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Molecular analyses of conjugative, gentamicin-resistance plasmids from staphylococcal clinical isolates
1School of Basic Life Sciences, University of Missouri, Kansas City 64110.
Abstract:
Gentamicin-resistant Staphylococcus aureus and Staphylococcus epidermidis strains which were isolated from infants with staphylococcal bacteremia were analyzed for the presence of self-transmissible gentamicin-resistance (Gmr) plasmids. Conjugative GMr plasmids of approximately 43.8-63 kilobases (kb) were found in all S. aureus strains. Inter- and intra-species transfer of Gmr plasmids by conjugation was observed from S. aureus to S. aureus and to S. epidermidis recipient strains. However, neither inter- nor intra-species transfer of gentamicin resistance by conjugation was observed with nine out of nine S. epidermidis donor strains which were mated with either S. epidermidis or S. aureus recipient strains. These conjugative Gmr plasmids were unable to comobilize a smaller (15-kb) plasmid present in all but two S. aureus clinical isolates. Many of the conjugative Gmr plasmids also carried genetic determinants for kanamycin, tobramycin, neomycin, and ethidium bromide resistance, and for beta-lactamase synthesis. EcoRI restriction endonuclease digests of the S. aureus Gmr conjugative plasmids revealed three different digestion patterns. Four EcoRI restriction endonuclease digestion fragments of 15, 11.4, 6.3, and 4.6 kb in size were common to all plasmids. These plasmids and conjugative Gmr staphylococcal plasmids from other geographical regions shared restriction digestion fragments of similar molecular weights. DNA hybridization with biotinylated S. aureus plasmid pIZ7814 DNA revealed a high degree of homology among these plasmids. A 50.9-kb plasmid from one of the nonconjugative S. epidermidis clinical isolates showed homology with the probe DNA but lacked a portion of a 6.3-kb fragment which was present in all conjugative plasmids and believed to carry much genetic information for conjugation.
Insights
Gentamicin-resistant Staphylococcus aureus strains harbor conjugative plasmids that transfer gentamicin resistance to other bacteria. However, Staphylococcus epidermidis strains in this study did not transfer resistance, indicating limited plasmid mobility.
Area of Science:
- Microbiology
- Molecular Biology
- Genetics
Background:
- Staphylococcal bacteremia in infants is a significant clinical concern.
- Antibiotic resistance, particularly to gentamicin, complicates treatment.
- Understanding the genetic basis of resistance, specifically plasmid-mediated transfer, is crucial.
Purpose of the Study:
- To investigate the presence and characteristics of gentamicin-resistance (Gmr) plasmids in Staphylococcus aureus and Staphylococcus epidermidis.
- To determine the conjugative transfer capabilities of these Gmr plasmids between staphylococcal species.
- To analyze the genetic content and relatedness of identified Gmr plasmids.
Main Methods:
- Isolation and characterization of gentamicin-resistant Staphylococcus aureus and Staphylococcus epidermidis from infant bacteremia cases.
- Conjugation experiments to assess inter- and intra-species transfer of Gmr plasmids.
- Plasmid analysis using EcoRI restriction endonuclease digestion and DNA hybridization.
Main Results:
- All analyzed Staphylococcus aureus strains contained self-transmissible, conjugative Gmr plasmids (43.8-63 kb).
- Gmr plasmids were successfully transferred from S. aureus to S. aureus and S. epidermidis recipients.
- No gentamicin resistance transfer was observed from nine S. epidermidis donor strains.
- Conjugative plasmids frequently carried additional resistance genes (kanamycin, tobramycin, neomycin, ethidium bromide) and beta-lactamase synthesis genes.
- Three distinct EcoRI digestion patterns were identified among S. aureus Gmr plasmids, with common fragments suggesting conserved genetic elements.
- High homology was observed among S. aureus Gmr plasmids and with plasmids from other regions.
- A nonconjugative S. epidermidis plasmid showed homology but lacked a key fragment associated with conjugation.
Conclusions:
- Conjugative Gmr plasmids are prevalent in S. aureus causing infant bacteremia and can transfer resistance to other staphylococci.
- S. epidermidis strains in this cohort appear to be poor donors of gentamicin resistance via conjugation.
- The identified Gmr plasmids share conserved genetic features, suggesting a common origin or successful dissemination of specific resistance elements.