微囊藻毒素对水环境的影响研究进展

微囊藻毒素是富营养化淡水水体中最常见的藻类毒素,是湖泊蓝藻产生的一类肽类毒素,它的产生受到藻类的遗传和环境因素的共同影响。由于其毒性大,分布广,结构稳定,从而成为水环境中的潜在危害物质。有关微囊藻毒素性质、毒理毒性、在环境中的迁移、转化以及控制预防已成为关注热点。在总结国内外研究的基础上,综述了微囊藻毒素的性质、产生机理以及其与水环境、水生生物(水生植物、鱼类、无脊椎动物)间的相互作用,讨论了微囊藻毒素对水生生物的影响以及水生生物对微囊藻毒素的降解作用,为水体中微囊藻毒素的防治提供科学的依据。     

扇贝毒素pectenotoxins(PTXs)研究进展

扇贝毒素(pectenotoxins,PTXs)是一类聚醚大环内酯结构的脂溶性海洋生物毒素,是由海洋甲藻中的鳍藻属Dinophysisspp.的几个种产生的,1984年首次从日本的养殖扇贝Patinopecten yessoensis中发现鉴定,具有很高的小鼠腹腔注射致死毒性。近年发现的地理区域不断扩大,我国尚属空白。就这一毒素的结构、来源生物、毒性、携带生物、地理分布、降解代谢及风险评估等研究现状作一系统综述,并分析展望了今后我国藻毒素研究的重点方向。     

我国常见的几种脂溶性贝毒的研究进展

近年我国沿海有毒赤潮的发生频次和规模呈明显的上升趋势,产生的藻毒素沿食物链传递并在贝类体内累积转化生成更为复杂的贝毒素,威胁贝类水产品食用安全和消费者健康。其中大部分贝毒的极性较低,易溶于脂类化合物,被称为脂溶性贝毒(lipophilic shellfish toxins,LST)。目前我国的养殖贝类正面临多种脂溶性贝毒的污染,主要包括大田软海绵酸(okadaic acid,OA)、蛤毒素(pectenotoxin,PTX)、虾夷扇贝毒素(yessotoxin,YTX)、氮杂螺环酸(azaspiracid,AZA)及环亚胺类毒素(cyclic imine,CI)中的gymnodimine(GYM)等几大类毒素。现就这些脂溶性贝毒的化学结构、来源、毒理学特征、食用安全限量及检/监测方法等内容做了系统总结,同时,对我国脂溶性贝毒的来源与贝类染毒状况做了简要概括,并对今后我国脂溶性贝毒的研究方向提出了建议。     

基于酵母双杂交的微囊藻毒素检测系统初探

微囊藻毒素是由淡水微囊藻产生的次生代谢物,结构为环状七肽,分子量约1000Da,对生物机体具有多种毒理作用,是一类较强的肿瘤促进剂,目前没有比较简单便宜的藻毒素检测方法,本文探索建立利用酵母双杂交系统进行藻毒素检测的方法.前期实验已用噬菌体表面展示肽库技术从随机12和7肽库中筛选获得若干与藻毒素有相互作用的多肽.本文分别挑选其中亲和力较高的3个多肽,构建到酵母双杂交系统中,将不同的肽段分别融合在含有Active Domain和Binding Domain的表达载体中,共转化酵母菌株AH109,然后测定β-半乳糖苷酶活性来反映下游报告基因半乳糖苷酶的表达情况,从而通过反映培养基里面是否含有藻毒素,进而利用此方法来比较方便地检测藻毒素.     

海洋卡盾藻(香港株)溶血毒素的提取和分离

海洋卡盾藻(Chattonella marina)是我国南方沿海主要的鱼毒性赤潮生物,近年来由该藻形成的赤成已造成多起近岸养殖鱼类大量死亡的事件。为了弄清该藻毒素的基本成分特征,本文研究了室内培养条件下海洋卡盾藻(香港株)溶血毒素的提取方法,观察了溶血毒素对红细胞的溶血过程,采用薄层色谱法对溶血成分进行了初步分析。结果表明:海洋卡盾藻藻细胞的超声波破碎最适条件为功率400W,4℃下处理15min;通过显微镜观察,证实提取的毒素对血红细胞膜具破坏作用;海洋卡盾藻合成的溶血毒素至少含有4种组分,其中1种可能为为脂类,3种为糖脂类。该研究成果有助于我国今后进一步开展鱼毒性赤潮生物毒素的研究。     

