Protection by natural human immunoglobulin M antibody to meningococcal serogroup B capsular polysaccharide in the infant rat protection assay is independent of complement-mediated bacterial lysis

Abstract

Neisseria meningitidis, an important cause of bacterial meningitis and septicemia worldwide, is associated with high mortality and serious sequelae. Natural immunity against meningococcal disease develops with age, but the specificity and functional activity of natural antibodies associated with protection are poorly understood. We addressed this question by using a selected subset of prevaccination sera (n = 26) with convergent or discrepant serum bactericidal activity (SBA) and infant rat protective activity (IRPA) against the serogroup B meningococcal strain 44/76-SL (B:15:P1.7,16) from Icelandic teenagers. The sera were analyzed by opsonophagocytic activity (OPA) assay, immunoblotting, immunoglobulin G (IgG) quantitation against live meningococcal cells by flow cytometry, and enzyme immunosorbent assay (EIA). High levels of SBA and OPA were reflected in distinct IgG binding to major outer membrane proteins and/or lipopolysaccharide in immunoblots. However, we could not detect any specific antibody patterns on blots that could explain IRPA. Only IgM antibody to group B capsular polysaccharide (B-PS), measured by EIA, correlated positively (r = 0.76, P < 0.001) with IRPA. Normal human sera (NHS; n = 20) from healthy Finnish children of different ages (7, 14, and 24 months and 10 years) supported this finding and showed an age-related increase in IRPA that coincided with the acquisition of B-PS specific IgM antibody. The protection was independent of complement-mediated bacterial lysis, as detected by the inability of NHS to augment SBA in the presence of human or infant rat complement and the equal protective activity of NHS in rat strains with fully functional or C6-deficient complement.

FIG. 1.

Summary “box-and-whiskers” plots based on the median, quartile, and extreme antibody values for the prevaccination sera from Icelandic teenagers evaluated for functionality (IRPA [a], SBA [b], OPA [c]) and quantity of meningococcal specific antibodies ([d to f]). On basis of the convergence or discrepancy of the IRPA and SBA data with the 44/76-SL (B:15:P1.7,16) strain, sera were divided into four different categories: category I, IRPA-positive, SBA-positive sera; category II, IRPA-negative, SBA-positive sera; category III, IRPA-positive, SBA-negative sera; and category IV, IRPA-negative, SBA-negative sera. The designated cutoff value (PI = 10) for IRPA is indicated by the horizontal dotted line.

FIG. 2.

Correlations between IRPA and functional (SBA [a] and OPA [b]) or quantitative antibody (c to f) assays for the prevaccination sera from Icelandic teenagers (n = 26). Strain 44/76-SL (B:15:P1.7,16) was used in IRPA, SBA, OPA, and preparation of the OMVs, and in measurement of IgG binding to live cells. (a) SBA titers versus protection indices (PIs); (b) OPA titers versus PIs; (c) anti-OMV IgG concentrations versus PIs; (d) IgG binding to live 44/76-SL cells versus PIs; (e) anti-B-PS IgG concentrations versus PIs; (f) anti-B-PS IgM concentrations versus PIs.

FIG. 3.

Age-related increase in IRPA against strain 44/76-SL (B:15:P1.7,16). Sera collected from Finnish children of the four indicated ages were used (n = 5/group). Horizontal lines, GMs of PIs at different ages; horizontal dotted line, designated cutoff value (PI = 10) for IRPA. Datum points are separated by 10% (i.e., jittered) to show individual datum points.

FIG. 4.

Correlation between anti-B-PS IgM antibody concentrations and PIs for sera from Finnish children (n = 20) of four different age groups (○, 7 months; ▵, 14 months; •, 24 months; ▴, 10 years). The horizontal dotted line indicates the cutoff value (PI = 10) for IRPA.

FIG. 5.

Influence of B-PS antibody depletion on passive protection in complement component C6-deficient rats. A serum from a 10-year-old Finnish child was absorbed with Al-B-PS or Al-C-PS. The results are shown as the GM CFU values per ml of blood ± the 95% CI from n = 5 rats. ND, not determined.

FIG. 6.

Correlations between IRPA and functional (SBA) (a) or quantitative antibody (b to d) assays for prevaccination sera from Icelandic teenagers (n = 26). Strain Cu385 (B:4:P1.19,15) was used in IRPA and SBA analyses and for preparing the OMVs. (a) SBA titers versus PIs; (b) anti-OMV IgG concentrations versus PIs; (c) anti-B-PS IgG concentrations versus PIs; (d) anti-B-PS IgM concentrations versus PIs.