氮磷营养因子对赤潮异弯藻生长的影响

研究了N、P营养浓度对赤潮异弯藻(Heterosigma akashiwo)生长的影响.结果表明,该藻的生长速率与N、P营养因子浓度的关系符合Monod公式.在NO₃^--N浓度达到7.5 mg·L^-1时,赤潮异弯藻开始生长;浓度为3.75～75 mg·L^-1时,赤潮异弯藻的比生长速率与NO₃^--N浓度成正比关系.N营养充足时,赤潮异弯藻的最大生长速率μ_m-n=0.3475·d^-1,K_s-n=18.91 mg·L^-1.PO₄^--P浓度为0～1.0 mg·L^-1时,赤潮异弯藻的比生长速率与P浓度成正比关系;P营养充足时,赤潮异弯藻的最大生长速率μ_m-p=0.3024·d^-1,K_s-p=0.4086 mg·L^-1.N/P达到25后藻细胞浓度达到最大,表明N/P为25时最适合赤潮异弯藻生长.赤潮异弯藻最适合在N 37.5～225.0 mg·L^-1、P 5.0～50.0 mg·L^-1、N/P=25条件下生长.     

The role of Azadinium spinosum (Dinophyceae) in the production of azaspiracid shellfish poisoning in mussels

Azaspiracids (AZAs) are a group of lipophilic polyether compounds first detected in Ireland which have been implicated in shellfish poisoning incidents around Europe. These toxins regularly effect shellfish mariculture operations including protracted closures of shellfish harvesting areas for human consumption. The armoured dinoflagellate Azadinium spinosum Elbrächter et Tillmann gen. et sp. nov. (Dinophyceae) has been described as the de novo azaspiracid toxin producer; nonetheless the link between this organism and AZA toxin accumulation in shellfish has not yet been established. In August 2009, shellfish samples of blue mussel (Mytilus edulis) from the Southwest of Ireland were analysed using liquid chromatography–tandem-mass spectrometry (LC–MS/MS) and were found to be above the regulatory limit (0.16 μg g⁻¹ AZA-equiv.) for AZAs. Water samples from this area were collected and one algal isolate was identified as A. spinosum and was shown to produce azaspiracid toxins. This is the first strain of A. spinosum isolated from Irish waters. The Irish A. spinosum is identical with the other two available A. spinosum strains from Scotland (3D9) and from Denmark (UTHE2) in its sequence of the D1–D2 regions of the LSU rDNA.A 24 h feeding trial of blue mussels (M. edulis) using an algal suspension of the Irish A. spinosum culture at different cell densities demonstrated that A. spinosum is filtered, consumed and digested directly by mussels. Also, LC–MS/MS analysis had shown that AZAs were accumulating in the shellfish hepatopancreas. The toxins AZA1 and -2 were detected in the shellfish together with the AZA analogues AZA3, AZA6, AZA17 and -19 suggesting that AZA1 and -2 are metabolised in the shellfish within the first 24 h after ingestion of the algae. The levels of AZA17 detected in the shellfish hepatopancreas (HP) were equivalent to the levels of AZA1 but in the remainder tissues the levels of AZA17 were four to five times higher than that of AZA1, only small quantities of AZA3 and -19 were present with negligible amounts of AZA6 detected after the 24 h period. This could have implications in the future monitoring of these toxins given that at present according to EU legislation only AZA1–AZA3 is regulated for. This is the first report of blue mussels’ (M. edulis) feeding on the azaspiracid producing algae A. spinosum from Irish waters.     

Environmental and nutritional factors which regulate population dynamics and toxin production in the dinoflagellate Alexandrium catenella

