Capsular Antibodies Research Articles

Natural acquired humoral immunity against serotype-specific group B Streptococcus rectovaginal colonization acquisition in pregnant women

Group B Streptococcus (GBS) rectovaginal colonization in pregnant women is associated with invasive GBS disease in newborns, preterm delivery and stillbirths. We studied the association of GBS serotype-specific capsular polysaccharide (CPS) antibody on new acquisition and clearance of rectovaginal GBS colonization in pregnant women from 20 weeks until 37 to 40 weeks' gestation. Serum serotype-specific CPS IgG antibody concentration was measured by multiplex enzyme-linked immunosorbent assay and opsonophagocytic activity (OPA) titres. Rectovaginal swabs were evaluated for GBS colonization, using standard culture methods and serotyping by latex agglutination, at five to six weekly intervals. Higher serotype III CPS antibody concentration was associated with lower risk of rectovaginal acquisition of serotype III during pregnancy (p 0.009). Furthermore, serotype-specific OPA titres to Ia and III were higher in women who remained free of GBS colonization throughout the study compared to those who acquired the homotypic serotype (p <0.001 for both serotypes). Serum CPS IgG values of ≥1μg/mL for serotype V and ≥3μg/mL for serotypes Ia and III were significantly associated with protection against rectovaginal acquisition of the homotypic serotype. A GBS vaccine that induces sufficient capsular antibody in pregnant women, including high OPA titres, could protect against rectovaginal colonization during the latter half of pregnancy.

HIV-1 Is Associated With Lower Group B Streptococcus Capsular and Surface-Protein IgG Antibody Levels and Reduced Transplacental Antibody Transfer in Pregnant Women.

Human immunodeficiency virus (HIV)-exposed infants are at increased risk of invasive Group B Streptococcus (GBS) disease; however, the reason for this increased susceptibility has not been characterized. We compared GBS capsular and surface-protein maternal immunoglobin G antibody concentrations and cord-maternal ratios between HIV-infected and HIV-uninfected mother-newborn dyads. Median capsular antibody concentrations (µg/mL) were lower in HIV-infected than HIV-uninfected women for serotypes Ib (P = .033) and V (P = .040); and for pilus island (PI)-1 (P = .016), PI-2a (P = .015), PI-2b (P = .015), and fibrinogen-binding protein A (P < .001). For serotypes Ia and III, cord-maternal ratios were 37.4% (P < .001) and 32.5% (P = .027) lower in HIV-infected compared to HIV-uninfected mother-newborn dyads. The adjusted odds of having capsular antibody concentration ≥2 µg/mL when comparing HIV-infected to -uninfected women were 0.33 (95% confidence interval [CI], .15-.75) and 0.34 (95% CI, .12-1.00) for serotypes Ia and III, respectively. Antibody levels and cord-maternal ratios were independent of CD4(+) lymphocyte counts or HIV-1 viral load. The lower GBS antibody concentrations and reduced transplacental antibody transfer in HIV-infected women, which likely contribute to their infants being at heightened susceptibility for invasive GBS disease, could possibly be mitigated by vaccination with a GBS conjugate vaccine currently under clinical development.

Assessment of clinical and immunological efficacy and safety of 23-valent pneumococcal vaccine in patients with rheumatoid arthritis

The objective of this trial was to assess clinical efficacy, immunogenicity, and safety of 23valent pneumococcal vaccine (PPV23) in patients with rheumatoid arthritis (RA). Methods . The trial enrolled 95 patients (75 women and 20 men aged 23 to 70 years) including 68 patients with RA and 27 subjects without systemic inflammatory rheumatic diseases (a control group) who had a recent history of ≥ 2 episodes of lower respiratory tract infection. At enrollment, all the patients received antiinflammatory therapy with methotrexate (MT) (n = 48), leflunomide (LEF) (n = 10), or MT + tumor necrosis factorα (TNFα) inhibitors (n = 10). A single 0.5ml dose of PPV23 (Pneumo23, Sanofi Pasteur) was administered subcutaneously under the regular therapy with MT or LEF or 3–4 weeks before TNFα inhibitor administration. During followup (1 and 3 months and 1 year after the vaccination), the patients underwent physical examination and routine clinical and laboratory investigations. Antipneumococcal capsular polysaccharide antibodies were measured in the serum using ELISA assay with commercial VaccZymeTM AntiPCP IgG Enzyme Immunoassay Kit (The Binding Site Group Ltd, Birmingham, UK). Results. Noone case of pneumonia was registered in vaccinated patients during 12month followup. A twofold increase in antipneumococcal antibody level was found 1 year after the vaccination in RA patients and the control group. The vaccine was well tolerated by 63 patients (66%); 25 patients (26%) experienced pain, swelling and hyperemia at the area of vaccine injection and 7 patients (8%) had lowgrade fever. Neither RA exacerbation nor new autoimmune disorders were diagnosed during the followup. Conclusion. The findings suggest that PPV23 showed good clinical efficacy, adequate immunogenicity and good tolerability in RA patients.

