Molecular and Genomic Based Insights Into The Evolution of Enterococcus Faecium From Commensal to Hospital-Adapted Pathogen

<p>The basis for the recent transition of <em>Enterococcus faecium</em> from a primarily commensal organism to one of the leading causes of hospital-acquired infections in the United States is not yet understood. To address this, the first part of my project assessed isolates from early outbreaks in the USA and South America using sequence analysis, colony hybridizations, and minimal inhibitory concentrations (MICs) which showed clinical isolates possess virulence and antibiotic resistance determinants that are less abundant or lacking in community isolates. I also revealed that the level of ampicillin resistance increased over time in clinical strains. By sequencing the <em>pbp5</em> gene, I demonstrated an ~5% difference in the <em>pbp5 </em>gene between strains with MICs <4ug/ml and those with MICs >4µg/ml, but no specific sequence changes correlated with increases in MICs within the latter group. A 3-10% nucleotide difference was also seen in three other genes analyzed, which suggested the existence of two distinct subpopulations of <em>E. faecium</em>. This led to the second part of my project analyzing concatenated core gene sequences, SNPs, the 16S rRNA, and phylogenetics of 21 <em>E. faecium</em> genomes confirming two distinct clades; a community-associated (CA) clade and hospital-associated (HA) clade. Molecular clock calculations indicate that these two clades likely diverged ~ 300,000 to > 1 million years ago, long before the modern antibiotic era. Genomic analysis also showed that, in addition to core genomic differences, HA <em>E. faecium</em> harbor specific accessory genetic elements that may confer selection advantages over CA <em>E. faecium</em>. The third part of my project discovered 6 <em>E. faecium</em> genes with the newly identified “WxL” domain. My analyses, using RT-PCR, western blots, patient sera, whole-cell ELISA, and immunogold electron microscopy, indicated that <em>E. faecium</em> WxL genes exist in operons, encode bacterial cell surface localized proteins, that WxL proteins are antigenic in humans, and are more exposed on the surface of clinical isolates versus community isolates (even though they are ubiquitous in both clades). ELISAs and BIAcore analyses also showed that proteins encoded by these operons bind several different host extracellular matrix proteins, as well as to each other, suggesting a novel cell-surface complex. In summary, my studies provide new insights into the evolution of <em>E. faecium</em> by showing that there are two distantly related clades; one being more successful in the hospital setting. My studies also identified operons encoding WxL proteins whose characteristics could also contribute to colonization and virulence within this species.</p>

Name thesis:degree_name

Doctor of Philosophy (PhD)

Level thesis:degree_level

Dissertation (PhD)

Year dc:date.available

2013

Author dc:creator

• Galloway-Pena, Jessica R

Contributors dc:contributor

• Barbara E. Murray, M.D.
• Steven J. Norris, Ph.D.
• Danielle Garsin, Ph.D.

• E. faecium
• MLST
• pbp5
• WxL
• Bacteriology
• Genomics
• Medicine and Health Sciences
• Molecular Genetics
• Pathogenic Microbiology

Repository record dc:identifier

https://digitalcommons.library.tmc.edu/utgsbs_dissertations/334

OAI identifier oai:identifier

oai:digitalcommons.library.tmc.edu:utgsbs_dissertations-1370