Enhanced antibody response in Venezuelan infants immunized with Haemophilus influenzae type b-tetanus toxoid conjugate vaccine

The safety and immunogenicity of primary immunization at 2, 4 and 6 months of age with Haemophilus influenzae type b capsular polysaccharide conjugated to tetanus toxoid (PRP-T; Act-HIB) were evaluated in infants in Valencia, Venezuela. In order better to assess reactions to PRP-T, subjects received their initial PRP-T vaccine a mean of 6.5 days after their initial diphtheria-tetanus-pertussis (DTP) vaccine. The PRP-T vaccine was well tolerated. Serum was obtained at ages 2 and 7 months (before the first and 1 month after the third PRP-T dose). Antibody responses were compared with those from Nashville infants who had received PRP-T and DTP simultaneously in a previous trial. The preimmunization titers in the Venezuelan and Nashville infants did not differ. The geometric mean postimmunization titer in the Venezuelan infants was 37.9 micrograms/ml, as compared with 3.63 micrograms/ml in the Nashville infants (P < 0.00001). Possible explanations for the exceptional antibody response of these Venezuelan infants to PRP-T include carrier priming caused by prior DTP immunization, synergy associated with the specific DTP vaccine used, preimmunization immunologic experience that differed from their United States counterparts and genetic differences that altered response to the vaccines. Further studies are proposed to evaluate these possibilities.     

The use of Haemophilus influenzae type b-tetanus toxoid conjugate vaccine mixed with diphtheria-tetanus-pertussis vaccine in Gambian infants

In preparation for an efficacy trial of PRP-T Haemophilus influenzae type b conjugate vaccine, 251 Gambian infants were randomized to receive three doses of PRP-T and diphtheria-tetanus-pertussis (DTP) vaccines at 2, 3 and 4 months of age, either by separate injections, or combined in the same syringe. One month after the third dose, there was no difference between anti-PRP levels in those infants who received the vaccines separately (GMT 5.83 micrograms ml-1), and those who received the vaccines combined (GMT 5.57 micrograms ml-1). The proportions achieving levels of 1.0 microgram ml-1 were 89% and 92% in the "separate" and "combined" vaccine groups, respectively. There were no significant differences between groups in levels of antibody to diphtheria or tetanus. Geometric mean titres of antibody directed against pertussis antigens in the "separate" and "combined" groups were as follows: pertussis toxin 14.2 and 13.1 ELISA units (EU) ml-1; filamentous haemagglutinin 12.2 and 9.7 EU ml-1; pertactin 17.2 and 9.0 EU ml-1 (P < 0.05), fimbrial 2/3 antigens 449 and 364 EU ml-1. The combination of PRP-T and DTP in the syringe prior to administration is safe and immunogenic. The lower levels of anti-pertussis antibody are of unknown clinical significance.     

Immunogenicity study of Haemophilus influenzae type B conjugate vaccine in Indian infants

OBJECTIVE: To assess the immunogenicity in Indian infants to Haemophilus influenzae b oligosaccharide conjugate vaccine (HbOC). DESIGN: Prospective multicenter study. SETTING: Pediatric Out Patient Department of general hospitals in Pune and Mumbai. SUBJECTS: 124 full term healthy infants brought for routine DPT/OPV immunization. METHODS: Infants were administered 3 doses of 0.5 ml of HbOC, on the same day as their DPT/OPV immunization, injected intramuscularly on the limb opposite to that where DPT vaccine was administered. Data on local reactions and general symptoms was collected for three days after every dose. The children had their blood collected for assay of anti PRP (polyribosil ribitol phosphate) antibody titers, along with the first injection and one month after the third injection. One hundred and three infants completed the study protocol with two blood collections. RESULTS: The initial geometric mean titers (GMT) of 0.124 mcg/ml rose by 37 times to 4.552 mcg/ml. Ninety eight children (95.1%) had a final titer of > or = 0.15 mcg/ml, the minimum level associated with protection, and 77 children (74.8%) had a final level of > or = 1.0 mcg/ml, a level associated with long term protection. CONCLUSION: HbOC is immunogenic in Indian infants when used as per the locally recommended DPT/OPV immunization schedule.     

Opposite effects of actively and passively acquired immunity to the carrier on responses of human infants to a Haemophilus influenzae type b conjugate vaccine

Recent advances in the molecular modelling of carbohydrates have brought this technique to a level comparable with that of protein and nucleic acid simulations. After a brief introduction to the techniques used in the computer simulation of carbohydrates and carbohydrate interactions, an overview of applications in the field of carbohydrate-related drug discovery is presented.     

