Uptake and depuration of paralytic shellfish toxins in the green-lipped mussel, Perna viridis: a dynamic model

Uptake and depuration of paralytic shellfish toxins in the green-lipped mussel, Perna viridis, were investigated by exposing the mussels to dinoflagellates (Alexandrium tamarense, ACTI01) under laboratory conditions for 8 d, then depurating them in clean seawater for 14 d. First-order linear differential equations were set up for five tissue compartments: Viscera, gill, hepatopancreas, adductor muscle, and foot. The solutions to these equations were used to fit the experimental data. We then estimated the parameters governing the model, which depend on the elimination rate from each compartment and the transfer coefficient between compartments. An assumption of the model is that the gills transport the dinoflagellates directly to the mouth and then to the viscera, where the ingested cells are broken down, releasing the toxins. The toxins absorbed are transferred to other tissues. During the uptake phase, the transfer coefficients from viscera to gill, hepatopancreas, adductor muscle, and foot were 0.03, 0.24, 0.01, and 0.004 per day, respectively. During the depuration phase, the transfer coefficients were 0.01, 0, 0.01, and 0.003 per day, respectively. In terms of the anatomical distribution of N-sulfocarbamoyl-11-hydroxysulfate (C2) toxins in various tissues, viscera and hepatopancreas contained the highest percentages (47-74% and 8-41%, respectively). Together, these two tissue compartments accounted for 71 to 96% of all C2 toxins present. The biokinetic model allows a quantitative prediction of C2 toxins in whole ussel as well as individual tissue compartments based on the density estimates and toxin load of dinoflagellate cells in the surrounding waters over time.     

广州市售贝类麻痹性贝毒和腹泻性贝毒污染状况分析

目的对广州市售双壳经济贝类麻痹性贝毒(PSP)和腹泻性贝毒(DSP)污染状况进行为期一年的抽样调查,了解其食用安全性。方法采用AOAC推荐的小鼠生物检测法进行PSP和DSP的毒力测定,采用HPLC进行PSP成分分析,根据FAO、日本和欧盟水产食品卫生要求及我国渔政渔港监督管理局制订的贝类安全食用标准对贝类水产品的食用安全性进行评价。结果在所调查的7种贝类中,有2种染有PSP,毒素含量在安全食用范围内,毒力大小随季节而变化,春冬两季含量相对较高;7种贝类中有6种共计36个样品染有DSP,有10个样品毒素含量超出安全食用标准,春冬两季染毒率较高。结论广州市售贝类PSP含量和检出率整体水平较低;DSP检出率稍高,毒素含量也较高,应引起有关部门的关注,加强DSP监测工作。     

麻痹性贝毒分析技术的研究进展

至今,麻痹性贝毒已对海洋经济造成了难以估计的损失,对人类健康也构成了潜在的威胁。鉴于传统的小白鼠生物分析法存在许多难以克服的局限性,近１０年来发展了许多新的分析技术,使麻性贝毒的检测能达到灵敏,高效,快速的要求,本文系统介绍了麻痹性贝毒的检测技术及进展。     

麻痹性贝类毒素细胞检测法的建立

目的:麻痹性贝类毒素的N-2a细胞检测方法的建立并确定可能的检测限值。方法:使用不同浓度毒素标准品结合藜芦定和乌本苷共同作用对数生长期的N-2a细胞,通过cck-8试剂盒检测细胞毒性,确立细胞检测方法的检出限及比较各毒素作用大小,并同步利用小鼠生物法进行验证,通过精确记录小鼠死亡时间,比较各毒素的毒作用大小。结果:麻痹性贝类毒素能明显降低藜芦定和乌本苷的细胞毒性,且具有剂量-反应关系,通过比较得出毒素各成分毒性大小为:neoSTX>STX>dcSTX>GTX1,4>GTX2,3>dcGTX2,3;小鼠生物法的毒作用平均死亡时间为:neoSTX组(6.5 min),GTX1,4组(8.0 min),STX组(9.0 min),dcSTX组(15 min);GTX2,3组和dcGTX2,3组未见动物死亡。结论:细胞检测与小鼠生物法具有较好的一致性,表明所建立的细胞检测法可行,且细胞毒性试验检测方法具有较高的灵敏度,最低检出限值可达到10-9mol/L剂量水平,其中浓度在10-6~10-8mol/L间具有较好的线性。     

