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The incidence of invasive group A streptococcal disease has increased in Norway since the 1980s. Analysis of 100 isolates recovered from 1988 to 2003 showed an increased genotypic diversity over time, while the prevalence of the strain that dominated in 1988, sequence type (ST)-28/emm-1, decreased. Necrotizing fasciitis was often associated with ST-15/emm-3.

There is currently no standardized serum bactericidal antibody (SBA) assay for evaluating immune responses to meningococcal outer membrane vesicle or protein vaccines. Four laboratories, Manchester Health Protection Agency (MC HPA), New Zealand Institute of Environmental Science and Research Limited (NZ ESR), Norwegian Institute of Public Health (NIPH), and Chiron Vaccines (Chiron), measured SBA titers in the same panel of human sera (n=76) from laboratory staff (n=21) vaccinated with MenBvac. Blood samples were collected prevaccination, prior to each of the three doses of MenBvac given at 6-week intervals, and 6 weeks following the third dose. Initial results showed a number of discrepancies in results between the four participating laboratories. The greatest effect on titers appeared to be due to differences among laboratories in the maintenance of the meningococcal serogroup B test strain, 44/76-SL. A repeat study was conducted using the same frozen isolate (meningococcal serogroup B test strain 44/76-SL), freshly distributed to all four laboratories. Using SBA titers from the tilt method for all samples, and using MC HPA as the comparator, the results were as follows for NZ ESR, NIPH, and Chiron, respectively, using log(10) titers: correlation coefficients (r) were 0.966, 0.967, and 0.936; intercepts were 0.08, 0.15, and 0.17; and slopes were 0.930, 0.851, and 0.891. In both prevaccination and postvaccination samples from 15 subjects assayed by all four laboratories, similar increases in SBA (fourfold or greater) were observed (for 11, 11, 9, and 9 subjects for MC HPA, NZ ESR, NIPH, and Chiron, respectively), and similar percentages of subjects with SBA titers of>or=4 p revaccination and 6 weeks following each dose were found. The SBA assay has been harmonized between the four different laboratories with good agreement on seroconversion rates, n-fold changes in titers, and percentages of subjects with SBA titers of >or=4.

Neisseria meningitidis bacteria of serogroup A are causing recurring meningitis epidemics on the African continent. An outer membrane vesicle (OMV) vaccine against serogroup A meningococci made from a subgroup III serogroup A meningococcal strain was previously shown to induce antibodies with serum bactericidal activity (SBA) in mice. We have here further investigated the properties of OMV vaccines made from five different subgroup III serogroup A meningococcal strains grown in a synthetic medium with low iron content. In addition to the major outer membrane proteins (PorA, PorB, RmpM, Opa and OpcA), small amounts of the NadA, TdfH, Omp85, FetA, FbpA and NspA outer membrane proteins, as well as lipooligosaccharides, were detected in the vaccines. The OMV vaccines were used to immunise mice. Anti-meningococcal IgG antibodies in the mouse sera were analysed by immunoblotting and by enzyme-linked immunosorbent assay against OMVs, and against live meningococcal cells in SBA and a flow-cytometric assay. The vaccines induced antibodies with high SBA and opsonophagocytic activity. The strongest IgG responses were directed against PorA. Significant SBA responses were also observed against a subgroup III strain, which did not express PorA, whereas no SBA was observed against a clone IV-1 serogroup A strain. An OMV vaccine from serogroup A meningococci may be an alternative to polysaccharide and conjugate polysaccharide vaccines for Africa.

