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Mini-Mu insertions in the tetracycline resistance determinant from Proteus mirabilis
1Departamento de Microbiologia, Imunologia e Parasitologia, Escola Paulista de Medicina, Universidade Federal de Sö Paulo, Brasil.
Abstract:
The inducible tetracycline resistance determinant isolated from Proteus mirabilis cloned into the plasmid pACYC177 was mutagenized by insertion of a mini-Mu-lac phage in order to define the regions in the cloned sequences encoding the structural and regulatory proteins. Three different types of mutants were obtained: one lost the resistance phenotype and became Lac+; another expressed the resistance at lower levels and constitutively; the third was still dependent on induction but showed a lower minimal inhibitory concentration. The mutant phenotypes and the locations of the insertions indicate that the determinant is composed of a repressor gene and a structural gene which are not transcribed divergently as are other known tetracycline determinants isolated from Gram-negative bacteria.
Insights
Researchers investigated the Proteus mirabilis tetracycline resistance determinant using mini-Mu-lac phage mutagenesis. This revealed a unique genetic structure, differing from other Gram-negative bacteria, with repressor and structural genes not transcribed divergently.
Area of Science:
- Microbiology
- Molecular Biology
- Genetics
Background:
- Tetracycline resistance is a significant concern in bacterial infections.
- Understanding the genetic basis of antibiotic resistance is crucial for developing new treatments.
- The Proteus mirabilis tetracycline resistance determinant's genetic organization was previously unclear.
Purpose of the Study:
- To define the regions encoding structural and regulatory proteins of the Proteus mirabilis tetracycline resistance determinant.
- To elucidate the transcriptional organization of this resistance determinant.
Main Methods:
- Cloning the tetracycline resistance determinant from Proteus mirabilis into plasmid pACYC177.
- Mutagenesis of the cloned determinant using mini-Mu-lac phage insertion.
- Phenotypic analysis of resulting mutants to assess tetracycline resistance and Lac+ activity.
Main Results:
- Three distinct mutant types were generated, characterized by altered tetracycline resistance levels and/or Lac+ expression.
- Mutant analysis indicated the presence of both repressor and structural genes within the determinant.
- Insertion locations correlated with specific phenotypic changes, aiding in gene region definition.
Conclusions:
- The Proteus mirabilis tetracycline resistance determinant comprises a repressor and a structural gene.
- Unlike other known determinants in Gram-negative bacteria, these genes are not transcribed divergently.
- This finding provides new insights into the genetic regulation of tetracycline resistance mechanisms.