双壳贝类麻痹性贝毒抗性机制的研究

麻痹性贝毒(PSP)是一类神经肌肉麻痹剂,可以阻断电压门控钠离子通道,造成神经系统传输障碍而产生麻痹作用。某些双贝壳类对PSP毒素具有耐受性和抗性,其抗性机制可能与代谢速率、生物转化和钠通道突变等有关,其中钠通道突变是双壳贝类对PSP毒素产生抗性的最重要的分子机制。     

一起贝类毒素引发的食物中毒

目的 全面了解深圳市一起因食用海产品角螺所致食物中毒的发生原因和致病因素,以采取有效的预防控制措施.方法 采用现场流行病学调查采集可疑食品进行检测.结果 18例病例主要症状为恶心、呕吐、腹痛;实验室检测结果:生角螺的螺肉中腹泻性贝类毒素(DSP)含量为147.04 μg/100 g,肠腺中为88.51 μg/100 g;熟角螺螺肉中为86.85 μg/100 g,肠腺中为62.78 μg/100 g均超标.结论 该起事件是由于食用被DSP污染的角螺引起的食物中毒事件.市场监督管理部门应对该批角螺进行溯源调查.推广预防DSP污染的养殖技术,建立贝毒污染预警,加强贝毒研究、监测、管理、控制,防止类似事件发生.     

麻痹性贝毒在广州市售经济贝类中污染状况分析

目的通过对广州市黄沙海产品批发市场7种经济贝类为期一年的麻痹性贝毒(PSP)污染状况的调查分析,了解海产品食用的安全性。方法毒性测定按照AOAC小白鼠法进行,成份分析利用高效液相色谱(HPLC),安全性评价采用联合国粮农组织(FAO)及我国渔政渔港监督管理局制定的贝类安全食用标准(4MUg肉)。结果在调查的84份贝样中,染毒贝整体毒力(消化腺与肉的加权平均毒力)低于4MUg肉,毒素最高含量仅为184MUg肉,所有贝类均在安全食用范围之内。染毒贝类主要为栉孔扇贝(ChlamysMimachlamysnobilis)和嵌条扇贝(Pectenalbicans)。毒素在2个种9份样品的消化腺中检出,某些样品肌肉组织出现小白鼠毒性反应。一份样品消化腺中毒素含量高达1452MUg,高效液相色谱(HPLC)分析表明,该样品毒素成份主要为B1、GTX23、GTX14及C类。结论广州市市售鲜贝PSP含量和检出率整体水平均较低;贝中消化腺毒素含量及检出率明显高于肌肉,个别腺体毒素含量超出标准。毒素分布存在季节性差异,春季毒素比较强,夏秋季检出率高。因此有针对性的加强贝毒监测非常必要。     

应用细菌回复突变试验研究贻贝中麻痹性贝毒的致突变性

目的 了解贻贝中麻痹性贝类毒素的致突变性。方法 采用TA97、TA98、TAl00、TAl02测试菌株,在-S9和+S9条件下分别对0.0029～1.83 μg STX/皿剂量范围内的贻贝中麻痹性贝类毒素进行检测。结果 在2种不同的条件下,各剂量水平的4种测试菌株都能正常生长;所有剂量的回变菌落数与对照组相比均无1倍及以上的显著增加,且无剂量-反应关系。结论 在该试验条件下,贻贝中麻痹性贝毒未见致突变性作用。     

