Antigenic and structural variation of the lipopolysaccharide of a Legionella pneumophila serogroup 1 strain and the effect on virulence and serum sensitivity.

To investigate the role of lipopolysaccharide in Legionella pneumophila virulence, specific mutants were isolated using an in situ enzyme immunoassay for screening. L. pneumophila has smooth lipopolysaccharide and is resistant to complement-mediated killing when exposed to normal human serum. The reactivities of three monoclonal antibodies (mAbs) were tested against the progeny of a single Legionella pneumophila serogroup 1 Los Angeles strain. The mAbs used were MAB2 and 4E9 (which define subgroups 1 and 2 respectively), and 1C5, which is a serogroup 1-specific mAb. All three mAbs bound to the lipopolysaccharide molecule isolated from reactive strains in immunoblots. The parent strain was 1C5$\sp+$, 4E9$\sp-$, MAB2$\sp+$ but spontaneously mutates to 1C5$\sp-$ or 4E9$\sp+$ at a frequency = 10$\sp{-4}$-10$\sp{-3}$. Three of four 4E9$\sp+$ mutants were also MAB2$\sp-$, but MAB2$\sp-$ strains could not be isolated directly from the parental strain. The epitopes for these mAbs are in the core region, and when altered, either do not cause detectable changes in the structure of the molecule (1C5$\sp-$ strains) or appear to reduce the molecular weight of the molecule (4E9$\sp+$ strains) when examined by SDS-PAGE. All mutants were stable when screened for revertants. When tested in a U937 cell model of infectivity, neither 1C5$\sp-$ or 4E9$\sp+$ mutant strains showed a substantial reduction in virulence. The MAB2$\sp-$ strains became serum-sensitive but when serum sensitivity was complemented to resistance in trans with a cosmid library, the mutant remained MAB2$\sp-$. By Southern hybridizations, it appeared that the mutation in the serum-sensitive, MAB2$\sp-$ strain was as a result of either gene conversion or deletion. The results of this study indicate that (1) mAb reactivity to lipopolysaccharide of L. pneumophila can change spontaneously at a high frequency and the mutations are stable, (2) loss of binding of these mAbs resulted in structural alterations that originate in the core, (3) the virulence of these mutants is not altered in macrophages in vitro, but certain mutants do become serum-sensitive, and (4) that serum-resistance in L. pneumophila is not determined by the MAB2 epitope, but the two phenotypes may be linked genetically.PhDMicrobiology and ImmunologyUniversity of Michigan, Horace H. Rackham School of Graduate Studieshttp://deepblue.lib.umich.edu/bitstream/2027.42/103835/1/9409792.pdfDescription of 9409792.pdf : Restricted to UM users only