentration added to the test device) for G. mosseae and 7 mg L^–1 for P2 culture, corresponding to effective Cd concentrations of approximately 50–70 g L^–1 and 200–500 g L^–1, respectively. The extract of the metal-polluted soil P2 decreased germination of spores from the reference G. mosseae but not from P2 culture. However, the extracts of two unpolluted soils with different physico-chemical characteristics did not affect G. mosseae, whereas germination of P2 spores was markedly decreased in the presence of one of the extracts. These results indicate a potential adaptation of AM fungi to elevated metal concentrations in soil. The tested spores may be considered as metal-tolerant ecotypes. Spore germination results in presence of soil extracts show the difficulty of assessing the ecotoxic effect of metals on AM fungi without considering other soil factors that may interfere in spore germination and hyphal extension.     

Ulva rigida (Ulvales,Chlorophyta) tank culture as biofilters for dissolved inorganic nitrogen from fishpond effluents

Ulva rigida was cultivated in 7501 tanks at different densities with direct and continuous inflow (at 2, 4, 8 and 12 volumes d^–1) of the effluents from a commercial marine fishpond (40 metric tonnes, Tm, of Sparus aurata, water exchange rate of 16 m³ Tm^–1) in order to assess the maximum and optimum dissolved inorganic nitrogen (DIN) uptake rate and the annual stability of the Ulva tank biofiltering system. Maximum yields (40 g DW m^–2 d^–1) were obtained at a density of 2.5 g FW 1^–1 and at a DIN inflow rate of 1.7 g DIN m^–2 d^–1. Maximum DIN uptake rates were obtained during summer (2.2 g DIN M^–2 d^–1), and minimum in winter (1.1 g DIN m^–2 d^–1) with a yearly average DIN uptake rate of 1.77 g DIN m^–2 d^–1 At yearly average DIN removal efficiency (2.0 g DIN m^–2 d^–1, if winter period is excluded), 153 m² of Ulva tank surface would be needed to recover 100% of the DIN produced by 1 Tm of fish.Abbreviations DIN= dissolved inorganic nitrogen (NH _inf4 ^sup+ + NO _inf3 ^sup– + NO _inf2 ^sup– ); - FW= fresh weight; - DW= dry weight; - PFD= photon flux density; - V= DIN uptake rate