固相萃取-高效液相色谱-串联质谱法检测贝类产品7种脂溶性贝类毒素

目的:建立固相萃取、高效液相色谱串联质谱法测定贝类产品中AZA1、YTX、OA、PTX2、GYM、SPX1、DTX-1 7种脂溶性贝类毒素残留的方法。方法:贝类样品用甲醇提取后,Strata-X固相萃取柱净化,甲醇洗脱。采用Waters Sun-FireTMC18(100 mm×2.1 mm,3.5μm)色谱柱,以乙腈-2 mmol/L乙酸铵水(80:20,v/v)作为流动相,流速0.2 ml/min,采用电喷雾质谱电离,多反应监测模式(MRM)对目标化合物定性及定量分析,外标法定量。结果:7种贝类毒素线性相关系数均大于0.998;方法定量下限(LOQ)为0.1μg/kg~2.7μg/kg;高、中、低3个添加水平的平均加标回收率在72.2%~101.9%,相对标准偏差为2.0%~13.6%。结论:该方法灵敏度高、操作简单高效,适用于贝类样品中脂溶性贝类毒素的定量及确证分析。     

Paralytic shellfish poisoning toxin profiles in green mussels from Trinidad and Venezuela

Paralytic shellfish poisoning (PSP) toxins were determined in green mussels (Perna viridis) from the Gulf of Paria between Trinidad and Venezuela, as well as from the northern coast of Venezuela. Aqueous extracts of samples were purified by passage through SPE C-18 and carboxylic columns, and oxidized by periodate and peroxide reagents, before analysis by LC with fluorescence detection. Spike and recovery of mixed toxins from uncontaminated mussel tissues, with % recoveries from 83 to 103, validated the method of analysis. Saxitoxin (STX), gonyautoxins (GTX), their analogs, and C toxins were identified in mussels from both countries. In the Gulf of Paria, PSP toxin profiles in mussels from the north were similar to each other, as were those from the south. However, the toxin profiles and concentrations varied widely among geographic areas, with GTX analogs and C1,2 toxins predominating. The highest total PSP concentrations (468 μg/100 g), as well as the largest number of individual toxins identified (7 of 8), were found at Playa Patilla on the northern coast of Venezuela, followed by Chaguaramas (244 μg/100 g; 6 of 8) and Güiria (154 μg/100 g; 5 of 8) in the northern Gulf of Paria in Trinidad. While the calculated STX equivalents in each sample were below the maximum permissible level (80 μg/100 g) permitted by Venezuela and other countries, its very detection prohibits their consumption in Trinidad under existing legislation.     

Microbiological monitoring of mussels and clams collected from the shellfish-growing marine areas in Rimini Province

A 5 years survey (1996-2000) was performed on the microbiological quality of shellfish (mussels and clams) collected from authorized shellfish-growing area of the North Adriatic Sea (province of Rimini). 7.0% of mussel samples (33/474) and 21.9% of clam samples (218/996) exceeded the legal limits for faecal indicators (Dlgs 530/92). The faecal contamination of shellfish was related to the organic pollution arriving from inland surface waters. In fact shellfish harvested near the coast were more contaminated, as well as shellfish grown in the surface layers of the open sea, where the fresh waters of the streams tend to stratify. Faecal contamination was also directly correlated with rainfall, probably because the atmospheric precipitation, increasing the flow of the streams, favoured the transport of organic substances and micro-organisms deriving from the soil washing and the municipal sewage effluents. Furthermore, when rainfall was particularly plentiful, sewage inflow could also exceed the hydraulic capacity of the treatment plants; thus making necessary to discharge untreated waste directly into surface waters. Salmonellae were found in 0.7% of clam samples. Although this percentage is very low, it shows that pathogenic micro-organisms are present in this area of sea. This finding, together with the high variability of shellfish pollution due to occasional factors as rainfall, emphasise the importance of the systematic monitoring of the microbiological quality of shellfish.     

