大西洋三文鱼和虹鳟鱼双重PCR检测方法的建立及应用

为了实现大西洋三文鱼和虹鳟鱼的物种掺假快速检测,本研究建立了一种基于双重PCR（普通和荧光）的物种鉴定方法。通过序列分析,分别基于线粒体COⅠ和Cyt b基因设计大西洋三文鱼和虹鳟鱼特异性引物,验证引物的特异性并对双重PCR反应体系进行优化,建立大西洋三文鱼和虹鳟鱼的双重PCR快速鉴定方法。利用本研究设计的引物,仅大西洋三文鱼和虹鳟鱼可以分别扩增出108 bp和207 bp的特异性条带,而其他23种非目标物种均未有扩增条带。经验证,在双重PCR反应体系中,大西洋三文鱼和虹鳟鱼引物的最佳添加量分别为0.1和0.4 μmol/L,熔解温度分别约为81.5 ℃和85 ℃。利用该方法从29份市售“三文鱼”样品中检测到6份大西洋三文鱼和3份虹鳟鱼,且与DNA条形码比对结果一致。本研究建立的大西洋三文鱼和虹鳟鱼双重PCR（普通和荧光）鉴定方法具有良好的特异性和适用性,为大西洋三文鱼和虹鳟鱼物种鉴定提供了可靠的技术手段。     

从真假三文鱼说起

众所周知，三文鱼大多从挪威进口，都是用冰鲜船装运至我国；而用三文鱼制作的生鱼片，确实称得上是上品，这在烹饪界早已得到了公认。然而你想不到吧，如今市场上还有鲜活的“国产三文鱼”卖呢，你更想不到，这种国内养殖三文鱼原来是经过“染色”的虹鳟鱼呢!     

基于蛋白质/肽组学的三文鱼物种溯源

不同三文鱼在外观和物种亲缘关系上都十分相近,本研究旨在建立一种大西洋鲑鱼、大马哈鱼和虹鳟鱼的物种溯源方法。基于质谱鸟枪法蛋白质组学,在95%置信水平上共鉴定得到912个蛋白质。经过对不同三文鱼特征蛋白质相应肽段的检测特异性和物种特异性筛选分析,得到潜在的物种特征肽段生物标志物。通过多反应监测的靶向肽组学,进一步验证物种特征肽段生物标志物,得到了大西洋鲑鱼10条物种特征肽段、大马哈鱼2条物种特征肽段和虹鳟鱼3条物种特征肽段,并对市售的商品三文鱼产品进行了方法应用。结果显示,该方法操作方便、准确可靠、特异性好、适用性强,可在常规分析实验室对商品三文鱼产品进行有效的鉴别分析。     

广东省三文鱼淡水寄生虫感染情况调查与分析

目的对广东省养殖及市售各环节三文鱼(本文泛指鲑科鱼类)的淡水寄生虫感染状况进行调查,分析广东省居民生食三文鱼感染淡水寄生虫的风险。方法在广州、深圳及佛山市的市售环节共采集三文鱼生155份;在广东省2个养殖场、青海省1个养殖场共采集鳟鱼肉样品84份并对养殖场进行环境卫生调查。对采集的三文鱼肉样品,用酶消化法检测致病性淡水寄生虫。采用SPSS20.0软件进行统计学分析。结果采自养殖、流通及餐饮环节的三文鱼样品共239份,使用酶消化法均未检出华支睾吸虫囊蚴等致病性寄生虫。养殖环境调查显示,3家养殖场环境总体良好,均无人为的粪便污染渠道,其中1家周边可见猫、狗自由出入。结论目前广东省生食三文鱼淡水寄生虫感染风险较低,但仍需持续开展监测及相关研究工作,掌握淡水三文鱼养殖及消费的远期寄生虫感染风险;通过推广三文鱼规范化养殖和鱼生冷冻灭虫处理、加强养殖卫生监管以及饮食卫生宣教等措施,最大程度降低生食三文鱼淡水寄生虫感染风险。     

双重实时荧光PCR法鉴别大西洋鲑鱼和虹鳟鱼

目的建立双重实时荧光PCR法同时鉴别大西洋鲑鱼和虹鳟鱼的检测方法。方法在现有单一物种检测方法的基础上,应用多重实时荧光PCR检测技术对大西洋鲑鱼和虹鳟鱼同时进行检测,并对PCR反应时间和反应温度进行优化。结果该方法通过1管PCR反应实现对大西洋鲑鱼和虹鳟鱼的同时鉴别,反应时间缩短至1 h以内,方法的特异性好,灵敏度可达到0.1%(DNA质量分数)。结论该方法检测效果好、时间短、成本低,可为监管部门解决三文鱼市场监管问题提供技术手段。     

广东省植物源性饮料掺假情况摸底调查

目的对广东省植物源性饮料中掺假情况进行调查,为监管部门提供决策依据。方法利用食品补充检验方法植物蛋白饮料中植物源性成分鉴定(BJS 201707),对广东省植物源性饮料掺假情况进行摸底调查。结果 20批次样品中,共发现7批次样品标识的植物源性成分未检出,未检出率为35%,并且都是标识的核桃源性成分未检出。结论植物源性饮料掺假情况较为突出,食品安全监管部门应加大监管力度。     

工业乳粉糊精掺假初步调查

对十余家乳粉生产企业、代加工企业和乳粉供应商的工业乳粉(大包乳粉)样品进行了多次糊精掺假检测调查.结果发现糊精检出率较高.     

