Determination of avidity of IgG against protein antigens from Streptococcus pneumoniae: assay development and preliminary application in clinical settings.

The measurement of antibody levels is a common test for the diagnosis of Streptococcus pneumoniae infection in research. However, the quality of antibody response, reflected by avidity, has not been adequately evaluated. We aimed to evaluate the role of avidity of IgG against eight pneumococcal proteins in etiologic diagnosis. Eight pneumococcal proteins (Ply, CbpA, PspA1 and 2, PcpA, PhtD, StkP-C, and PcsB-N) were used to develop a multiplex bead-based avidity immunoassay. The assay was tested for effects of the chaotropic agent, multiplexing, and repeatability. The developed assay was applied to paired samples from children with or without pneumococcal disease (n = 38 for each group), determined by either serology, polymerase chain reaction (PCR), or blood culture. We found a good correlation between singleplex and multiplex assays, with r ≥ 0.94.The assay was reproducible, with mean inter-assay variation ≤ 9% and intra-assay variation < 6%. Children with pneumococcal disease had lower median avidity indexes in the acute phase of disease for PspA1 and 2 (p = 0.042), PcpA (p = 0.002), PhtD (p = 0.014), and StkP-C (p < 0.001). When the use of IgG avidity as a diagnostic tool for pneumococcal infection was evaluated, the highest discriminative power was found for StkP-C, followed by PcpA (area under the curve [95% confidence interval, CI]: 0.868 [0.759-0.977] and 0.743 [0.607-879], respectively). The developed assay was robust and had no deleterious influence from multiplexing. Children with pneumococcal disease had lower median avidity against five pneumococcal proteins in the acute phase of disease compared to children without disease.

Detection of antibody responses against Streptococcus pneumoniae, Haemophilus influenzae, and Moraxella catarrhalis proteins in children with community-acquired pneumonia: effects of combining pneumococcal antigens, pre-existing antibody levels, sampling interval, age, and duration of illness.

We evaluated the effects of combining different numbers of pneumococcal antigens, pre-existing antibody levels, sampling interval, age, and duration of illness on the detection of IgG responses against eight Streptococcus pneumoniae proteins, three Haemophilus influenzae proteins, and five Moraxella catarrhalis proteins in 690 children aged <5 years with pneumonia. Serological tests were performed on acute and convalescent serum samples with a multiplexed bead-based immunoassay. The median sampling interval was 19 days, the median age was 26.7 months, and the median duration of illness was 5 days. The rate of antibody responses was 15.4 % for at least one pneumococcal antigen, 5.8 % for H. influenzae, and 2.3 % for M. catarrhalis. The rate of antibody responses against each pneumococcal antigen varied from 3.5 to 7.1 %. By multivariate analysis, pre-existing antibody levels showed a negative association with the detection of antibody responses against pneumococcal and H. influenzae antigens; the sampling interval was positively associated with the detection of antibody responses against pneumococcal and H. influenzae antigens. A sampling interval of 3 weeks was the optimal cut-off for the detection of antibody responses against pneumococcal and H. influenzae proteins. Duration of illness was negatively associated with antibody responses against PspA. Age did not influence antibody responses against the investigated antigens. In conclusion, serological assays using combinations of different pneumococcal proteins detect a higher rate of antibody responses against S. pneumoniae compared to assays using a single pneumococcal protein. Pre-existing antibody levels and sampling interval influence the detection of antibody responses against pneumococcal and H. influenzae proteins. These factors should be considered when determining pneumonia etiology by serological methods in children.

Specific antibody deficiency in children with recurrent respiratory infections: a controlled study with follow-up.

