Effects of temperature,salinity and their interaction on growth of benthic dinoflagellates Ostreopsis spp. from Thailand

Benthic dinoflagellates Ostreopsis spp. are known as producers of palytoxin and its analogs, resulting occasionally in human health problems worldwide. Although distribution of Ostreopsis spp. along the Thai coasts has been reported, little is known about their growth characteristics. To discuss the bloom dynamics of Ostreopsis spp. in Thailand, first we tested four kinds of media to optimize growth conditions and then clarified the effects of temperature, salinity and temperature–salinity interaction on the growth of strains of the O. cf. ovata Thailand subclade, O. cf. ovata South China Sea subclade, Ostreopsis sp. 6 and Ostreopsis sp. 7. We showed that the f/2 medium was a suitable medium which gave the highest cell yields for all the strains tested. The strains of the O. cf. ovata Thailand subclade, O. cf. ovata South China Sea subclade and Ostreopsis sp. 6 grew in the temperature range 20–32.5 °C, whereas the strain of Ostreopsis sp. 7 grew in 20–30 °C. The semi-optimal temperature ranges (≧80% of the maximal growth rate) for the former three strains were 22.7–27.4 °C, 27.9–30.8 °C and 23.5–26.4 °C, respectively, whereas that of the latter strain was 23–27.2 °C. The optimal temperature for the O. cf. ovata South China Sea subclade was 30 °C, whereas for the others it was 25 °C. All the Ostreopsis strains tested could grow in a salinity range of 20–40. The semi-optimal salinities for the O. cf. ovata Thailand subclade, O. cf. ovata South China Sea subclade Ostreopsis sp. 6 and Ostreopsis sp. 7 were 28.7–35, 23.8–30.8, 29.8–36 and 28–36, respectively. The optimal salinities for the O. cf. ovata Thailand subclade and O. cf. ovata South China Sea subclade were 30 and 25, respectively, whereas for Ostreopsis sp. 6 and Ostreopsis sp. 7 it was 35. In this study, our results suggested that the optimal and tolerable temperature–salinity conditions differ among the Thai Ostreopsis species/clades/subclades. Tolerances of the O. cf. ovata Thailand subclade, O. cf. ovata South China Sea subclade and Ostreopsis sp. 6 to the high temperature of 32.5 °C may allow these organisms to be distributed in the tropical areas, where the water temperature often reaches >30 °C.     

Ostreopsis cf. ovata in the Gulf of Gabès (south-eastern Mediterranean Sea): morphological,molecular and ecological characterization

In the last few decades, the frequency of the toxic benthic dinoflagellate Ostreopsis cf. ovata proliferation has increased in the Mediterranean Sea. These blooms are associated with harmful effects on human health and the environment. The present work provides the first long term study on the spatio-temporal distribution of O. cf. ovata in relation to physical parameters in the Gulf of Gabès coastal waters (south-eastern Mediterranean Sea), as well as its morphological, molecular and physiological features. The strains of O. cf. ovata were identified morphologically by light and epifluorescence microscopy. The morphology and the size range of cultured strains were similar to those described regarding O. cf. ovata isolated from the Mediterranean Sea. The ultrastructural analysis of O. cf. ovata cells using the transmission electron microscopy showed the presence of numerous vesicles (VE) containing spirally coiled fibers (SCFs) connected to the mucus canal (CH). The phylogenetic tree based on the internal transcribed spacer region containing the 5.8S rDNA (ITS-5.8S rDNA) revealed that O. cf. ovata strains were placed into the Mediterranean/Atlantic clade. In addition, O. cf. ovata toxicity was evaluated by the mouse bioassay and a dose level ≥ 4 × 10⁴ cells was found to be lethal to mice. The examination of the O. cf. ovata occurrence in the Gulf of Gabès at a large temporal scale (1997–2012) revealed a clear seasonal pattern with dominance from midsummer (July) to late autumn (November). Furthermore, a positive correlation was found between the abundance of O. cf. ovata and salinity, whereas no correlation was found as regards temperature. The occurrence of O. cf. ovata was only detected at salinity above 35 and the highest concentrations were observed at 45. Laboratory experiments confirmed such a result and showed that isolated O. cf. ovata strains had optimal growth at salinity ranging between 35 and 45, with its peak at 40.     

Temperature and sex dependent effects on cardiac mitochondrial metabolism in Atlantic cod (Gadus morhua L.)

