Removal of Prymnesium parvum (Haptophyceae) cells under different nutrient conditions by clay

In this study, we have shown that the ichthyotoxic Prymnesium parvum (haptophyte) can be successfully removed by spraying the surface with a phosphatic clay/polyaluminium chloride (PAC) mix (final concentration, 4 g l⁻¹ and 5 ppm, respectively). The alga was grown in non-axenic batch cultures with nitrogen deficient, phosphorus deficient and nutrient sufficient media. Sub-samples of the nutrient sufficient culture were diluted to obtain cell abundances equal to those in the nutrient deficient cultures. Clay/PAC removed up to 100% of the cells in the low cell nutrient sufficient treatment after 72 h, but removal in the other treatments was lower (up to 84%). The nitrogen deficient cells were found to be the most toxic, measured as haemolytic activity of the cells (HE₅₀), just prior to the start of the experiment. However, the toxicity of the cells in all treatments was found to fluctuate during the incubation time with a general increase in toxicity towards the end, suggesting that the cells became stressed during sedimentation and/or when trapped in the clay. But the amount of released toxins was always below the detection limit of the haemolytic assay, and the abundance of free-living bacteria, derived from flow cytometer counts, increased throughout the experiment. This suggests that released toxins were either trapped by the clay particles or effectively degraded by the bacterial community. The study shows that the clay method can be efficient in mitigation of P. parvum blooms, but further studies have to be conducted where optimum clay concentrations are determined to ensure that the efficiency is high against nutrient deficient cells and at high cell abundances, i.e. conditions found during blooms in eutrophic coastal waters, and to determine the fate of the cells and their toxins during and after sedimentation.     

Removal of Prymnesium parvum (Haptophyceae) and its toxins using clay minerals

Laboratory experiments were conducted to examine the ability of several clay minerals from Sweden to remove the fish-killing microalga, Prymnesium parvum Carter, from suspension. In their commercial form (i.e. after incineration at 400 °C), seawater slurries (salinity = 26) of the three minerals tested were generally ineffective at removing P. parvum from culture within a range of 0.01 to 0.50 g/L, and after 2.5 h of flocculation and settling. Dry bentonite (SWE1) displayed the highest removal efficiency (RE) at 17.5%, with 0.50 g/L. Illite (SWE3) averaged only 7.5% RE between 0.10 to 0.50 g/L, while kaolinite (SWE2) kept the cells suspended instead of removing them. Brief mixing of the clay-cell suspension after SWE1 addition improved RE by a factor of 2.5 (i.e. 49% at 0.50 g/L), relative to no mixing. The addition of polyaluminum chloride (PAC, at 5 ppm) to 0.50 g/L SWE1 also improved RE to 50% relative to SWE1 alone, but only minor improvements in RE were seen with SWE2 and SWE2 combined with PAC. In further experiments, P. parvum grown in NP-replete conditions were removed in greater numbers than cells in N- or P-limited cultures, at 0.10–0.25 g/L of SWE1 and 5 ppm PAC. With 0.50 g/L, RE converged at 40% for all three culture conditions. The toxin concentration of NP-replete cultures decreased from 24.2 to 9.2 μg/mL (60% toxin RE) with 0.10–0.50 g/L SWE1 treatment and 5 ppm PAC. A strong correlation was found between cell and toxin RE (r²=0.995). For N-limited cultures, toxin RE ranged between 21 and 87% with the same clay/PAC concentrations, although the correlation between cell and toxin removal was more moderate (r²=0.746) than for NP-replete conditions. Interestingly, the toxin concentration within the clay-cell pellet increased dramatically after treatment, suggesting that clay addition may stimulate toxin production in N-stressed cells. For P-limited cultures, toxin concentration also decreased following clay/PAC treatment (i.e. 36% toxin RE), but toxin removal was poorly correlated to cell removal (r²=0.462). To determine whether incineration affected SWE1’s removal ability, a sample of its wet, unprocessed form was tested. The RE of wet bentonite (SWE4) was slightly better than that of SWE1 (31% versus 17%, respectively, at 0.50 g/L), but when 5 ppm PAC was added, RE increased from 10 to 64% with 0.05 g/L of SWE4, and increased further to 77% with 0.50 g/L. There were no significant differences in RE among NP-replete, N-limited and P-limited cultures using PAC-treated SWE4. Finally, RE varied with P. parvum concentration, reaching a maximum level at the lowest cell concentration (1×10³ cells/mL): 100% RE with 0.10 and 0.50 g/L SWE4 + 5 ppm PAC. RE dropped as cell concentration increased to 1×10⁴ and 5×10⁴ cells/mL, but rose again when concentration increased to 1×10⁵ cells/mL, the concentration used routinely for the removal experiments above. Based on these results, SWE4 with PAC was the most effective mineral sample against P. parvum. Overall, these studies demonstrated that clay flocculation can be effective at removing P. parvum and its toxins only under certain treatment conditions with respect to cell concentration, clay type and concentration, and physiological status.     