蓝藻毒素去除方法研究进展

蓝藻水华中产生的次生代谢产物蓝藻毒素对人类及陆生动物具有严重毒性和致癌作用,同时会危害水域生态系统的结构和功能。有毒藻华的频繁发生及其引发的环境和健康问题已经成为全球环境科学领域的研究热点。现有的净水技术去除饮用水中藻毒素效率不高,因此寻找高效的藻毒素去除方法是目前亟待解决的问题。本文首先阐述了蓝藻毒素的产生和释放机理,将蓝藻毒素的去除分为除藻和去除水体中溶解性藻毒素两类;其次,本文在国内外大量研究成果的基础上综述了当前蓝藻毒素的降解途径,并详细介绍了活性炭吸附法、高级氧化降解法、微生物降解法等具有实际应用前景的去除方法;最后,本文总结了当前蓝藻毒素去除方法的局限性,指出了未来的研究应向高效率、低能耗、无污染、实用性强的方向发展。     

球形棕囊藻对五种水生动物的急性毒性作用

利用室内半静水法研究球形棕囊藻培养液、藻细胞悬浮液、去藻细胞滤液、藻细胞内容物、藻细胞碎片以及溶血毒素粗提物对卤虫(Artemia sinica)、褶皱臂尾轮虫(Brachionus plicatilis)、蒙古裸腹潘(Moina mongolia)、赤点石斑鱼幼鱼(Epinephelus akaara)和南美白对虾(Penaeus vannamei)5种水生动物的影响,并对其致毒途径进行分析.结果表明,与对照相比,培养液、藻细胞悬浮液处理卤虫,其运动速度下降,培养液、藻细胞悬浮液与毒素粗提物处理轮虫72 h,存活率显著降低(P＜0.05),卤虫、轮虫消化道中充满藻细胞,提示摄食球形棕囊藻可能是引起卤虫、轮虫中毒的主要途径.培养液、滤液和藻细胞内容物对赤点石斑鱼幼鱼均有显著的致死作用,而藻细胞碎片和溶血毒素粗提物对幼鱼的存活无明显影响,表明球形棕囊藻细胞表面并不存在毒性物质,球形棕囊藻可能会分泌有毒物质进入水体,进而影响鱼类的生长.培养液、藻细胞悬浮液、去藻细胞滤液、毒素粗提物处理蒙古裸腹溞72 h虽有死亡,但存活率与对照无显著差异;南美白对虾的情况与蒙古裸腹溞类似,表明球形棕囊藻及相关组分对蒙古裸腹溞和对虾的影响较小.这些结果表明,球形棕囊藻对不同生物的毒害作用不同;摄食和分泌有毒、有害物是球形棕囊藻引致生物中毒的主要途径;除了溶血毒素以外,球形棕囊藻还可能产生其他鱼毒性物质.     

环境水体藻毒素生物处理技术研究进展

近年来,随着水体富营养化程度加剧,蓝藻水华现象时有发生,蓝藻及其释放的藻毒素对生态环境和人类健康构成严重威胁.本文概述了常见藻毒素的分类及其主要理化性质,总结了生物接触氧化、生物滤池和生物-生态耦合等工艺对藻毒素的去除性能与机制,分析了生物处理工艺的反应条件(温度、pH和水力停留时间)、原水性质及营养限制等影响因素,并对藻毒素去除机理、新工艺研发等方面进行了展望.     

土壤污染的生态毒理诊断研究进展

土壤生态毒理学是污染土壤生态风险评价以及土壤污染控制的理论依据,土壤环境中有毒物质的生态毒理效应及其分子机制的研究是土壤生态毒理学的核心内容.土壤污染生态毒理诊断集合了土壤污染的全部信息,提供了土壤的整体毒性效应.因此,开展土壤污染生态毒理诊断研究有重要的理论意义和现实意义.本文在参考国内外土壤污染生态毒理诊断研究成果的基础上,介绍了常用土壤污染生态毒理效应诊断方法和指标,并探讨了不同方法的利弊.传统毒理诊断方法往往只能表征胁迫的程度,不能解释损伤和响应发生分子机理.污染土壤毒性评估的生物标记物和土壤污染分子诊断技术,可深入探讨外源胁迫下生物的解毒机制.本文对其进行了较为系统的介绍,并对未来的研究趋势进行展望.     