ASSESSMENT OF THE IMMUNOGENICITY AND SAFETY OF 23-VALENT POLYSACCHARIDE PNEUMOCOCCAL VACCINE IN PATIENTS WITH RHEUMATIC DISEASES

Objective: to investigate the immunogenicity and safety of 23-valent polysaccharide pneumococcal vaccine in patients with rheumatic diseases (RD). Subjects and methods. The prospective open-label comparative study enrolled 133 people (102 (76.7%) women and 31 (23.3%) men) aged 23 to 76 years, including 79 patients with rheumatoid arthritis (RA), 16 with systemic sclerosis, and 7 with dermatomyositis/polymyositis, as well as 31 subjects without systemic inflammatory RD (a control group), who had a recent history of at least two cases of lower respiratory tract infections (bronchitis, pneumonia). At their inclusion, all the patients with RD were receiving ant-inflammatory therapy, including 52 taking methotrexate (MT), 14 – leflunomide (LEF), and 13 – MT + tumor necrosis factor-α (TNF-α) inhibitors. The 23-valent polysaccharide pneumococcal vaccine Pneumo-23 (Sanofi Pasteur, France) was administered in a single dose of 0.5 ml subcutaneously during continuous MT or LEF therapy for the underlying disease or 3–4 weeks before the use of TNF-α inhibitors. Clinical examinations of the patients and conventional laboratory studies were performed during control visits (1, 3, and 12 months after vaccination). The serum levels of anti-pneumococcal capsular polysaccharide antibodies were measured in 102 patients by enzyme immunoassay using commercial VaccZymeTM Anti-PCP IgG Enzyme Immunoassay kits (The Binding Site Group Ltd, United Kingdom). Results and discussion. No clinical and radiological symptoms of pneumonia were recorded in any case during the follow-up period of 12 months. The patients with RD and the control group showed a significant, more than double increase in anti-pneumococcal antibodies 12 months following vaccination. Vaccination was well tolerated: 90 (68%) patients displayed no adverse events; 37 (28%) had pain, cutaneous swelling and hyperemia up to 2 cm in diameter at the site of injection for vaccination;6 (4%) had low-grade fever. There were no episodes of a RD exacerbation or any new autoimmune disorders during the follow-up period. Conclusion. The findings were suggestive of the sufficient immunogenicity and good tolerability of 23-valent pneumococcal vaccine in patients with RD.

Evaluation and validation of a serum bactericidal antibody assay for Haemophilus influenzae type b and the threshold of protection