广东省牛羊肉及其制品中掺杂掺假情况的调查分析

目的对广东省内牛、羊肉及其制品中掺杂掺假情况进行风险监测,从而为监管部门的后续监督管理提供指导性意见。方法使用特异性引物和探针,对广东省市场上出售的牛、羊肉及其制品进行动物源性成分鉴定。并与标签明示肉源进行比对,确认掺假类别。结果共检验50份样品,其中牛肉25份,羊肉8份,混合肉类17份。检出10份掺假肉食品,总掺假率为20.0%。掺假样品均为牛肉制品,羊肉制品为未发现掺假情况。结论用猪肉和鸡肉进行肉类的掺假是目前主要的掺假手段。混合肉类制品在标签明示肉类成分方面比较混乱,部分样品检出标签未标示的肉类成分。进一步开展肉制品的掺假检测具有重要的社会意义。     

2013年苏州地区肉及其制品掺假情况调查

了解苏州地区肉及其制品的掺假情况,通过对肉类种源与标签明示肉源进行比对,鉴别摻假食品,为加强食品标签管理提供依据。方法 运用自建的动物源性食品种源判定Taqman实时荧光PCR检测体系对苏州地区的肉及其制品进行种源判定,与标签明示肉源进行比对,鉴别摻假食品。结果 本次调查共检验涉及32个生产单位的90份样品,总不符合率为25.6%(23/90)。检测的44份牛肉及其制品中有12份与标签不符,8份用猪肉部分替代牛肉,1份以鸭肉部分代替牛肉进行销售;此外有3份不含有牛肉成分,存在猪、鸡、鸭源性肉类之外的肉类成分。共检测羊肉及其制品16份,有2份用鸭肉代替羊肉出售,3份羊肉样品中掺入了部分猪成分,其中1份样品还存在单个样品掺杂两种外源肉类的现象(猪源性和鸭源性)。检测猪肉及其制品19份,其中2份样品含有标签未注明的鸡肉成分。在所检测的11份混合肉类样品中有4份成分与标签不符,主要是以廉价的鸡肉取代/部分取代相对高价的牛肉和猪肉。结论 肉制品掺假情况明显,用猪肉、鸭肉部分代替牛肉和羊肉仍是主要的掺假手段,牛肉掺假样品主要是熟制牛肉制品,而火锅食用羊肉卷样品则是羊肉掺假高危品,开展肉制品摻假检测对规范肉制品市场具有积极意义。此外,3份未知种源成分的牛肉样品提示在现有检测基础上还需扩大检测范围,防患于未然。     

三文鱼全垒打

三文鱼常以刺身的面目来行走江湖,以至于人们往往忽略了——它毕竟是一条鱼呀,与所有的鱼类一样,有鱼头、鱼尾、鱼脊骨,当它的主体被盛上冰盘之后,其余的组成该做何处理？     

PCR-RFLP of the mitochondrial cytochrome oxidase gene: a simple method for discrimination between Atlantic salmon (Salmo salar) and rainbow trout (Oncorhynchus mykiss)

A DNA-based method (PCR-RFLP) has been developed for discrimination between Atlantic salmon (Salmo salar) and rainbow trout (Oncorhynchus mykiss). The polymerase chain reaction (PCR) was used for amplification of a 464 bp fragment of the mitochondrial cytochrome oxidase subunit II (COII) gene. Digestion of the products with endonucleases Nci I and Sau 3AI, followed by agarose gel electrophoresis of the digested products, yielded specific restriction profiles that enabled direct visual identification of the species analysed. This PCR-RFLP methodology allowed clear discrimination of Atlantic salmon and rainbow trout samples both in raw and smoked products. © 1999 Society of Chemical Industry     

Immunostick Colorimetric ELISA Assay for the Identification of Smoked Salmon,Trout and Bream

A colorimetric ELISA using immunostick tubes has been developed for the rapid identification of smoked salmon (Salmo salar), trout (Oncorhynchus mykiss) and bream (Brama raii). The assay uses polyclonal antibodies raised in rabbits against muscle-soluble proteins of smoked salmon (anti-SSP), trout (anti-TSP) and bream (anti-BSP) and rendered species-specific by blocking them with the heterologous species of smoked fish. The blocked antibodies were used for the immunorecognition of the smoked fish samples bound to the paddles of immunostick tubes and the immunocomplex detected with peroxidase-conjugated goat anti-rabbit immunoglobulins. The blue colour developed by conversion of the peroxidase substrate allows unequivocal identification of S salar, O mykiss and B raii. © 1997 SCI.     

