[1]王李宝,黎 慧,于志君,等.四种南美白对虾常见病原微流控芯片联检方法的测试[J].江苏农业学报,2023,(02):471-478.[doi:doi:10.3969/j.issn.1000-4440.2023.02.020]
　WANG Li-bao,LI Hui,YU Zhi-jun,et al.Testing of microfluidic chip for joint detection of four common pathogens of Litopenaeus vannamei[J].,2023,(02):471-478.[doi:doi:10.3969/j.issn.1000-4440.2023.02.020]
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四种南美白对虾常见病原微流控芯片联检方法的测试()
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江苏农业学报[ISSN:1006-6977/CN:61-1281/TN]

卷:
期数:
2023年02期
页码:
471-478
栏目:
畜牧兽医·水产养殖
出版日期:
2023-04-30

文章信息/Info

Title:
Testing of microfluidic chip for joint detection of four common pathogens of Litopenaeus vannamei
作者:
王李宝1; 黎 慧1; 于志君1; 2; 史文军1; 冯卫1; 赵然1; 2; 沈辉1; 成婕1; 管小平3; 秦亚兵4; 万夕和1
（1.江苏省海洋水产研究所，江苏南通226007；2.上海海洋大学水产科学国家级实验教学示范中心，上海201306；3.伊犁悦然生态农业有限公司，新疆伊犁835300；4.如东县大豫镇综合服务中心，江苏南通226412）
Author(s):
WANG Li-bao1; LI Hui1; YU Zhi-jun1; 2; SHI Wen-jun1; FENG Wei1; ZHAO Ran1; 2; SHEN Hui1; CHENG Jie1; GUAN Xiao-ping3; QIN Ya-bing4; WAN Xi-he1
（1.Jiangsu Institute of Oceanology and Marine Fisheries, Nantong 226007 China;2.National Demonstration Center for Experimental Fisheries Science Education, Shanghai Ocean University， Shanghai 201306, China;3.Yili Yueran Ecological Agriculture Co., Ltd. Yili 835300, China;4.Comprehensive Service Center of Dayu Town, Rudong County, Nantong 226412， China）
关键词:
南美白对虾; 病原; 微流控芯片; 联检
Keywords:
Litopenaeus vannamei; pathogens; microfluidic chip; joint detection
分类号:
S945.4
DOI:
doi:10.3969/j.issn.1000-4440.2023.02.020
文献标志码:
A
摘要:
为评估微流控芯片法在虾肝肠胞虫（EHP）、致急性肝胰腺坏死病副溶血性弧菌（VpAHPND）、十足目虹彩病毒1（DIV1）和白斑综合征病毒（WSSV）4种南美白对虾常见病原联检中的各项性能，设计针对上述4种病原扩增反应的引物，对已有芯片进行新的功能划分，同时优化该基因芯片的反应体系及恒温扩增条件。结果表明，优化的联检方法可同时检测上述4种病原，特异性好，30 min反应时间即可给出病原联检结果。EHP、 DIV1、WSSV、VpAHPND指标检出限分别为1 μl 0.57拷贝、0.43拷贝、2.12拷贝 、3.18拷贝，已达到甚至超过了环介导等温扩增技术(LAMP)检测的灵敏性。本次测试中的南美白对虾4种病原微流控芯片联检方法所用的检测试剂盒具有操作简便、检测灵敏、特异性强、检测时间短、设备要求简单等特点，适合整个养殖过程中对南美白对虾EHP、VpAHPND、DIV1和WSSV的监测与筛查工作。
Abstract:
To evaluate the performance of microfluidic chip method in the joint detection of four common pathogens of Litopenaeus vannamei including Enterocytozoon hepatopenaei (EHP), acute hepatopancreas necrosis disease caused by Vibrio parahaemolyticus (VpAHPND), Decapod iridescent virus 1 (DIV1) and white spot syndrome virus (WSSV), primers for the amplification reactions of the above four pathogens were designed, new functional divisions were made for the existing chips, and the reaction system and constant temperature amplification conditions of the microfluidic chip were optimized. The results showed that, the optimized combined detection method could detect the above four pathogens simultaneously with good specificity, and the joint detection results could be obtained within 30 min reaction time. The detection limits of EHP, DIV1, WSSV and VpAHPND were 0.57 copies/μl, 0.43 copies/μl, 2.12 copies/μl and 3.18 copies/μl, respectively. Some of the limits had reached or even exceeded the testing sensitivity of loop-mediated isothermal amplification (LAMP) method. The detection kit used in the joint detection method of microfluidic chip for the four pathogens of L. vannamei in this test is characterized by simple operation, sensitive detection, strong specificity, short detection time and simple equipment requirements, and is suitable for the monitoring and screening of EHP, VpAHPND, DIV1 and WSSV of L. vannamei in the whole breeding process.

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[1]宋居易,刘昱辰,张苏楠,等.南美白对虾4种病原体可视化快速检测试剂盒（生物芯片法）的研发[J].江苏农业学报,2020,(03):656.[doi:doi:10.3969/j.issn.1000-4440.2020.03.018]
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备注/Memo

备注/Memo:
收稿日期：2022-07-20 基金项目：江苏省农业科技自主创新基金项目[CX（21）3160]；江苏省种业振兴揭榜挂帅项目[JBGS（2021）122]；江苏省农业重大新品种创制项目（PZCZ201747）；江苏现代农业产业技术体系项目[JATS（2022）419、JATS（2022）164];苏北科技专项先导性项目（SZ-YC202114）作者简介：王李宝(1978-)，男，江苏南通人，硕士，正高级工程师，主要从事海水健康养殖研究。（E-mail）zebra_w@126.com。黎慧为共同第一作者。通讯作者：万夕和，（Tel）13338089869；（E-mail）wxh1708@163.com
更新日期/Last Update: 2023-05-12