The effects of environmental and nutritional factors on population dynamics and toxin production were examined in Alexandrium catenella, maintained in enriched K media in laboratory cultures. Starting with a density of 50 cell ml⁻¹, the dinoflagellate population typically showed a lag phase and an exponential growth phase which lasted 14 days each, and then entered the stationary phase, with a maximal capacity of 12–18,000 cell ml⁻¹-. Population densities showed distinct diurnal patterns, with population growth beginning 2–4 hours in darkness. The optimal physical conditions for growth were pH 8.5,salinity of 30–35‰, temperature of 20–25°C, and photoperiod of 14//10D to 16L/8D. The cell cycle was determined by flow cytometry on synchronized batch cultures maintained at optimal pH, salinity, temperature and under 5 different photoperiod regimes. It was found that the G₁ phase was timed to end at approximately 3 h after onset of darkness, and the G₂/M phase had begun at 4 hours. Nutrient supply markedly affected population growth. Under optimal physical conditions, the optimal concentrations for macronutrients and micronutrients were: NH⁺⁻⁴- 0.025–0.2 mM,NO⁻³ 0.22–8.83 mM, glycerophosphate0.04–0.06 mM, silicate 0.1–0.54 mM; FeEDTA 0.07–0.11 mM;Co 0.1 μM, Cu 0.005–0.04 μM; Mn 0.22–7.2 μM;Mo 0.03–0.6 μM; Se 0.02–0.1 μM; Zn 0.04–1.6μM; thiamin 0.075–6 μM; vitamin B₁₂0.0004–0.004 μM; biotin 0.007–0.015 μM; EDTA5–40 μM. The toxin profile of A. catenella was determined by HPLC and found to include in descending order: GTX-4, GTX-3, GTX-1, B2, neosaxitoxin, saxitoxin. Toxin content per cell was highest in cell populations in the early exponential phase. The highest toxin per litre medium was recorded at 20°C at the beginning of the stationary phase,when cell density was highest and toxin/cell was still relatively high. At10°C, the cell density was low while the amount of toxin/cell was high;while at 30°C, the population at full capacity was low and the toxin/cell was also low. The population and toxin data thus provided an explanation for the peak level of PSP contamination in shellfish during the months of March–April around the eastern and southern side of Hong Kong and a minor peak extending to the western side in September–October, when the physical conditions of the seawater provided the right environment for toxin accumulation. Toxin content in the dinoflagellate reached its maximum during the S-phase of the cell cycle. Nitrogen restriction in the medium reduced population growth and toxin production, while phosphorus restriction reduced only population growth but enhanced toxin accumulation in the cells. This revised version was published online in August 2006 with corrections to the Cover Date.     

Effect of silicon on the growth and phosphorus uptake of rice

A pot experiment was conducted to measure the effect of silicon on phosphorus uptake and on the growth of rice at different P levels. Rice (Oryza sativa L. cv. Akebono) was cultured in Kimura B nutrient solution without and with silicon (1.66 mM Si) and with three phosphorus levels (0.014 mM P, low; 0.21 mM, medium; and 0.70 mM, high).Shoot dry weight with Si (+Si) in solution increased with increasing P level, while shoot weight without Si (–Si) was maximum at 0.21 mM P, suggesting that +Si raised the optimum P level for rice. +Si increased shoot weight more when P was low or high than when P was medium.The concentration and amount of inorganic P in shoots increased with increasing P level. +Si did not significantly decrease P uptake by rice at 0.014 mM P, however, uptake at 0.21 and 0.70 mM P was 27 and 30 percent less than uptake with –Si, respectively. In –Si with 0.21 and 0.70 mM P, inorganic P in shoots was more than double the concentration in shoots grown in +Si solutions.The Si concentration in shoots decreased slightly with increasing P level, although Si uptake was not significantly affected by P. +Si decreased the uptake of Fe and Mn by an average of 20 and 50 percent, respectively, thus P/Mn and P/Fe ratios increased in the shoot when P was low.From the results above, the beneficial effect of Si on the growth of rice was clearly shown when P was low or high. This effect may have resulted from decreased Mn and Fe uptake, and thus increased P availability within P deficient plants, or from reduced P uptake when P was high.     

Dependence of Pseudomonas aeruginosa continous culture biosurfactant production on nutritional and environmental factors

Summary Continuous culture studies with Pseudomonas aeruginosa were performed in order to establish nutritional and environmental conditions necessary for high production of biosurfactants. Empirical adjustments of the mineral medium formulation showed that better yields of the active compounds, rhamnolipids, are obtained by minimizing the concentration of the respective salts of magnesium, calcium, potassium, sodium and the trace elements. Improvements in performance were more evident when the intial substrate concentration, glucose, was increased up to 73 gl^-1. Further, the ranges for pH (6.2 to 6.4) and temperature (32° to 34°C) that yield high biosurfactant biosynthesis were established. Concerning the physiological state of the microorganism, rhamnolipid formation was restricted to specific growth rates lower than D=0.14 h^-1. By applying the conditions elaborated up to 300 mg rhamnose l^-1 h^-1 (equivalent to 685 mg rhamnolipid l^-1 h^-1) were obtained in a continuous production process.     

