The Vsa Shield of Mycoplasma pulmonis Is Antiphagocytic

Abstract

The infection of mice with Mycoplasma pulmonis is a model for studying chronic mycoplasmal respiratory disease. Many in vivo and in vitro studies have used the organism to gain a better understanding of host-pathogen interactions in chronic respiratory infection. The organism's Vsa proteins contain an extensive tandem repeat region. The length of the tandem repeat unit varies from as few as 11 amino acids to as many as 19. The number of tandem repeats can be as high as 60. The number of repeats varies at a high frequency due to slipped-strand mispairing events that occur during DNA replication. When the number of repeats is high, e.g., 40, the mycoplasma is resistant to lysis by complement but does not form a robust biofilm. When the number of repeats is low, e.g., 5, the mycoplasma is killed by complement when the cells are dispersed but has the capacity to form a biofilm that resists complement. Here, we examine the role of the Vsa proteins in the avoidance of phagocytosis and find that cells producing a protein with many tandem repeats are relatively resistant to killing by macrophages. These results may be pertinent to understanding the functions of similar proteins that have extensive repeat regions in other microbes.

CpG underrepresentation and the bacterial CpG-specific DNA methyltransferase M.MpeI

“…Most but not all predicted CpG methyltransferases contain such dinucleotide repeat tracts and are therefore probably phase variable (SI Appendix, Table S5). We note that slipped strand mispairing at repetitive DNA is common in bacteria (16,17) and has been described in Mycoplasms in the context of variations of the repertoire of potentially immunogenic surface proteins (18).…”

EPS-I polysaccharide protectsMycoplasma pulmonisfrom phagocytosis

“…This antibiotic was chosen because it is effective at inhibiting the growth of M. pulmonis and because studies had shown that chloramphenicol at concentrations up to 200 μg mL −1 does not diminish the ability of macrophages to phagocytose and kill bacteria (van den Broek, ). In our previous study (Shaw et al ., ), chloramphenicol was added to the reactions at a final concentration of 30 μg mL −1 . This concentration of chloramphenicol inhibits the growth of mycoplasma strains CTG38 and CTG1701.…”

“…The exact location of Tn4001T in the genome of CTG1701 and details of the construction of the complemented strain have been described (Daubenspeck et al, 2009). All of the mycoplasma strains used in this study produce a VsaG protein of identical size, containing 36 tandem repeats, and are thought to differ phenotypically only in the production of polysaccharide (Shaw et al, 2012). Mycoplasmas were grown in mycoplasma broth, suspended in freezing medium, sonicated to break up aggregates and assayed for CFU on mycoplasma agar as described previously (Shaw et al, 2012).…”

“…MH-S cells were prepared as described previously (Shaw et al, 2012). The cells were activated with interferon gamma and lipopolysaccharide followed by three washes in assay buffer (Hank's balanced salt solution buffered with 25 mM HEPES, pH 7.4, supplemented with 5% foetal bovine serum).…”

“…Size variation occurs in the tandem-repeat region of the protein and results from slipped-strand mispairing during DNA replication. Vsa size variation has a critical role in cytoadherence, biofilm formation and the avoidance of killing from complement and alveolar macrophages (Simmons & Dybvig, 2003, 2007Simmons et al, 2004;Shaw et al, 2012). Not all species of mycoplasma produce proteins analogous to the Vsa proteins, and other types of antiphagocytic molecules should exist.…”