Cryptosporidium parvum: Radiation-induced alteration of the oocyst proteome

Cryptosporidium parvum is a waterborne protozoan parasite that is found intracellularly in host animals, including humans, and causes severe diarrhea, which can lead to the death of an immunocompromised individual. Previously, we found that this organism is highly radioresistant as it can productively infect mice after exposure to a 10-kGy dose of γ-radiation.To understand how C. parvum avoids radiation damage, we characterized its protein expression patterns 6, 24, and 48 h after a 10-kGy dose of γ-radiation using two-dimensional PAGE. The gels showed 10 silver-stained spots that increased or decreased in size following γ-irradiation. Five proteins contained in these spots were identified using MALDI-TOF MS peptide fingerprinting, and two of these showed an increase in expression after γ-irradiation. These proteins were identified by LC–MS/MS as proteasome subunit alpha type 4 (NTN hydrolase fold) and thioredoxin peroxidase-like protein. The roles of these two upregulated proteins as related to the radioresistance of C. parvum remain to be evaluated.     

The effect of blueberry extracts on Giardia duodenalis viability and spontaneous excystation of Cryptosporidium parvum oocysts, in vitro

The protozoan parasites Giardia duodenalis and Cryptosporidium parvum are common causes of diarrhoea, worldwide. Effective drug treatment is available for G. duodenalis, but with anecdotal evidence of resistance or reduced compliance. There is no effective specific chemotherapeutic intervention for Cryptosporidium. Recently, there has been renewed interest in the antimicrobial properties of berries and their phenolic compounds but little work has been done on their antiparasitic actions. The effect of various preparations of blueberry (Vaccinium myrtillus) extract on G. duodenalis trophozoites and C. parvum oocysts were investigated. Pressed blueberry extract, a polyphenolic-rich blueberry extract, and a commercially produced blueberry drink (Bouvrage) all demonstrated antigiardial activity. The polyphenol-rich blueberry extract reduced trophozoite viability in a dose dependent manner. At 167 μg ml⁻¹, this extract performed as well as all dilutions of pressed blueberry extract and the Bouvrage beverage (9.6 ± 2.8% live trophozoites remaining after 24 h incubation). The lowest dilution of blueberry extract tested (12.5% v/v) contained >167 μg ml⁻¹ of polyphenolic compounds suggesting that polyphenols are responsible for the reduced survival of G. duodenalis trophozoites. The pressed blueberry extract, Bouvrage beverage and the polyphenolic-rich blueberry extract increased the spontaneous excystation of C. parvum oocysts at 37 °C, compared to controls, but only at a dilution of 50% Bouvrage beverage, equivalent to 213 μg ml⁻¹ gallic acid equivalents in the polyphenolic-rich blueberry extract. Above this level, spontaneous excystation is decreased. We conclude that water soluble extracts of blueberries can kill G. duodenalis trophozoites and modify the morphology of G. duodenalis and C. parvum.     

