氮磷对水华束丝藻生长及生理特性的影响

从滇池分离得到水华束丝藻藻株,以BG11为基础培养基,在(25±1)℃,光照强度20μE/m2.s,光/暗周期为16h:8h的培养条件下,水华束丝藻的生长周期约为28d,以光密度(OD665)为指标的生长曲线符合典型的"S"型;在实验浓度范围内,N、P的浓度变化均对水华束丝藻的生长产生影响,在一定范围内([NO3--N]:1.6—245.1 mg/L;[PO43--P]:0.3—1.4 mg/L),高浓度的N、P有利于水华束丝藻的生长,但浓度过高([NO3--N]>245.1 mg/L;[PO43--P]>6.9 mg/L)时,则对其生长产生抑制作用;N、P在缺乏和浓度很低时可对水华束丝藻光系统Ⅱ活性(Fv/Fm)产生显著影响(p<0.01);本实验结果还表明,N、P浓度的改变可影响水华束丝藻硝酸还原酶、酸性磷酸酶和碱性磷酸酶的活性;在低N、P浓度下,水华束丝藻的硝酸还原酶和酸性磷酸酶活性升高,显著高于正常BG11培养时的活性(p<0.05)。这一结果表明,在低N、P营养环境中,水华束丝藻可通过调节自身的酶系统来提高对营养的吸收和利用,从而满足自身生长和细胞增殖的需要。     

GROWTH RESPONSE OF A NITROGEN FIXER (ANABAENA FLOS-AQUAE,CYANOPHYCEAE) TO LOW NITRATE1

Anabaena flos-aquae (Lyngb.) Bréb. was grown in varying concentrations of nitrate. Specific growth rates, as estimated in batch culture, were constant and approached the maximum rate at all concentrations of NO₃^?-N tested bewteen 0 and 400 μ/L. Steady-state biomass, as determined in semicontinuous culture, did not vary with NO₃^? at slower dilution rates. However at a faster dilution rate, significantly less biomass occurred in intermediate concentrations of NO₃^? than in either higher or lower concentrations. The results indicate that both growth rate and standing crop are maximized by either N₂ fixation or NO₃^? assimilation, but extracellular NO₃^? reduces the rate of N₂ fixation. Consequently, at very low NO₃^? concentrations, growth is virtually maximized by N₂ fixation alone, and at high concentrations of NO₃^?, N₂ fixation is inhibited but growth is maximized by assimilation of NO₃^?. At intermediate concentrations of NO₃^?, growth becomes a function of NO^3? assimilation augmented by N₂ fixation. In this case, full growth potential is realized only if hydraulic residence time is sufficiently long to compensate for the reduced rate of N₂ fixation. Growth rate and standing crop are not diminished in response to the large amount of energy allocated to N₂ fixation. Instead, other cellular processes are probably affected negatively during N₂ fixation.     

STORAGE OF PHOSPHORUS IN NITROGEN-FIXING ANABAENA FLOS-AQUAE (CYANOPHYCEAE)1

P accumulation and metabolic pathway in N₂-fixing Anabaena flos-aquae (Lyngb.) Bréb were investigated in P-sufficient (20 μMP) and P-limited (2 μMP) turbidostats in combined N-free medium. The cyanobacterium grew at its maximum rate (μ_max, 1.13 d^?1) at the high P concentration and at 65% of μ_max under P limitation, with total cell P concentrations (Q_P) at steady states of 12.0 and 5.2 fmol·cell^?1, respectively. At steady state, polyphosphates (PP_i) accounted for only 3% of Q_P (0.4 fmol·cell^?1) in P-rich cells. Its concentration in P-limited cells was 5.8% (0.3 fmol·cell^?1). On the other hand, sugar P was very high at 22% of Q_P in P-rich cells and was undetectable in P-limited cells. Pulse chase experiments with ³²P showed that P-rich cells initially incorporated the labeled P into the acid-soluble PP_i fraction within the first few minutes and to a lesser extent into nucleotide P. Radioactivity in the PP_i then declined rapidly with concomitant increases in sugar P and nucleotide P fractions. In contrast, in P-limited cells, no radiolabel was detected in acid-soluble PP_i, and ³²P was initially incorporated into nucleotide P, sugar P, and ortho P fractions. The latter two fractions then subsequently declined. Therefore, under N₂-fixing conditions the cyanobacteria appeared to store P as sugar P and also utilize P through different pathways under P-rich and -limited conditions. When nitrate was supplied as the N source under P-sufficient conditions, PP_i accounted for about 15% of steady-state Q_P, but no sugar P was detected. Therefore, the same organism stored P in different cell P fractions depending on its N sources.     