环境因子对东海原甲藻生长及脲酶活性的影响

以我国东南沿海大规模赤潮原因种东海原甲藻为实验材料,研究了环境因子对其生长及脲酶活性的调控作用。结果表明,东海原甲藻的适宜生长温度为20—25℃,而25℃下脲酶活性最高。在光强2μmol m-2s-1条件下细胞密度显著下降(P0.05),但仍能维持较高的脲酶活性(9.405 fmol h-1个-1)。在盐度20—40范围内,东海原甲藻能够维持快速生长和较高的脲酶活性。氮源组成对东海原甲藻生长无显著影响,但对脲酶活性影响较大。具体而言,东海原甲藻脲酶活性与尿素浓度呈显著正相关关系,而无机氮源NH+4和NO-3对其脲酶活性具有显著的抑制作用,在氮缺乏条件下脲酶活性明显增强。东海原甲藻脲酶活性对环境温度、光照、盐度和营养的响应特征可能是在长期进化中形成的生态适应策略,使其在无机氮源不足时得以转而利用有机氮源,从而在资源竞争中占据有利地位。     

Influence of environmental factors on the production of Potamothrix alatus paravanicus Poddubnaya & Pataridze (Tubificidae) in different areas of Lake Sevan

A modification of the Hynes' method was used to estimate production of Potamothrix alatus paravanicus Poddubnaya & Pataridze (Tubificidae), the dominant species of benthic invertebrate in Lake Sevan. Maximum production (5–24 g m^t\-2 wet weight) occurred in the sublittoral, while maximum biomass was found (8–17 g m^–2 wet weight) in the profundal. The turnover ratio of the average cohort varied between 2.4 and 5.0. The annual turnover ratio (P/B) decreased from 2.1–3.1 in the littoral to 0.1–0.4 in the profundal zone.A logarithmic correlation was found between P/B and temperature and oxygen regime, and depth and average size of clitellate specimens.     

Effect of culture conditions on the NADH/NAD ratio and total amounts of NAD(H) in chemostat cultures of Enterococcus faecalis NCTC 775

Abstract Enterococcus faecalis was grown in chemostat culture on various energy sources at dilution rates ranging from 0.05 h⁻¹ to 0.5 h⁻¹, under both aerobic and anaerobic conditions. NADH/NAD ratios and total nicotinamide adenine dinucleotide pool size (NAD(H)) were determined. It was found that the NADH/NAD ratio was controlled by the steady state product concentrations rather than by the degree of reduction of the energy source. Highest ratios were observed when NADH was reoxidized via ethanol formation, whereas in aerobic cultures, in which predominantly acetate was produced and oxidation of NADH occurred via the NADH oxidase, ratios were lowest. Addition of ethanol to the medium resulted in an increase of the NADH/NAD ratio, both aerobically and anaerobically. The total amount of NAD(H) was found to be influenced by the culture conditions. Under anaerobic conditions, the NADH oxidation (NAD reduction) rate appeared to correlate with the total amount of nicotinamide nucleotides. In contrast, no effect of the culture conditions on the total amount of NAD(H) was observed in aerobically grown cells.     

Influence of environmental and nutritional factors in the production of astaxanthin from Haematococcus pluvialis

Astaxanthin extracted from green algae is desirable in the food and pharmaceutical industries due to its antioxidant properties. The green unicellular clear water microalga Haematococcus pluvialis has a high production rate of astaxanthin; indeed, it contains more than 80% astaxanthin content in its cells. This remarkable astaxanthin production is commonly obtained under stress conditions such as nutrient deficiency (N or P), high NaCl concentrations, variations of temperature, and other factors. In this vein, a great research effort has been oriented to determine optimal conditions for astaxanthin production by H. pluvialis.The objective of the present study was the analysis of environmental factors, such as light intensity, aeration and nutrients on the growth and astaxanthin production of H. pluvialis. Maximum growth of H. pluvialis obtained was 3.5x10(5) cells/ml in BBM medium at 28 degrees C under continuous illumination (177 micromol photon m(-2)s(-1)) of white fluorescent light, with continuous aeration (1.5 v.v.m.). Meanwhile, maximal astaxanthin production was 98 mg/g biomass in BAR medium with continuous illumination (345 micromol photon m(-2)s(-1)), with 1 g/l of sodium acetate and without aeration.     