Seasonal and interannual dynamics of polyphenols in Myriophyllum verticillatum and their allelopathic activity on Anabaena variabilis

In 4 successive years, we investigated the seasonal and interannual variability of the total polyphenolic pool and of the individual polyphenolic compounds in Myriophyllum verticillatum, as well as their allelopathic activity in a small eutrophic lake. We tested whether nutrient availability explained interannual and seasonal changes in the production of polyphenols. There were no strong interannual variations in plant tissue carbon, nitrogen and phosphorus concentrations, while total phenolic compounds (TPC) significantly differed between years, especially in apical meristems (range: 38–122 mg g⁻¹  dry weight (DW)). Seasonal patterns, with maxima between May and July, changed between years. Partially confirming the carbon-nutrient balance hypothesis sensu Bryant et al. [Bryant, J.P., Chapin III, F.S., Klein, D.R., 1983. Carbon/nutrient balance of boreal plants in relation to vertebrate herbivory. Oikos 40, 357–368], we found correlations between TPC and the C/N (carbon/nitrogen) ratio in some but not all years, especially in apical meristems. Plant tissue phosphorus content accounted also for the variability in TPC in some years. Crude extracts of apical meristems always inhibited the growth of Anabaena variabilis, used as a target cyanobacterium. Plant TPC concentration and allelopathic activity were significantly correlated in all years except in 2005. Bioassay-directed fractionation of M. verticillatum extracts coupled with LC–MS analyses of the respective fractions revealed several isomers of HHDP-di- and -tri-galloylglucose apparently responsible for the allelopathic effects. The individual active compounds revealed a more distinct seasonal pattern compared to the pool of phenolic compounds in M. verticillatum, with a clear maximum in May, the ecologically most relevant period for inhibitory effects of submerged macrophytes on phytoplankton.     

Phosphate solubilization potential and stress tolerance of Eupenicillium parvum from tea soil

Eupenicillium parvum was recorded for first time during isolation of phosphate-solubilizing microorganisms from the tea rhizosphere. The fungus developed a phosphate solubilization zone on modified Pikovskaya agar, supplemented with tricalcium phosphate. Quantitative estimation of phosphate solubilization in Pikovskaya broth showed high solubilization of tricalcium phosphate and aluminium phosphate. The fungus also solubilized North Carolina rock phosphate and Mussoorie rock phosphate, and exhibited high levels of tolerance against desiccation, acidity, salinity, aluminium, and iron. Solubilization of inorganic phosphates by the fungus was also observed under high stress levels of aluminium, iron, and desiccation, though the significant decline in phosphate solubilization was marked in the presence of aluminium than iron. The fungal isolate showed 100 % identity with E. parvum strain NRRL 2095 ITS 1, 5.8S rRNA gene and ITS 2, complete sequence; and 28S rRNA gene, partial sequence.     

Susceptibility of Cydia pomonella to Bacillus thuringiensis under laboratory and field conditions

The delta-endotoxin of 12 strains in 10 subspecies of Bacillus thuringiensis Berliner was highly active against Cydia pomonella (L.) when assayed under laboratory conditions on artificial diet. These results could not be confirmed in the field.The disappointing results obtained under field conditions are due to the behaviour of the target insect. C. pomonella larvae do not ingest food during penetration of the fruit. The larva bites pieces of the epidermis and deposits them without ingestion on top of the entry hole.

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Zusammenfassung Das delta-Endotoxin von B.t. war in Laborversuch auf Kunstmedium gegenüber den Larven des Apfelwicklers, C. pomonella, sehr aktiv. Die hohe Aktivität konnte aber unter Feldbedingungen nicht mehr bestätigt werden.Es wurde nachgewiesen, dass die unbefriedigenden Resultate von B. thuringiensis unter Feldbedingungen auf das Verhalten der Junglarven zurückzuführen sind: Die Larven nehmen während dem Eindringen in den Apfel keine Nahrung auf, sondern deponieren die herausgebissenen Epidermistücke über der Einbohrstelle.

