Related Experiment Videos
Efficient transfer of the pheromone-independent Enterococcus faecium plasmid pMG1 (Gmr) (65.1 kilobases) to
1Department of Microbiology, Gunma University School of Medicine, Maebashi, Gunma, Japan. yasuike@sb.gunma-u.ac.jp
Abstract:
Plasmid pMG1 (65.1 kb) was isolated from a gentamicin-resistant Enterococcus faecium clinical isolate and was found to encode gentamicin resistance. EcoRI restriction of pMG1 produced five fragments, A through E, with molecular sizes of 50.2, 11.5, 2.0, 0.7, and 0.7 kb, respectively. The clockwise order of the fragments was ACDEB. pMG1 transferred at high frequency to Enterococcus strains in broth mating. pMG1 transferred between Enterococcus faecalis strains, between E. faecium strains, and between E. faecium and E. faecalis strains at a frequency of approximately 10(-4) per donor cell after 3 h of mating. The pMG1 transfers were not induced by the exposure of the donor cell to culture filtrates of plasmid-free E. faecalis FA2-2 or an E. faecium strain. Mating aggregates were not observed by the naked eye during broth mating. Small mating aggregates of several cells in the broth matings were observed by microscopy, while no aggregates of donor cells which had been exposed to a culture filtrate of E. faecalis FA2-2 or an E. faecium strain were observed, even by microscopy. pMG1 DNA did not show any homology in Southern hybridization with that of the pheromone-responsive plasmids and broad-host-range plasmids pAMbeta1 and pIP501. These results indicate that there is another efficient transfer system in the conjugative plasmids of Enterococcus and that this system is different from the pheromone-induced transfer system of E. faecalis plasmids.
Insights
A novel gentamicin resistance plasmid, pMG1, was identified in Enterococcus faecium. This plasmid efficiently transfers between Enterococcus species via a non-pheromone-induced conjugation system, distinct from known mechanisms.
Area of Science:
- Microbiology
- Molecular Biology
- Genetics
Background:
- Gentamicin resistance is a significant concern in clinical settings.
- Enterococcus species are common causes of hospital-acquired infections.
- Understanding plasmid-mediated resistance and transfer mechanisms is crucial for infection control.
Purpose of the Study:
- To characterize the gentamicin resistance plasmid pMG1 isolated from Enterococcus faecium.
- To investigate the transfer efficiency and mechanism of pMG1 between Enterococcus strains.
- To compare the transfer system of pMG1 with known plasmid conjugation systems.
Main Methods:
- Plasmid isolation and characterization (EcoRI restriction digestion, molecular size determination).
- Conjugation experiments (broth mating) to assess transfer frequency between Enterococcus strains.
- Southern hybridization to investigate DNA homology with known plasmids.
- Microscopy to observe mating aggregates.
Main Results:
- Plasmid pMG1 (65.1 kb) confers gentamicin resistance and was isolated from a clinical Enterococcus faecium isolate.
- pMG1 demonstrated high-frequency transfer (approx. 10(-4) per donor cell) to various Enterococcus species (E. faecalis, E. faecium) via broth mating.
- The transfer mechanism was not induced by pheromones and showed no homology to known pheromone-inducible or broad-host-range plasmids, indicating a novel conjugation system.
Conclusions:
- Plasmid pMG1 represents a novel conjugative plasmid in Enterococcus.
- It possesses an efficient, non-pheromone-induced transfer system distinct from previously described mechanisms.
- This finding highlights the diversity of mobile genetic elements contributing to antibiotic resistance spread in Enterococcus.