For evaluation of serum bactericidal activity (SBA) as surrogate for the efficacy of outer membrane vesicle (OMV) vaccines against Neisseria meningitidis serogroup B disease, we have reanalyzed data from a randomized double blind placebo-controlled efficacy trial involving 172000 secondary school students (aged 13-14 years) in Norway (1988-1991). A cohort of the efficacy trial consisting of 880 individuals was selected for immunogenicity studies. An efficacy of 87% was calculated for a 10-month observation period. However, after an observation period of 29 months, the estimated efficacy against group B disease induced by vaccination was 57%. The immunogenicity study showed that the SBA geometric mean titer (GMT) for the vaccinees was 2.4 before vaccination and 19.0 six weeks after the second vaccine dose. One year after vaccination the GMT was reduced to 2.8. A separate three-dose study with 304 adolescents showed that with a third dose at 10 months after the second dose (i.e. when cases of disease started to appear) a strong booster response was induced. Ten months after the second dose the SBA was reduced to near pre-immunization level. Following the third dose the SBA geometric mean titer of 2.7 increased to 62.3. One year after the third dose, the GMT was markedly higher than 6 weeks after the second dose (12.6 versus 8.8). Thus, protection after vaccination corresponds with the level of SBA. In order to reach lasting protective levels of SBA in a population, three vaccine doses are probably required. Measurements of SBA are likely to be useful for evaluating various upcoming formulations and improvements of immunization regimens for OMV vaccines.

We have studied the ability of outer membrane vesicle (OMV) vaccines from Neisseria meningitidis serogroup B to induce vaccine-specific antibody and spleen cell proliferative responses in mice after being administered intranasally (i.n.) and/or subcutaneously (s.c.). A series of four weekly i.n. doses (25 microg) without adjuvant or a single s.c. dose (2.5 microg) with aluminum hydroxide was followed 2 months later by secondary i.n. or s.c. immunizations. After i.n. priming, both immunoglobulin G (IgG) antibody responses in serum, measured by enzyme-linked immunosorbent assay, and IgA antibodies in saliva and extracts of feces were significantly boosted by later i.n. immunizations. The IgG antibody responses in serum were also significantly augmented by secondary s.c. immunization after i.n. as well as s.c. priming. Sera from mice immunized i.n. reached the same level of bactericidal activity as after s.c. immunizations. The s.c. immunizations alone, however, had no effect on mucosal IgA antibody responses, but could prime for booster antibody responses in secretions to later i.n. immunizations. The i.n. immunizations also led to marked OMV-specific spleen cell proliferation in vitro. Both serum antibody responses and spleen cell proliferation were higher after i.n. priming and later s.c. immunizations than after s.c. immunizations alone. There was thus no evidence that i.n. priming had induced immunological tolerance within the B- or T-cell system. Our results indicate that a nonproliferating meningococcal OMV vaccine given i.n. can induce immunological memory and that it may be favorably combined with similar vaccines for injections.

The infant rat infection model previously developed to evaluate protective ability of passively administered murine antibodies to group B meningococcal (MenB) surface antigens was adapted for human sera. Several challenge doses were tested, aiming at sensitive detection of protection with little interassay variability. Doses of 10(5) and 10(6) colony forming units of strain IH5341 (MenB:15:P1.7,16) injected intraperitoneally gave consistently high levels of bacteremia and meningitis developed in 6 h in 50-100% of the pups. A monoclonal antibody mAb735 to the MenB capsule, injected 1-2 h before bacterial challenge, gave full protection at a dose of 2 microg/pup. Sera from adult volunteers immunized with a MenB outer membrane vesicle vaccine reproducibly reduced bacterial counts in the blood and cerebrospinal fluid, whereas a normal human serum, lacking bactericidal and opsonophagocidal activity, was unprotective.

Antibodies against the class 4 outer membrane protein (OMP) from Neisseria meningitidis have been purified from sera from vaccinees immunized with the Norwegian meningococcal group B outer membrane vesicle vaccine. The human sera and purified antibodies reacted strongly with the class 4 OMP in immunoblots, whereas experiments with whole bacteria showed only weak reactions, indicating that the antibodies mainly reacted with parts of the class 4 molecule that were not exposed. The purified human anti-class 4 OMP antibodies and the monoclonal antibodies (MAbs) were neither bactericidal nor opsonic against live meningococci. Three new MAbs against the class 4 OMP were generated and compared with other, previously described MAbs. Three linear epitopes in different regions of the class 4 OMP were identified by the reaction of MAbs with synthetic peptides. The MAbs showed no blocking effect on bactericidal activity of MAbs against other OMPs. However, one of the eight purified human anti-class 4 OMP antibody preparations, selected from immunoblot reactions among sera from 27 vaccinees, inhibited at high concentrations the bactericidal effect of a MAb against the class 1 OMP. However, these antibodies were not vaccine induced, as they were present also before vaccination. Therefore, this study gave no evidence that vaccination with a meningococcal outer membrane vesicle vaccine containing the class 4 OMP induces blocking antibodies. Our data indicated that the structure of class 4 OMP does not correspond to standard beta-barrel structures of integral OMPs and that no substantial portion of the OmpA-like C-terminal region of this protein is located at the surface of the outer membrane.

