Generation of reactive oxygen species in different fractions of the coelomocytes of holothurian Eupentacta fraudatrix in response to the thermostable toxin of Yersinia pseudotuberculosis in vitro

Pure fraction (92%–95%) of phagocytes (FP) and a mixture of amoebocytes (62%) and morula cells (38%)-FPMC- of the holothurianEupentacta fraudatrix' (Holothuroidea, Dendrochirota) were obtained by using ficoll-verographine step gradient. Basal production of reactive oxygen species (ROS) in FP quantified by using reduction of nitroblue tetrazolium (NBT) was more than twice that in FPMC. Thermostable toxin ofYersinia pseudotuberculosis (TST) at different concentrations (0.2; 0.5; 2.5 μg/ml, but not 0.1 μg/ml) stimulated NBT reduction in FPMC after 24 h incubation. In FP, TST at concentrations of 0.1 and 0.2 μg/ml inhibited and at concentrations of 0.5 and 2.5 μg/ml stimulated NBT reduction after 24 h incubation. Maximal effect was observed in FP and FPMC at TST concentrations of 0.5 and 0.2 μg/ml, respectively. Addition of catalase (0.7 μg/ml) to the cells treated with TST (2.5 μg/ml) was followed by a decrease in NBT reduction compared to that under toxin treatment alone. TST stimulated superoxide dismutase activity in concentration-dependent manner (maximum at 0.5 μg/ml concentration in FP) after 24 h treatment, and this stimulation was prevented by a commercial catalase. Plant lectin concanavalin A stimulated NBT reduction more than 5-fold in FPMC compared to the control. With addition of TST, lectin stimulated ROS to lesser extent than that with lectin alone. When catalase, TST, and lectin were added into the FPMC simultaneously, ROS increase was similar to that under lectin treatment alone. On the whole, data obtained indicated that ROS generation in holothurian coelomocytes especially occurs in both stimulated and not stimulated phagocytes, and that changes in ROS production by these cells may be one of the mechanisms of antibacterial protection of holothurians. This work was financially supported by the Russian Foundation for Fundamental Research Grant (No. 00-04-48949).     

Satellite detection of red tide in Ariake Sound, 1998–2001

High resolution SeaWiFS data was used to detect red tide events that occurred in the Ariake Sound, Japan, a small embayment known as one of the most productive areas in Japan. SeaWiFS chlorophyll data clearly showed that a large red tide event, which damaged seaweed (Nori) cultures, started early in December 2000 in Isahaya Bay, expanded to the whole sound and persisted to the end of February 2001. The monthly average of SeaWiFS data from May 1998 to December 2001 indicated that the chlorophyll peaks appeared twice a year, in early summer and in fall, after the peaks of rain and river discharge. The SeaWiFS data showed that the red tide event during 2000–2001 winter was part of the fall bloom; however, it started later and continued significantly longer than other years. Satellite ocean color data is useful to detect the red tide; however the algorithms require improvement to accurately estimate chlorophyll in highly turbid water and in red tide areas.     

利玛原甲藻腹泻性毒素的生物学测定

１９９４年和１９９５年分季度从海南省三亚海区采集附生于褐藻上的利玛原甲藻Ｐｒｏｒｏｃｅｎｔｒｕｍｌｉｍａ（Ｅｈｒｅｎｂｅｒｇ）Ｄｏｄｇｅ，经丙酮提取。提取液用ｐＨ＝３的水除去麻痹性贝毒（ＰＳＰ），取残留液注入小白鼠（ＢａＢ／Ｃ纯系），有腹泻性贝毒（ＤｉａｒｒｈｅｔｉｃＳｈｅｌｌｆｉｓｈＰｏｉｓｏｎｉｎｇ即ＤＳＰ）中毒的症状出现。初步确定此种藻中提取的毒素为ＤＳＰ。     

