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Comparison of the transposon-like structures encoding clindamycin resistance in Bacteroides R-plasmids
Abstract:
The R-plasmids pBF4, pBFTM10, and pBI136 encode transmissible clindamycin resistance (Ccr) in Bacteroides spp. These plasmids are distinct replicons but the regions implicated in Ccr share some homology and appear to have a transposon-like structure. To better understand the mechanism of dissemination and to locate the Ccr determinant(s), the genetic and structural properties of the Ccr regions of each plasmid were compared and contrasted. For this work a single EcoRI restriction fragment containing the Ccr region from each plasmid was cloned into pBR322 in Escherichia coli. Results of restriction mapping and heteroduplex experiments showed that the pBF4 EcoRI-D and pBFTM10 EcoRI-B fragments shared more than 90% base sequence homology but that the EcoRI-C fragment of pBI136 had diverged significantly. The pBI136 fragment also did not confer tetracycline resistance in E. coli as shown for the pBF4 EcoRI-D fragment (D.G. Guiney, P. Hasegawa, and C. E. Davis, 1984, Plasmid 11, 248-252). Heteroduplex experiments showed that the pBI136 EcoRI-C and pBF4 EcoRI-D fragments shared a 1.2-kb region of homology attributed to a directly repeated sequence which bounds the Ccr region. Southern hybridization studies indicated that an additional 0.85 kb of the pBI136 EcoRI-C fragment was homologous to the EcoRI-D fragment of pBF4. This region was characterized by its sequential restriction endonuclease sites for HindIII, AvaII, and DdeI, and it is proposed that the Ccr gene(s) resides in this area.
Insights
Researchers investigated clindamycin resistance (Ccr) genes in Bacteroides plasmids. They found distinct genetic structures, with some regions showing homology, suggesting a common origin for Ccr dissemination.
Area of Science:
- Molecular Biology
- Microbiology
- Genetics
Background:
- R-plasmids pBF4, pBFTM10, and pBI136 confer transmissible clindamycin resistance (Ccr) in Bacteroides species.
- These plasmids are distinct replicons, but Ccr regions exhibit homology and transposon-like structures, indicating potential for dissemination.
Purpose of the Study:
- To elucidate the mechanism of Ccr dissemination in Bacteroides spp.
- To genetically and structurally characterize the Ccr regions of plasmids pBF4, pBFTM10, and pBI136.
- To pinpoint the location of the Ccr determinant(s).
Main Methods:
- Cloning of EcoRI restriction fragments containing Ccr regions into pBR322 in Escherichia coli.
- Restriction mapping and heteroduplex experiments to compare DNA sequences.
- Southern hybridization to assess homology between fragments.
Main Results:
- pBF4 EcoRI-D and pBFTM10 EcoRI-B fragments showed >90% base sequence homology.
- pBI136 EcoRI-C fragment diverged significantly but shared a 1.2-kb homologous region with pBF4 EcoRI-D, attributed to a directly repeated sequence.
- An additional 0.85 kb on pBI136 EcoRI-C was homologous to pBF4 EcoRI-D, containing specific restriction sites (HindIII, AvaII, DdeI), likely harboring Ccr genes.
Conclusions:
- The Ccr regions of these R-plasmids share structural similarities and homologous sequences, suggesting a common evolutionary origin.
- The Ccr determinant(s) are likely located within a specific 0.85-kb region on the pBI136 EcoRI-C fragment, characterized by sequential restriction endonuclease sites.
- Understanding these genetic structures is crucial for comprehending the dissemination of clindamycin resistance in Bacteroides.