     

Effect of the fungus Pochonia chlamydosporia and fosthiazate on the multiplication rate of potato cyst nematodes (Globodera pallida and G. rostochiensis) in potato crops grown under UK field conditions

This is the first report of the successful use of Pochonia chlamydosporia as a biological control agent against potato cyst nematodes (PCN) (Globodera pallida and G. rostochiensis) in potato crops grown under commercial field conditions and represents an important step in the development of biological control for PCN. Two field experiments were established in consecutive years (2006 and 2007) at different field sites in Shropshire, England. Treatments comprised of (1) untreated control, (2) P. chlamydosporia, (3) P. chlamydosporia with the nematicide fosthiazate and (4) fosthiazate alone. In both experiments, significant reductions in the nematode multiplication rate (Pf/Pi) for P. chlamydosporia treated plots were observed (48% and 51% control, respectively). The P. chlamydosporia treatment did not differ significantly from both fosthiazate treatments in terms of Pf/Pi in spite of the trend towards increased control particularly in Experiment 1. P. chlamydosporia therefore provided similar levels of nematode population control as fosthiazate. The combined treatment did not provide any additional reduction in Pf/Pi but demonstrated that P. chlamydosporia was compatible with fosthiazate. Over the different developmental stages of the juvenile nematodes, there was evidence of parasitism of adult females on the plant root by P. chlamydosporia. Root colonization by P. chlamydosporia was higher in the P. chlamydosporia treatment due to increased levels of nematodes in plant roots. Results from both experiments demonstrated the efficacy of P. chlamydosporia as a biological control agent of PCN and indicate its potential for use as part of an integrated pest management strategy.     

Isolation of polyketides from Prymnesium parvum (Haptophyta) and their detection by liquid chromatography/mass spectrometry metabolic fingerprint analysis

Prymnesium parvum is a microalga that forms blooms coupled with the presence of potent exotoxins; however, no chemical standards are currently available for the toxins. Streamlined methods are presented for the separation and enrichment of polyketide toxins, prymnesin-1 (prym1) and prymnesin-2 (prym2). Prymnesins were separated by reversed-phase chromatography and detected by positive-mode electrospray ionization MS to generate a unique metabolic fingerprint. More than 10 ions were detected and mass assignments were in agreement with predicted isotopic distributions for the intact compounds and related fragments; ions occurred as multiply protonated species and with common salt adducts. The most prevalent ion was observed at 919.88 m/z, which represents the aglycone [prym_agly + 2H]²⁺ backbone structure common to both molecules. Expanded mass spectra for this and related ions were in excellent agreement (<0.5 ppm) with empirically derived spectra based on elemental composition and naturally occurring isotopes. These investigations have confirmed the isolation of polyketide prymnesins from whole cells, which heretofore has not been reproduced since their original characterization. Moreover, this study represents the first time these compounds have been verified in aqueous materials. These tools should allow the direct identification and analysis of polyketide prymnesins, which will greatly improve our understanding of these toxins in P. parvum.     

Rejoining of gamma-ray-induced DNA damage in Cryptosporidium parvum measured by the comet assay

Cryptosporidium parvum is a well-known waterborne intracellular protozoan that causes severe diarrheal illness in immunocompromised individuals. This organism is highly resistant to harsh environmental conditions and various disinfectants, and it exhibits one of the highest known resistances to gamma irradiation. We investigated rejoining of gamma-ray-induced DNA damage in C. parvum by neutral comet assay. Oocysts were gamma irradiated at various doses (1, 5, 10, and 25 kGy) and were incubated for various periods (6-96 h) after exposure to 10 kGy. The comet tail moment showed that the number of DNA double-strand breaks increased concomitantly with the gamma irradiation dose. When investigating rejoining after irradiation at 10 kGy, double-strand breaks peaked at 6 h postirradiation, and rejoining was highest at 72 h postirradiation. The observed rejoining pattern suggests that repair process occurs slowly even when complex DNA double-strand breaks in C. parvum were induced by high dose irradiation, 10 kGy.     

