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KMID : 0357319950300030325
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Journal of the Korean Society for Microbiology
1995 Volume.30 No. 3 p.325 ~ p.344
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Screening of TEM-1 Beta-Lactamase Gene by Dot Blot Hybridization
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조동택
최관수/이제철/김정민/이상화/이유철/김정완/설성용
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Abstract
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The efficiency of dot blot hybridization combined with plasmid profile analysis was evaluated for the molecular epidemiology of nosocomial infections or various outbreaks of infectious diseases.
Fifty strains of gram-negative organisms which had been confirmed to be nosocomial agents by CDC(Centers for Disease Control, Atalnta, Georgia, U.S.A.) were subjected to the study of the validity of identifying method for the above purpose.
Non-isotopic labeled TEM-1 beta-lactamase gene(digoxigenin labeled) was used for dot blot hybridization and Southern hybridization. Time required for the detection of TEM-1 gene by dot blot hybridization found to be one or two days less than
those
of
Southern hybridization and also yielded quite a specific test result.
Strains which showed positive TEM-1 gene in dot blot hybridization were further studied by Southern hybridization. In most strains of E. coli, TEM-1 gene was encoded on plasmid of 13.9 kilobase.
The plasmids encoding TEM-1 gene of Klebsiella pneumoniae were much larger than those of E. coli strains, and two strains of Klebsiella pneumoniae carried multiple TEM-1 gene which was encoded in the plasmid of various size.
Among many techniques of plasmid profile analysis, Bimboim and Doly method and Portnoy method have been known to be most simple and effective especially in case of gram-negative organisms. Portnoy method was able to save at least one day compared
to
Bimboim and Doly method and the quality of data was not so bad as those obtained by the latter method.
Dot hybridization method by using non-isotopic labeled gene probe was found to be essential and most excellent primary test for the rational approach to investigate molecular epidemiology of nosocomial outbreaks and could be applied and performed
in
ordinary clinical laboratory because of its simplicity and safety of technique without any difficult problem due to the handling of radioisotope of p32 or S35.
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KEYWORD
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