Characterisation of a phase-variable restriction-modification system in Enterococcus faecalis
The gastrointestinal colonizer Enterococcus faecalis is an opportunistic, multi-drug resistant pathogen and a leading cause of nosocomial infections. Vancomycin- resistant enterococci (VRE) are of particular concern and have been selected for by agricultural use of antibiotics. A single E. faecalis VRE clone was shown to historically dominate and persist in poultry and disseminate to clinical reservoirs in NZ. Recent sequencing of this dominant clonal strain has revealed the presence of a phase- variable type I restriction-modification (RM) system. RM systems are genome defence mechanisms that differentiate between self and non-self DNA through methylation of the genome. The system’s enzymes are directed to specific DNA motifs by the sequence specificity subunits (HsdS) of the system. The presence of two hsdS genes in this E. faecalis RM system that share sequence homology indicates potential for recombination evenrts to generate multiple hsdS alleles able to redirect the systems enzymes. This has the potential to confer differential methylation of the genome which may 1) alter intrinsic gene expression and 2) confer great genome defence. This project aimed to identify the prevalence of RM in E. faecalis, analyse the phase- variability of the previously identified RM system and to understand the functional role this RM systems in the persistence of the clonal E. faecalis strain. To carry this out a combination of bioinformatic analyses, deep sequencing and conformational laboratory techniques were used. RM systems were fairly variable in E. faecalis with least conservation seen in the hsdS subunits. Recombination events between hsdS genes in this strain was confirmed and shown to correlate with changes in gene expression. Although exposure to antibiotic stress did not appear to influence recombination between the hsdS subunits in these systems, it was observed to regulate the expression of the enzymes of a similar type I RM system in E. faecalis JH2- 2. Recombination of this system likely generates different physiological states of these strains as seen in Streptococcus pneumonaie that may have helped in its historical persistence and survival in multiple hosts by creating diversity within a clonal population. Additionally, regulation of this system in response to antibiotic stress may allow increased mobile genetic element (MGE) acquisition whilst still enabling the ability of the system to confer immune defence. Further testing is required, but this preliminary investigation indicates the system is functional and could confer benefits that may have contributed to persistence of this E. faecalis strain.
Advisor: Morgan, Xochitl; Cook, Greg
Degree Name: Bachelor of Biomedical Sciences with Honours
Degree Discipline: Microbiology and Immunology
Publisher: University of Otago
Keywords: E. faecalis; Enterococcus; Restriction Modification; Phase Variation; New Zealand; Persistance
Research Type: Thesis