Fermentation of salmon fillets with a variety of lactic acid bacteria

The influence of five strains of lactic acid bacteria (four Lactobacillus and one Carnobacterium) on the quality of fermented salmon fillets was studied. Best starter growth (increase of more than 1 log in 3 days) and acidification of muscle (e.g. pH reduction of approximately 0.7 units in 5 days) were achieved with the two commercial strains L. sake LAD and L. alimentarius BJ33. pH reduction was consistently lower (e.g. reduction of 0.2 units in 5 days) with C. piscicola 85. Protein breakdown as observed on SDS-PAGE gels was similar for all strains. In contrast, the starter strain did influence texture and colour changes. Fast acidifying strains L. sake LAD and L. alimentarius BJ33 brought about a firmer overall texture and a lighter colour, while softening of flesh occurred in samples processed with C. piscicola 85. Sensory evaluations indicated that samples processed with fast acidifying strains were preferred. L. sake LAD and L. alimentarius BJ33 are regarded as suitable starters for fermentation of salmon fillets.     

麻哈鱼掺假鉴定及制度防范措施和建议

目的建立麻哈鱼掺假感官识别方法及DNA序列储备库,并提出掺假行为在制度上的防范措施和建议。方法制作麻哈鱼简略检索表,对麻哈鱼整条鱼进行形态对比,并对常见麻哈鱼产品进行感官对比。同时按照设定的对应引物和检测方法,列出主要麻哈鱼DNA序列,针对可能发生掺杂施假的麻哈鱼实施DNA检测,可以将得到的序列与建立的基因库序列进行比对。结果感官鉴定并不能单独准确判定麻哈鱼是否掺假,需要结合DNA检测方法,实现快速鉴定。结论该方法可以快速对麻哈鱼进行感官及DNA掺假鉴定,为麻哈鱼检验提供理论依据,促进麻哈鱼加工业的健康发展。     

Lipid distribution and composition of commercially farmed atlantic salmon (salmosalar)

Skin, red and white muscle, belly flap, dorsal fat depot, backbone, head, visceral tissue and liver of commercially farmed Atlantic salmon (Salmo salar) were analysed for total lipid content, lipid classes and fatty acid composition. The fat is deposited in a number of tissues and organs with the highest level in the dorsal fat depot (38.4% of wet weight), red muscle (27.2%) and belly flap (28.1%). The fat content in white muscle is 9.6% of wet weight. The lipid class distribution is nearly uniform throughout the fish body. The belly flap contains the highest amount of triacylglycerols (98.8% of total lipid) and white muscle and visceral tissue have the lowest levels, 93.3% and 93.1%, respectively. The variation in total saturates, monoenes, polyenes, n-6 and n-3 fatty acids was much less between tissues than the variation in total lipid content. In the edible part, the belly flap (16.7%) contained significantly lower levels of n-3 fatty acids than red (18.4%) and white muscle (19.7%) mainly due to lower level of 22: 6n-3 and 20: 5n-3 fatty acids.     

Sensory evaluation of Atlantic salmon fed three types of herring-based diet

Atlantic salmon (Salmo salar) were fed three different types of commercial and experimental herring-based feeds and, at the end of the feeding trials, sensory panels evaluated the acceptability of the salmon fillets. The feeds were all made from Atlantic herring but using different procedures: silage feeds were made from fermented herring, moist salmon feeds from fresh herring, and dry feeds from herring meal. Silage feeds were compared with either one or two of the other feeds in three feeding trials. Growth rates of animals fed a commercial silage feed and two dry feeds were similar, although growth on the commercial silage feed tended to be better. However, growth on commercial or experimental silage feeds was significantly lower (P < 0·001) than on either commercial or experimental moist feeds. Analysis of covariance indicated that there were no significant tank effects. At the end of the feeding trials, sensory evaluation of fish fillets by triangle test indicated no significant difference (P > 0·05) in fillet flavours or appearance between fish fed silage and fish fed the other diets.     

Age of Farmed Atlantic Salmon at Seawater Transfer Affects Muscle Collagen Content and Solubility at Harvest

Salmon released to seawater as different smolt types differed significantly in the total collagen content and the solubility of the collagen present. A 11/2 smolt contained more total collagen than 1/2 and 1 smolt types. Less insoluble collagen and more of the pepsin soluble collagen was generally present in 11/2 smolt type as compared with both 1/2 and 1 types, with the exception of prerigor type 1 salmon. No differences between smolt types in acetic soluble collagen were present. Fish with the higher amount of collagen also had the lowest end pH. Although the bled body weight differed, no correlation between body weight and collagen content or its solubility was found. Fish with higher end pH had lower total collagen and pepsin soluble collagen (r = ‐0.77, P = 0.0001 and r = ‐0.51, P = 0.04, respectively, and more insoluble collagen (r = 0.72, P = 0.002).