氮磷钾配比与用量对湘杂油763产量和经济效益的影响

为了筛选出冬油菜新品种"湘杂油763"在湖南油菜主产区推广的适宜氮磷钾配比及用量,于2008—2010年连续进行了2次该品种的氮磷钾配比及用量的大田小区试验,研究了不同氮磷钾配比及用量对其生物产量、籽粒产量、油分产量和经济效益的影响。结果表明:氮磷钾配比(N:P2O5:K2O)为1:0.50:0.50(处理B)和氮磷钾配比(N:P2O5:K2O)为1:0.43:0.57(处理G)的处理籽粒产量较高,分别达到2231和2065kg.hm-2,处理B显著高于除G外的其他处理;氮磷钾3种养分中,以氮肥对提高油菜籽粒产量的作用最大,其次为磷肥,作用最小者为钾肥;油分产量及植株生物产量变化规律与籽粒产量基本一致;按照处理B配比,籽粒产值为9816.97元.hm-2、施肥仅1641.23元.hm-2、施肥效益为6741.34元.hm-2,产投比达4.11;研究区域内N:P2O5:K2O=1:0.50:0.50,用量分别180(N)、90(P2O5)和90kg.hm-2(K2O)为较适宜施肥比例及用量。     

The effects of major environmental factors and nutrient limitation on growth and encystment of planktonic dinoflagellate Akashiwo sanguinea

The bloom-forming dinoflagellate Akashiwo sanguinea is commonly observed in estuarine and coastal waters around the world. Annually recurrent blooms of this species have been observed in the coastal waters of China, particularly in the Sishili Bay, Yantai since 2004. However, limited studies have been conducted on the recurrence mechanism of A. sanguinea other than periodical monitoring of its population dynamics and associated environmental variables. Thus, to further explore the bloom and succession mechanisms of A. sanguinea in the field, we studied the effects of major nutritional components on the growth and encystment of A. sanguinea, as well as the effects of key environmental factors on the growth of A. sanguinea through a series of laboratory trials. Our results indicated that A. sanguinea was able to grow well within the temperature range of 20–25 °C, salinity range of 20 - 30, with the maximum laboratory irradiance of 78.14 μE m⁻² s⁻¹, and was able to survive and grow in low nutrient. However, lower concentrations of nutrients (e.g., nitrate, phosphate) and higher ammonium exerted different degrees of limiting effects on the growth of A. sanguinea, and induced 2.3–21.24% of vegetative cells to form resting cysts simultaneously in laboratory cultures. On the other hand, very limited or no cyst formation was observed in nutrient-replete or extremely low nutrient cultures, indicating the threshold effect of nutritional stress on the encystment of A. sanguinea. The physiological strategy of encystment of A. sanguinea in nutrient-limiting environment facilitates the survival and succession of A. sanguinea species in fluctuating seawaters, and provides seed sources for reoccurring algal blooms under favorable environmental conditions.     

Multiple environmental controls on phytoplankton growth strategies determine adaptive responses of the N : P ratio

The controls on the ‘Redfield’ N : P stoichiometry of marine phytoplankton and hence the N : P ratio of the deep ocean remain incompletely understood. Here, we use a model for phytoplankton ecophysiology and growth, based on functional traits and resource‐allocation trade‐offs, to show how environmental filtering, biotic interactions, and element cycling in a global ecosystem model determine phytoplankton biogeography, growth strategies and macromolecular composition. Emergent growth strategies capture major observed patterns in marine biomes. Using a new synthesis of experimental RNA and protein measurements to constrain per‐ribosome translation rates, we determine a spatially variable lower limit on adaptive rRNA:protein allocation and hence on the relationship between the largest cellular P and N pools. Comparison with the lowest observed phytoplankton N : P ratios and N : P export fluxes in the Southern Ocean suggests that additional contributions from phospholipid and phosphorus storage compounds play a fundamental role in determining the marine biogeochemical cycling of these elements.     