To test the hypothesis that impaired mitochondrial respiration limits cardiac performance at warm temperatures, and examine if any effect(s) are sex-related, the consequences of high temperature on cardiac mitochondrial oxidative function were examined in 10 °C acclimated, sexually immature, male and female Atlantic cod. Active (State 3) and uncoupled (States 2 and 4) respiration were measured in isolated ventricular mitochondria at 10, 16, 20, and 24 °C using saturating concentrations of malate and pyruvate, but at a submaximal (physiological) level of ADP (200 µM). In addition, citrate synthase (CS) activity was measured at these temperatures, and mitochondrial respiration and the efficiency of oxidative phosphorylation (P:O ratio) were determined at [ADP] ranging from 25–200 µM at 10 and 20 °C. Cardiac morphometrics and mitochondrial respiration at 10 °C, and the thermal sensitivity of CS activity (Q₁₀=1.51), were all similar between the sexes. State 3 respiration at 200 µM ADP increased gradually in mitochondria from females between 10 and 24 °C (Q₁₀=1.48), but plateaued in males above 16 °C, and this resulted in lower values in males vs. females at 20 and 24 °C. At 10 °C, State 4 was ~10% of State 3 values in both sexes [i.e. a respiratory control ratio (RCR) of ~10] and P:O ratios were approximately 1.5. Between 20 and 24 °C, State 4 increased more than State 3 (by ~70 vs. 14%, respectively), and this decreased RCR to ~7.5. The P:O ratio was not affected by temperature at 200 μM ADP. However, (1) the sensitivity of State 3 respiration to increasing [ADP] (from 25 to 200 μM) was reduced at 20 vs. 10 °C in both sexes (K_m values 105±7 vs. 68±10 μM, respectively); and (2) mitochondria from females had lower P:O values at 25 vs. 100 μM ADP at 20 °C, whereas males showed a similar effect at 10 °C but a much more pronounced effect at 20 °C (P:O 1.05 at 25 μM ADP vs. 1.78 at 100 μM ADP). In summary, our results demonstrate several sex-related differences in ventricular mitochondrial function in Atlantic cod, and suggest that myocardial oxidative function and possibly phosphorylation efficiency may be limited at temperatures of 20 °C or above, particularly in males. These observations could partially explain why cardiac function in Atlantic cod plateaus just below this species׳ critical thermal maximum (~22 °C) and may contribute to yet unidentified sex differences in thermal tolerance and swimming performance.     

Proliferation of the toxic dinoflagellate Ostreopsis cf. ovata in relation to depth,biotic substrate and environmental factors in the North West Mediterranean Sea

In recent decades, the North West Mediterranean Sea has been seriously affected by the development of the toxic benthic dinoflagellate Ostreopsis cf. ovata, which is associated with harmful effects on human health and the environment. The present work aims to provide a large overview of the multiple environmental factors assumed to regulate or influence the growth of Ostreopsis. An intensive sampling campaign over a full annual cycle was performed along the French and Italian coasts (in six sites from Cassis to Genoa), to determine patterns of temporal and spatial distributions of both O. cf. ovata epiphytic and planktonic cells. Results highlighted substantial seasonal variations in the abundance of Ostreopsis. These variations correlated to seawater temperature, with an optimum growth temperature ranging from 23 °C to 27.5 °C. Phosphate concentration, rather than nitrogen or silicate, was also positively associated with Ostreopsis. Decreases in oxygen and increases in chlorophyll a concentrations were recorded during the summer blooming period. The maximal Ostreopsis epiphytic abundance was generally higher on Dictyota spp. than on the other two sampled macroalgae (up to 8.54 × 10⁶ cells g⁻¹ FW), even though statistical analysis did not support a clear substrate preference. Epiphytic abundances were significantly higher at a very shallow depth (0.5 m), than at 1 and/or 3 m depths. High anthropogenic pressure (related to population density) seems to have promoted the occurrence of blooms in urbanized areas, which could partly explain the strong demarcation in Ostreopsis development between Western and Eastern sampling sites. The ecological niche of Ostreopsis cf. ovata needs precise definition, which will require further in situ and in vitro experimental studies, to determine the relative importance of distinct environmental parameters.     