Potential allelopathic effect of neo-clerodane diterpenes from Teucrium chamaedrys (L.) on stenomediterranean and weed cosmopolitan species

The allelopathic effects of neo-clerodane diterpenes, isolated from Teucrium chamaedrys (L.), have been evaluated on the seed germination and seedling growth of four coexisting Mediterranean species (Dactylis hispanica, Petrorhagia velutina, Phleum subulatum and Petrorhagia saxifraga) and two cosmopolitan weeds (Amaranthus retroflexus and Avena fatua). All of the structures have been elucidated on the basis of their spectroscopic features. The bioassays data, analyzed by principal component analysis, showed more negative effects on weeds respect to coexisting species. Moreover D. hispanica, P. velutina, P. subulatum showed both stimulating or inhibiting effects depending on the type of metabolite and the concentration used in the test.     

Latitudinal differentiation in the effects of the toxic dinoflagellate Alexandrium spp. on the feeding and reproduction of populations of the copepod Acartia hudsonica

Blooms of the dinoflagellate Alexandrium spp. increase in their frequency, toxicity and historical presence with increasing latitude from New Jersey (USA) to the Gaspé peninsula (Canada). Biogeographic variation in these blooms results in differential exposure of geographically separate copepod populations to toxic Alexandrium. We hypothesize that the ability of copepods to feed and reproduce on toxic Alexandrium should be higher in copepods from regions that are frequently exposed to toxic Alexandrium blooms. We tested this hypothesis with factorial common environment experiments in which female adults of the copepod Acartia hudsonica from five separate populations ranging from New Jersey to New Brunswick were fed toxic and non-toxic strains of Alexandrium, and the non-toxic flagellate Tetraselmis sp. Consistent with the hypothesis, when fed toxic Alexandrium we observed significantly higher ingestion and egg production rates in the copepods historically exposed to toxic Alexandrium blooms relative to copepods from regions in which Alexandrium is rare or absent. Such differences among copepod populations were not observed when copepods were fed non-toxic Alexandrium or Tetraselmis sp. These results were also supported by assays in which copepods from populations both historically exposed and naïve to toxic Alexandrium blooms were fed mixtures of toxic Alexandrium and Tetraselmis sp. Two-week long experiments demonstrated that when copepods from populations naïve to toxic Alexandrium were fed a toxic strain of Alexandrium they failed to acclimate, such that their ingestion rates remained low throughout the entire two-week period. The differences observed among populations suggest that local adaptation of populations of A. hudsonica from Massachusetts (USA) to New Brunswick (Canada) has occurred, such that some populations are resistant to toxic Alexandrium.     

Oviposition in Culicoides impunctatus under laboratory conditions

We present a laboratory-based examination of oviposition preference in the Scottish biting midge, Culicoides impunctatus (Diptera: Ceratopogonidae). A variety of oviposition substrates were screened in no-choice and choice bioassays for efficacy in eliciting an egg-laying response. Both upper-layer photosynthetic Sphagnum spp. moss and Juncus articulatus infusions were identified as producing increased oviposition under no-choice conditions. During choice trials against a control of damp cottonwool, upper-layer Sphagnum spp. moss produced a significantly greater egg-laying response. These conclusions are interpreted in terms of possible cues involved in oviposition site selection and assessed for future use in colonisation of this troublesome species.     

Prorocentrum minimum(clone Exuv) is nutritionally insufficient, but not toxic to the copepod Acartia tonsa

