title: Visual Detection of Enterohemorrhagic Escherichia coli O157:H7 Using Loop-Mediated Isothermal Amplification.
abstract: BACKGROUND
  Escherichia coli O157:H7, an important foodborne pathogen, can cause serious renal damage, which can also lead to mortality. Since a rapid and sensitive method is needed to identify this pathogenic agent, we evaluated Loop-Mediated Isothermal Amplification Assay (LAMP) to detect Escherichia coli O157:H7.
  
  
  METHODS
  We used six primers that specifically identified the rfbE gene. To examine the sensitivity of the method, different dilutions were subjected to the LAMP reaction. Other bacterial strains also were investigated to determine the specificity of the test. The turbidity of the amplified products was assayed by visual detection. The amplified products were detected by addition of SYBR Green II to the reaction tubes.
  
  
  RESULTS
  Amplification products were observed as a ladder-like pattern on the agarose gel. A white turbidity emerged in the positive tubes. Under UV light, the positive samples were green, whereas the negative samples were orange. The detection limit of the LAMP was 78 pg/tube, and this indicated that it was 100 times more sensitive than PCR for the detection of EHEC. No LAMP products were detected when template DNA of non-EHEC strains were used, suggesting high specificity of the LAMP assay.
  
  
  CONCLUSIONS
  The results indicated that the LAMP assay is a valuable diagnostic assay to identify EHEC O157:H7. In addition, the simplicity, sensitivity, specificity, and rapidity of this assay make it a useful method to diagnose pathogens in primary labs without any need for expensive equipment or specialized techniques.
authors: Ph.D. of Medical Bacteriology, Professor, Molecular Biology Research Center, Baqiyatallah University of Medical Sciences, Tehran, Iran.
authors: Reza
authors: R
authors: Ranjbar
authors: M.Sc. of Biotechnology, Department of Agriculture and Plant Breeding, Faculty of Agriculture, Zanjan University, Zanjan, Iran.
authors: Maryam
authors: M
authors: Erfanmanesh
authors: Ph.D. of Medical Bacteriology, Department of Pathobiology, School of Public Health, Tehran University of Medical Sciences, Tehran, Iran.
authors: Davoud
authors: D
authors: Afshar
authors: Ph.D. of Pharmaceutical Biotechnology, Assistant Professor, Department of Pharmaceutical Biotechnology, Faculty of Pharmacy, Lorestan University of Medical Sciences, Khorramabad, Iran.
authors: Mohsen
authors: M
authors: Mohammadi
authors: Ph.D. Candidate of Pharmaceutical Biotechnology, Department of Pharmaceutical Biotechnology, Tehran University of Medical Sciences, Tehran, Iran.
authors: Omar
authors: O
authors: Ghaderi
authors: Department of Agriculture and Plant Breeding, Faculty of Agriculture, Zanjan University, Zanjan, Iran.
authors: Ali
authors: A
authors: Haghnazari
keywords: Enterohemorrhagic Escherichia coli O157:H7; Loop-Mediated Isothermal Amplification; SYBR Green II; rfbE gene