Limited role of antibody in clearance of Streptococcus pneumoniae in a murine model of colonization

Abstract

Colonization is the first step in the interaction between Streptococcus pneumoniae and its human host. To better understand the mechanisms contributing to natural carriage, a mouse model of pneumococcal colonization was developed with a clinical isolate of S. pneumoniae previously characterized in experimental colonization of humans. Similar to carriage events in humans, colonization of mice was self-limited and there was no evidence of lower respiratory tract or invasive disease. Carriage induced a serum antibody response to whole pneumococci that was associated temporally with clearance of colonization in three inbred strains of mice. Individual mice, however, did not demonstrate a correlation between the density of colonization and amounts of serum or of mucosal antibodies, including antibodies of different isotypes and antigenic specificities. The role of antibody in the clearance of carriage was then examined in mice with genetic defects in humoral immunity. xid mice, which have deficient responses to polysaccharide antigens, cleared colonization at the same rate as the parent strain. Finally, we showed that microMT mice, which lack mature B cells and fail to produce antibody, were unaffected in the density or duration of colonization. These results demonstrate that antibody is not required for clearance of pneumococcal colonization in mice.

FIG. 1.

Levels of pneumococcal colonization in comparison with serum antibody titer over time. The density of pneumococcal colonization, expressed as log CFU per milliliter of upper respiratory tract lavage fluid, was determined for BALB/c (A), C57BL/6 (B), and CBA/J (C) mice at the postinoculation day indicated. Whole bacteria (P1121) and strain-specific PspA total immunoglobulin titers were determined by ELISA. Values shown are the mean CFU per milliliter or titers ± standard errors of the means.

FIG. 2.

Level of colonization as a function of antibody titer in individual BALB/c mice. The density of pneumococcal colonization, expressed as log CFU per milliliter of upper respiratory tract lavage fluid, was determined for BALB/c mice at day 35 postinoculation. Total whole bacteria (P1121) and strain-specific PspA serum immunoglobulin titers (A), P1121-specific serum IgA, IgG, and IgM titers (B), P1121-specific immunoglobulin titers in upper respiratory tract lavage fluids (C), and P1121-specific IgA titers in NALT and anterior cervical lymph node culture supernatants (D) were determined by ELISA. Values shown are the mean titers from three independent determinations and log CFU per milliliter for seven individual BALB/c mice.

FIG. 3.

Comparison of the density of colonization in CBA/N (A) and μMT (B) mice over time with their respective immunocompetent parent strains. Values shown are the mean log CFU per milliliter ± standard errors of the means for groups of five mice each. The asterisks indicates a significant decrease in the density of colonization compared to the first time point at which colonization was determined (unpaired two-tailed t test, P ≤ 0.05). No significant differences in the densities of colonization were observed between CBA/N and CBA/J or μMT and C57BL/6 mice.

FIG. 4.

Effect of prior colonization on subsequent challenge with the same strain. CBA/N or CBA/J were initially inoculated with P1121 or PBS and subsequently challenged with P1121str^r or PBS 6 weeks later. Values shown are the mean log CFU per milliliter ± standard errors of the means for five mice per group. The asterisk indicates a P of ≤0.05 compared to second challenge with PBS (unpaired two-tailed t test).

FIG. 5.

Effect of immunization on the density of colonization. BALB/c mice were immunized with either strain-specific PspA or sham immunized with adjuvant alone. PspA-specific antibody titers were determined by ELISA (A). Values shown are mean total antibody titers ± standard errors of the means for upper respiratory tract lavage fluids. The asterisks indicates a P of ≤0.05 compared to control (unpaired two-tailed t test). The density of colonization (B) was determined by plating serial dilutions of upper respiratory tract lavage fluids for colony counting. Values shown are the mean log CFU per milliliter ± standard errors of the means.