Influence of nutritional and environmental factors on polysaccharide production by Azotobacter vinelandii cultured on 4-hydroxybenzoic acid

The capacity of 4-hydroxybenzoic acid to support exopolysaccharide (EPS) biosynthesis was investigated. Carbon source concentration, nitrogen supplementation, and other nutritional and environmental factors were optimized to obtain maximal EPS recovery. Higher EPS yields were obtained in nitrogen-free media amended with 20–30 mM 4-hydroxybenzoic acid. In general, modifications in inorganic salt concentration did not alter EPS production, except in the case of magnesium ions. Increased levels of this cation were correlated to greater EPS yields. Production was strongly influenced by certain environmental factors. Optimal values of 34°C, 80 rpm and neutral or slightly basic conditions were selected. Under these conditions, more than 25% of the carbon source supplied was converted to EPS and the production was improved about 42% in comparison to that observed in the initial media. Journal of Industrial Microbiology & Biotechnology (2001) 27, 5–10. Received 05 November 2000/ Accepted in revised form 30 April 2001     

Plant composition and diversity in a semi-natural Mediterranean island landscape: the importance of environmental factors

Abstract

Islands crucially contribute to the Mediterranean Basin’s high floristic diversity, which, however, is at risk facing climate and land-use changes. Besides the identification of highly diverse areas, the knowledge about factors favouring diversity is of great importance. We analysed plant species diversity and composition related to environmental factors over varied vegetation units on a former Italian prison island in the northwest of Sardinia. Due to a long history of land use with grazing and later abandonment the nowadays protected island features a semi-natural landscape and can serve as an example for strongly anthropogenic altered insular ecosystems. Floristic composition, soil properties, microclimate and ungulate abundance were assessed. Relationships of vegetation composition and diversity with abiotic variables were examined by Canonical Correspondence Analysis, which indicated the importance of air temperature, soil moisture, slope gradient and C/N ratio for floristic differentiation. Most important abiotic factors for plant species richness were relative air humidity and soil moisture, while floristic diversity was mainly determined by air temperature and pH. Furthermore, observation data pointed to an adverse influence of ungulate abundance for plant species diversity. Regarding nature conservation, grazing intensity thus must be critically taken into account, especially for sensitive vegetation units like the coastal garrigue.     

Effects of limiting nutrients and N:P ratios on the phytoplankton growth in a shallow hypertrophic reservoir

The purpose of this study was to evaluate the effects of limiting nutrients and the N:P ratios on the growth of phytoplankton (mainly cyanobacteria) in a shallow hypertrophic reservoir between November 2002 and December 2003. Nutrient enrichment bioassays (NEBs) were conducted, along with analyses of seasonal ambient nutrients and phytoplankton taxa, in the reservoir. The average DIN:TDP and TN:TP mass ratios in the ambient water were 90 (range: 17–187) and 34 (13–60), respectively, during the study period. The dissolved inorganic phosphorus showed seasonal variation, but less than that of inorganic nitrogen. The TN:TP ratios ranged from 13 to 46 (mean: 27 ± 6) during June–December when the cyanobacteria, Microcystis, dominated the phytoplankton composition. The NEBs showed that phytoplankton growth was mainly stimulated by the phosphorus (all of total 17 cases), rather than the nitrogen concentration (8 of 17 cases). The rapid growth rate of cyanobacteria was evident with TN:TP ratios less than 30. According to the results of the NEBs with different N concentrations (0.07, 0.7 and 3.5 mg l⁻¹), but the same N:P ratios and when the nitrogen concentration was highest, the cyanobacterial growth reached a maximum at N:P ratios <1. Overall, the response of cyanobacterial growth was a direct function of added phosphorus in the NEBs, and was greater with increased N concentrations. Thus, cyanobacterial blooms favored relatively low N:P ratios in this hypertrophic reservoir system. An erratum to this article is available at .     

杀灭土壤中线虫对小麦生长和吸收N、P的影响

盆栽条件下研究了施用杀线剂(克线磷,67mg·kg^-1干土)和干热(105℃,2h)两种杀线措施对小麦生长和N、P养分吸收的影响。杀线剂对土壤中线虫的平均杀灭率约为80%, 干热处理的杀灭率为100%. 在杀线剂处理中,苗期至抽穗期小麦生物量、拔节期至成熟期植株含N量、全生育期植株吸N量以及抽穗和成熟期吸P量均显着低于对照。土壤干热处理后抽穗和成熟期小麦的生物量、含N量及N、P吸收量也比对照显着降低。两种杀线处理植株地上部生物量和N、P吸收量与相应处理全株变化趋势基本一致。但杀灭线虫对植株含P量影响较小。分析杀线虫后小麦生长和养分吸收受抑主要与土壤有机氮的矿化作用减弱、微生物活动产生的植物生长促进物质降低有关     