鲢、鲤和鲫肝细胞原代培养

微囊藻毒素(Microcystins)是一类淡水水体中危害很严重的生物毒素(Biotoxin),由微囊藻毒素所引发的环境问题及其对人类健康的危害正日益受到科学家的关注1.已知微囊藻毒素作用的靶器官为肝脏,以往的研究多集中在微囊藻毒素对动物肝脏组织的损伤,如口服或腹腔注射毒素,引起肝组织结构破坏、肝出血甚至肝坏死,但用整体实验动物或器官研究微囊藻毒素毒理学较难深入,因此建立毒理学实验模型十分重要.肝脏作为动物体内最重要的解毒器官,是研究微囊藻毒素毒理学的主要对象.一般毒理学实验都采用肝脏原代培养细胞,因为原代培养细胞生理生化及遗传特性稳定,适于研究外界毒物的毒性、毒理及肝细胞对毒物的应答和解毒机理.本实验通过对鲢(Hypophthalmichthy molitrix Carier et Valencienines)、鲤(Cyprinus carpio Linnaeus)和鲫(Carassius auratus Linnaeus)肝脏原代细胞培养,以建立稳定的毒理学实验模型,为微囊藻毒素毒理学研究奠定基础.       

Positive feedbacks between bottom-up and top-down controls promote the formation and toxicity of ecosystem disruptive algal blooms: A modeling study

Harmful algal blooms that disrupt and degrade ecosystems (ecosystem disruptive algal blooms, EDABs) are occurring with greater frequency and severity with eutrophication and other adverse anthropogenic alterations of coastal systems. EDAB events have been hypothesized to be caused by positive feedback interactions involving differential growth of competing algal species, low grazing mortality rates on EDAB species, and resulting decreases in nutrient inputs from grazer-mediated nutrient cycling as the EDAB event progresses. Here we develop a stoichiometric nutrient–phytoplankton–zooplankton (NPZ) model to test a conceptual positive feedback mechanism linked to increased cell toxicity and resultant decreases in grazing mortality rates in EDAB species under nutrient limitation of growth rate. As our model EDAB alga, we chose the slow-growing, toxic dinoflagellate Karenia brevis, whose toxin levels have been shown to increase with nutrient (nitrogen) limitation of specific growth rate. This species was competed with two high-nutrient adapted, faster-growing diatoms (Thalassiosira pseudonana and Thalassiosira weissflogii) using recently published data for relationships among nutrient (ammonium) concentration, carbon normalized ammonium uptake rates, cellular nitrogen:carbon (N:C) ratios, and specific growth rate. The model results support the proposed positive feedback mechanism for EDAB formation and toxicity. In all cases the toxic bloom was preceded by one or more pre-blooms of fast-growing diatoms, which drew dissolved nutrients to low growth rate-limiting levels, and stimulated the population growth of zooplankton grazers. Low specific grazing rates on the toxic, nutrient-limited EDAB species then promoted the population growth of this species, which further decreased grazing rates, grazing-linked nutrient recycling, nutrient concentrations, and algal specific growth rates. The nutrient limitation of growth rate further increased toxin concentrations in the EDAB algae, which further decreased grazing-linked nutrient recycling rates and nutrient concentrations, and caused an even greater nutrient limitation of growth rate and even higher toxin levels in the EDAB algae. This chain of interactions represented a positive feedback that resulted in the formation of a high-biomass toxic bloom, with low, nutrient-limited specific growth rates and associated high cellular C:N and toxin:C ratios. Together the elevated C:N and toxin:C ratios in the EDAB algae resulted in very high bloom toxicity. The positive feedbacks and resulting bloom formation and toxicity were increased by long water residence times, which increased the relative importance of grazing-linked nutrient recycling to the overall supply of limiting nutrient (N).     

Antibiotic treatment enhances C2 toxin production by Alexandrium tamarense in batch cultures

The effects of a mixture of penicillin G and streptomycin on the growth and C2 toxin production of a marine dinoflagellate, Alexandrium tamarense CI01, were investigated to determine if antibiotic treatment would increase the toxin yield of the cultured algae in batch cultures. Algal growth and toxin production were both enhanced markedly when the culture was supplemented with the antibiotics, each at an initial concentration of 100 unit ml⁻¹ in medium,² but were severely inhibited when the concentration was 500 unit ml⁻¹ or higher. Short-term pretreatment of algal inocula with the antibiotics at 100, 500, and 1000 unit ml⁻¹ all produced the enhancing effects on the algal cultures in an autoclaved medium. A prolonged antibiotic pretreatment of the algal culture followed by repeated sterile cultivation resulted in an algal culture free of cultivable bacteria. This “drug-treated” culture became more resistant to the toxicity and more responsive to the enhancing effects of the antibiotics. Our results indicated that the antibiotics can enhance growth and C2 toxin productivity not only through their inhibition of the growth of bacteria that compete for nutrients with the coexisting algae, but also through their direct effects on the physiology of the algae. Supplementation of the two antibiotics therefore is an efficient way to increase the yield of C2 toxin in the production cultures of A. tamarense CI01.     