Ten adult volunteers, with low prevaccination levels of serum IgG antibodies against meningococcal antigens (< 1 microg ml(-1)), received three doses of the Norwegian group B meningococcal outer membrane vesicle (OMV) vaccine intramuscularly at weeks 0, 6 and 46. Anti-OMV IgG subclass responses were measured and compared with serum bactericidal activity (SBA) and opsonic activity against the vaccine strain 44/76. All vaccinees showed an IgG1 antibody response after each vaccine dose. The vaccine-induced median serum IgG1 antibody levels were 16, 17 and 18 microg ml(-1) 2-6 weeks after the first, second and third dose, respectively. Three vaccinees showed a weak IgG3 response after the first dose, whereas 8 and 9 showed a response after the second (median = 10 microg ml(-1)) and third dose (median = 10 microg ml(-1)), respectively. Low levels of anti-OMV IgG2 antibodies were found, whilst specific IgG4 antibodies were only detected for one vaccinee. The vaccine induced at least a fourfold increase in SBA titre in 8 vaccinees after the first dose, in 9 vaccinees after 2 doses and in all vaccinees after 3 doses. A positive correlation was found between IgG1 subclass antibody levels and SBA (r = 0.62, P < 0.0001). Elevated opsonophagocytic activity, measured as respiratory burst (RB), was observed in all vaccinees after one vaccine dose and usually increased after 2 and 3 doses. A strong positive correlation was found between IgG1 antibody levels and RB (r = 0.76, P < 0.0001). In conclusion, we have shown that systemic meningococcal OMV vaccination in adult vaccinees mainly induced IgG1 antibodies which correlated with bactericidal and opsonic activity, but also a considerable amount of IgG3 antibodies, which, in contrast to the IgG1 response, was induced only after 2 or 3 vaccine doses and declined more rapidly.

Sera from vaccinees and controls who contracted serogroup B meningococcal disease during the blinded and open parts of a two-dose protection trial in Norway were compared for antigen-specific and bactericidal antibodies against vaccine strain 44/76 (B:15:P1.7,16). From 16 of 20 (80%) vaccinees and 26 of 35 (74%) controls, one or more serum samples (n = 104) were collected during the acute phase (1 to 4 days), early convalescent phase (5 to 79 days), and late convalescent phase (8 to 31 months) after onset of disease. Binding of immunoglobulin G (IgG) to the major outer membrane antigens (80- and 70-kDa proteins, class 1, 3, and 5 proteins, and lipopolysaccharide [LPS]) on immunoblots was measured by digital image analysis. Specific IgG levels in vaccinees increased from acute to early convalescent phases, followed by a decline, while controls showed a small increase over time. Vaccinees had significantly higher levels than controls against class 1 and 3 porins and LPS in acute sera, against all antigens during early convalescence, and against class 1 and 3 porins in the later sera. Vaccinees who were infected with strains expressing subtype P1.7,16 proteins demonstrated a level of IgG binding to protein P1.7,16 with early-convalescent-phase sera that was fourfold higher than that of those infected with other strains. Bactericidal titers in serum against the vaccine strain were 192-fold higher for vaccinees than those for controls during early convalescence, but similarly low levels were found during late convalescence. A vaccine-induced anamnestic response of specific and functional antibody activities was thus shown, but the decrease in protection over time after vaccination indicated that two vaccine doses did not induce sufficient levels of long-term protective antibodies.