Characterization of saxitoxin production and release and phylogeny of sxt genes in paralytic shellfish poisoning toxin-producing Aphanizomenon gracile

The freshwater cyanobacterium Aphanizomenon gracile is one of the most widely distributed producers of the potent neurotoxins saxitoxin (STX) and its derivatives (paralytic shellfish poisoning toxins, PSP toxins). However, the phylogeny of STX biosynthesis genes and the regulation of STX production and release remain poorly studied in the genus Aphanizomenon. In this study, two A. gracile strains from Spanish freshwaters were grown in semi-continuous cultures under three temperatures (15, 20 and 28 °C) and their STX production and release were determined by Enzyme-Linked ImmunoSorbent Assay (ELISA). STX production was stable along the temperature range, with 1.4–2.3-fold shifts in biomass-standardized STX contents, and maxima of 0.22 μg equivalent STX mg⁻¹ dry weight 15.3 fg equiv STX cell⁻¹ and 15.1 μg equiv STX mg⁻¹ Chl a. The extracellular fraction was remarkably high (13.6–35.3%), not clearly affected by temperature but with nitrate-depleted medium (BG11₀) inducing a 2-fold increase in extracellular content. STX production and release were not directly related to growth rates. The 16S rRNA phylogenetic analyses in sixteen A. gracile strains from Spanish and German freshwaters showed that PSP-producing A. gracile grouped within a monospecific and highly supported cluster, together with PSP-producing Aphanizomenon sp. NH-5 and clearly separated from a monospecific Aphanizomenon flos-aquae cluster. The sixteen A. gracile strains formed also monospecific and highly supported clusters for PSP-biosynthesis genes (sxtG, sxtI, sxtH and sxtX) together with Aphanizomenon sp. NH-5. This study evidences an elevated extracellular proportion of STX in A. gracile with importance for risk assessment, and supports the identification of Aphanizomenon sp. NH-5 as A. gracile.     

Physiological variables determined under laboratory conditions may explain the bloom of Aphanizomenon ovalisporum in Lake Kinneret

Response of Aphanizomenon ovalisporum to certain environmental parameters was studied to gain a better understanding of the conditions which may have stimulated its autumnal bloom in Lake Kinneret. Optimal temperature for A. ovalisporum growth was 26–30?°C, resulting in growth rates of 0.2–0.3?day^?1, similar to those observed in the lake. Maximal rate of CO₂ fixation (assimilation numbers of 6–8?μg?C?μg^?1?Chl?h^?1) was obtained at low irradiances (I _k of 40–100?μmol?photons?m^?2?s^?1), 200?μM Pi and low N:Pi ratios. Growth was strongly affected by phosphorus availability, reaching a maximum at Pi concentrations above 40?μM. The high demand for phosphorus was indicated by an increase in alkaline phosphatase activity. The relative abundance of Pi in the cells increased by 4-fold in Pi-rich compared with Pi-limited cultures. Uptake of Pi was faster in Pi-depleted compared with Pi-sufficient cells. Maximal photosynthetic rates and K_1/2(HCO₃ ^?) were 140–220?μmol?O₂?mg^?1?Chl?h^?1 and 10–24?μM, respectively. At pH 7.0 the K _1/2(CO₂) was 2.2 and fell to 0.04?μM at pH 9.0. These data indicated that A. ovalisporum is a HCO₃ ^? user, and can explain its high photosynthetic rates during the bloom, under high pH and low dissolved CO₂ conditions. Na⁺ concentrations of about 5?mM were essential for A. ovalisporum growth at high pH approaching values in the lake.     

MORPHOLOGICAL AND 16S rRNA GENE EVIDENCE FOR RECLASSIFICATION OF THE PARALYTIC SHELLFISH TOXIN PRODUCING APHANIZOMENON FLOS‐AQUAE LMECYA 31 AS APHANIZOMENON ISSATSCHENKOI (CYANOPHYCEAE)1

A taxonomic reevaluation of the paralytic shellfish toxin (saxitoxins) producing cyanobacterium Aphanizomenon flos‐aquae Ralfs ex Born. & Flah. LMECYA31 was done using morphology and 16S rRNA gene sequences. We found that strain LMECYA31 was incorrectly identified as Aph. flos‐aquae based on (a) lack of bundle formation in trichomes, (b) shape of terminal cells in the trichomes, (c) lower similarity (<97.5%) in the 16S rRNA gene sequences relative to those of Aph. flos‐aquae, and (d) comparison within a phylogenetic tree of 16S rRNA gene sequences. The shape of the terminal trichome cells and the shape and size of the vegetative cell, heterocyst, and akinete in strain LMECYA31 match characters of Aph. issatschenkoi (Ussachew) Proschkina‐Larvernko. 16S rRNA gene sequences and phylogenetic clusters constructed from 16S rRNA gene sequences support our conclusion that strain LMECYA31 should be Aph. issatschenkoi.     