Antibody responses to tetanus toxoid and Haemophilus influenzae type b conjugate vaccines following autologous peripheral blood stem cell transplantation (PBSCT)

OBJECTIVE: Although there are many anecdotal reports that psychological intervention is effective in enhancing adjustment to spinal cord injury (SCI), there are little data to support this assertion. To date, reports of few longitudinal-based controlled trials that assessed psychological outcomes for SCI persons have been published. This study was conducted to determine long-term efficacy of cognitive behavior therapy during rehabilitation. DESIGN: The study employed a nonrandomized controlled trial, and measures were taken on three occasions: before, immediately after, and 12 months after treatment. SETTING, OUTCOME MEASURES, AND INTERVENTION: Anxiety, depressive mood, and self-esteem were assessed in 28 SCI persons consecutively selected on admission to hospital, who participated in specialized group cognitive behavior therapy (CBT) during rehabilitation. CONTROLS: The intervention group's responses on the measures were compared with a control group of 41 SCI persons who only received traditional rehabilitation services during their hospitalization. RESULTS: There were no overall group differences on anxiety, depressive mood, and self-esteem, although there was a trend for the treatment group to have greater levels of improvement in depression scores across time in comparison to the control group. However, those in the treatment group who reported high levels of depressive mood before the CBT treatment were significantly less depressed 1 year after injury, compared to similar persons in the control group. CONCLUSIONS: While it appears not everyone who experiences SCI needs CBT, at least in the hospital phase of their rehabilitation, those who report high levels of depressive mood benefited greatly from CBT.     

Preclinical studies on a recombinant group B meningococcal porin as a carrier for a novel Haemophilus influenzae type b conjugate vaccine

In anticipation of future combination vaccines, a recombinant class 3 porin (rPorB) of group B meningococci was evaluated as an alternative carrier protein for a Haemophilus influenzae type b (Hib) polyribosylribotol phosphate (PRP) conjugate vaccine. The use of rPorB may avoid undesirable immunologic interactions among vaccine components, including epitopic suppression from conventional carriers (e.g. tetanus toxoid [TT]), as well as provide desirable immunomodulatory effects. Rats were found to be more reliable and consistent than mice or guinea pigs for studying antibody responses to the Hib conjugates. Different Hib conjugates, Hib-TT and Hib-rPorB, consisting of PRP conjugated by reductive amination to TT or rPorB, were compared in rats. Commercially available, licensed vaccines, HbOC (HibTITER) and PRP-T (OmniHib), were used as reference controls. Maximum geometric mean ELISA IgG titers were obtained in rats after only two doses, showing booster effects for all. However, Hib-rPorB immunization consistently resulted in responses that were 1-2 orders of magnitude greater than those for the other conjugates, including the licensed control vaccines. A maximum 4600-fold rise was observed for Hib-rPorB after two doses, and, unlike the other conjugates, a 100% response rate was always achieved without adjuvant. These results warrant further investigation of Hib-rPorB in combination with DTaP.     

A routine high-performance size-exclusion chromatography to determine molecular size distribution of Haemophilus influenzae type b conjugate vaccines

High-performance size exclusion chromatography has been used to determine the molecular size distribution of Haemophilus influenzae type b (Hib) conjugate vaccines. Both high molecular weight preparations of native Hib capsular polysaccharide coupled to tetanus toxoid and low molecular weight vaccines with Hib oligosaccharides linked to the CRM197 nontoxic mutant diphtheria protein were analysed. Columns with different fractionation ranges were used for the two kinds of vaccines. This method showed to be rapid, accurate and reproducible for different lots of Hib vaccine of different composition produced by various manufacturers. It could replace more time-consuming chromatographic methods enabling control authorities to employ a single methodological approach for different Hib vaccines.     

Comparison of multiple immunization schedules for Haemophilus influenzae type b-conjugate and tetanus toxoid vaccines following bone marrow transplantation

Antibody concentrations to vaccine-preventable diseases decline following BMT and an optimal schedule for vaccination after transplant has not been established. We examined antibody responses to tetanus toxoid (TT) and Haemophilus influenzae type b-conjugate (HIB) vaccines of BMT patients immunized at 6, 12 and 24 months (6 month group, n = 21) and compared them to those previously reported for patients immunized at 3, 6, 12 and 24 months (3 month group, n = 74) or at 12 and 24 months (12 month group, n = 17) following transplantation. Geometric mean total anti-HIB and IgG anti-TT concentrations were significantly higher after the 12 month dose in the 3 and 6 month immunization groups compared to the group who received their first dose at 12 months. Although HIB antibody concentrations were higher in the 3 month and 6 month groups 12 to 24 months after BMT, the proportion of patients with protective levels was not significantly different from the proportion protected in the 12 month group. Following the 24 month immunizations, geometric mean antibody concentrations to HIB and TT were similar for all three immunization groups. The proportion of patients in each group with protective levels of HIB antibody after the 24 month dose was > or = 80%. A two dose schedule of HIB and TT vaccines at 12 and 24 months after BMT should afford protection.     