To study how the different immunoglobulin (Ig)G subclass antibodies may confer protection against systemic meningococcal disease, we isolated IgG1, IgG2 and IgG3 antibodies from plasma from vaccinees immunized with the Norwegian meningococcal outer membrane vesicle vaccine. Four IgG1, one IgG2 and four IgG3 preparations were purified. The IgG2 and IgG3 subclass preparations were free from contaminating subclasses, whereas the IgG1 preparations contained from 0 to 14% of IgG2 and/or IgG3. Immunoblotting against whole-cell meningococcal antigens showed broad specificities of the various preparations, both within and between subclasses. These subclass preparations were tested for opsonophagocytic and bactericidal activity. As targets we used two different variants of the meningococcal vaccine strain, with low (44/76-SL) and high (44/76-1) expression of the outer membrane protein Opc. Using polymorphonuclear leucocytes as effector cells in the presence of human complement, all three IgG subclass preparations revealed high, and similar, opsonophagocytic activities against 44/76-SL, whereas against 44/76-1 the IgG2 preparation showed a reduced activity and most IgG3 preparations were slightly more active than the IgG1 preparations. Regarding bactericidal activity, all the three subclasses were highly active against 44/76-SL. Against 44/76-1 the bactericidal activities were somewhat more varied: all IgG1 and three IgG3 preparations exhibited higher activities than against 44/76-SL. Due to the low concentration in the IgG2 preparations, only a weak activity was seen against 44/76-1. One IgG3 preparation that was highly opsonophagocytic revealed no bactericidal activity against either of the two bacterial variants examined. In conclusion, we have shown that the IgG subclass effector functions differ from person to person, but that antibodies of IgG1, IgG2 and IgG3 subclasses, judged by their behaviour in the functional tests, may all contribute to protection against meningococcal disease.

Outer membrane vesicle (OMV) vaccines were made from Neisseria meningitidis strain 44/76 and its two short-chain lipopolysaccharide (LPS) mutants, Mu-1 and Mu-4. Only the 44/76 vaccine contained LPS with the host antigen lacto-N-neotetraose. The protein composition of the vaccines was similar. The LPS carbohydrate chain length proved to influence drastic changes in the LPS immunogenicity as well as the outer membrane proteins (OMPs) ability to elicit functional antibodies in mice. Only LPS in the Mu-1 and Mu-4 vaccines were immunogenic, and the 44/76 vaccine differed also by not inducing antibodies to the class 4 OMP. The Mu-1 vaccine, with a LPS carbohydrate chain comprising only two residues of 2-keto-3-deoxy-octonic acid, induced lower bactericidal activity and less antibodies to the class 1 OMP, compared to the two other vaccines. This indicates that LPS of a certain carbohydrate chain length is required for adequate exposure of the class 1 OMP epitopes essential for inducing bactericidal antibodies.

An ELISA method was developed to quantitate gravimetrically (microgram/ml) the IgG subclass response against a Norwegian vaccine composed of outer membrane vesicles (OMV) isolated from a Neisseria meningitidis B:15:P1.7,16 epidemic strain. Chimeric mouse-human anti-hapten NIP (5-iodo-4-hydroxy-3-nitrophenacetyl) antibodies of each subclass were used for calibration purposes. Before vaccination, low amounts of IgG1 and IgG2 antibodies against OMV were detectable in all vaccinees, whereas IgG3 was only detectable in one of the 21 vaccinees. After vaccination, IgG1 antibodies dominated the response followed by IgG3 and low to moderate levels of IgG2 antibodies. IgG4 was only detectable at very low levels in a few vaccinees. All sera showed close to parallel dose-response curves to each other for IgG1 and IgG3, whereas the IgG2 curves were not parallel to chimeric IgG2 and could thus not be quantitated gravimetrically. For IgG3, 1/3 of the vaccinee sera showed non-parallel dose-response curves to the rest of the vaccinee sera and to chimeric IgG3 and could not be gravimetrically quantitated. The rest of the sera showed parallel dose-response curves with the chimeric IgG3 and gravimetric quantitation was possible. This study illustrates that chimeric antibodies can be used as calibrators to quantitate IgG subclass antibody responses against OMV in gravimetric units and that the vaccine mainly induces IgG1 and IgG3 antibodies in humans.