This study tested whether the dinoflagellate Prorocentrum minimum is nutritionally insufficient or toxic to the copepod Acartia tonsa. Experiments were carried out with adult female A. tonsa and the P. minimum clone Exuv, both isolated from Long Island Sound. Initially, the functional and numerical responses of A. tonsa feeding on exponentially growing P. minimum cells were characterized. These experiments revealed that A. tonsa readily ingested P. minimum cells, up to the equivalent of 200% of body carbon day⁻¹, but egg production was relatively low, with a maximum egg production rate of 22% of body carbon day⁻¹. Hence, the egg production efficiency (egg carbon produced versus cell carbon ingested) was low (10%). In a separate experiment, ingestion and egg production rates were measured as a function of food concentration with cells in different growth stages (early-exponential, late-exponential/early-stationary, and late-stationary growth phase) to simulate conditions during a bloom. There was no indication that cells in the stationary phase resulted in lower ingestion or egg production rates relative to actively growing cells. Egg hatching success remained high (>80%) and independent of the cell growth phase. In a third experiment specifically designed to test the hypothesis that P. minimum is toxic, ingestion, egg production and egg hatching success were measured when females were fed mixtures of P. minimum and the diatom Thalassiosira weissflogii, but in which total food concentration was held constant and the proportion of P. minimum in the mixed diet varied. A. tonsa readily ingested P. minimum when it was offered in the mixed diet, with no detrimental effects on egg production or egg hatching observed. Supplementing P. minimum with T. weissflogii increased both the egg production rate and the egg production efficiency. It is concluded that P. minimum is nutritionally insufficient, but not toxic to A. tonsa. Finally, it is estimated that in the field grazing by A. tonsa is approximately equivalent to 30% of the maximum daily growth rate of P. minimum. Hence, copepod grazing cannot be ignored in field and modeling studies of the population dynamics of P. minimum.     

Biological control of brown rot (Moniliana laxa Ehr.) on apricot (Prunus armeniaca L. cv. Hacıhaliloğlu) by Bacillus, Burkholdria, and Pseudomonas application under in vitro and in vivo conditions

In 2002 and 2003, a study was conducted to determine the effect of bacterial strains, Burkholdria OSU 7, Bacillus OSU 142, and Pseudomonas BA 8, on biological control of brown rot disease (Monilinia laxa Ehr.) on apricot cv. Hacıhaliloğlu in Malatya province of Turkey. Apricot orchard at full blooming stage was inoculated with conidial suspension (1 × 10⁶ spores/ml) of M. laxa Ehr. After inoculation, two apricot trees for each application were treated with each of the three biological control agents (Burkholdria gladii OSU 7, Bacillus subtilis OSU 142, and Pseudomonas putida BA 8) by spraying (1 × 10⁹ cfu/ml) on inoculated branches. Disease incidence was evaluated for untreated (control 1) and four different treatment groups including commercial disease management (control 2, positive control: 3% Bourdox in fall, 50% Cupper at pink flower, 30 g/100 l Corus at first blooming, and 300 g/100 l Captan at last blooming stage) and treatments including each of the three bacterial strains (OSU 7, OSU 142, and BA 8). The results showed that disease incidence for negative control (control 1) was 9.94, which was significantly higher than disease incidence for commercial application (2.57%) or bacterial treatments (2.82–5.00%) in the first year. In 2003, the lowest disease incidence observed in OSU 7 treatment (6.80%), while disease incidence rate for positive control and negative control were 9.45% and 28.46%, respectively. This result may suggest that OSU 7 has potential to be used as biopesticide for effective management of brown rot disease on apricot.     

Light acclimatisation of Elodea nuttallii grown under ambient DIC conditions

Light acclimatisation capabilities of Elodea nuttallii at nearly ambient DIC conditions were investigated by determining growth characteristics, main photosynthetic parameters and pigmentation of plants incubated at 5 different irradiances (10–146 μmol photons m⁻² s⁻¹). Positive net growth was observed under all light treatments tested. Maximum ratio root versus shoot (r:s) of 1.86 was achieved at medium irradiances (72–94 μmol photons m⁻² s⁻¹), whereas at low (10 μmol photons m⁻² s⁻¹) and high irradiances (146 μmol photons m⁻² s⁻¹) r:s was significantly lower (0.39 and 1.05, respectively). With respect to main photosynthetic parameters, an increase of light compensation points (E _c), attended by decreasing ratios of light saturation points of photosynthesis (E _k)/irradiance were observed. E _c values were comparable to other low-light adapted macrophytes, which indicate that E. nuttallii can be regarded as a low-light adapted plant, under photorespiratory conditions. This was also confirmed by maximum E _k values of just 73 μmol photons m⁻² s⁻¹. Further support was achieved from pigmentation and non-photochemical quenching (NPQ) data, both indicating rather limited acclimatisation ability at light treatments above 90 μmol photons m⁻² s⁻¹. These results are discussed with respect to the competitive abilities of E. nuttallii under nearly ambient (photorespiratory) DIC conditions, especially in dense stands and turbid phytoplankton-dominated waters.     