Specific antibody deficiency (SAD) to unconjugated pneumococcal vaccine (PPV) is an established primary B cell immunodeficiency. The occurrence and natural history of SAD in children is unclear. We conducted an observational study to identify SAD in children with recurrent respiratory infections. Ninety-nine children, mean age 5·9 (range 2-16) years, with recurrent or severe infections were vaccinated with PPV; serum antibody concentrations for serotypes 4, 6B, 9V, 14, 18C, 19F and 23F were measured before and 2 weeks after vaccination with enzyme immunoassay. The retrospective control group consisted of 89 healthy children matched for age and gender. No children had received previous conjugated pneumococcal vaccine (PCV) or PPV. The structured history of infectious diseases of all participants was collected. Ten of 91 (11%) children (eight excluded due to immunoglobulin G subclass deficiency) with recurrent respiratory infections had SAD. In the control group, three children (3%) responded inadequately to PPV (P = 0·05). Most children with SAD also had many other minor immune defects. After 0·5-5 years (medium 3·8), eight children with SAD were revaccinated with PPV; five responded adequately and three inadequately. Two SAD children were revaccinated with PCV, one developed an adequate and one an inadequate response. Two children with SAD received treatment with intravenous immunoglobulin; the remaining eight children recovered without replacement therapy during the follow-up. SAD is common in young children with recurrent respiratory infections, but it is often transient and resolves itself within a few years without specific treatment.

Immunization against pneumococcal disease in HIV-infected patients: conjugate versus polysaccharide vaccine before or after reconstitution of the immune system (CTN-147).

To investigate whether patients should be immunized immediately or delay immunization until after reconstitution of the immune system and whether a conjugate or polysaccharide vaccine results in a better immunologic response. Seventy-nine patients were randomly assigned, utilizing a two by two factorial design to receive immediate immunization or delay immunization. Baseline characteristics were similar for the four arms: 78% men, median age 41 years, median time since HIV diagnosis 0.3 years, median CD4 60 cells/mm(3) and median HIV viral load 5.02 log copies/mL. Results in favour of delayed immunization were observed for those serotypes showing a response. The proportional odds ratios for delayed versus immediate immunization were 0.341 (P = 0.04) and 0.204 (P = 0.004) at months 6 and 12, respectively. No differences in immunological response were observed between the two individual vaccines for the shared serotypes studied. HIV-infected adults produced a higher immunological response to pneumococcal vaccine after reconstitution of the immune system.

Antibody response to Haemophilus influenzae type-b conjugate vaccine in children and young adults with congenital asplenia or after undergoing splenectomy.

Absence of the spleen constitutes a risk of infection caused by encapsulated bacteria. The aim of our study was to determine the immune response to Haemophilus influenzae type-b (Hib) conjugate vaccine (HibCV) in asplenic individuals, considering the cause of asplenia, the age when splenectomy was carried out, and previous Hib vaccinations. Twenty asplenic patients, aged five to 25 years, were immunized with a single dose of HibCV. The specific antibody concentrations against HibCV were measured by enzyme-linked immunosorbent assay. Before vaccinations, the geometric mean antibody concentration (GMC) had an average value of 3.21 µg/ml and was comparable for all of the patients, regardless of the causes of asplenia. After vaccinations, the GMC was significantly higher, with an average of 6.78 µg/ml. Further, 4.5 years after vaccinations, the GMC was comparable to that of previously unvaccinated children. Moreover, 17/20 patients had GMC ≥ 1.0 µg/ml, which included all of the children with congenital asplenia, children splenectomized before the age of six years, and only 57% of children splenectomized after that age. HibCV gives asplenic patients long-term protection. Hence, HibCV should be administered regardless of previous vaccinations and time from splenectomy, even if antibody evaluation is not available.

Immune response to the 7-valent pneumococcal conjugate vaccine in 30 asplenic children.

The aim of the study was to determine the concentration of pneumococcal antibodies after a dose of 7-valent pneumococcal conjugate vaccine (PCV7) in 30 asplenic children between 4 months and 19 years of age. Fifteen children had received pneumococcal polysaccharide vaccine (PPV) approximately 5 years prior to vaccination with PCV7. The antibody concentrations against serotypes 4, 6B, 9V, 14, 18C, 19F, and 23F were measured by ELISA before and after the PCV7 vaccination. Before vaccination with PCV7, the antibody concentrations were similar in children who had or had not received PPV previously. A dose of PCV7 stimulated a good immune response in asplenic patients. Prior immunization with PPV did not affect the antibody concentration after the vaccination with PCV7. In conclusion, asplenic children vaccinated with PPV may need revaccination with PPV earlier than the recommended 3-5 years after the first dose. PCV7 induces a satisfactory immune response in asplenic patients and should be considered as an alternative vaccine in that patient group.

