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Isolation and characterization of two plasmids that mediate macrolide resistance in Escherichia coli: transferability
J Katayama1, H Okada, K O'Hara
1Department of Microbiology, School of Pharmacy, Tokyo University of Pharmacy and Life Science, Hachioji, Japan.
Abstract:
Escherichia coli BM2506 is highly resistant to macrolide antibiotics; it produces macrolide 2'-phosphotransferase II [MPH(2')II] which inactivates such drugs. We investigated the localization and the transfer of the macrolide-resistance determinant that encoded the mphB gene for MPH(2')II in strain BM2506. Although we detected no clear band of plasmid DNA after agarose gel electrophoresis, transformation analysis using satellite DNA that corresponded to plasmid DNA after CsCl-ethidium bromide gradient centrifugation and restriction analysis of plasmid DNA in transformants showed that strain BM2506 harbored two plasmids, pTZ3721 (84 kb) and pTZ3723 (24 kb), that specified resistance to macrolides, ampicillin, streptomycin, tetracycline and sulfonamide and to macrolides and ampicillin, respectively. Southern hybridization showed that the mphB gene hybridized to both plasmids. Furthermore, pTZ3721 was transferred by conjugation to another strain of E. coli and pTZ3723 was mobilized with a self-transferable plasmid RP1 to other strains of E. coli. Therefore, it appears that the mphB gene is located on two plasmids in BM2506 and can be transferred to other strains of E. coli by conjugation or mobilization.
Insights
The mphB gene, responsible for macrolide antibiotic resistance in Escherichia coli BM2506, is located on two plasmids. These plasmids can be transferred to other bacterial strains via conjugation or mobilization.
Area of Science:
- Microbiology
- Molecular Biology
- Genetics
Background:
- Escherichia coli BM2506 exhibits high resistance to macrolide antibiotics.
- This resistance is mediated by macrolide 2'-phosphotransferase II (MPH(2')II), an enzyme that inactivates these drugs.
Purpose of the Study:
- To investigate the localization of the mphB gene, which encodes MPH(2')II.
- To determine the transferability of the macrolide-resistance determinant in E. coli BM2506.
Main Methods:
- Plasmid DNA isolation and analysis using CsCl-ethidium bromide gradient centrifugation.
- Transformation assays and restriction enzyme digestion of plasmid DNA.
- Southern hybridization to confirm gene localization.
- Conjugation and mobilization experiments for gene transfer.
Main Results:
- Strain BM2506 harbors two plasmids, pTZ3721 (84 kb) and pTZ3723 (24 kb), conferring various antibiotic resistances.
- The mphB gene was detected on both plasmids.
- Plasmid pTZ3721 was transferred by conjugation, and pTZ3723 was mobilized to other E. coli strains.
Conclusions:
- The mphB gene is located on two distinct plasmids within E. coli BM2506.
- This macrolide resistance gene can be disseminated to other bacterial strains through horizontal gene transfer mechanisms.