Isolation, characterization and pathology of the toxin from a Microcystis aeruginosa (= Anacystis cyanea) bloom.

The nature of the toxicity of a bloom of blue-green alga, M. aeruginosa (= Anacystis cyanea), that occurred in a man-made lake was investigated. Crude algal bloom extracts were toxic to laboratory mice when injected intraperitoneally. The lethal dose (LD100) of these extracts was 15-30 mg of lyophilized algal bloom per kilogram body weight. The toxin was purified by a procedure that included ammonium sulphate fractionation, solvent extraction, acid precipitation, Sephadex G25 and DEAE-Sephadex chromatography, and high-voltage electrophoresis at pH 6.5. The preparation gave a single spot on high-voltage electrophoresis at pH 9.0, had no free amino group, and was characterized by a simple amino acid composition of equimolar quantities of L-methionine, L-tyrosine, D-alanine, D-glutamic acid, erythro beta-methyl aspartic acid and methylamine. The LD50 for the purified toxin was estimated to be 0.056 mg/kg of mice, and the approximate LD100 is 0.070 mg/kg, based on the total material found from amino acid analysis. Parenteral administration of the purified toxin to mice produced extensive liver lobular haemorrhage and death within 1-3 h. Repeated inoculation of sublethal doses daily over some weeks produced progressive hepatocyte degeneration and necrosis and the development of fine hepatic fibrosis.     

Toxic substance from a natural bloom of Microcystis aeruginosa.

A toxic substance contained in the blue-green alga Microcystis aeruginosa was purified and partially characterized. Toxic algal cells were collected from a highly eutrophic lake in Japan, and the toxin was purified by homogenization, ultrafiltration, gel filtration, and ion-exchange chromatography. The final preparation gave a single peak on high-performance liquid chromatography. The toxicity was somewhat less than that reported for other toxins from this alga. The water extract of 6.7 mg (dry weight) of cells and 72 microgram of the purified protein was required to kill a mouse (1 mouse unit). The main amino acids of the toxin were glutamic acid, asparatic acid, alanine, glycine, arginine, and leucine. The molecular weight of the toxin was 2,950 as determined by high-performance liquid chromatography.     

Modelling the Stoichiometric Regulation of C-Rich Toxins in Marine Dinoflagellates

Toxin production in marine microalgae was previously shown to be tightly coupled with cellular stoichiometry. The highest values of cellular toxin are in fact mainly associated with a high carbon to nutrient cellular ratio. In particular, the cellular accumulation of C-rich toxins (i.e., with C:N > 6.6) can be stimulated by both N and P deficiency. Dinoflagellates are the main producers of C-rich toxins and may represent a serious threat for human health and the marine ecosystem. As such, the development of a numerical model able to predict how toxin production is stimulated by nutrient supply/deficiency is of primary utility for both scientific and management purposes. In this work we have developed a mechanistic model describing the stoichiometric regulation of C-rich toxins in marine dinoflagellates. To this purpose, a new formulation describing toxin production and fate was embedded in the European Regional Seas Ecosystem Model (ERSEM), here simplified to describe a monospecific batch culture. Toxin production was assumed to be composed by two distinct additive terms; the first is a constant fraction of algal production and is assumed to take place at any physiological conditions. The second term is assumed to be dependent on algal biomass and to be stimulated by internal nutrient deficiency. By using these assumptions, the model reproduced the concentrations and temporal evolution of toxins observed in cultures of Ostreopsis cf. ovata, a benthic/epiphytic dinoflagellate producing C-rich toxins named ovatoxins. The analysis of simulations and their comparison with experimental data provided a conceptual model linking toxin production and nutritional status in this species. The model was also qualitatively validated by using independent literature data, and the results indicate that our formulation can be also used to simulate toxin dynamics in other dinoflagellates. Our model represents an important step towards the simulation and prediction of marine algal toxicity.     

