Yutong Li, Meng Qu, Enchong Liu, Shuo Wan, Yanhua Jiang, Yingying Guo, Wenjia Zhu, Na Li, Lianzhu Wang, Lin Yao
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This method enabled the simultaneous identification of <i>M. piceus</i>, <i>C. idella</i>, <i>H. molitrix</i>, and <i>H. nobilis</i> in marine fish surimi. The designed primers and probes demonstrated high specificity, with no cross-reactivity to 24 other fish species in the qPCR reaction. The detection limits of the method were 0.0005 ng μL<sup>−1</sup> for <i>M. piceus</i> and <i>H. nobilis</i> and 0.005 ng μL<sup>−1</sup> for <i>C. idella</i> and <i>H. molitrix</i>. The method detected <i>M. piceus</i> at 0.1%, <i>C. idella</i> at 0.01%, and both <i>H. molitrix</i> and <i>H. nobilis</i> at 0.001% in fish mixtures. Among 50 commercial samples, 31 tested positive for one or more of these species.</p><h3>Conclusion</h3><p>The developed qPCR method specifically detects <i>M. piceus</i>, <i>C. idella</i>, <i>H. molitrix</i>, and <i>H. nobilis</i> in marine fish surimi and has potential for use in routine quality control by food regulators.</p></div>","PeriodicalId":561,"journal":{"name":"Food Analytical Methods","volume":"18 10","pages":"2373 - 2383"},"PeriodicalIF":3.0000,"publicationDate":"2025-08-15","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":"{\"title\":\"Development of a Real-Time PCR Method for Identifying Four Major Freshwater Fish in Marine Fish Surimi\",\"authors\":\"Yutong Li, Meng Qu, Enchong Liu, Shuo Wan, Yanhua Jiang, Yingying Guo, Wenjia Zhu, Na Li, Lianzhu Wang, Lin Yao\",\"doi\":\"10.1007/s12161-025-02870-3\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<div><h3>Background</h3><p>The adulteration of marine fish surimi with freshwater fish without consumer disclosure is a growing concern in China. Thus, a rapid detection method is needed for four major freshwater species: black carp (<i>Mylopharyngodon piceus</i>), grass carp (<i>Ctenopharyngodon idella</i>), silver carp (<i>Hypophthalmichthys molitrix</i>), and bighead carp (<i>Hypophthalmichthys nobilis</i>).</p><h3>Methods and Results</h3><p>Mitochondrial NADH dehydrogenase subunit 5 (ND5) sequences were aligned and analyzed to design species-specific primers and probes for a real-time quantitative polymerase chain reaction (qPCR) assay. This method enabled the simultaneous identification of <i>M. piceus</i>, <i>C. idella</i>, <i>H. molitrix</i>, and <i>H. nobilis</i> in marine fish surimi. The designed primers and probes demonstrated high specificity, with no cross-reactivity to 24 other fish species in the qPCR reaction. The detection limits of the method were 0.0005 ng μL<sup>−1</sup> for <i>M. piceus</i> and <i>H. nobilis</i> and 0.005 ng μL<sup>−1</sup> for <i>C. idella</i> and <i>H. molitrix</i>. 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引用次数: 0
摘要
在中国，淡水鱼掺入海鱼鱼糜而不向消费者披露的问题日益受到关注。因此，需要对淡水鱼（Mylopharyngodon piceus）、草鱼（Ctenopharyngodon idella）、鲢鱼（Hypophthalmichthys molitrix）和鳙鱼（Hypophthalmichthys nobilis）四种主要淡水鱼类进行快速检测。方法与结果对线粒体NADH脱氢酶亚基5 （ND5）序列进行比对和分析，设计物种特异性引物和探针，用于实时定量聚合酶链反应（qPCR）检测。该方法可同时鉴定海鱼鱼糜中的piceus、C. idella、H. molitrix和H. nobilis。所设计的引物和探针具有较高的特异性，在qPCR反应中与其他24种鱼类无交叉反应。方法的检出限分别为0.0005 ng μL−1和0.005 ng μL−1。该方法在混合鱼中分别检测出0.1%的青壁棘球蚴、0.01%的idella棘球蚴和0.001%的molitrix和nobilis棘球蚴。在50份商业样本中，31份检测出一种或多种这些物种呈阳性。结论所建立的qPCR方法可在海鱼鱼糜中特异性检测出青piceus、C. idella、H. molitrix和H. nobilis，在食品监管部门的日常质量控制中具有一定的应用价值。
Development of a Real-Time PCR Method for Identifying Four Major Freshwater Fish in Marine Fish Surimi
Background
The adulteration of marine fish surimi with freshwater fish without consumer disclosure is a growing concern in China. Thus, a rapid detection method is needed for four major freshwater species: black carp (Mylopharyngodon piceus), grass carp (Ctenopharyngodon idella), silver carp (Hypophthalmichthys molitrix), and bighead carp (Hypophthalmichthys nobilis).
Methods and Results
Mitochondrial NADH dehydrogenase subunit 5 (ND5) sequences were aligned and analyzed to design species-specific primers and probes for a real-time quantitative polymerase chain reaction (qPCR) assay. This method enabled the simultaneous identification of M. piceus, C. idella, H. molitrix, and H. nobilis in marine fish surimi. The designed primers and probes demonstrated high specificity, with no cross-reactivity to 24 other fish species in the qPCR reaction. The detection limits of the method were 0.0005 ng μL−1 for M. piceus and H. nobilis and 0.005 ng μL−1 for C. idella and H. molitrix. The method detected M. piceus at 0.1%, C. idella at 0.01%, and both H. molitrix and H. nobilis at 0.001% in fish mixtures. Among 50 commercial samples, 31 tested positive for one or more of these species.
Conclusion
The developed qPCR method specifically detects M. piceus, C. idella, H. molitrix, and H. nobilis in marine fish surimi and has potential for use in routine quality control by food regulators.
期刊介绍：
Food Analytical Methods publishes original articles, review articles, and notes on novel and/or state-of-the-art analytical methods or issues to be solved, as well as significant improvements or interesting applications to existing methods. These include analytical technology and methodology for food microbial contaminants, food chemistry and toxicology, food quality, food authenticity and food traceability. The journal covers fundamental and specific aspects of the development, optimization, and practical implementation in routine laboratories, and validation of food analytical methods for the monitoring of food safety and quality.
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