The effect of environmental parameters and cyanobacterial blooms on phytoplankton dynamics of a Portuguese temperate Lake

The increasing occurrence of cyanobacterial blooms in freshwaters is of great concern due to the ability of many cyanobacteria to produce cyanotoxins. In the present work, the eutrophied Vela Lake (Central Portugal), used for recreational purposes and as a water source for agriculture, was monitored every fortnight between 2000 and 2001. Phytoplankton diversity and densities were measured and correlated to environmental parameters. A seasonal phytoplanktonic succession was observed and it was mainly correlated with conductivity, temperature, total suspended solids and nutrients availability (particularly phosphorus). Diatoms were dominant during winter months (inferior temperatures and higher nutrients availability) followed by green algae in early spring and then cyanobacteria from late spring until early autumn (less nutrient availability and higher temperatures). A massive cyanobacterial bloom of Aphanizomenon flos-aquae occurred early in May 2001 and was preceded by the lowest nitrogen levels measured in the water during all the study period. At the time of this bloom senescence, dissolved oxygen was severely depleted and a massive death of ichthyofauna was recorded. A Microcystis aeruginosa bloom was also detected in July 2001 and it occurred following a rapid decrease in abundance of green algae and diatoms. By considering not only the environmental parameters but also the occurrence of cyanobacterial blooms as explanatory variables in a canonical correspondence analysis, the variance explained for the phytoplanktonic assemblage during the study period was increased in about 7% achieving a total of 61.0%, indicating a correlation that may be due to the known competitive advantage and/or allelopathy of the bloom-forming cyanobacteria towards microalgae.     

溶藻细菌L7对水华鱼腥藻氮代谢的影响

【目的】进一步探明藻菌关系,研究溶藻细菌对藻类氮代谢的影响及其作用机制。【方法】将水华鱼腥藻和溶藻细菌L7按两种比例接种入BG11培养液中,在室内进行共培养(藻细胞初始密度为1.21×108cells/L;溶藻细菌L7初始密度分别为1.75×107、1.75×108CFU/mL)。连续7 d测定藻细胞数、异形胞频率和藻细胞内的硝酸还原酶(NR)活性、谷氨酰胺合成酶(GS)活性、谷氨酸合成酶(GOGAT)活性、蛋白质含量、丙二醛(MDA)含量。【结果】低密度溶藻细菌L7能够促进藻生长(第7天藻细胞密度是对照组的1.58倍),增加异形胞频率(第7天高于对照组66.67%);高密度则会抑制藻生长(第7天藻细胞密度相比对照组下降98.84%),降低异形胞频率(第7天为0)。在藻细胞内氮代谢关键酶活性方面,接种后2 5 d,两处理组中藻细胞内NR和GOGAT活性均极显著高于对照组(P<0.01);接种后0 5 d,高密度处理组的GS活性极显著高于对照组(P<0.01),而低密度处理组的则在大部分时间内极显著低于对照组(P<0.01)。在整个实验期内,低密度处理组中藻细胞内蛋白质含量一直极显著高于对照组(P<0.01);而在高密度处理组中,除第5天外,细胞内蛋白质含量则全部极显著低于对照组(P<0.01)。接种后2 4 d,高密度处理组中藻细胞内MDA含量呈现上升趋势,并极显著高于其余两组(P<0.01)。【结论】低密度溶藻细菌L7能够提高水华鱼腥藻对氮源的需求,加速蛋白质合成,促进氮代谢;而高密度溶藻细菌L7会对藻细胞产生过氧化伤害,阻碍蛋白质合成和氮代谢过程。     

Characterization of human monocyte activation by a water soluble preparation of Aphanizomenon flos-aquae

Aphanizomenon flos-aquae (AFA) is a fresh-water microalgae that is consumed as a nutrient-dense food source and for its health-enhancing properties. The current research characterizes the effect of a water soluble preparation from AFA on human monocyte/macrophage function and compares the effect of AFA with responses from known agents that modulate the immune system. At 0.5 μg/ml the AFA extract robustly activated nuclear factor kappa B (NF-kappa B) directed luciferase expression in THP-1 human monocytic cells to levels at 50% of those achieved by maximal concentrations (10 μg/ml) of bacterial lipopolysaccharide (LPS). In addition, the AFA extract substantially increased mRNA levels of interleukin-1β(IL-1β) and tumor necrosis factor-α(TNF-α), and enhanced the DNA binding activity of NF-kappa B. The effects of AFA water soluble preparation were similar to the responses displayed by LPS, but clearly different from responses exhibited by tetradecanoyl phorbol acetate (TPA) and interferon-gamma (INF-γ). Pretreatment of THP-1 monocytes with factors known to induce hyporesponsiveness suppressed both AFA-dependent and LPS-dependent activation. These results suggest that the macrophage-activating properties of the AFA water soluble preparation are mediated through pathways that are similar to LPS-dependent activation.