Seasonal dynamics and spatial distribution of epiphytic dinoflagellates in Peter the Great Bay (Sea of Japan) with special emphasis on Ostreopsis species

Studies of epiphytic dinoflagellates in Peter the Great Bay, Sea of Japan in 2008–2011 revealed the presence of 13 species. Five of the species are known as potentially toxic: Amphidinium carterae, A. operculatum, Ostreopsis cf. ovata, O. cf. siamensis and Prorocentrum lima. The maximum species richness and abundance of epiphytic dinoflagellates were observed in autumn (from September to October). Ostreopsis spp. were most widely distributed and predominated, amounting to 99% of the total density of dinoflagellates. Multi-year seasonal dynamics of Ostreopsis spp. in Peter the Great Bay showed that these cells appear as epiphyton in August after maximum warming of surface waters (22–24 °С) and disappear in early November, when the water temperature decreases below 7 °С. Ostreopsis spp. proliferation occurred in September, when the water temperature was 17.2–21.0 °C. The highest densities of Ostreopsis spp. were recorded on September 9, 2010 on the rhodophyte Neorhodomela aculeata – 230 × 10³ cells g⁻¹ DW or 52 × 10³ cells g⁻¹ FW. The spatial distribution of epiphytic dinoflagellates was investigated in the near-shore areas of Peter the Great Bay during the second half of September 2010 to evaluate the role of hydrodynamic conditions. Epiphytic dinoflagellates were not found in sheltered sites having weak mixing hydrodynamics. However, the abundances of Ostreopsis spp. were significantly higher at sites having moderate turbulence compared to biotopes experiencing strong wave action. Densities of Ostreopsis spp. were not significantly different on macrophytes with branched thallus of all taxonomic divisions. However, the average cell densities of Ostreopsis spp. on green algae with branched thallus were significantly higher than on green algae having laminar thallus.     

Variability of sea-surface temperature and sea-ice cover in the Fram Strait over the last two millennia

A sediment core located on the West Spitzbergen margin in the Fram Strait (78°54.931′N, 6°46.005′E, water depth: 1497 m) was analyzed for its dinocyst content in order to reconstruct hydroclimatic variations of the last 2500 years. The relative abundance of dinocyst taxa and principal component analysis show a major transition at about 300 cal. years BP. It is characterized by the disappearance of thermophilic taxa Spiniferites mirabilis-hyperacanthus and Impagidinium sphaericum and the increase of polar–subpolar taxa Impagidinium pallidum and Pentapharsodinium dalei. Sea-surface temperature (SST) estimates suggest warmer conditions than present (anomaly～+2 °C) averaging at 7 °C in summer until 300 cal. years BP, although cooling pulses are recorded around 1700, 1500, 1200 and 800 cal. years BP. The last 300 years were marked by a cooling from 7.6 to 3.5 °C and sea-ice cover increasing up to 7 months/yr. The results demonstrate that the Fram Strait area is sensitive to hydroclimatic variations, notably with respect to sea-ice and SSTs, which are linked to the relative strength of northward flow of North Atlantic waters to the East and southward outflow of cold and fresh waters from the Arctic Ocean. Based on our data, the warmest part of our record around 1320 cal. years BP is the only interval of the last 2500 years that provides a possible analogue for the modern post-AD 2000 interval, which is characterized by sea-ice free conditions.     

Growth and bioactive secondary metabolites of arctic Protoceratium reticulatum (Dinophyceae)

Harmful algal blooms are mainly caused by marine dinoflagellates and are known to produce potent toxins that may affect the ecosystem, human activities and health. Such events have increased in frequency and intensity worldwide in the past decades. Numerous processes involved in Global Change are amplified in the Arctic, but little is known about species specific responses of arctic dinoflagellates. The aim of this work was to perform an exhaustive morphological, phylogenetical and toxinological characterization of Greenland Protoceratium reticulatum and, in addition, to test the effect of temperature on growth and production of bioactive secondary metabolites. Seven clonal isolates, the first isolates of P. reticulatum available from arctic waters, were phylogenetically characterized by analysis of the LSU rDNA. Six isolates were further characterized morphologically and were shown to produce both yessotoxins (YTX) and lytic compounds, representing the first report of allelochemical activity in P. reticulatum. As shown for one of the isolates, growth was strongly affected by temperature with a maximum growth rate at 15 °C, a significant but slow growth at 1 °C, and cell death at 25 °C, suggesting an adaptation of P. reticulatum to temperate waters. Temperature had no major effect on total YTX cell quota or lytic activity but both were affected by the growth phase with a significant increase at stationary phase. A comparison of six isolates at a fixed temperature of 10 °C showed high intraspecific variability for all three physiological parameters tested. Growth rate varied from 0.06 to 0.19 d⁻¹, and total YTX concentration ranged from 0.3 to 15.0 pg  YTX cell⁻¹ and from 0.5 to 31.0 pg YTX cell⁻¹ at exponential and stationary phase, respectively. All six isolates performed lytic activity; however, for two isolates lytic activity was only detectable at higher cell densities in stationary phase.     