Neisserial porins, the major protein constituents of the outer membrane capable of inducing antibody responses in humans, are considered to be meningococcal vaccine candidates, so it is important to map the relevant B-cell epitopes. For B-cell epitope analyses of the serotype 15 PorB protein in Neisseria meningitidis, paired sera from selected patients with systemic meningococcal disease (SMD) were screened with synthetic 12mer peptides spanning the PorB protein sequence, and/or its variable region 1 (VR1). A 'SMD-related' linear B-cell epitope was found within the VR1 region consisting of 14 residues (17svFHQNGQVTEvtt30). A 23mer soluble peptide (D63b2) that covered the VR1 region, including the complete 17svFHQNGQVTEvtt30 sequence, was recognized, whereas no detectable binding was observed to a 16mer peptide (D63a1) containing most of the essential sequence (19FHQNGQVTEvtt30). A low frequency of IgG responses specific for the PorB linear epitopes was found in convalescent-phase sera from 132 SMD patients studied, as judged from both immunoblotting studies (24/132; 18.2%) and reactivity with peptide D63b2 (18/132; 13.6%). Peptide D63b2 significantly inhibited IgG binding to the denatured PorB protein on immunoblots, suggesting that this B-cell epitope was one of the main linear epitopes on the PorB protein recognized by sera from some SMD patients.

The antibody kinetics in sera from 27 adults after three doses of the Norwegian group B meningococcal outer membrane vesicle (OMV) vaccine was studied. The vaccinees received the third dose 4 to 5 years after the first two. Antibody responses against outer membrane proteins (OMPs) and lipopolysaccharides were studied by enzyme-linked immunosorbent assay and immunoblotting and with serum bactericidal assays (SBA) with three variants of the vaccine strain, 44/76. Six weeks after the second injection, the geometric mean (GM) of the levels of immunoglobulin G (IgG) against OMVs was about sevenfold higher than that of prevaccination levels, and 74% of the vaccinees developed a greater-than-twofold rise in SBA titer. After 6 months, the GM of IgG levels declined to about threefold higher, and after 4 to 5 years it declined to about twofold higher, than that before vaccination. The third dose induced a rapid increase in SBA titers in 96% of the vaccinees, and the GM of levels of IgG against OMVs rose to about 14-fold the prevaccination level. One year later, the IgG antibody levels had dropped to 4.6-fold the prevaccination level, but 88% of the vaccinees still showed bactericidal activity. The response after the two first doses was higher in individuals with prevaccination antibodies, but no such effect was found after three doses. The use of defined mutants in SBA and linear multiple regression analyses indicated that among the major OMPs, antibodies to the Opc and class 1 proteins made the most important single contributions to the bactericidal activity against the vaccine strain, but it also demonstrated the importance of antibodies against other antigens. After three doses, 68% of the vaccinees showed a significant SBA response against a strain lacking both the Opc and the class 1 proteins. Three doses converted almost all subjects to SBA responders and gave higher antibody levels and relatively less serosubtype-specific bactericidal activity than did two doses, probably indicating a broader cross-protection against heterologous strains.

Based on differences in reaction pattern with monoclonal antibodies against the P1.16 epitope, a new variant of the class 1 protein in Neisseria meningitidis serogroup B was identified in Norway. A single amino acid deletion was revealed when the part of the gene region encoding the second variable region of the protein was sequenced. This new variant was designated P1.16c. About 5% of the B:15:P1.7,16 strains in Norway from the time period 1987-1991 were P1.16c. In a localized area in Southern Norway, 5/8 (62%) of the P1.7,16 strains were P1.16c. The P1.16b mutant, recently described in England, was not found among the Norwegian meningococcal isolates. Strains carrying the P1.16c mutation showed increased resistance to bactericidal killing, not only by P1.16-specific monoclonal antibodies, but also by the sera from individuals immunized with a vaccine based on outer membranes from a B:15:P1.7,16 strain.

