Characterization of free exopolysaccharides secreted by Mycoplasma mycoides subsp. mycoides

Contagious bovine pleuropneumonia is a severe respiratory disease of cattle that is caused by a bacterium of the Mycoplasma genus, namely Mycoplasma mycoides subsp. mycoides (Mmm). In the absence of classical virulence determinants, the pathogenicity of Mmm is thought to rely on intrinsic metabolic...

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Published inPloS one Vol. 8; no. 7; p. e68373
Main Authors Bertin, Clothilde, Pau-Roblot, Corinne, Courtois, Josiane, Manso-Silván, Lucía, Thiaucourt, François, Tardy, Florence, Le Grand, Dominique, Poumarat, François, Gaurivaud, Patrice
Format Journal Article
LanguageEnglish
Published United States Public Library of Science 15.07.2013
Public Library of Science (PLoS)
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Summary:Contagious bovine pleuropneumonia is a severe respiratory disease of cattle that is caused by a bacterium of the Mycoplasma genus, namely Mycoplasma mycoides subsp. mycoides (Mmm). In the absence of classical virulence determinants, the pathogenicity of Mmm is thought to rely on intrinsic metabolic functions and specific components of the outer cell surface. One of these latter, the capsular polysaccharide galactan has been notably demonstrated to play a role in Mmm persistence and dissemination. The free exopolysaccharides (EPS), also produced by Mmm and shown to circulate in the blood stream of infected cattle, have received little attention so far. Indeed, their characterization has been hindered by the presence of polysaccharide contaminants in the complex mycoplasma culture medium. In this study, we developed a method to produce large quantities of EPS by transfer of mycoplasma cells from their complex broth to a chemically defined medium and subsequent purification. NMR analyses revealed that the purified, free EPS had an identical β(1->6)-galactofuranosyl structure to that of capsular galactan. We then analyzed intraclonal Mmm variants that produce opaque/translucent colonies on agar. First, we demonstrated that colony opacity was related to the production of a capsule, as observed by electron microscopy. We then compared the EPS extracts and showed that the non-capsulated, translucent colony variants produced higher amounts of free EPS than the capsulated, opaque colony variants. This phenotypic variation was associated with an antigenic variation of a specific glucose phosphotransferase permease. Finally, we conducted in silico analyses of candidate polysaccharide biosynthetic pathways in order to decipher the potential link between glucose phosphotransferase permease activity and attachment/release of galactan. The co-existence of variants producing alternative forms of galactan (capsular versus free extracellular galactan) and associated with an antigenic switch constitutes a finely tuned mechanism that may be involved in virulence.
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Conceived and designed the experiments: PG CB. Analyzed the data: PG F. Tardy CPR JC. Project conception: FP F. Thiaucourt LMS. Electron microscopy: PG DLG. Monomer composition and NMR analysis: CPR JC. All other experiments: PG CB. Drafted the manuscript: F. Tardy PG CPR LMS. Critically reviewed the manuscript and approved the final version: CB CPR JC LMS F. Thiaucourt F. Tardy DLG FP PG.
Current address: Institut National de Recherche Agronomique, UMR1309 CMAEE, Montpellier, France
Competing Interests: The authors have declared that no competing interests exist.
Current address: Centre International de Recherche en Agronomie pour le Développement, UMR CMAEE, Montpellier, France
ISSN:1932-6203
1932-6203
DOI:10.1371/journal.pone.0068373