Impact of nutrients and herbivory by Eccritotarsus catarinensis on the biological control of water hyacinth,Eichhornia crassipes

Many water hyacinth infestations in South Africa are the symptom of eutrophication, and as a result, biological control of this weed is variable. This study examined the effects of herbivory by the mirid, Eccritotarsus catarinensis, on water hyacinth grown at high, medium and low nitrogen (N) and phosphorus (P) nutrient concentrations. Water nutrient concentration appears to be the overriding factor affecting plant growth parameters of water hyacinth plants—at high nutrient concentrations, leaf and daughter plant production were more than double than at low nutrient concentrations, while stem length was twice as great at high nutrient concentrations compared to low concentrations. Chlorophyll content was also twice as high at high nutrient concentrations than low concentrations. Conversely, flower production at high nutrient concentrations was less than half that at low concentrations. Herbivory by E. catarinensis did not have as great an effect on water hyacinth vigour as nutrient concentration did, although it significantly reduced the production of daughter plants by 23 ± 9%, the length of the second petiole by 13 ± 5%, and chlorophyll content of water hyacinth leaves by 15 ± 6%. In terms of insect numbers, mirids performed better on plants grown under medium nutrient conditions (99 ± 28 S.E.), compared to high nutrient concentrations (52 ± 27 S.E.), and low nutrient concentrations (25 ± 30 S.E.). Thus, these results suggest that the fastest and most significant reduction in water hyacinth proliferation would be reached by lowering the water nutrient concentrations, and herbivory by E. catarinensis alone is not sufficient to reduce all aspects of water hyacinth vigour, especially at very high nutrient concentrations.     

Purification and characterization of an extracellular laccase from a Pseudomonas sp. LBC1 and its application for the removal of bisphenol A

The Pseudomonas sp. LBC1 produced extracellular laccase when grown in the nutrient broth. The enzyme was purified using acetone precipitation and an anion-exchange chromatography. The molecular weight of the purified laccase was estimated as 70 kDa by sodium dodecyl sulfate polyacrylamide gel electrophoresis. An enzyme showed maximum substrate specificity towards o-tolidine than other substrates of laccase including 2,2′-azinobis, 3-ethylbenzothiazoline-6-sulfonic acid, hydroquinone, N,N′-dimethyl phenylene diamine, syringic acid and veratryl alcohol. The optimum pH and temperature for the laccase activity were 4.0 and 40 °C, respectively. Cyclic voltammogram revealed the redox potential of purified enzyme as 0.30 V. The laccase was stable up to 40 °C and within pH range 6.0–8.0. Sodium azide and EDTA strongly inhibited laccase activity. The purified laccase completely degraded the higher concentration of bisphenol A within 5 h. Biodegradation metabolites of bisphenol A were characterized by using FTIR, HPLC and GC–MS.     

Rising temperatures may increase growth rates and microcystin production in tropical Microcystis species

Rising temperatures (1.4–6 °C) due to climate change have been predicted to increase cyanobacterial bloom occurrences in temperate water bodies; however, the impacts of warming on tropical cyanobacterial blooms are unknown. We examined the effects of four different temperatures on the growth rates and microcystin (MC) production of five tropical Microcystis isolates (M. ichthyoblabe (two strains), M. viridis, M. flos-aquae, and M. aeruginosa). The temperature treatments are based on current temperature range in Singapore's reservoirs (27 °C and 30 °C), as well as projected mean (33 °C) and maximum temperatures (36 °C) based on tropical climate change estimates of +6 °C in air temperature. Increasing temperatures did not significantly affect the maximum growth rates of most Microcystis strains. Higher growth rates were only observed in one M. ichthyoblabe strain at 33 °C and M. flos-aquae at 30 °C where both were isolated from the same reservoir. MC-RR and MC-LR were produced in varying amounts by all four species of Microcystis. Raised temperatures of 33 °C were found to boost total MC cell quota for three Microcystis strains although further increase to 36 °C led to a sharp decrease in total MC cell quota for all five Microcystis strains. Increasing temperature also led to higher MC-LR:MC-RR cell quota ratios in M. ichthyoblabe. Our study suggests that higher mean water temperatures resulting from climate change will generally not influence growth rates of Microcystis spp. in Singapore except for increases in M. ichthyoblabe strains. However, toxin cell quota may increase under moderate warming scenarios depending on the species.     

