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Characterization of a plasmid codifying the synthesis of a beta-lactamase produced by Shigella flexneri
V Echeverría1, M A Mondaca, R Montoya
1Department of Molecular Biology, Faculty of Biological Sciences, University of Concepción, Chile.
Abstract:
The resistance to beta-lactam antibiotics shown by a strain of Shigella flexneri was plasmid-coded. This plasmid, pMAM-1, when transferred to Escherichia coli K-12 by conjugation, presented the same molecular weight (100 kbp) and conferred the same high level of resistance to ampicillin in the transconjugant as in the wild type strains (MIC, 2048-4096). Restriction analysis of the plasmid in transconjugants revealed various restrictive sites to some endonucleases (i.e. Bam HI, Eco RI, Pst I, Nco I, Cla I, Sf I and Sau 3AI, Nhe and Hin dIII), and no restrictive sites at all for other endonucleases (such as Xho I, Dra I, Kpn I, and Sal I). Some restricted DNA fragments were appropriate for cloning and isolation of the beta-lactamase gene present in Shigella flexneri UCSF 129. This work provides the first step in this direction.
Insights
A plasmid from Shigella flexneri confers high ampicillin resistance to E. coli. This study characterizes the plasmid, pMAM-1, aiding in the isolation of its beta-lactamase gene.
Area of Science:
- Microbiology
- Molecular Biology
- Genetics
Background:
- Antibiotic resistance is a growing global health threat.
- Beta-lactam antibiotics, like ampicillin, are crucial for treating bacterial infections.
- Shigella flexneri strains can develop resistance to these vital drugs.
Purpose of the Study:
- To investigate the genetic basis of beta-lactam antibiotic resistance in Shigella flexneri.
- To characterize the plasmid responsible for ampicillin resistance.
- To lay the groundwork for isolating the beta-lactamase gene.
Main Methods:
- Plasmid transfer via bacterial conjugation from Shigella flexneri to Escherichia coli K-12.
- Determination of plasmid molecular weight (100 kbp).
- Restriction enzyme analysis of the plasmid DNA using various endonucleases.
Main Results:
- The plasmid pMAM-1 successfully conferred high-level ampicillin resistance (MIC 2048-4096) to E. coli K-12 transconjugants.
- Restriction analysis revealed specific cleavage sites for several endonucleases and no sites for others.
- Identified suitable DNA fragments for potential cloning of the beta-lactamase gene.
Conclusions:
- Plasmid pMAM-1 is responsible for ampicillin resistance in the studied Shigella flexneri strain.
- The characterized restriction sites provide a foundation for molecular cloning strategies.
- This research is a critical first step towards understanding and potentially combating beta-lactamase-mediated resistance.