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Conjugal transfer of plasmid-borne multiple antibiotic resistance in Streptococcus faecalis var. zymogenes
Abstract:
A strain of Streptococcus faecalis var. zymogenes, designated JH1, had high-level resistance to the antibiotics streptomycin, kanamycin, neomycin, erythromycin, and tetracycline. These resistances were lost en bloc from approximately 0.1% of cells grown in nutrient broth at 45 C. The frequency of resistance loss was not increased by growth in the presence of the "curing" agents acriflavine or acridine orange, but after prolonged storage in nutrient agar 17% of cells became antibiotic sensitive. Covalently closed circular deoxyribonucleic acid (DNA) molecules were isolated from the parental strain and from antibiotic-sensitive segregants by using cesium chloride-ethidium bromide gradients. DNA molecular species were identified by using neutral sucrose gradients. Strain JH1 contained two covalently closed circular DNA species of molecular weights 50 x 10(6) and 38 x 10(6). An antibiotic-sensitive segregant, strain JH1-9, had lost the larger molecular species. A second sensitive segregant, strain JH1-5, had also lost the larger molecular species but a new molecular species of approximate molecular weight 6 x 10(6) was present. The antibiotic resistances that were curable from the parental strain were transferred to antibiotic-sensitive strains of S. faecalis and to strain JH1-9, during mixed incubation in nutrient broth at 37 C. Data to be described are interpreted to suggest that the transfer is by a conjugal mechanism. Analysis of the plasmid species in recipient clones showed that all had received the plasmid of molecular weight 50 x 10(6). Strain JH1-5 was not a good recipient. Analysis of one successful recipient clone of JH1-5 revealed that it had gained the 50 x 10(6) molecular weight plasmid but lost the 6 x 10(6) molecular weight species. These data are interpreted to mean that the multiple antibiotic resistance is borne by a transferable plasmid of 50 x 10(6) molecular weight, and that in clone JH1-5 this plasmid suffered a large deletion leaving only a 6 x 10(6) remnant which was incompatible with the complete replicon.
Insights
Multiple antibiotic resistances in Streptococcus faecalis were lost and transferred via a conjugal mechanism. These resistances are linked to a 50 x 10^6 molecular weight plasmid, which can be deleted or lost.
Area of Science:
- Microbiology
- Molecular Biology
- Genetics
Background:
- Streptococcus faecalis var. zymogenes strain JH1 exhibits high-level resistance to multiple antibiotics.
- Antibiotic resistance can be lost spontaneously or under specific conditions.
Purpose of the Study:
- To investigate the genetic basis of multiple antibiotic resistance in S. faecalis.
- To determine the mechanism of antibiotic resistance loss and transfer.
Main Methods:
- Isolation and characterization of covalently closed circular deoxyribonucleic acid (DNA) using cesium chloride-ethidium bromide and neutral sucrose gradients.
- Induction of antibiotic resistance loss through heat treatment and prolonged storage.
- Conjugation experiments to transfer antibiotic resistance to sensitive strains.
Main Results:
- Strain JH1 contains two circular DNA species (50 x 10^6 and 38 x 10^6 molecular weight).
- Antibiotic-sensitive segregants (JH1-9, JH1-5) lost the larger (50 x 10^6) DNA species, with JH1-5 retaining a smaller remnant (6 x 10^6).
- Antibiotic resistance was transferable via conjugation, with recipients acquiring the 50 x 10^6 molecular weight plasmid.
Conclusions:
- Multiple antibiotic resistance in strain JH1 is mediated by a transferable plasmid of 50 x 10^6 molecular weight.
- The 6 x 10^6 molecular weight species in JH1-5 represents a deleted remnant of the resistance plasmid, incompatible with the complete replicon.
- Conjugal transfer is the mechanism responsible for the spread of this multi-drug resistance plasmid.