Prior to routine immunisation, Haemophilus influenzae serotype b (Hib) was a major cause of serious bacterial infections, particularly in young children. In the United Kingdom, introduction of the Hib conjugate vaccine into the national childhood immunisation schedule has led to a sustained decline in invasive Hib disease across all age-groups. Evaluation of the immune response to Hib conjugate vaccines involves measurement of serum IgG antibodies against the capsular polyribosyl-ribitol-phosphate (PRP) polysaccharide by enzyme-linked immunosorbent assay (ELISA), with accepted short-term and long-term protective thresholds of ≥0.15μg/mL and ≥1.0μg/mL, respectively. These levels were derived by passive immunisation or immunisation with pure polysaccharide, and their relevance for protection following immunisation with conjugate vaccines remains unclear. This study aimed to modify and optimise a serum bactericidal antibody (SBA) assay to evaluate the functional activity of Hib antibodies generated following Hib conjugate vaccination. Validation of the Hib SBA assay was deemed acceptable for all assay parameters tested. A strong correlation between anti-PRP IgG concentrations and SBA titres was observed in vaccinated adults (r=0.81), as well as infants after primary immunisation at 2, 3, and 4 months (r=0.635) and after the 12-month booster (r=0.746). The assay identified some children with high anti-PRP IgG but low SBA activity and vice versa. The predictive protective SBA titre corresponding to a post-booster anti-PRP IgG of 1.0μg/mL was 8. Thus, the optimised Hib SBA assay was specific and reproducible and correlated with anti-PRP IgG. Such assays may have a role in evaluating immune responses to conjugate vaccines in addition to measuring capsular antibodies.

Colloidal gold-based immunochromatographic strip test compromising optimised combinations of anti-S. suis capsular polysaccharide polyclonal antibodies for detection of Streptococcus suis

A rapid diagnosis kit that detects Streptococcus suis (S. suis) antigens from urine with an immunochromatographic stripe (ICS) test was developed in this study. The ICS test was produced using colloidal gold coated with polyclonal antibodies (pAbs) against S. suis. The pAbs were developed from rabbits immunised with S. suis serotype 2 capsular polysaccharides (CPS). Development of the pAbs was investigated to establish their binding to CPS and to determine the maximum sensitivity of two combination antibodies for the ICS test. The results of the ICS optimisation revealed that the combinations of pAb C-N1 and pAb C-N2 had the highest sensitivity to CPS. The minimum limitation of ICS sensitivity indicated 1.0×104 colony forming units (CFU) and a CPS concentration of 0.05µg. The assay time for detection of S. suis antigens is less than 15min, which is suitable for rapid detection. A cross-reactive test was also conducted, and it detected no other bacteria (Streptococcus pneumoniae, Streptococcus agalactiae, Staphylococcus aureus, Enterococcus faecalis, Escherichia coli, Pseudomonas aeruginosa, and Klebsiella pneumoniae). The cross-reactivity of other serotypes in S. suis was also investigated, and tests for serotypes of 1, 1/2, 3, 4, 5, 6, 7, 8, 9, 14, and 16 were positive. This study presents the first report of a development of an ICS that enables the quantitative detection of streptococcal antigens. The S. suis ICS provides several advantages over other methods, including the speed and simplicity of use.

A Burkholderia pseudomallei Outer Membrane Vesicle Vaccine Provides Protection against Lethal Sepsis

The environmental Gram-negative encapsulated bacillus Burkholderia pseudomallei is the causative agent of melioidosis, a disease associated with high morbidity and mortality rates in areas of Southeast Asia and northern Australia in which the disease is endemic. B. pseudomallei is also classified as a tier I select agent due to the high level of lethality of the bacterium and its innate resistance to antibiotics, as well as the lack of an effective vaccine. Gram-negative bacteria, including B. pseudomallei, secrete outer membrane vesicles (OMVs) which are enriched with multiple protein, lipid, and polysaccharide antigens. Previously, we demonstrated that immunization with multivalent B. pseudomallei-derived OMVs protects highly susceptible BALB/c mice against an otherwise lethal aerosol challenge. In this work, we evaluated the protective efficacy of OMV immunization against intraperitoneal challenge with a heterologous strain because systemic infection with phenotypically diverse environmental B. pseudomallei strains poses another hazard and a challenge to vaccine development. We demonstrated that B. pseudomallei OMVs derived from strain 1026b afforded significant protection against septicemic infection with B. pseudomallei strain K96243. OMV immunization induced robust OMV-, lipopolysaccharide-, and capsular polysaccharide-specific serum IgG (IgG1, IgG2a, and IgG3) and IgM antibody responses. OMV-immune serum promoted bacterial killing in vitro, and passive transfer of B. pseudomallei OMV immune sera protected naive mice against a subsequent challenge. These results indicate that OMV immunization provides antibody-mediated protection against acute, rapidly lethal sepsis in mice. B. pseudomallei-derived OMVs may represent an efficacious multivalent vaccine strategy against melioidosis.