固相吸附毒素跟踪技术(SPATT)在浙江南麂海域的应用

浙江南麂岛海域是我国近岸贝类的重点养殖区,近年来一直遭受脂溶性贝类毒素(lipophilic shellfish toxins,LSTs)的污染,威胁着人类健康。为了建立有效的食品安全预警方法,本研究利用固相吸附毒素跟踪技术(solid phase adsorption toxin tracking,SPATT)在该海域进行了为期一年的野外监测和LSTs毒素分析。结果表明:7种LSTs毒素在SPATT(DIAION?HP20)中被检出,分别是大田软海绵酸(okadaic acid,OA)、鳍藻毒素-1(dinophysistoxin-1,DTX-1)、虾夷扇贝毒素(yessotoxin,YTX)及其衍生物homo YTX、扇贝毒素-2(pectenotoxin-2,PTX-2)及其衍生物7-epi-PTX2sa和环亚胺毒素(gymnodimine,GYM);有8种毒素在厚壳贻贝(Mytilus galloprovincialis)中被检出,分别是OA、DTX1、homo YTX、PTX2、7-epi-PTX2sa、GYM、原多甲藻酸贝类毒素-3(azaspiracid-3,AZA-3)和虾夷扇贝毒素衍生物45-OH-homo YTX。整体而言,SPATT(HP20)中吸附的毒素种类与贝肉中监测出的毒素种类大体上一致,且两者间OA、DTX1和PTX2的浓度在时间上具有较好的相关性,可见SPATT(HP20)对这些LSTs毒素具有较高的灵敏度,有望作为水体中LSTs毒素的预警监测材料。南麂岛海域LSTs分布特征明显,夏季最高,其次是春秋季,冬季最低,其中2014年夏季贝肉中OA毒素含量高达77.19ng/g贝肉,超出国家限量标准(45ng/g贝肉)比例达11.76%,为浙江南麂海域贻贝的食用安全带来隐患。     

2004年春季东海赤潮高发区COD分布及其与赤潮关系的初步研究

2004年4～5月初在东海赤潮高发区暴发的特大规模原甲藻赤潮前期和暴发初期对该海域进行的现场调查,并对该海域COD的分布特征进行了探讨。结果表明,赤潮暴发前COD为0．295-1.836mg／L,主要受陆源输入影响。根据其在局部海区底层出现的异常升高结合其他参数分析可对特定海区潜在赤潮暴发的可能性进行评估。赤潮暴发时COD为0．36～3．14mg／L,表层和中层与叶绿素存在显著正相关关系,表明其主要受生物影响。富营养化指数表明赤潮暴发前近一半海域已经处于富营养化状态,但COD对富营养化的贡献不如营养盐重要。     

海水养殖真鲷弧菌病病原菌外毒素的分离、纯化及生物学活性

1996年6月初，对福建省平潭县真鲷网箱养殖场发生的弧菌病进行研究，并确定该病病原菌为最小弧菌（Vibrio mimicus）。采用硫酸铵沉淀、DEAE－纤维素层析及Sephadex-G100层析等方法对最小弧菌产生的外毒素进行研究。结果表明，该外毒素为单一的多肽，分子量为30．456kD，对真鲷的LD50为4．48μg，对真鲷等养殖动物的溶血性最强，对Vero细胞的CD50值为0．48μg，肠毒性弱阳性。该外毒素与最小弧菌产生的其他毒力因子相比，性质独特，是一类新的毒力因子，建议命名为Vm-Pm毒素。     

南麂列岛自然保护区海域红色裸甲藻赤潮及其成因分析

2002年9月16～19日南麂列岛海域发生3km2 红褐色赤潮 ,其原因种为红色裸甲藻(GymnodiniumsanguineumHirasaka) ,密度高达4.6×105 个/L。赤潮发生时伴随细菌的大量繁殖 ,江浙沿岸流所携带的营养物质是本次赤潮发生的基础 ,森拉克台风是本次赤潮的重要诱因。南麂海域的赤潮多发季节为4～5月份 ,9～11月份发生赤潮的可能性也较大 ,大沙岙、马祖岙和火岙等水体交换相对较弱的岙口为赤潮多发区。为保护南麂列岛国家级海洋自然保护区珍稀海洋贝藻资源 ,需协调好保护和开发之间日益尖锐的矛盾 ,控制养殖业的发展规模和游客的数量。     