A randomized study on donor immunization with tetanus-diphtheria, Haemophilus influenzae type b and inactivated poliovirus vaccines to improve the recipient responses to the same vaccines after allogeneic bone marrow transplantation.

The HLA-identical sibling donors of 111 bone marrow transplantation (BMT) recipients were randomised to receive or not to receive tetanus-diphtheria (T-d), Haemophilus influenzae type b (Hib), and inactivated poliovirus (IPV) vaccines 2-10 weeks before BM harvest. Fifty-three (DV+ group) recipients received the graft from a vaccinated donor and 58 (DV- group) from an unvaccinated donor. All recipients were vaccinated with the T-d, Hib and IPV vaccines at 3, 6 and 12 months after BMT. Diphtheria and Hib antibody concentrations were consistently higher in the DV+ than in the DV- group from 6 months post transplantation onwards. The differences were significant at 6 and 13 months for diphtheria and at 12 months for Hib antibody concentrations. Tetanus, PV1, PV2 and PV3 antibody levels were similar in both groups. Patients transplanted from donors with high tetanus, diphtheria and Hib antibody concentrations had higher respective antibody concentrations after BMT than those transplanted from donors with low antibody concentrations. Especially patients whose donors have low-specific antibody concentrations may benefit from donor vaccination with protein and conjugate vaccines.

Subclass distribution of natural salivary IgA antibodies against pneumococcal capsular polysaccharide of type 14 and pneumococcal surface adhesin A (PsaA) in children.

A number of studies have shown that the ratio of IgA1 and IgA2 subclasses in secretions can depend upon the nature of the antigen inducing their production. In order to evaluate the effect of the nature of the antigen on the subclass distribution of the naturally occurring salivary IgA antibodies against Streptococcus pneumoniae, we used enzyme immunoassay to measure the levels of natural IgA, IgA1 and IgA2 antibodies to pneumococcal capsular polysaccharide type 14 (PS14) and pneumococcal surface adhesin A (PsaA) in saliva of children during their first 2 years of life. The sum of anti-PS14 and anti-PsaA IgA1 and IgA2 correlated significantly with the antigen-specific total IgA, which showed that IgA1 and IgA2 add up to IgA. IgA1 was the predominant subclass for both antigens. The median of anti-PS14 and anti-PsaA IgA1 was higher than that of IgA2, and the antigen-specific IgA1 was found in a larger proportion of samples than IgA2. The ratio of IgA1 to IgA2 (IgA1/IgA2 ratio) was lower for anti-PS14 than for anti-PsaA, suggesting that the PS antigen induced more IgA2 than the protein antigen. The possible impact of the IgA subclass distribution on protection of mucosal surfaces by natural or vaccine-induced antibodies needs to be determined.

Maternal immunization with pneumococcal polysaccharide vaccine in the Philippines.

A randomized, controlled study was conducted to evaluate the immunogenicity and reactogenicity of the 23-valent pneumococcal (Pnc) polysaccharide (PS) vaccine among pregnant women and to ascertain the transfer of anti-Pnc antibody (Ab) from mother to infant. One hundred and sixty women received either one dose of Pnc PS vaccine, Haemophilus influenzae type b conjugate vaccine and tetanus toxoid (TT) (Pnc vaccine group, N=106) or TT only (control group, N=54). Sera were obtained from all mothers prior to vaccination and 4 weeks after from the vaccinated group. Cord blood was obtained in 75% of deliveries. Anti-Pnc Ab for serotypes 1, 5, 6B, 14, 18C and 19F was determined using enzyme immunoassay. The Pnc vaccine and control groups were comparable in terms of age, parity, gravidity, prior doses of TT, and pre-vaccination geometric mean concentration (GMC in microg/ml) of anti-Pnc Ab. Between 66 and 87% of the mothers had type-specific Ab prior to vaccination. There was a significant rise in anti-Pnc Ab (varying from 3.3- to 9.1-fold for the individual serotypes) between the pre and post-vaccination samples. Adverse reactions were mild and required no treatment. The level of anti-Pnc Ab in cord blood was significantly lower in the control group compared to the Pnc vaccine group. Vaccination of pregnant women with Pnc Ps vaccine induces good immune response and Ab can be transferred to their infants via cord blood thus providing enhanced protection.