Antibody responses in serum and lung to intranasal immunization with Haemophilus influenzae type b polysaccharide conjugated to cholera toxin B subunit and tetanus toxoid

AIM: Cholera toxin B subunit (CTB) has previously been used as a mucosal carrier for various vaccine candidate antigens. The objective of this study was to see if coupling a bacterial polysaccharide, Haemophilus influenzae type b capsular polysaccharide (HibCPS), to CTB, either directly or through prior coupling to tetanus toxoid (TT), would improve the immunogenicity of HibCPS after nasal immunization. METHODS: HibCPS was conjugated to CTB, TT or via TT to CTB, using glutaraldehyde or 1-ethyl-3(3-dimethylaminopropyl)-carbodiimide (EDAC) and N-succinimidyl 3-(2-pyridyldithio) propionate (SPDP). The conjugates were characterized and used for intranasal (IN) and subcutaneous (SC) immunizations of mice. The anti-Hib, -TT and -CTB antibody titers in serum and lungs after the immunizations were measured with ELISA. RESULTS: The HibCTB was poorly immunogenic both given IN and SC compared with HibTT and HibTTCTB, probably because of inefficient coupling. In contrast, the conjugation of CTB to the HibTT conjugate resulted in a preparation which was superior both to the HibTT and the HibCTB conjugates in inducing local IgA and IgG anti-HibCPS antibodies in the lungs. The anti-HibCPS serum IgG titers after IN immunization with the HibTTCTB conjugate were similar to the titers after IN immunization with HibTT, or SC immunization with a commercial HibCRM conjugate vaccine. In contrast to the other conjugates, the HibTTCTB conjugate also gave rise to anti-Hib serum IgA titers. CONCLUSION: We conclude that appropriate conjugation to CTB increases the mucosal immunogenicity of HibCPS, and that intranasal immunization with such a conjugate can give rise to both local and systemic anti-HibCPS antibody responses.     

Interlaboratory study evaluating quantitation of antibodies to Haemophilus influenzae type b polysaccharide by enzyme-linked immunosorbent assay

An interlaboratory study was conducted to determine whether an enzyme-linked immunosorbent assay (ELISA) with an antigen preparation composed of various-sized fragments of Haemophilus influenzae type b polysaccharide conjugated to human serum albumin could be standardized across laboratories and whether the ELISA-derived results from different laboratories are equivalent to those obtained by the standard radioactive antigen binding assay (RABA) for quantitation of anti-H, influenzae type b polysaccharide antibodies. Twenty coded human serum samples were quantitated by ELISA in 11 laboratories and by RABA in 5 laboratories. The mean RABA-derived values served as the basis for all comparisons. While the overall correspondence of antibody values between the two methods was good, significant differences were found among some of the 11 ELISA data sets and among the mean RABA values. Seven laboratories generated higher ELISA antibody values for low-titered sera. Four laboratories generated antibody concentrations that were not statistically different between the two assay methods. The results therefore indicate that the ELISA can tolerate substantial variations in protocol, such as the use of different plates and different antibody reagents, without affecting the quantitation of serum antibodies. However, attention should be focused on low-titered sera, as some assay conditions may yield spurious results. This ELISA is a serologic assay which can serve as an alternative to the RABA for quantitation of antibodies to H. influenzae type h polysaccharide.     

MF59 adjuvant enhances antibody responses of infant baboons immunized with Haemophilus influenzae type b and Neisseria meningitidis group C oligosaccharide-CRM197 conjugate vaccine