The 5C protein is expressed by the strain of Neisseria meningitidis (44/76) used for production of the Norwegian meningococcal group B outer membrane vesicle vaccine and is included in the final formulation of this vaccine. The immunoglobulin G antibody response to 5C stimulated by vaccination, systemic meningococcal disease, and carriage was measured using ELISAs with synthetic liposomes as antigen and by immunoblotting. Increased levels of IgG were found in paired sera from all three groups. The antibodies were bactericidal to meningococci of serogroups A and B that expressed large amounts of 5C but not to meningococci expressing smaller amounts. There was a linear correlation between bactericidal titer and units of IgG to 5C.

Antibody responses after vaccination with three different formulations of a new meningococcal group B outer membrane vesicle (OMV) vaccine have been studied with the ELISA technique using four different antigens. Sera from about 1200 vaccinees participating in steps 1, 2, 3 and 6 of the phase II clinical trials in Norway were analysed. The effects of non-covalently complexing the OMV antigen to group C polysaccharide (C-PS) and of adsorbing OMV (with and without C-PS) to aluminium hydroxide (AH) were studied. All three vaccine formulations were highly immunogenic in humans. Adsorption of the vaccine to AH had a relatively small effect on the immune response, but the results indicated that the booster response was stronger with the adsorbed than with the unadsorbed vaccines. Some increase in the immune response against OMV was also observed by non-covalent complexing OMV with C-PS, particularly after the second dose. In most of the vaccinees the antibody levels were significantly reduced 6 to 12 months after vaccination. Adsorption of the vaccine to AH had no effect on the antibody response against C-PS. Comparison with bactericidal activity of the same sera was done. A highly significant correlation was observed between the bactericidal titres and the levels of IgG antibodies against OMV and class 5C protein, whereas the correlation between antibody levels against lipopolysaccharide and the bactericidal activity was poor.

A placebo controlled, double blind efficacy trial with a new outer membrane vesicle vaccine against systemic meningococcal disease of serogroup B, has been conducted in Norwegian secondary schools. The study was randomized at school level (1335 schools) and 171,800 students volunteered. The study started in October 1988 and the code was opened in June 1991. Out of the thirty-six proven cases of acute, severe, systemic disease caused by serogroup B meningococci among the participants, twelve occurred in eleven schools given vaccine, twenty-four in twenty-four schools given placebo. twenty-four cases were recorded among secondary school students who did not participate in the study. The protection rate was calculated to 57.4% with a p-value of 1.2% and lower limit of confidence (95%) to 27.7%. The results have initiated research towards an improved outer membrane vesicle vaccine against this disease.

An outer membrane vesicle vaccine against acute, systemic disease caused by meningococci of serogroup B has been developed. The vaccine has been tested consecutively in phase I and phase II clinical trials including more than 5000 volunteers. These trials provided data on safety, immunogenicity and reactogenicity and possible effect on carriage of meningococci in the throat, and consequently formed the basis for two major protection trials; one in secondary school students and one among military recruits. The aims, design and major results of phase I and phase II studies are described as well as the design and organization of the protection trials.

Results from the serum bactericidal assay (BA) after immunization of human volunteers with the Norwegian serogroup B meningococcal outer membrane vesicle vaccine are surveyed. In the phase II trials with adults we found very high seroconversion rates (greater than 98%) against the vaccine strain in the BA. Details in the antigenic composition of the inoculum used in the BA seem very important as shown here by finding lower bactericidal titres with teenager sera when tested with a variant of the standard inoculum. The present preliminary report corresponds to the presentation given at the Report Meeting on the Norwegian Meningococcal Vaccine Trial, Oslo, 12 September, 1991.