Proteomic analysis of the adaptation to warming in the Antarctic bacteria Shewanella frigidimarina

Antarctica is subjected to extremely variable conditions, but the importance of the temperature increase in cold adapted bacteria is still unknown. To study the molecular adaptation to warming of Antarctic bacteria, cultures of Shewanella frigidimarina were incubated at temperatures ranging from 0 °C to 30 °C, emulating the most extreme conditions that this strain could tolerate. A proteomic approach was developed to identify the soluble proteins obtained from cells growing at 4 °C, 20 °C and 28 °C. The most drastic effect when bacteria were grown at 28 °C was the accumulation of heat shock proteins as well as other proteins related to stress, redox homeostasis or protein synthesis and degradation, and the decrease of enzymes and components of the cell envelope. Furthermore, two main responses in the adaptation to warm temperature were detected: the presence of diverse isoforms in some differentially expressed proteins, and the composition of chaperone interaction networks at the limits of growth temperature. The abundance changes of proteins suggest that warming induces a stress situation in S. frigidimarina forcing cells to reorganize their molecular networks as an adaptive response to these environmental conditions.     

Development and characterization of a new Bombyx mori cell line for protein expression

A Bombyx mori continuous cell line, designated DZNU-Bm-17, was established from larval ovaries. The cells were initially grown in MGM-448 insect cell culture medium supplemented with 10% fetal bovine serum and 3% heat inactivated B. mori hemolymph at 25 ± 1 °C and later adapted gradually to TNM-FH medium. Partially adhered refractive cells were the predominant cell type in the culture. The cells took about 1055 days to complete 100 passages in TNM-FH medium. The population doubling time of the cell line was about 30–34 h at 25 ± 1 °C. The cell population was largely diploid, but a few triploids and tetraploids were also observed. DNA profiles using simple sequence repeat loci established the differences between the DZNU-Bm-1, Bm-5, DZNU-Bm-12, DZNU-Bm-17, and BmN cell lines. The cell line was susceptible to budded virus of B. mori nucleopolyhedrovirus (BmNPV), and 85–92% of the cells harbored BmNPV with an average of 15 occlusion bodies/infected cell. The cells expressed the luciferase and green fluorescent proteins using the BmNPV bacmid vector. We suggest the usefulness of the DZNU-Bm-17 cell line for BmNPV-based baculoviral expression studies.     

In vivo temperature limitations of photosynthesis in Phaseolus vulgaris L

The purpose of this study was to evaluate the temperature response of photosynthesis in two common bean genotypes differing in crop yield when grown under warm conditions. The cultivar Nobre is sensitive to high temperatures, whereas Diplomata shows better crop yield under high temperatures. Plants were grown in a greenhouse prior to transferring to a controlled environment cabinet for the temperature treatments. In a first experiment, 30 days-old plants were subjected to a short exposure (1 day) at temperatures that varied from 9 °C to 39 °C. Diplomata had lower net CO₂ assimilation rate (A) at 15 °C and 21 °C, but higher from 27 °C to 39 °C. Photosynthetic parameters calculated from modeling the response of A to the intercellular CO₂ concentration suggested that the different temperature responses of the two genotypes are caused by different rates of diffusion of CO₂ to the assimilation site, not by differences in biochemical limitations of photosynthesis. While stomatal conductance (g_s) did not differ between the genotypes, mesophyll conductance (g_m) was slightly greater for Nobre at 15 °C, but much higher in Diplomata from 21 °C to 39 °C. In a second experiment, no difference was observed in biomass accumulation between the two genotypes after growth for 24 days under a 35/20 °C (day/night) regime. Hence, the differences in photosynthesis did not cause variation in plant growth at the vegetative stage. The differential genotypic response of g_m to temperature suggests that g_m might be an important limitation to photosynthesis in Nobre, the common bean genotype sensitive to elevated temperature. However, more studies are needed employing other methods for g_m evaluation to validate these results.     