SYMBIODINIUM SPP. ISOLATES FROM STONY CORAL: ISOLATION,GROWTH CHARACTERISTICS AND EFFECTS OF UV IRRADIATION

Rengefors, K.¹ & Legrand, C.^{2 1}Limnology, Department of Ecology, Ecology Building, Lund University, SE-223 62 Lund, Sweden; ²Department of Biology and Environmental Science, University of Kalmar, Barlastgatan 1, SE-391 82 Kalmar, Sweden Freshwater dinoflagellates may form dense blooms during winter in ice-covered lakes. Unlike their marine counterparts freshwater dinoflagellates are rarely considered as potential toxin producers. Here we tested if the winter species Peridinium aciculiferum produces a toxin(s)and investigated the potential adaptive function of that toxin - i.e. predator defense or inhibition of competitors (allelopathy). We found that P. aciculiferum produces a toxin(s). Using traditional toxicity bioassays (Artemia toxicity test and haemolytic activity assay), we detected the production of a toxic substance by P. aciculiferum cells from both the field and from laboratory cultures. Cultures deprived of phosphorus showed most toxicity. Potential predators, such as Daphnia galeata (Cladocera) and Eudiaptomus graciloides (Copepoda), were apparently not harmed by P. aciculiferum toxicity. However, a competitor, the flagellate Rhodomonas lacustris (Cryptophyceae), which co-occurs with P. aciculiferum was killed (allelopathy) by the P. aciculiferum toxin. The toxin(s) caused the cells of R. lacustris to swell and burst. We concluded that allelopathy may be an adaptive strategy of winter dinoflagellates, allowing them to outcompete other phytoplankton species, and thereby dominate the algal biomass.     

霍乱毒素佐剂的研究进展

霍乱毒素（CT）是霍乱弧菌分泌的一种不耐热肠毒素,具有很强的免疫原性和佐剂活性,是当今研究得最多且最深入的粘膜免疫佐剂之一,CT本身有很强的毒副作用,而一定的毒性又似乎是发挥佐剂作用所必须的。通过各种改造,使之具备优良的佐剂活性而没有显的毒副作用是当前研究的主要目标。本从CT作为佐剂的作用机理出发,概述当前对其进行改造的几种研究方向。     

赤潮藻毒素生物合成研究进展

合成毒素是赤潮藻类的一个常见特征,已知能够产生毒素的微藻有70多种。作为次级代谢产物,藻毒素的产生可能是一种压制或清除其它藻类竞争者的一种反应,在群落演替、种间竞争中发挥重要作用。目前,人们对藻毒素生物合成机理依然知之甚少,相关基因的研究仍无明显突破。利用环境因子诱导毒素生成变化进而分离差异表达基因或者比较不同产毒藻株间基因表达的差异,从中克隆藻毒素生物合成基因似乎是一种极具潜力的研究方向。     

Microelectrode array (MEA) platform as a sensitive tool to detect and evaluate Ostreopsis cf. ovata toxicity

In the last decade, the occurrence of harmful dinoflagellate blooms of the genus Ostreopsis has increased both in frequency and in geographic distribution with adverse impacts on public health and the economy. Ostreopsis species are producers of palytoxin-like toxins (putative palytoxin and ovatoxins) which are among the most potent natural non-protein compounds known to date, exhibiting extreme toxicity in mammals, including humans. Most existing toxicological data are derived from in vivo mouse assay and are related to acute effects of pure palytoxin, without considering that the toxicity mechanism of dinoflagellates can be dependent on the varying composition of complex biotoxins mixture and on the presence of cellular components.In this study, in vitro neuronal networks coupled to microelectrode array (MEA)-based system are proposed, for the first time, as sensitive biosensors for the evaluation of marine alga toxicity on mammalian cells. Toxic effect was investigated by testing three different treatments of laboratory cultured Ostreopsis cf. ovata cells: filtered and re-suspended algal cells; filtered, re-suspended and sonicated algal cells; conditioned growth medium devoid of algal cells. The great sensitivity of this system revealed the mixture of PTLX-complex analogues naturally released in the growth medium and the different potency of the three treatments to inhibit the neuronal network spontaneous electrical activity. Moreover, by means of the multiparametric analysis of neuronal network activity, the approach revealed a different toxicity mechanism of the cellular component compared to the algal conditioned growth medium, highlighting the potential active role of the first treatment.     

环境汞污染对藻类的毒性效应及其影响因素

综述了汞污染对藻类的毒性效应及影响因素。水环境中汞主要以元素汞、无机汞和有机汞3种形式存在。藻类吸附汞主要分为胞外的快速吸附和胞内的缓慢富集,在安全浓度内,金属汞对藻生长有一定的促进作用,随着浓度增大,抑制藻生长或致死。汞进入藻体细胞后,藻类为了存活会产生一系列保护机制。藻类对汞的排斥和排出作用可能就是对汞耐性的一种重要机制。藻类也可以通过多种方式减少汞进入藻类细胞,以及通过与其他物质结合汞使其排出胞外。温度、pH、生物学因素等影响重金属对藻类的毒性作用。并就藻类对汞耐性和适应机理、利用藻类修复和监测重金属污染、藻类响应汞胁迫的信号转导途径及其保护机制等未来研究领域进行了展望。