Maternal pneumococcal capsular IgG antibodies and transplacental transfer are low in South Asian HIV-infected mother-infant pairs

BackgroundOur understanding of the mother-to-child transfer of serotype-specific pneumococcal antibodies is limited in non-immunized, HIV-positive women. MethodsWe compared geometric mean antibody concentrations (GMCs), geometric mean transplacental cord:maternal ratios (GMRs) and proportions of samples with protective antibody concentration (≥0.35μg/ml) to serotypes 1, 4, 5, 6B, 9V, 14, 18C, 19F, 23F between 74 HIV-infected and 98 HIV-uninfected mother-infant pairs who had not received pneumococcal immunization in South Asia. Multivariable analysis was performed to assess the influence of HIV on protective antibody concentrations. ResultsHIV-infected mothers and their infants exhibited lower GMCs and GMRs than their uninfected counterparts. This was significant for all serotypes except maternal GMC to serotype 1 and GMR for serotype 6B. In multivariate analysis, HIV was significantly associated with reduced odds of having protective pneumococcal IgG levels; 56–73% reduction for 3 maternal serotypes (4, 5, 23F) and 62–90% reduction for all cord samples except serotype 6B. ConclusionsMaternal HIV infection is associated with lower levels of maternal pneumococcal antibodies and disproportionately lower cord antibodies, relative to maternal antibodies, suggesting that HIV infection compromises transplacental transfer. Reassessment of maternal and/or infant pneumococcal immunization strategies is needed in HIV-infected women and their infants.

The differential impact of coadministered vaccines, geographic region, vaccine product and other covariates on pneumococcal conjugate vaccine immunogenicity.

Background:Antipneumococcal capsular polysaccharide antibody concentrations are used as predictors of vaccine efficacy against vaccine serotype (ST) pneumococcal disease among infants. While pneumococcal conjugate vaccines (PCV) are recommended globally, factors associated with optimal PCV immune response are not well described. We aimed to systematically assess local setting factors, beyond dosing schedule, which may affect PCV antibody levels.Methods:We conducted a literature review of PCV immunogenicity, abstracting data from published reports, unpublished sources, and conference abstracts from 1994 to 2010 (and ad hoc 2011 reports). Studies included in this analysis evaluated ≥ 2 primary doses of PCV before 6 months of age in non–high-risk populations, used 7-valent or higher PCV products (excluding Aventis-Pasteur and Merck products) and provided information on geometric mean concentration (GMC) for STs 1, 5, 6B, 14, 19F or 23F. Using random effects meta-regression, we assessed the impact of geographic region, coadministered vaccines and PCV product on postprimary GMC, adjusting for dosing schedule and ELISA laboratory method.Results:Of 12,980 citations reviewed, we identified 103 vaccine study arms for this analysis. Children in studies from Asia, Africa and Latin America had significantly higher GMC responses compared with those in studies from Europe and North America. Coadministration with acellular pertussis DTP compared with whole-cell DTP had no effect on PCV immunogenicity except for ST14, where GMCs were higher when coadministered with acellular pertussis DTP. Vaccine product, number of PCV doses, dosing interval, age at first dose and ELISA laboratory method also affected the GMC.Conclusions:PCV immunogenicity is associated with geographic region and vaccine product; however, the associations and magnitude varied by ST. Consideration of these factors is essential when comparing PCV immunogenicity results between groups and should be included in the evidence base when selecting optimal PCV vaccine schedules in specific settings.

Development and Characterization of a Multiplex Bead-Based Immunoassay To Quantify Pneumococcal Capsular Polysaccharide-Specific Antibodies

Enzyme-linked immunosorbent assay (ELISA), the traditional antibody quantification technique, has several limitations, especially when used to evaluate multivalent and/or infant vaccines. We have developed a multiplex bead-based antibody quantification assay (MBIA) to measure antibody response to multiple pneumococcal (Pn) serotypes (St) in a single assay. MBIA was compared with the WHO ELISA using a WHO panel of 12 international calibration sera for 7 Pn Sts. An agreement of 75 to 92% was obtained for all 7 Sts. MBIA exhibited good robustness, with the assay variability at ≤ 16%. A major contributor to MBIA variability was the cell wall polysaccharide (CWPs) content in Pn St-specific capsular Ps. This necessitated careful CWPs (20 μg/ml) preadsorption of sera. MBIA is specific, robust, and reproducible and offers high throughput. The use of MBIA will greatly reduce the cost and time required to evaluate the immune response to multiple Pn Sts and could help promote the licensure of future Pn and other multivalent vaccines.

