Appl Microbiol Biotechnol DOI 10.1007/s00253-014-5855-8
APPLIED GENETICS AND MOLECULAR BIOTECHNOLOGY
Improved PCR assay for the specific detection and quantitation of Escherichia coli serotype O157 in water Min Seok Cho & Kiseong Joh & Tae-Young Ahn & Dong Suk Park
Received: 9 April 2014 / Revised: 11 May 2014 / Accepted: 23 May 2014 # Springer-Verlag Berlin Heidelberg 2014
Abstract Escherichia coli serotype O157 is still a major global healthcare problem. However, only limited information is now available on the molecular and serological detection of pathogenic bacteria. Therefore, the development of appropriate strategies for their rapid identification and monitoring is still needed. In general, the sequence analysis based on stx, slt, eae, hlyA, rfb, and fliCh7 genes is widely employed for the identification of E. coli serotype O157; but there have been critical defects in the diagnosis and identification of E. coli serotype O157, in that they are also present in other E. coli serogroups. In this study, NCBI-BLAST searches using the nucleotide sequences of the putative regulatory protein gene from E. coli O157:H7 str. Sakai found sequence difference at the serotype level. The specific primers from the putative regulatory protein gene were designed and investigated for their sensitivity and specificity for detecting the pathogen in environment water samples. The specificity of the primer set was evaluated using genomic DNA from 8 isolates of E. coli serotype O157 and 32 other reference strains. In addition, the sensitivity and specificity of this assay were confirmed by successful identification of E. coli serotype O157 in
Electronic supplementary material The online version of this article (doi:10.1007/s00253-014-5855-8) contains supplementary material, which is available to authorized users. M. S. Cho : D. S. Park (*) National Academy of Agricultural Science, Rural Development Administration, Suwon 441-707, Republic of Korea e-mail: [email protected] K. Joh Department of Bioscience and Biotechnology, Hankuk University of Foreign Studies, Yongin, Gyeonggi 449-791, Republic of Korea M. S. Cho : T.
APPLIED GENETICS AND MOLECULAR BIOTECHNOLOGY
Improved PCR assay for the specific detection and quantitation of Escherichia coli serotype O157 in water Min Seok Cho & Kiseong Joh & Tae-Young Ahn & Dong Suk Park
Received: 9 April 2014 / Revised: 11 May 2014 / Accepted: 23 May 2014 # Springer-Verlag Berlin Heidelberg 2014
Abstract Escherichia coli serotype O157 is still a major global healthcare problem. However, only limited information is now available on the molecular and serological detection of pathogenic bacteria. Therefore, the development of appropriate strategies for their rapid identification and monitoring is still needed. In general, the sequence analysis based on stx, slt, eae, hlyA, rfb, and fliCh7 genes is widely employed for the identification of E. coli serotype O157; but there have been critical defects in the diagnosis and identification of E. coli serotype O157, in that they are also present in other E. coli serogroups. In this study, NCBI-BLAST searches using the nucleotide sequences of the putative regulatory protein gene from E. coli O157:H7 str. Sakai found sequence difference at the serotype level. The specific primers from the putative regulatory protein gene were designed and investigated for their sensitivity and specificity for detecting the pathogen in environment water samples. The specificity of the primer set was evaluated using genomic DNA from 8 isolates of E. coli serotype O157 and 32 other reference strains. In addition, the sensitivity and specificity of this assay were confirmed by successful identification of E. coli serotype O157 in
Electronic supplementary material The online version of this article (doi:10.1007/s00253-014-5855-8) contains supplementary material, which is available to authorized users. M. S. Cho : D. S. Park (*) National Academy of Agricultural Science, Rural Development Administration, Suwon 441-707, Republic of Korea e-mail: [email protected] K. Joh Department of Bioscience and Biotechnology, Hankuk University of Foreign Studies, Yongin, Gyeonggi 449-791, Republic of Korea M. S. Cho : T.
