Genome Modification in Enterococcus faecalis OG1RF Assessed by Bisulfite Sequencing and Single-Molecule Real-Time...

Area of Science:

• Microbiology
• Genetics
• Molecular Biology

Background:

• Enterococcus faecalis is a Gram-positive bacterium causing life-threatening infections.
• Multidrug-resistant (MDR) strains pose a significant public health threat due to limited treatment options.
• Mobile genetic elements (MGEs) contribute to antibiotic resistance and virulence in MDR E. faecalis.

Purpose of the Study:

• To investigate DNA modification systems (restriction-modification or R-M) in E. faecalis for self vs. nonself DNA discrimination.
• To determine the role of R-M systems in modulating the conjugative transfer of antibiotic resistance plasmids.
• To understand how genome defense mechanisms impact MGE acquisition and antibiotic resistance dissemination.

Main Methods:

• Genome resequencing of the oral E. faecalis isolate OG1RF.
• Single-molecule real-time (SMRT) and bisulfite sequencing to identify DNA modifications.
• Analysis of R-M system components and their impact on DNA methylation and gene transfer.

Main Results:

• OG1RF exhibits global 5-methylcytosine (m5C) methylation at 5'-GCWGC-3' motifs, conferred by a type II R-M system.
• Disruption of this R-M system affected OG1RF's electrotransformability and conjugative transfer of an antibiotic resistance plasmid.
• A second DNA methyltransferase conferred N(4)-methylcytosine (m4C) methylation at 5'-CCGG-3' motifs.

Conclusions:

• Restriction-modification systems act as a barrier against MGE acquisition in E. faecalis.
• Differential genome modification influences horizontal gene transfer frequencies.
• R-M systems likely play a crucial role in the dissemination of antibiotic resistance genes within the E. faecalis species.