Serotype-independent pneumococcal experimental vaccines that induce cellular as well as humoral immunity

For prevention of Streptococcus pneumoniae (pneumococcus) infections in infancy, protein-conjugated capsular polysaccharide vaccines provide serotype-specific, antibody-mediated immunity but do not cover all of the 90+ capsule serotypes. Therefore, microbiologists have sought protective noncapsular antigens common to all strains. Alternatively, we investigated killed cells of a noncapsulated strain, which expose many such common antigens. Given to mice intranasally, this vaccine elicits antibody-independent, CD4+ T lymphocyte-dependent accelerated clearance of pneumococci of various serotypes from the nasopharynx mediated by the cytokine IL-17A. Such immunity may reproduce the natural resistance that develops in infants before capsular antibodies arise. Given by injection, the killed cell vaccine induces bifunctional immunity: plasma antibodies protective against fatal pneumonia challenge, as well as IL-17A–mediated nasopharyngeal clearance. Human testing of this inexpensive candidate vaccine by intramuscular injection is planned. Bacterial cellular vaccines are complex—a challenge for reproducibility. However, when several known protective antigens were deleted, the killed pneumococcal vaccine was still protective. This antigenic redundancy may prevent vaccine escape variants by recombinational loss, which is frequent in pneumococcus. Biochemically defined immunogens with bifunctional activity have also been devised. These immunogens are three-component conjugates in which cell wall teichoic acid (a common antigen capable of T cell activation) is coupled to a genetic fusion of two common pneumococcal proteins: a protective surface antigen and a derivative of pneumolysin, which provides TLR4 agonist activity and induces antitoxic immunity. Such constructs induce accelerated clearance when given intranasally and induce both immune mechanisms when injected. The defined composition permits analysis of structure-function activity.

A Serotype 3 Pneumococcal Capsular Polysaccharide-Specific Monoclonal Antibody Requires Fcγ Receptor III and Macrophages To Mediate Protection against Pneumococcal Pneumonia in Mice

Antibodies to pneumococcal capsular polysaccharide (PPS) are required for PPS-based vaccine-mediated protection against Streptococcus pneumoniae. Previous work established that 1E2, a mouse IgG1 to PPS3 that does not induce serotype 3 (ST3) S. pneumoniae killing by phagocytes in vitro, protects mice from death after intranasal infection with ST3, but its efficacy was abrogated in FcγR (F common gamma receptor)-deficient mice. In this study, we determined whether 1E2 efficacy against pulmonary ST3 infection requires FcγRIII. 1E2 did not protect FcγRIII-deficient (FcγRIII(-/-)) mice. Studies of the mechanism of 1E2-mediated effects showed that it resulted in a marked reduction in lung inflammation in ST3-infected wild-type (Wt [C57BL/6]) mice that was abrogated in FcγRIII(-/-) mice. 1E2 had no effect on early bacterial clearance in the lungs of ST3-infected Wt, FcγRIIB(-/-), or FcγRIII(-/-) mice, but it reduced levels of bacteremia and serum macrophage inflammatory protein-2) (MIP-2), interleukin-6 (IL-6), and tumor necrosis factor alpha (TNF-α) in Wt and FcγRIIB(-/-) mice, strains in which it is protective. As previous work showed that neutrophils were dispensable for 1E2 efficacy, we investigated whether macrophages are required for 1E2 efficacy against intranasal infection with ST3 and found that its efficacy was abrogated in Wt mice depleted of macrophages intranasally. In vitro studies revealed that1E2 promoted ST3 internalization by naïve alveolar macrophages but did not induce early intracellular killing. Macrophages from 1E2-treated ST3-infected mice studied ex vivo exhibited more apoptosis than those from FcγRIII(-/-) mice. These findings suggest that 1E2 mediates protection against ST3 in mice by affecting the inflammatory response, perhaps in part via macrophage apoptosis, rather than by inducing early bacterial clearance.