For more than 15 years, Norway has had the highest incidence of meningococcal disease in northern Europe, with 80% of cases being due to serogroup B meningococci. The case-fatality has remained high, at about 10%. In this study, an outer membrane vaccine, which had previously been shown to induce an increase in bactericidal antibodies to the parent strain, was assessed in a large-scale, randomised, double-blind trial. From October, 1988, 171,800 students in secondary schools volunteered to take part in a double-blind, placebo-controlled, efficacy trial with school as the randomisation unit. Hospitals and clinics that routinely receive patients with infectious disease were asked to report urgently all cases of suspected meningitis and/or septicaemia in 13-21-year-old students in Norway. These cases were registered and further investigated according to a detailed protocol. 89 out of the 221 cases investigated by June 3, 1991, were shown to be severe systemic disease due to group B meningococci. 36 cases in 35 schools took part in the trial (11 schools with vaccinated students and 24 with students given placebo). The calculated rate of protection was thus 57.2% (p = 0.012, one-sided test). The findings suggest that, although the vaccine conferred protection against group B meningococcal disease, the effect was insufficient to justify a public vaccination programme.

To elucidate pathogenic aspects and serodiagnostic possibilities for meningococcal disease, we investigated levels of specific antimeningococcal immunoglobulin G (IgG), IgA, and IgM in serum by using an enzyme-linked immunosorbent assay with outer membrane antigen prepared from a Neisseria meningitidis B:15:P1.16 strain. Serum samples were drawn on hospital admission as well as during convalescence from patients suspected of purulent meningitis or meningococcal septicemia, and single samples were drawn from population controls. A total of 637 samples were examined blindly. On admission, the average antimeningococcal immunoglobulin levels were about the same in the meningococcal disease patients as in the population controls. Septicemic patients, however, had significantly lower values. During one week the mean specific immunoglobulin levels in meningococcal-disease patients increased 6 times for IgG, 14 times for IgA, and 5 times for IgM. Children younger than 1 year showed a modest and more slowly developing antibody response. There were no statistically significant differences in average antibody responses among patients infected with meningococci of different serotypes. At 100% specificity, the increase in IgG, IgA, and IgM yielded diagnostic sensitivities for meningococcal disease of 84, 52, and 66%, respectively. One of seven serum pairs from the patient control group with unknown etiology was positive for meningococcal disease in this assay. The patients with meningococcal disease originally diagnosed only by clinical signs and symptoms showed a slightly lower rate of seroconversion than the patients in whom the diagnosis was supported by test results showing a systemic Neisseria meningitidis isolate.

Antibody responses of adult volunteers given a vaccine containing meningococcal capsular polysaccharides (serogroups A, C, Y, and W-135) noncovalently complexed with serotype 2b:P1.2 and 15:P1.16 outer membrane proteins have been studied. Sera were analyzed by enzyme-linked immunosorbent assay methods for immunoglobulin G (IgG), IgM, and IgA antibodies and for bactericidal activities against the homologous strains. The vaccination was performed as a double-blind experiment with 47 volunteers, of whom 23 received the protein-polysaccharide vaccine and 24 received the control preparation containing the polysaccharides only. Ten additional persons volunteered for the protein-polysaccharide vaccine. Before vaccination, carriers of meningococci had significantly higher levels of specific IgG and IgA and also higher bactericidal activities than noncarriers. At 2 weeks postvaccination we found significant IgG and bactericidal antibody responses against both the 2b:P1.2 and 15:P1.16 strains in about 70% of the protein-polysaccharide vaccinees. The immune response induced by disease was compared with that induced by vaccination by analyzing paired sera from 13 survivors of serogroup B serotype 15 meningococcal disease. We found that the mean specific IgG level in acute-phase sera was lower than average in prevaccination sera from the vaccinees but similar to that of healthy noncarriers before vaccination. The convalescent-phase sera showed IgG responses similar to those of the vaccinees, but the IgM response to disease was significantly higher than after vaccination. The immune response to disease caused by serogroup B serotype 15 meningococci was found by enzyme-linked immunosorbent assay analysis to be about the same with outer-membrane antigens from a serotype 2b strain as it was with antigens from a serotype 15 strain.