Enterococcal Polysaccharide Antigen (EPA) rhamnan backbone contributes to cell wall architecture and is essential for antimicrobial resistance, innate immune evasion and phage infection
- 1The University of Sheffield School of Biosciences
Abstract
Enterococci are opportunistic pathogens classified by the World Health Organization as high-priority microorganisms. They cause a broad spectrum of infections, and their intrinsic and acquired resistance to antimicrobials makes these infections particularly difficult to treat and eradicate. In Enterococcus faecalis, the most frequently isolated enterococcal pathogen in humans, antimicrobial resistance and innate immune evasion are largely driven by the Enterococcal Polysaccharide Antigen (EPA). This surface polymer underpins key virulence traits, including resistance to host defence mechanisms, reduced susceptibility to multiple classes of antimicrobials, and susceptibility to bacteriophage infection. EPA consists of a rhamnan backbone decorated with strain-specific substituents that are essential for its biological activity. Here, we show that epaB encodes the enzyme responsible for the first committed step in assembling the EPA rhamnan chain. Using NMR spectroscopy, we demonstrate that E. faecalis lacking epaB produces an EPA polymer composed solely of decorations directly anchored to the peptidoglycan, with no detectable rhamnan backbone. The absence of this rhamnan moiety profoundly alters cell wall architecture, as revealed by atomic force microscopy of the mutant cell walls. The epaB mutation also abolishes innate immune evasion and virulence in the zebrafish infection model, while conferring resistance to bacteriophages. Collectively, these findings demonstrate that both the rhamnan backbone and its decorations are required for EPAs full biological activity, establishing the structural and functional interdependence of these two components.
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BibTeX
@misc{mesnage2026enterococcal,
title = {{Enterococcal Polysaccharide Antigen (EPA) rhamnan backbone contributes to cell wall architecture and is essential for antimicrobial resistance, innate immune evasion and phage infection}},
author = {S. Mesnage and l. Kupcova and N. Nathoo and B. J. Michno and K. S. Chellappa and T. Lawson and M. McNeil and J. L. Davis and P. Manivannan and J. S. Norwood and R. E. Smith and E. Maes and l. Pasquina-Lemonche and T. K. Prajsnar and M. L. Rowe and H. C. Dorfmueller and G. P. Stafford and M. P. Williamson},
howpublished = {bioRxiv},
year = {2026},
month = aug,
doi = {10.64898/2026.08.24.746643},
url = {https://www.biorxiv.org/content/10.64898/2026.08.24.746643v1},
note = {CC BY-NC-ND},
}
RIS
TY - UNPB
TI - Enterococcal Polysaccharide Antigen (EPA) rhamnan backbone contributes to cell wall architecture and is essential for antimicrobial resistance, innate immune evasion and phage infection
AU - S. Mesnage
AU - l. Kupcova
AU - N. Nathoo
AU - B. J. Michno
AU - K. S. Chellappa
AU - T. Lawson
AU - M. McNeil
AU - J. L. Davis
AU - P. Manivannan
AU - J. S. Norwood
AU - R. E. Smith
AU - E. Maes
AU - l. Pasquina-Lemonche
AU - T. K. Prajsnar
AU - M. L. Rowe
AU - H. C. Dorfmueller
AU - G. P. Stafford
AU - M. P. Williamson
PY - 2026
DA - 2026/08/24
PB - bioRxiv
DO - 10.64898/2026.08.24.746643
UR - https://www.biorxiv.org/content/10.64898/2026.08.24.746643v1
AB - Enterococci are opportunistic pathogens classified by the World Health Organization as high-priority microorganisms. They cause a broad spectrum of infections, and their intrinsic and acquired resistance to antimicrobials makes these infections particularly difficult to treat and eradicate. In Enterococcus faecalis, the most frequently isolated enterococcal pathogen in humans, antimicrobial resistance and innate immune evasion are largely driven by the Enterococcal Polysaccharide Antigen (EPA). This surface polymer underpins key virulence traits, including resistance to host defence mechanisms, reduced susceptibility to multiple classes of antimicrobials, and susceptibility to bacteriophage infection. EPA consists of a rhamnan backbone decorated with strain-specific substituents that are essential for its biological activity. Here, we show that epaB encodes the enzyme responsible for the first committed step in assembling the EPA rhamnan chain. Using NMR spectroscopy, we demonstrate that E. faecalis lacking epaB produces an EPA polymer composed solely of decorations directly anchored to the peptidoglycan, with no detectable rhamnan backbone. The absence of this rhamnan moiety profoundly alters cell wall architecture, as revealed by atomic force microscopy of the mutant cell walls. The epaB mutation also abolishes innate immune evasion and virulence in the zebrafish infection model, while conferring resistance to bacteriophages. Collectively, these findings demonstrate that both the rhamnan backbone and its decorations are required for EPAs full biological activity, establishing the structural and functional interdependence of these two components.
ER -