Development response of Spodoptera exigua to eight constant temperatures: Linear and nonlinear modeling

Temperature-dependent development of Spodoptera exigua (Hübner) were evaluated at eight constant temperatures of 12, 15, 20, 25, 30, 33, 34 and 36 °C with a variation of 0.5 °C on sugar beet leaves. No development occurred at 12 °C and 36 °C. Total developmental time varied from 120.50 days at 15 °C to 14.50 days at 33 °C. As temperature increased from 15 °C to 33 °C, developmental rate (1/developmental time) of S. exigua increased but declined at 34 °C. The lower temperature threshold (T_min) was estimated to be 12.98 °C and 12.45 °C, and the thermal constant (K) was 294.99 DD and 311.76 DD, using the traditional and Ikemoto–Takai linear models, respectively. The slopes of the Ikemoto–Takai linear model for different immature stages were different, violating the assumption of rate isomorphy. Data were fitted to three nonlinear models to predict the developmental rate and estimate the critical temperatures. The T_min values estimated by Lactin-2 (12.90 °C) and SSI (13.35 °C) were higher than the value estimated by Briere-2 (8.67 °C). The estimated fastest development temperatures (T_fast) by the Briere-2, Lactin-2 and SSI models for overall immature stages development of S. exigua were 33.4 °C, 33.9 °C and 32.4 °C, respectively. The intrinsic optimum temperature (T_Φ) estimated from the SSI model was 28.5 °C, in which the probability of enzyme being in its native state is maximal. The upper temperature threshold (T_max) values estimated by these three nonlinear models varied from 34.00 °C to 34.69 °C. These findings on thermal requirements can be used to predict the occurrence, number of generations and population dynamics of S. exigua.     

Temperature modulates hepatic carbohydrate metabolic enzyme activity and gene expression in juvenile GIFT tilapia (Oreochromis niloticus) fed a carbohydrate-enriched diet

The effects of rearing temperature on hepatic glucokinase (GK), glucose-6-phosphatase (G6Pase) and Glucose-6-phosphate dehydrogenase (G6PD) activity and gene expression were studied in GIFT (genetically improved farmed tilapia) tilapia fed a high carbohydrate diet containing 28% crude protein, 5% crude lipid and 40% wheat starch. Triplicate groups of fish (11.28 g initial body weight) were fed the diet for 45 days at 22 °C, 28 °C or 34 °C. At the end of the trial, final body weight of juvenile at 28 °C (59.12 g) was higher than that of the fish reared at 22 °C (27.13 g) and 34 °C (43.17 g). Feed intake, feed efficiency and protein efficiency ratio were also better at 28 °C. Liver glycogen levels were higher at 28 °C, while plasma glucose levels were higher in the 22 °C group. Significant (P<0.05) effects of water temperature on enzymes activities and gene expression were observed. Hepatic GK activity and mRNA level were higher at 28 °C than at 34 °C. Higher G6Pase and G6PD activity and gene expression were observed at 22 °C. Overall, the data show that juveniles reared at 28 °C exhibited enhanced liver glycolytic capacity. In contrast, hepatic gluconeogenesis and lipogenesis were increased by low temperature (22 °C).     

Growth and domoic acid content of Pseudo-nitzschia australis isolated from northwestern Baja California,Mexico, cultured under batch conditions at different temperatures and two Si:NO3 ratios