The ability of the adjuvant MF59 to enhance the immunogenicity of polysaccharide-protein conjugate vaccines was investigated in infant baboons. MF59 consists of stable droplets (<250 nm) of the metabolizable oil squalene and two surfactants, polyoxyethylene sorbitan monooleate and sorbitan trioleate, in an oil-in-water emulsion. In humans, MF59 is well tolerated and enhances the immunogenicity of recombinant protein subunit or particle vaccines. Its effect on the immunogenicity of polysaccharide-protein conjugate vaccines is unknown. Baboons 1 to 4 months of age were immunized intramuscularly with Neisseria meningitidis group C and Haemophilus influenzae type b (Hib) oligosaccharide-CRM197 conjugate vaccines. The lyophilized vaccines were reconstituted with phosphate-buffered saline (PBS), Al(OH)3 (alum), or MF59. Groups of five animals each were given three injections of the respective formulations, with one injection every 4 weeks. Four weeks after each immunization, the MF59 group had up to 7-fold-higher geometric mean anticapsular-antibody titers than the alum group and 5- to 10-fold-higher N. meningitidis group C bactericidal-antibody titers. Twenty-one weeks after the third immunization, the MF59 group still showed 5- to 10-fold-higher anticapsular-antibody titers. The antibody responses of the animals given the vaccines reconstituted with PBS were low at all times measured. Both the MF59 and alum groups, but not the PBS group, showed booster antibody responses to unconjugated Hib and N. meningitidis group C polysaccharides, results consistent with induction of memory B cells. Thus, MF59 may be useful for accelerating and augmenting immunity to polysaccharide-protein conjugate vaccines in infants.     

The immunogenicity of three Haemophilus influenzae type B conjugate vaccines after a primary vaccination series in Philippine infants

Serum antibody responses to three Haemophilus influenzae type b (Hib) capsular polysaccharide-protein conjugate vaccine (PRP-OMP, PRP-T, and HbOC) were evaluated in 174 Philippine infants after a primary vaccination series. Children were randomized to receive one of the Hib vaccines (Hib groups) or into a control group. Vaccination was carried out at six, 10 and 14 weeks of age based on the local Expanded Program of Immunization schedule. Sera were collected at six weeks of age for the Hib groups and one month after the third dose for all subjects. Anti-Hib concentrations were determined by the Farr-type radioimmunoassay. There were no significant differences (P = 0.3626) in the prevaccination anti-Hib geometric mean concentration (GMC) among the three Hib groups. Differences in the GMC after the primary series of three doses were significant (P < 0.0001); GMC was highest for PRP-T (6.62 micrograms/ml), followed by HbOC (1.9 micrograms/ml), then PRP-OMP (1.06 micrograms/ml), and lowest for the control group (0.11 microgram/ml). We conclude that all three Hib conjugate vaccines (PRP-T, HbOC, and PRP-OMP) were immunogenic after three primary doses among Philippine infants.     

Conjugate vaccines and the carriage of Haemophilus influenzae type b

Pharyngeal carriage of Haemophilus influenzae type b (Hib) is important in the transmission of Hib organisms, the pathogenesis of Hib disease, and the development of immunity to the bacterium. The remarkable success of current vaccination programs against Hib has been due in part to the effect of conjugate Hib vaccines in decreasing carriage of Hib. This review explores evidence for this effect, and discusses the possible mechanisms of the mucosal influence of Hib conjugate vaccines.     

Antibody responses of three Haemophilus influenzae type b conjugate vaccines after one, two and three doses in Filipino children

Differences in the magnitude of antibody response after one, two or three doses of Haemophilus influenzae type b conjugate vaccines have been reported which may influence decision-making regarding which vaccine should be used. This is of particular importance in developing countries where children may not receive a full immunization series and the vaccination schedule may be delayed. Serum antibody responses to three Hib capsular polysaccharide protein conjugate vaccines (PRP-OMP, HbOC and PRP-T) were evaluated in 102 Filipino infants. Vaccination was carried out at 6, 10 and 14 weeks of age based on the national Expanded Programme on Immunization (EPI) schedule together with diphtheria-tetanus-pertussis, hepatitis B and oral poliomyelitis vaccines. Sera were collected at 6 weeks and 1 month after each vaccination. Anti-Hib polysaccharide antibody concentrations were determined by Farrtype radioimmunoassay (RIA) and enzymeimmunoassay (EIA), Following the first dose, the geometric mean concentrations (GMC, micrograms ml-1) for PRP-OMP, HbOC and PRP-T were 0.69, 0.27 and 0.38, respectively. After two doses, there was a significant response (P < 0.05) to PRP-OMP and PRP-T (0.89 and 1.47) but not for HbOC (0.37). Differences in the GMC after the primary series were significant (pairwise P < 0.05): GMC was highest for PRP-T (4.0), followed by HbOC (1.6) and PRP-OMP (1.1). All three Hib vaccines were immunogenic when given in the local EPI schedule in Filipino infants although significant differences in the kinetics and magnitude of antibody responses were noted. The anti-Hib antibody concentrations determined by RIA and EIA were also compared in order to validate the latter for use in laboratories where it is feasible. There was a good correlation (r2 = 76%; P = 0.0001) in the Hib antibody titres obtained by both assays.     

