Isolation, structural characterization, and immunological evaluation of a high-molecular-weight exopolysaccharide from Staphylococcus aureus.
Colonization of implanted medical devices by coagulase-negative staphylococci such as Staphylococcus epidermidis is mediated by the bacterial polysaccharide intercellular adhesin (PIA), a polymer of beta-(1-->6)-linked glucosamine substituted with N-acetyl and O-succinyl constituents. The icaADBC locus containing the biosynthetic genes for production of PIA has been identified in both S. epidermidis and S. aureus. Whereas it is clear that PIA is a constituent that contributes to the virulence of S. epidermidis, it is less clear what role PIA plays in infection with S. aureus. Recently, identification of a novel polysaccharide antigen from S. aureus termed poly N-succinyl beta-(1-->6)-glucosamine (PNSG) has been reported. This polymer was composed of the same glycan backbone as PIA but was reported to contain a high proportion of N-succinylation rather than acetylation. We have isolated a glucosamine-containing exopolysaccharide from the constitutive over-producing MN8m strain of S. aureus in order to prepare polysaccharide-protein conjugate vaccines. In this report we demonstrate that MN8m produced a high-molecular-weight (>300,000 Da) polymer of beta-(1-->6)-linked glucosamine containing 45-60% N-acetyl, and a small amount of O-succinyl (approx 10% mole ratio to monosaccharide units). By detailed NMR analyses of polysaccharide preparations, we show that the previous identification of N-succinyl was an analytical artifact. The exopolysaccharide we have isolated is active in in vitro hemagglutination assays and is immunogenic in mice when coupled to a protein carrier. We therefore conclude that S. aureus strain MN8m produces a polymer that is chemically and biologically closely related to the PIA produced by S. epidermidis.
Joyce
Joseph G
JG
Departments of Virus and Cell Biology, Merck Research Laboratories, WP16-107, P.O. Box 4, West Point, PA 19486, USA. joseph_joyce@merck.com
Abeygunawardana
Chitrananda
C
Xu
Qiuwei
Q
Cook
James C
JC
Hepler
Robert
R
Przysiecki
Craig T
CT
Grimm
Karen M
KM
Roper
Keith
K
Ip
Charlotte C Yu
CC
Cope
Leslie
L
Montgomery
Donna
D
Chang
Mason
M
Campie
Sherilyn
S
Brown
Martha
M
McNeely
Tessie B
TB
Zorman
Julie
J
Maira-Litrán
Tomas
T
Pier
Gerald B
GB
Keller
Paul M
PM
Jansen
Kathrin U
KU
Mark
George E
GE
D000818:Animals; D002236:Carbohydrate Conformation; D002850:Chromatography, Gel; D004797:Enzyme-Linked Immunosorbent Assay; D006386:Hemagglutination Tests; D007982:Levulinic Acids / Q000032:analysis / Q000737:chemistry; D009682:Magnetic Resonance Spectroscopy; D051379:Mice; D008970:Molecular Weight; D011135:Polysaccharides, Bacterial / Q000737:chemistry / Q000276:immunology* / Q000302:isolation & purification*; D013211:Staphylococcus aureus / Q000737:chemistry*
D007982:Levulinic Acids; D011135:Polysaccharides, Bacterial; C032246:levulinic acid