Domoic acid (DA) poisoning in the southern part of the California Current System has been associated typically with blooms of Pseudo-nitzschia australis. The environmental variables that promote growth and DA production in the Mexican part of this system have not been identified. The present study investigated the effect of temperature and two nutrient ratios on the growth characteristics and DA content of two (BTS-1, BTS-2) P. australis strains isolated from the Pacific coast of northern Baja California peninsula, México. Of the different temperatures assayed (10, 12, 14, 15, 18 and 20 °C), the maximum cell abundance was detected at 12 °C for BTS-2 and 14 °C for BTS-1. The highest maximum specific growth rate (1.69 day⁻¹) was measured at 15 °C for BTS-2. With the exception of cells maintained at 15 °C, growth characteristics were similar in P. australis cultured in a high Si:NO₃ (2.5) or low Si:NO₃ (0.5) ratio at each temperature. Dissolved (dDA) and cellular (cDA) DA content measured at the stationary phase of growth was similar in cells cultivated at the different temperatures. No difference in cDA (between 0.11 and 1.87 pg DA cell⁻¹) was observed in cells cultivated at the two nutrient ratios. To evaluate if P. australis accumulates DA (cDA + dDA) at different stages of the culture and not only during the stationary phase of growth, the BTS-1 strain was cultivated at 14 °C and the content of this toxin was measured during culture development. The cultures were maintained at high (HL; 200 μmol quanta m⁻² s⁻¹) and low light (LL; 30 μmol quanta m⁻² s⁻¹) and in the two nutrient ratios to evaluate the effect of these variables on DA content. The photosynthetic performance and pigment concentration were measured as indicators of the physiological condition of the cells. cDA was detected in all culture conditions and during the different stages of growth. The highest DA content was measured during the lag phase of growth and it was present mainly in the medium (dDA = 70.83 pg DA cell⁻¹). Cells cultivated at HL produced more DA than LL cultured cells. P. australis cultured in HL presented lower photosynthetic rates than LL cells and had similar concentrations of photoprotective pigments and the highest maximum photosynthetic rates were detected during the lag phase of growth in all culture conditions. The results demonstrate that P. australis from northern Baja California peninsula presents a narrow temperature range for optimal growth under batch culture conditions. P. australis produce DA at different stages of growth, and DA content was related to the light intensity at which the cells were cultivated.     

Mathematical modeling of Microcystis aeruginosa growth and [D-Leu1] microcystin-LR production in culture media at different temperatures

The effect of temperature (26 °C, 28 °C, 30 °C and 35 °C) on the growth of native CAAT-3-2005 Microcystis aeruginosa and the production of Chlorophyll-a (Chl-a) and Microcystin-LR (MC-LR) were examined through laboratory studies. Kinetic parameters such as specific growth rate (μ), lag phase duration (LPD) and maximum population density (MPD) were determined by fitting the modified Gompertz equation to the M. aeruginosa strain cell count (cells mL⁻¹). A 4.8-fold increase in μ values and a 10.8-fold decrease in the LPD values were found for M. aeruginosa growth when the temperature changed from 15 °C to 35 °C. The activation energy of the specific growth rate (Eμ) and of the adaptation rate (E₁/LPD) were significantly correlated (R² = 0.86). The cardinal temperatures estimated by the modified Ratkowsky model were minimum temperature = 8.58 ± 2.34 °C, maximum temperature = 45.04 ± 1.35 °C and optimum temperature = 33.39 ± 0.55 °C.Maximum MC-LR production decreased 9.5-fold when the temperature was increased from 26 °C to 35 °C. The maximum production values were obtained at 26° C and the maximum depletion rate of intracellular MC-LR was observed at 30–35 °C. The MC-LR cell quota was higher at 26 and 28 °C (83 and 80 fg cell⁻¹, respectively) and the MC-LR Chl-a quota was similar at all the different temperatures (0.5–1.5 fg ng⁻¹).The Gompertz equation and dynamic model were found to be the most appropriate approaches to calculate M. aeruginosa growth and production of MC-LR, respectively. Given that toxin production decreased with increasing temperatures but growth increased, this study demonstrates that growth and toxin production processes are uncoupled in M. aeruginosa. These data and models may be useful to predict M. aeruginosa bloom formation in the environment.     

The toxicity and intraspecific variability of Alexandrium andersonii Balech

The toxicity of Alexandrium andersonii Balech is unclear and its intraspecific variability has yet to be studied. To address these gaps in our knowledge, in the present work five strains of A. andersonii from four different localities were characterized. The results showed that despite genetic homogeneity in the 5.8-ITS (internal transcribed spacer) and large subunit (LSU) regions and similar growth rates, strains originating from different locations varied with respect to cell size, the ratios of certain pigments, and their growth patterns. Cultures of the strains grown at 20 °C were analyzed for toxicity using four different methodologies. The two officially established methods, mouse bioassay and high-performance liquid chromatography with fluorescence detection (HPLC-FLD) and post-column reaction analysis of PSP toxins, failed to show the toxicity of any strain. Strains grown at 14 °C were also negative for PSP toxins by HPLC-FLD. However, strains grown at 20 °C exhibited both a response characteristic of the presence of toxin-inhibiting voltage-gated sodium channels, as demonstrated in a neuroblastoma neuro-2a cell-based assay, as well as hemolytic activity in a sheep red blood cell assay.