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Characterization of two plasmids from Campylobacter jejuni isolates that carry the aphA-7 kanamycin resistance
F C Tenover1, C L Fennell, L Lee
1Seattle Veterans Affairs Medical Center, Washington 98108.
Abstract:
Two small plasmids of 11.5 and 9.5 kb, each carrying an aphA-7 kanamycin phosphotransferase gene, were studied. The MICs of kanamycin for the two human Campylobacter jejuni isolates harboring the plasmids were 10,000 and 5,000 micrograms/ml, while the MICs of amikacin were 32 and 8 micrograms/ml, respectively. The MICs of gentamicin and tobramycin were less than or equal to 2 micrograms/ml for both isolates. The restriction endonuclease maps of the plasmids were similar, with the larger plasmid showing two discrete regions of additional DNA. When the aphA-7 gene from each plasmid was cloned into pBR322, the aphA-7 gene expressed the kanamycin resistance phenotype in Escherichia coli. For transformants containing the cloned aphA-7 gene, kanamycin MICs were greater than or equal to 128 micrograms/ml. The aphA-7 gene was also subcloned from the plasmid pFKT4420 into the E. coli-Streptococcus shuttle vector pDL278 and was transformed into Streptococcus gordonii Challis. For streptococcal transformants containing the novel plasmid, kanamycin MICs were 4,000 micrograms/ml. In the presence of a tetracycline resistance plasmid, both small plasmids could be mobilized during conjugal matings to Campylobacter coli recipients.
Insights
Two Campylobacter jejuni plasmids confer high kanamycin resistance via the aphA-7 gene. This gene functions in E. coli and Streptococcus, demonstrating its broad applicability in antimicrobial resistance studies.
Area of Science:
- Microbiology
- Molecular Biology
- Genetics
Background:
- Campylobacter jejuni is a significant human pathogen.
- Antimicrobial resistance in bacteria is a growing public health concern.
- Plasmids are key vehicles for the dissemination of antibiotic resistance genes.
Purpose of the Study:
- To characterize two small plasmids from Campylobacter jejuni isolates.
- To investigate the function and expression of the aphA-7 kanamycin phosphotransferase gene.
- To assess the potential for plasmid mobilization and gene transfer.
Main Methods:
- Plasmid DNA isolation and characterization.
- Determination of minimum inhibitory concentrations (MICs) for various antibiotics.
- Restriction endonuclease mapping of plasmids.
- Gene cloning and expression studies in Escherichia coli and Streptococcus gordonii.
- Conjugal mating experiments for plasmid transfer.
Main Results:
- Two plasmids (11.5 and 9.5 kb) carrying the aphA-7 gene conferred high-level kanamycin resistance in C. jejuni.
- The aphA-7 gene expressed kanamycin resistance in E. coli and S. gordonii, with high MICs observed.
- Plasmids were mobilizable to Campylobacter coli recipients in the presence of a tetracycline resistance plasmid.
Conclusions:
- The aphA-7 gene is a significant contributor to kanamycin resistance in C. jejuni.
- This gene demonstrates functional expression across different bacterial species, highlighting its potential for broad dissemination.
- The mobilizable nature of these plasmids suggests a mechanism for spreading antibiotic resistance within Campylobacter populations.