Identification of Klebsiella Pneumoniae by Capsular Polysaccharide Polyclonal Antibodies

The aim of this study is to develop simple and rapid diagnostic method which can be utilized routinely for detection of Klebsiella pneumoniae. This study was done in India at DRDE, Gwalior. We have collected standard K.pneumoniae strain 3296 from Yamaguchi University, Japan and total fifty-nine clinical isolates of Klebsiella species from Armed Force Medical College, Pune, and Patel chest hospital, New Delhi. Bacteria were grown on trypticase soy broth and two Newzealand wistar white rabbit were immunized subcutaneously with purified capsular polysaccharide (CPS) of K.pneumoniae. Specificity of hyper immune sera raised against CPS was tested by counter current immunoelectrophoresis. We have generated four monoclonal antibodies, out of which two IgM producing clones (KP-1 & KP-2) and hyper immune sera against CPS antigen were used for agglutination test. KP-1, KP-2 and CPS hyperimmune sera showed different reactions. Out of the twenty biochemically confirmed K.pneumoniae clinical isolates, hyperimmune sera to CPS detected sixteen as positive whereas KP-1 reacted to eleven and KP-2 to thirteen of these isolates. One clinical Klebsiella species biochemically negative for K.pneumoniae and other Enterobacteriaceae organisms were negative by agglutination test to all the three antibody reagents. K.aerogenes, however, reacted to KP-2 and CPS hyperimmune sera. Standard K.pneumoniae was positive to all the three antibodies. The rapidity and ease of performance merited the hyper immune sera to CPS and IgM monoclonal antibody based agglutination test as the preliminary identification system for K.pneumoniae.

The efficacy of pneumococcal capsular polysaccharide-specific antibodies to serotype 3 Streptococcus pneumoniae requires macrophages

The efficacy of antibody immunity against Streptococcus pneumoniae stems from the ability of opsonic, serotype (ST)-specific antibodies to pneumococcal capsular polysaccharide (PPS) to facilitate killing of the homologous ST by host phagocytes. However, PPS-specific antibodies have been identified that are protective in mice, but do not promote opsonic killing in vitro, raising the question of how they mediate protection in vivo. To probe this question, we investigated the dependence of antibody efficacy against lethal systemic (intraperitoneal, i.p.) infection with Streptococcus pneumoniae serotype 3 (ST3) on macrophages and neutrophils for the following PPS3-specific monoclonal antibodies (MAbs) in survival experiments in mice using a non-opsonic human IgM (A7), a non-opsonic mouse IgG1 (1E2) and an opsonic mouse IgG1 (5F6). The survival of A7- and PPS3-specific and isotype control MAb-treated neutrophil-depleted and neutrophil-sufficient and macrophage-depleted and macrophage-sufficient mice were determined after i.p. challenge with ST3 strains 6303 and WU2. Neutrophils were dispensable for A7 and the mouse MAbs to mediate protection in this model, but macrophages were required for the efficacy of A7 and optimal mouse MAb-mediated protection. For A7-treated mice, macrophage-depleted mice had higher blood CFU, cytokines and peripheral neutrophil levels than macrophage-sufficient mice, and macrophage-sufficient mice had lower tissue bacterial burdens than control MAb-treated mice. These findings demonstrate that macrophages contribute to opsonic and non-opsonic PPS3-specific MAb-mediated protection against ST3 infection by enhancing bacterial clearance and suggest that neutrophils do not compensate for the absence of macrophages in the model used in this study.

Adenoidectomy and nasopharyngeal carriage of Streptococcus pneumoniae in young children

ObjectiveThe effect of adenoidectomy on nasopharyngeal colonisation of pathogens has not previously been evaluated. The authors studied the effect of adenoidectomy on nasopharyngeal colonisation by bacteria causing otitis media and...