ELISA快速测定带子中麻痹性贝类毒素含量

目的 分析测定导致中毒的带子及珠海市售的带子中麻痹性贝类毒素(PSP)含量.方法 依据SN/T1773-2006,采用美国Abraxiskits公司生产的SAXITOXIN (PSP) ELISA KIT (96T)试剂盒,测定食物中毒客户留样及市售的带子样品中麻痹性贝类毒素含量,并进行方法检出限、相对标准偏差(RSD)测定,以确保检测结果的有效性.结果 6份带子样品(1份食物中毒客户留样,5份市场抽检),其中5份检出麻痹性贝类毒素,含量为9.1～59.9 μg/kg.方法检出限为5.0 μg/kg、RSD为12.9％.结论 所检6份带子样品中有5份检出含有麻痹性贝类毒素(PSP).所检样品PSP的含量与流行病学调查较吻合.     

两种方法检测织纹螺中麻痹性毒素的分析比较

目的：了解织纹螺带毒情况，探讨织纹螺毒素特征。方法：用小鼠生物法测定织纹螺麻痹性毒素；用ELISA法测定STX，按织纹螺肉中毒素含量≥400MU／100g或STX≥80μg／100g判为对人有毒害的毒螺。结果：小鼠生物法测定检出有毒织纹螺51份，占总数的32．90％。以中毒样品检出毒螺的数量最多，占总数的23．23％，检出的毒素含量最高达21600 MU／100g肉，其毒性也是最强的；其次为监测点样品，毒螺的检出数量为9．03％，市场样品较少，而新产螺点样品未检出有毒织纹螺。ELISA法测定检出毒螺16份，带毒率为10．32％，两法同时检出毒螺的仅为29．57％，在毒素含量和检出毒螺的数量上ELISA法明显低于小鼠生物法，两者存在显著性差异P〈0．05。结论：宁波市织纹螺的毒性较强，小鼠生物法检出的毒性反映了织纹螺携带的实际毒力，ELISA法检出的是织纹螺中STX含量，说明织纹螺中携带的毒素不仅仅是PSP，该结论与以往报道的织纹螺携带的毒素主要是PSP不一致，提示我们为进一步控制织纹螺中毒，有深入探讨织纹螺麻痹类毒素必要。     

腹泻性贝类毒素的细胞F-肌动蛋白荧光检测法的建立

目的通过检测大田软海绵酸(OA)对HL-7702肝细胞的F-肌动蛋白的解聚作用,建立腹泻性贝类毒素(DSP)的荧光检测法。方法使用鬼比环肽标记F-肌动蛋白,多功能酶标仪检测荧光强度,通过荧光强度的变化分析样品中毒素的含量。比较荧光检测法和ELISA法对贝类样品的检测结果,分析所建荧光检测法的可靠性。结果 OA能明显破坏细胞F-肌动蛋白的聚合。随着作用浓度的增加,破坏程度也随之增加。在2.5～40 nmol/L范围内呈现明显的线性关系(R2=0.9931),检测限值可达到2.01μg/100g贝肉;进行样本检测时,加标回收率为92.76%～96.49%,并与ELISA法检测具有较好的线性相关(R2=0.830)。结论与现有的检测法相比,F-肌动蛋白荧光检测法具有较好的重复性和较低的检出限,可用于有毒贝类的筛查与检测。     

从暴发腹泻的贻贝中同时检出腹泻性贝类毒素和扇贝毒素

目的:2011年5月25日-5月27日,我市苍南县暴发了57例因食用贻贝引起的腹泻性贝类毒素(DSP)中毒事件,采集了19份样品进行检测。方法:采用超高效液相色谱-串联质谱方法对贻贝中5种脂溶性贝类毒素进行检测,根据欧盟贝类毒素的限量值进行评价。结果:15份样品检出大田软海绵酸(OA)、鳍藻毒素-1(DTX-1)和7-epi-pectenotoxin-2 seco acid(7-epi-PTX-2sa),16份检出pectenotoxin-2seco acid(PTX-2sa),7份检出pectenotoxin-2(PTX-2)。其中4份样品中游离OA和DTX-1含量超出欧盟限量值3倍多,它们的总OA约超出9倍。其余均低于欧盟限量值。首次在我国贝类中检出了PTX-2sa和7-epi-PTX-2sa。结论:鉴于部分贻贝样品DSP超标,今后应加强贝类上市前的监测工作。