The dynamics of the N/P ratio and stratification in a large nitrogen-limited estuary as a result of upwelling: a tendency for offshore Nodularia blooms

The effect of upwelling on the preconditions for noxious cyanobacterial blooms in a nitrogen-limited estuary is studied with Nodularia blooms in the Gulf of Finland especially in mind. An idealised physical–chemical model is derived and integrated. The physical development in the model agrees with a classical picture of upwelling for the scales of interest. The chemical component indicates a transient minimum of dissolved inorganic nitrogen to phosphorus ratio in the middle of the gulf, coinciding with the stratification maximum resulting from physical dynamics. A general tendency for offshore cyanobacterial blooms in similar physical and biological conditions is deduced from these results.     

Effect of environmental conditions on decomposition in eight woody species of a dry tropical forest

Litter decay is a significant part of carbon budget. Due to strong environmental control, the changes in the environment may drastically influence the litter decay rates. Litter decomposition of eight dry tropical woody species, viz. Shorea robusta, Buchanania lanzan, Diospyros melanoxylon, Lagerstroemia parviflora, Lannea coromandelica, Terminalia tomentosa, Holarrhena antidysenterica and Lantana camara was studied to document the effect of intra-annual changes in the environment. Litter decomposition was monitored at monthly intervals at five sites using litter bag technique over an annual cycle in a dry tropical deciduous forest of Vindhyan highland, India. Weight loss differed among species and through months, and ranged from 15.38% in L. camara at Kotwa site in January to 30.72% in T. tomentosa at Hathinala site in August. Peak weight loss occurred in August and averaged 46.2% across species and sites. Nitrogen and phosphorus mineralization rates also varied significantly from species to species. T. tomentosa having higher nitrogen content and lower C/N ratio than other species exhibited faster weight loss. Nitrogen and phosphorus contents of litter showed significant positive correlation with weight loss. C/N ratio was negatively related to decay constant, and the weight loss was positively related to the soil surface temperature as well as soil moisture content.     

The effect of temperature and salinity on growth rate and azaspiracid cell quotas in two strains of Azadinium poporum (Dinophyceae) from Puget Sound,Washington State

Azaspiracids (AZA) are novel lipophilic polyether marine biotoxins associated with azaspiracid shellfish poisoning (AZP). Azaspiracid-59 (AZA-59) is a new AZA that was recently detected in strains of Azadinium poporum from Puget Sound, Washington State. In order to understand how environmental factors affect AZA abundances in Puget Sound, a laboratory experiment was conducted with two local strains of A. poporum to estimate the growth rate and AZA-59 (both intra- and extracellular) cell quotas along temperature and salinity gradients. Both strains of A. poporum grew across a wide range of temperatures (6.7 °C to 25.0 °C), and salinities (15 to 35). Growth rates increased with increasing temperature up to 20.0 °C, with a range from 0.10 d⁻¹ to 0.42 d⁻¹. Both strains of A. poporum showed variable growth rates from 0.26 d⁻¹ to 0.38 d⁻¹ at salinities from 15 to 35. The percentage of intracellular AZA-59 in both strains was generally higher in exponential than in stationary phase along temperature and salinity gradients, indicating higher retention of toxin in actively growing cells. Cellular toxin quotas varied by strain in both the temperature and salinity treatments but were highest at the lowest growth rates, especially for the faster growing strain, NWFSC1011.Consistent with laboratory experiments, field investigations in Sequim Bay, WA, during 2016–2018 showed that A. poporum was detected when salinity and temperature became favorable to higher growth rates in June and July. Although current field data of A. poporum in Puget Sound indicate a generally low abundance, the potential of local A. poporum to adapt to and grow in a wide range of temperature and salinity may open future windows for blooms. Although increased temperatures, anticipated for the Puget Sound region over the next decades, will enhance the growth of A. poporum, these higher temperatures will not necessarily support higher toxin cell quotas. Additional sampling and assessment of the total toxicity of AZA-59 will provide the basis for a more accurate estimation of risk for azaspiracid poisoning in Puget Sound shellfish.