Microsoft Word 肖剑-二校 682

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1 6 2 Vol. 6 No Journal of Food Safety and Quality Feb., 2015 肖剑 1, 陈秀云 2, 梁美丹 1, 陈楷 1, 洪红 1, 焦红 3* (1., ; 2., ; 3., ) 摘要 : 目的 (simultaneous amplification and testing, SAT) 方法, SAT, 结果 4 SAT 100%, ; (94.2%)(99.5%) (100%)(100%), 9.6%, 94% 结论 SAT, 4 关键词 : SAT; ; ; Simultaneous amplification and testing application in detection of foodborne pathogenic microorganisms XIAO Jian 1, CHEN Xiu-Yun 2, LIANG Mei-Dan 1, CHEN Kai 1, HONG Hong 1, JIAO Hong 3* (1.Guangzhou Institute of Food Inspection, Guangzhou , China; 2. Guangzhou Yi-Jian Biomedical Technology Development Co. Ltd, Guangzhou , China 3. Guangdong Entry-Exit Inspection and Quarantine Bureau, Guangzhou , China) ABSTRACT: Objective To evaluate the application value of simultaneous amplification and testing (SAT) in detection of foodborne pathogenic microorganisms. Methods The sensitivity, specificity, false positive rate, false negative rate and accuracy of Salmonella spp., Staphylococcus aureus, Vibrio parahaemolyticus and Listeria monocytogenes were studied by using the methods of national standards and molecular sequencing. Meanwhile, the significant difference of positive rate was analyzed by the chi-square. Results The sensitivity of four pathogens were all 100%, and there were no false negative and misdetection rate. The specificity of Salmonella spp., Staphylococcus aureus, Vibrio parahaemolyticus and Listeria monocytogenes were 94.2%, 99.5%, 100%, and 100% respectively, and they all conformed to the requirements of the test. The false positive rates were all below 9.6% and the accuracy rates were superior to the standard (94%). Conclusion It is feasible to detect the four pathogens above with SAT for its high sensitivity, specificity and accuracy. KEY WORDS: simultaneous amplification and testing; food detection; sensitivity; specificity * 通讯作者 :,, jhciq1228@163.com *Corresponding author: JIAO Hong, Professor, Guangdong Entry-Exit Inspection and Quarantine Bureau, No.66, Zhujiang New City Huacheng Avenue, Guangzhou , China. jhciq1228@163.com

2 2, : 引言,, 30% [1],,, [2] 2013 ( ), 32.2%,, 60.4%,, [3],, ELISA [4] [5] [6] [7], DNA PCR [8,9], [10],,, (simultaneous amplification and testing, SAT),,, [11] [12] [13] [14] SAT (2~12 h),,,, SAT (Salmonella spp.) (Staphylococcus aureus) (Vibrio parahaemolyticus)(listeria monocytogenes), 4 SAT, SAT 2 材料与方法 2.1 样品, 1 表 1 试验用样品种类批次及检验项目 Table 1 The sample types, numbers and inspection items () 1 ( ) 30 2 () 30 3 (() ( ) ) 30 4 () ( ) 30 7 () 30 8 ( ( ) ) 30 9 (() ( ( )) ()

3 主要培养基及试剂 BPW (, ); 7.5% (); 3(); (); Baird-Parker ; LB (); (, ); ( ); SAT ( ); (()) 2.3 主要试验仪器 ( SP250); ( GR85DR); (ThermoFisher1300); VITEK2 ( ); (AB MagMAXTM Express 96); PCR (ABI 7500FAST) 2.4 试验方法及原理国标法 GB [15] GB [16] GB [17] GB [18] SAT 法 SAT 2.4.1, 200 μl, 10 μl, 200 μl 100 μl (), 60 5 min, 10 min, ( ), RNA, ;, 30 μl PCR, 10 μl SAT, PCR,, SAT, ( ), NCBI( Blast 统计分析 , SAT, [19] : 98%, 90.4%, <2%, <9.6%, 94%, <3.84, 2 表 2 统计分析表 Table 2 Statistical analysis table A B A+B C D C+D A+C B+D N=A+B+C+D A ; B, ; C, ; D 1) (p+)= A / (A+C) * 100% 2) (p-)= D / (B+D) * 100% 3) (relative accuracy)= (A+D) /N * 100% 4)(pf-)=(1- )* 100% 5)(pf+)=(1- )* 100% ,, McNemar s (χ 2 ), [19] : 2 2 ( a - b 1) χ a b : a ; b 3 结果 3.1 SAT 法检验结果 2.4.3, 4 SAT, 3

4 2, : 685 表 3 Table 3 4 种致病菌 SAT 法与国标法结果统计表 The results of four pathogens tested with SAT and national standard SAT 方法性能指标及显著性差异分析, SAT, ( 4) 100%, 0%,,,,, (χ 2 =40.0) ; 100%, 0%, ; 100%,, 100% [19], ;, ;, ; SAT,,, 3.3 SAT 法测序方法评价分析 SAT, 42 30, 12 ; 8 7, 1 ; 2,, 5, 3 SAT + ( 6), 2.4.3, : 100%, 0%,, 90.4%( ), 94%(); 5.8%, 0.5%, [19],,, 7 4 讨论与结论 SAT Gen-probe (transcription mediated amplification, TMA), RNA, RNA,,,,,, Table 4 表 4 4 种致病菌 SAT 法与国标法比较性能指标及显著性差异结果 The performance indexes and the significant differences of four pathogens with SAT and national standard (%) (%) (%) (%) (%) χ