Murine immune responses to Neisseria meningitidis group C capsular polysaccharide and a thymus-dependent toxoid conjugate vaccine

The polysaccharide (PS) capsules of many pathogenic bacteria are poor immunogens in infants and young children as a result of the delayed response to PS antigens during ontogeny. The development of polysaccharide-protein conjugate vaccines for Haemophilus influenzae type b, which have proven to be efficacious in this age group, has led to active development by a number of investigators of conjugate vaccines for other diseases. We describe here the response of several mouse strains to the capsular PS of Neisseria meningitidis group C (MCPS) conjugated to tetanus toxoid (MCPS-TT) and the same response in BALB/c mice as a model of the immune consequences of conjugate vaccine immunization. The use of a conjugate vaccine results in a shift in the isotype elicited in response to the MCPS, from immunoglobulin M (IgM) and IgG3 to primarily IgG1. A response to MCPS-TT is seen even among mouse strains which respond poorly to MCPS itself, emphasizing the importance of a strain survey when choosing a mouse model for a vaccine. The marked increase in IgG1 antibody titer was accompanied by a large increase in bactericidal activity of sera from these animals. Animals primed with the conjugate vaccine demonstrated a booster response after secondary immunization with either the MCPS or the conjugate. The ability to produce a boosted IgG1 anti-MCPS response to the MCPS can be transferred to adoptive recipients by B cells alone from mice primed with MCPS-TT but not mice primed with MCPS alone. These data indicate that in BALB/c mice a single immunization with MCPS-TT is sufficient to induce a shift to IgG1 and generate a memory B-cell population that does not require T cells for boosting.     

Antibody response to outer membrane protein of nontypeable Haemophilus influenzae in otitis-prone children

One of the major outer membrane proteins of nontypeable Haemophilus influenzae, P6, is highly conserved among strains, serves as a target for bactericidal antibody, and has been proposed as a possible vaccine candidate. The serum antibody response to P6 was studied in otitis-prone and normal children by an enzyme-linked immunosorbent assay. Of 20 otitis-prone children, 12 (60%) had a serum IgG antibody response to P6 after otitis media; however, the mean acute antibody level for the group, 4.6 micrograms/ml, was not significantly different from the convalescent level, 5.4 micrograms/ml. Anti-P6 antibody levels were also measured longitudinally for 10 to 25 months in 30 otitis-prone and 13 healthy children. Antibody levels increased sevenfold in the normal group compared with less than three-fold for the otitis-prone group and were significantly higher in the normal children after the age of 18 months (p < 0.05). Finally, otitis-prone children who had two or more episodes of otitis media with nontypeable H. influenzae did not have an anamnestic antibody response to P6. The failure to recognize P6 as a specific immunogen may account for recurrent infections. Moreover, the data suggest that otitis-prone children may not respond adequately to a vaccine containing P6.     

Isotypes and opsonophagocytosis of pneumococcus type 6B antibodies elicited in infants and adults by an experimental pneumococcus type 6B-tetanus toxoid vaccine

Streptococcus pneumoniae is a major respiratory pathogen of infants, children, and the elderly. Polysaccharide vaccines have been useful in adult populations but do not elicit protective immunity in infants and young children. To enhance their immunogenicity, vaccines of pneumococcal polysaccharides conjugated to proteins are being developed. In this study antibody levels and opsonic activities were compared in sera of infants and adults injected with pneumococcal polysaccharide type 6B (Pn6B) conjugated to tetanus toxoid (TT) (Pn6B-TT). Healthy infants were injected with Pn6B-TT; group A was injected at 3, 4, and 6 months of age, and group B was injected at 7 and 9 months of age. A booster injection was given at 18 months. Adults were injected once. Antibodies were measured by enzyme-linked immunosorbent assay and radioimmunoassay, and their functional activities were measured by opsonophagocytosis of radiolabelled pneumococci. In adults, increases in immunoglobulin M (IgM), IgG, IgA, IgG1, and IgG2 to Pn6B were observed. Infants reached adult levels of IgG1 anti-Pn6B after the primary injections. After the booster injection the infant groups had total IgG- and IgM-Pn6B antibody levels similar to those of adults. After the booster injection, IgG1 was the dominant infant anti-Pn6B isotype and at a level higher than in vaccinated adults, but IgA and IgG2 antibodies remained at very low levels. Opsonic activity increased significantly after Pn6B-TT injections; the highest infant sera showed opsonic activity comparable to that of vaccinated adults. Overall, opsonic activity correlated best with total and IgG anti-Pn6B antibodies (r = 0.741, r = 0.653, respectively; n = 35) and was highest in sera with high levels of all Pn6B antibody isotypes. The results indicate the protective potential of a pneumococcal 6B polysaccharide protein conjugate vaccine for young infants.