Najafi A, Qorbanalizadgan M, Tavakoli H, Ahmadi A. Rapid ِِ Diagnosis of Listeria monocytogenesby PCR Method with hlyAgene . Iran J Med Microbiol 2009; 3 (2 and 3) :9-14
URL:
http://ijmm.ir/article-1-151-en.html
1- Molecular Biology Research Center, Baghiatollah University of Medical Sciences, Tehran, Iran
2- Nutrition Department, Public Health Research Center, Baghiatollah University of Medical Sciences, Tehran, Iran. , h.tavakoli1344@yahoo.com
Abstract: (19388 Views)
Background and Objectives: Currently the conventional method for the diagnosis of Listeria
monocytogenes is culture method. However, the isolation rate is reduced in culture due to animals' food
which is contains antibiotics. Besides, culture method is time consuming and takes more than 36 hrs. So
more precise and rapid technique is needed. The aim of this study was application of PCR technique based
on hlyA gene fragment for rapid detection of Listeria monocytogenes in clinical samples.
Materials and Methods: hlyA gene was selected as a specific target sequence for detection of Listeria
monocytogenes. Listeria monocytogenes PTCC1163 was used as a standard organism for optimization
experiments. A range of bacterial pathogens was used for specificity test including Escherichia coli: ATCC
35218 Salmonella typhi PTCC: 1609. Phenol – chloroform method was used for DNA extraction. PCR
technique was performed as per standard protocol. Amplified product was detected by 1% gel agarose
electrophoresis, stained by ethidiome bromide.
Results: Provided data confirmed amplification of expected product. Specificity test proved no cross
reaction with tested organisms. Sensitivity test detected 500fg Listeria monocytogenesDNA as a final
detection limit.
Conclusion: Optimized experiment confirmed that the applied PCR protocol is quite fast with high
sensitivity and specificity performing in less than three hours.
Application of this method in clinical laboratories can help the rapid diagnosis of Listeria monocytogenesin
samples.
Type of Study:
Original Research Article |
Subject:
Medical Bacteriology Received: 2013/11/22 | Accepted: 2013/11/22 | ePublished: 2013/11/22