基于SPE与SPATT的水体中麻痹性贝类毒素检测方法构建与应用

为实现麻痹性贝类毒素(paralytic shellfish poisoning,PSP)的实时监控与提前预警,本研究构建了基于固相萃取技术(solid phase extraction,SPE)与固相吸附毒素跟踪技术(solid phase adsorption toxin tracking,SPATT)的水体中PSP检测方法,重点优化了吸附材料及前处理方法,评价了回收率、检出限等指标,并将方法应用于2019年春季秦皇岛山海关海域PSP消长过程的监测中,比较评估了两种方法的监控预警效果。结果表明:SPE方法选用ENVI-Carb 500mg/6mL固相萃取柱,过样体积为50mL,13种PSP组分的平均回收率为82.2%±10.0%、检出限为4.0-20.0ng/L;SPATT方法选用SP207大孔吸附树脂,洗脱时间为静置Id最佳,整体回收率约为9.2%;在实际应用中,结合产毒藻密度及贻贝富集毒素含量的变化,发现SPE方法的检测结果可实时表征海域PSP风险状况,对于贻贝中PSP的预警效果也显著优于SAPTT方法,后者不仅因监控方式相对滞后一个监测周期,且灵敏度及准确性均较差。对于秦皇岛海域,当SPE方法检测结果达到100ng STX eq/L时,该海域贻贝中PSP残留将具有潜在的食用安全风险,跟踪过程表明这一阈值可提前两周预警贻贝富集毒素含量超出我国限量标准(800μg STX eq/kg),这对于强化风险警示并制定防范措施具有积极作用。     

Growth of Daphnia magna males and females fed with the cyanobacterium Microcystis aeruginosa and the green alga Scenedesmus obliquus in different proportions

Laboratory experiments were used to study the sensitivity of both male and female Daphnia magna to a toxic cyanobacterium, Microcystis aeruginosa. Male and female D. magna were fed with M. aeruginosa and a green alga (Scenedesmus obliquus) in different mixtures that included 0%, 25%, 50%, 75% and 100% Microcystis. Growth of both males and females declined with increasing proportion of Microcystis in the diet. Males were slightly less sensitive to Microcystis than females with EC₅₀ for growth inhibition at 72% and 63% Microcystis in the diet, respectively. On monospecific Microcystis diet, mortality rates for males (0.16 d^–1) and females (0.17 d^–1) were similar. The reproductive rate of females was the most sensitive endpoint with an EC₅₀ of 42% Microcystis in the diet, whereas population growth rates were the least sensitive (EC₅₀ of 74% Microcystis). Microcystis in the diet severely depressed growth and reproduction in Daphnia and poses a severe threat to its survival in which only a switch in reproductive strategy might provide Daphnia a refuge to a Microcystis environment that gradually becomes uninhabitable.     

盐度胁迫下中肋骨条藻和东海原甲藻的生长及内源多胺含量的变化

在实验室内不同盐度梯度下培养常见的两种赤潮藻——中肋骨条藻(Skeletonema costatum s.l.)和东海原甲藻(Prorocentrum donghaiense),根据logistic生长模型获得了终止生物量Bf和最大生长速率μmax,并测定了藻体中过氧化物酶(POD)、二胺氧化酶(DAO)、多胺氧化酶(PAO)的活性和丙二醛(MDA)、腐胺(Put)、亚精胺(Spd)、精胺(Spm)的含量。结果表明,高、低盐度胁迫下,两种藻均会积累MDA,活性氧自由基的伤害增强,藻的生长受到了抑制。同时,两种藻都会提升多胺含量,通过多胺的调节作用来缓解胁迫的伤害,促进生长,但是在提升的多胺种类和形态上,两种藻存在着差异。高盐胁迫下,中肋骨条藻会提升多胺尤其是游离态多胺的含量来缓解伤害,东海原甲藻则依靠结合态的亚精胺和游离态的精胺的调节作用。低盐胁迫下,中肋骨条藻会提升游离态腐胺的含量,而东海原甲藻体内各形态的多胺都会上升。