Peroxidase mediated hydrogen peroxide production in barley roots grown under stress conditions

All applied metals (Co, Al, Cu, Cd) and NaCl inhibited barley root growth. No root growth inhibition was caused by drought exposure, in contrast to cold treatment. 0.01 mM H₂O₂ stimulated root growth and GA application did not affect root growth at all. Other activators and inhibitors of H₂O₂ production (SHAM, DTT, 10 mM H₂O₂, 2,4-D) inhibited root growth. Loss of cell viability was most significant after Al treatment, followed by Cd and Cu, but no cell death was induced by Co. Drought led to slight increase in Evans blue uptake, whereas neither NaCl nor cold influenced this parameter. DTT treatment caused slight increase in Evans blue uptake and significant increases were detected after 2,4-D and 10 mM H₂O₂ treatment, but were not induced by others stressors. Metal exposure increased guaiacol-POD activity, which was correlated with oxidation of NADH and production of H₂O₂. Exposure to drought caused a minor change in NADH oxidation, but neither H₂O₂ production nor guaiacol-POD activity was increased. Cold and NaCl application decreased all monitored activities. Increase in NADH oxidation and guaiacol-POD activity was caused by 10 mM H₂O₂ and 0.01 mM 2,4-D treatment, which also caused enhancement of H₂O₂ production. Slight inhibition of all activities was caused by 0.01 mM H₂O₂, GA, DTT; more pronounced inhibition was detected after SHAM treatment. The role of H₂O₂ production mediated by POD activity in relation to root growth and cell viability under exposure to some abiotic stress factors is discussed.     

Domoic acid production by Pseudo-nitzschia australis and Pseudo-nitzschia calliantha isolated from North Chile

The morphology and toxicity of the four ubiquitous species belonging to the genus Pseudo-nitzschia found in mixed blooms of phytoplankton from northern Chilean waters were studied. The phytoplankton samples and cultures obtained were identified by scanning electron microscopy, revealing the presence of Pseudo-nitzschia australis, P. calliantha, P. pseudodelicatissima and P. subfraudulenta. This is the first report of P. calliantha in northern Chile. Toxin analyses using the LC–MS method confirmed the presence of domoic acid in P. australis and P. calliantha. Domoic acid was not detected in cultures of P. subfraudulenta. This study therefore confirms P. australis and P. calliantha as an unequivocal source of domoic acid in Chilean waters. P. australis is probably the most important producer of amnesic shellfish toxin in view of its domoic acid content. However, more research is needed to evaluate the potential for toxin production in P. pseudodelicatissima.     

Effects of growth conditions on exopolysaccharide production by Cyanospira capsulata

Batch cultures of Cyanospira capsulata, a heterocystous cyanobacterium possessing a thick polysaccharidic capsule, were characterized by increasing viscosity owing to the continuous release of a soluble polysaccharide (EPS) into the culture medium. Both capsulated trichomes and solubilized EPS contributed to the flow properties of whole cultures. A typical pseudoplastic behaviour, the more marked the more aged were the cultures, was evidenced.The production of EPS was investigated under different growth conditions by changing some nutritional and physical parameters known to affect the synthesis of exopolysaccharides in algae and cyanobacteria. Among the factors tested (Ca²⁺, Mg²⁺ or PO₄⁻⁴ deficiencies, salinity and pH) only Mg²⁺ shortage caused a significant enhancement of the EPS production. Under continuous illumination in open ponds, the EPS productivity of batch cultures on standard mineral medium was about 5·8 g m⁻² day⁻¹, whereas under Mg²⁺ deficiency with a consequent increase of the cultures' viscosity