A Preliminary Study on Multiplex PCR for Detecting Animal-derived Escherichia coliO157:H7

doi:10.7668/hbnxb.2011.05.018

Abstract

Abstract: To develop a multiplex PCR method for rapid and specific detection of Escherichia coli O157∶ H7,two pairs of primers were designed based on the rfbE and fliC genes of Escherichia coli O157∶ H7 in GenBank,from which two sets of PCR were established specific to rfbE and fliC gene,respectively.And then they were combined to a multiplex PCR after being optimized.The multiplex PCR system was evaluated for its specificity and sensitivity,and used to detect the clinical isolates.The results showed that the assay could amplify the 327 bp and 247 bp regions of corresponding genes rfbE and fliC.The multiplex PCR method had the characteristics of good specificity and repeatability,the sensitivity being 102 cfu/mL of bacteria samples.The multiplex PCR was a rapid,specific and sensitive method for the detection of Escherichia coli O157∶ H7,and provided new method for detection of Escherichia coli O157∶ H7.

Key words: Escherichia coli O157∶ H7, Multiplex PCR, rfbE gene, fliC gene

CLC Number:

S854.4

CHEN Ya-jun, HU Hui, DUAN Zhi-gang, CUI Bao-an, ZHANG Long-xian, MENG Zhen-bei, PENG Xin-ran, CHEN Li-ying, WANG Ya-bin. A Preliminary Study on Multiplex PCR for Detecting Animal-derived Escherichia coliO157:H7[J]. ACTA AGRICULTURAE BOREALI-SINICA, 2011, 26(5): 87-91. doi: 10.7668/hbnxb.2011.05.018.

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References

[1] 叶菊莲,占利,陆群英,等.浙江省O157大肠杆菌监测及其分子生物学特性[J].中国人兽共患病学报,2008,24(3):248.
[2] 徐晓可,吴清平,周艳红,等.肉类中大肠杆菌O157:H7多重PCR检测方法的建立[J]微生物学通报,2008,35(4):619-622.
[3] Holland J L,Louie L,Simor A E,et al.,Detection of Escherichia coli O157:H7 directly from stools:evaluation of commercial extraction methods for purifying fecal DNA[J].J Clin Microbiology,2000,11:4108-4113.
[4] Slutsker L,Ries A A,Greene K D,et al.Escherichia coli O157:H7 diarrhea in the United States;clinical and epidemiologic features[J].Ann Intern Med,1997,126 (7):505-513.
[5] 潘群兴,王永山,刘洁,等.肠出血性大肠杆菌O157:H7 rfbE与fliC基因的表达及鉴定[J].江苏农业学报,2009,25(3):568-571.
[6] 吴家林,肖勇,凌霞,等.肠出血性大肠杆菌O157:H7多重PCR快速检测研究[J].现代预防医学,2009,36(1):117.
[7] 郑桂丽,廖绍安,李钏华,等.大肠杆菌O157特异基因的PCR检测方法[J].中国卫生检验杂志,2005,15(10):1183-1185.
[8] 宋宏新,李宏,邢佩.食品中大肠杆菌O157:H7的选择性培养与检测方法研究[J].陕西科技大学学报,2008,26(2):64-67.
[9] Udo Reischl,Mohammad T Y,Jochen K,et al.Real-Time fluorescence PCR assays for detection and characterization of Shiga toxin,intimin,and enterohemolysin genes from Shiga toxin-producing Escherichia coli[J].J Clin Microbiology,2004,42 (9):4092-4100.
[10] Rüttler M E,Yanzo C S,Cuiti o M J,et al.Evaluation of a multiplex PCR method to detect enteroaggregative Escherichia coli[J].Biocell,2006,30:301-308.
[11] Pass M,Odedra R,Batt R M.Multiplex PCRs for identication of Escherichia coli virulence genes[J].J Clin Microbiology,2000,5:2001-2004.
[12] 张雪寒,何孔旺,卢维彩,等.出血性大肠杆菌O157:H7 Stx2B-Tir-Stx1 B多价融合蛋白表达及免疫原性研究[J].华北农学报,2010,25(4):180-185.
[13] Bilge S S,Vary J C,Dowell S F,et al.Role of the Escherichia coli O157:H7 side chain in adherence and analysis of an rfb locus[J].Infection and Immunity,1996,64(11):4795-4801.
[14] Wang L,Reeves P R.Organization of Escherichia coli O157:H7O atigen gene cluster and identification of its specific genes[J].Infection and Immunity,1998,66:3545-3551.

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