5 686 6 Table 5 表 5 不同检测项目测序结果情况 The results of difference of inspection item SAT Table 6 表 6 几种致病菌 SAT 法与国标 + 测序法结果统计表 The results of several pathogens tested with SAT and national standard & sequencing SAT 表 7 4 种致病菌测序后 SAT 法与国标法性比较性能指标及显著性差异结果 Table 7 The performance indexes and the significant difference of four pathogens with SAT by national standard and sequencing (%) (%) (%) (%) (%) χ , [12], SAT,,, 4 100%, ;, 42, 30, (94.2%), (5.8%), (96.4%); 8 7, SAT 99.5%, 0.5%, 99.6%; 2, 100%, SAT,

6 2, : 687,,, ;, ;,,,, ;,, SAT,, 参考文献 [1] WHO. Background Paper: Developing A Food Safety Strategy Draft [Z] [2],,. [J]., 2011, 23 (11): , Sun XL, Jang DL, Zhang YZ. Biosensors used in the detection of foodborne pathogenic bacteria [J]. Chin J Microecol, 2011, 23 (11): , [3],,,. [J]., 2012, 33(21 ): Feng L, Huang JC, Liu X, et al. Research Progress on Rapid Detection of Food-borne Bacterial Pathogens [J]. Food Sci, 2012, 33(21): [4],,,. ELISA GICA [J]., 2007, 17(3): Wu QP, Wang DP, Yang J, et al. ELISA and GICA in detection of food-borne pathogens [J]. Chin J Health Lab Technol, 2007, 17(3): [5],. [J]., 2011, 1: Zhang X, Xian Y. Immune magnetic beads and which using in detection of food microbiological [J]. Qual and safety of Agro-products, 2011, 1: [6],. [J]., 2010, 35(3): Ren YN, Che ZM. The application of immunolabelling technique in food safety detection [J]. Chin Cond, 2010, 35(3): [7] Danscher G, Norgaard.JO. Light microscopic visualization of colloidal gold on resin-embeddent issue [J]. J Histochem Cytochem, 1983, 31(12): [8],,. [J]., 2010, 31(9): Xu WJ, Zhou W, Xu CX. Application of Molecular Technology in Detection of Food-borne Pathogens [J]. Anim Husbandry Feed Sci, 2010, 31(9): [9] Pandey BD, Poudel A, Yoda T, et al. Development of an in-house loop-mediated isothermal amplification(lamp) assay for detection of Mycrobacterium tuberculosis and evaluation insputum samples of Nepalese patients [J]. J Med Microbiol, 2008, 57 (4): [10],,,. [J]., 2008, 18(12): Wu YW, Wu XW, Wu QP, et al. Advance on methods for microbial testing slip of food hygiene detection [J]. Chin J Health Lab Technol, 2008, 18(12): [11],,,. (SAT) [J]., 2012, 34(6): Sha W, He Y, Jiang RH, et al. Diagnostic value of Simultaneous Amplification and Testing(SAT) in patients with pulmonary tuberculosis [J]. Chin J Antituberc, 2012, 34(6) : [12],,,. J., 2013, 24(2): Li LH, Chen LD, Liu WT, et al. Comparison of simultaneous amplification and testing and liquid culture in the detection of ureaplasma urealyticum [J]. Lett Biotechnol, 2013, 24(2): [13],,,. J., 2010, 28(4): Gu WM, Yang Y, Wu L, et al. Simultaneous Amplification and Testing in Chlamydia trachomatis infection of the urinary tract [J]. Chin J Clin Lab Sci, 2010, 28(4): [14],,. [J]., 2012, 33(4): Gao ZH, Jin Y, Chen F. Simultaneous Amplification and Testing

7 688 6 in Neisseria gonorrhoeae of urine [J]. Int J Lab Med, 2012, 33(4): [15] GB [S]. GB National food safety standard Food [19],. [M]., Li H, Lei ZW. Handbook of Validation and Confirmation of Food Microbiological Methods [M]. China Standard Publishing House, microbiological examination: Salmonella [S]. [16] GB [S]. GB National food safety standard Food microbiological examination: Staphylococcus aureus [S]. [17] GB [S]. GB National food safety standard Food microbiological examination: Vibrio parahaemolyticus [S]. [18] GB [S] GB National food safety standard Food microbiological examination: Listeria monocytogenes [S]. 作者简介 ( 责任编辑 : 白洪健 ),, xjhq521@163.com,, jhciq1228@163.com

Microsoft Word 王似锦-二校 681

Microsoft Word 王似锦-二校 681 6 2 Vol. 6 No. 2 2015 2 Journal of Food Safety and Quality Feb, 2015 VIDAS 王似锦 *, 高春 ( ( ), 100035) 摘要 : 目的 (VIDAS) 方法 21 BPW BPW, VIDAS, 结果 21, BPW ; BPW 20 VIDAS GB, 95.2% 结论 VIDAS 96 h, VIDAS 48 h VIDAS,,