Safety and Immunogenicity of a 13-Valent Pneumococcal Conjugate Vaccine Compared to Those of a 7-Valent Pneumococcal Conjugate Vaccine Given as a Three-Dose Series with Routine Vaccines in Healthy Infants and Toddlers

A 13-valent pneumococcal conjugate vaccine (PCV13) has been developed to improve protection against pneumococcal disease beyond that possible with the licensed 7-valent vaccine (PCV7). This study compared the safety and immunogenicity of PCV13 with those of PCV7 when given as part of the pediatric vaccination schedule recommended in Italy. A total of 606 subjects were randomly assigned to receive either PCV13 or PCV7 at 3, 5, and 11 months of age; all subjects concomitantly received diphtheria-tetanus-acellular pertussis-hepatitis B-inactivated polio-Haemophilus influenzae type B (DTaP-HBV-IPV/Hib) vaccine. Vaccine reactions were monitored. Antibody responses to DTaP-HBV-IPV/Hib antigens, serotype-specific anticapsular polysaccharide IgG responses, and antipneumococcal opsonophagocytic assay (OPA) activity were measured 1 month after the two-dose primary series and 1 month after the toddler dose. Overall, the safety profile of PCV13 was similar to that of PCV7. The response to DTaP-HBV-IPV/Hib antigens was substantially the same with both PCV13 and PCV7. PCV13 elicited antipneumococcal capsular IgG antibodies to all 13 vaccine serotypes, with notable increases in concentrations seen after the toddler dose. Despite a lower immunogenicity for serotypes 6B and 23F after the primary series of PCV13, responses to the seven common serotypes were comparable between the PCV13 and PCV7 groups when measured after the toddler dose. PCV13 also elicited substantial levels of OPA activity against all 13 serotypes following both the infant series and the toddler dose. In conclusion, PCV13 appeared comparable to PCV7 in safety profile and immunogenicity for common serotypes, demonstrated functional OPA responses for all 13 serotypes, and did not interfere with immune responses to concomitantly administered DTaP-HBV-IPV/Hib vaccine.

Elderly subjects do not show impaired pneumococcal capsular polysaccharide serotype-specific antibody responses as assessed by a multiplexed bead assay

Elderly subjects do not show impaired pneumococcal capsular polysaccharide serotype-specific antibody responses as assessed by a multiplexed bead assay

Phenotypic and genomic characterization of pneumococcus-like streptococci isolated from HIV-seropositive patients

Accurate differentiation between pneumococci and other viridans streptococci is essential given their differences in clinical significance. However, classical phenotypic tests are often inconclusive, and many examples of atypical reactions have been reported. In this study, we applied various phenotypic and genotypic methods to discriminate between a collection of 12 streptococci isolated from the upper respiratory tract of HIV-seropositive individuals in 1998 and 1999. Conventional phenotypic characterization initially classified these streptococci as Streptococcus pneumoniae, as they were all sensitive to optochin and were all bile soluble. However, they did not agglutinate with anti-pneumococcal capsular antibodies and were also far more resistant to antimicrobial agents than typeable pneumococci isolated in the same period. Genotypic characterization of these isolates and control isolates by both multilocus sequence analysis (MLSA) and comparative genomic hybridization (CGH) showed that only a single isolate was genetically considered to be a true S. pneumoniae isolate, and that the remaining 11 non-typable isolates were indeed distinct from true pneumococci. Of these, 10 most closely resembled a subgroup of Streptococcus mitis isolates genetically, while one strain was identified as a Streptococcus pseudopneumoniae isolate. CGH also showed that a considerable part of the proposed pneumococcal core genome, including many of the known pneumococcal virulence factors, was conserved in the non-typable isolates. Sequencing of part of the 16S rRNA gene and investigation for the presence of ply by PCR corroborated these results. In conclusion, our findings confirm the close relationship between streptococci of the Mitis group, and show that both MLSA and CGH enable pneumococci to be distinguished from other Mitis group streptococci.

Response Is Seen to Conjugate Vaccine but Not to Polysaccharide Vaccine Twelve Months After Rituximab.

Abstract 2728Poster Board II-704 Background:After therapy with the monoclonal antibody anti-CD20 rituximab ® (R), normal B-cells are depleted and their recovery in blood is delayed until six-twelve months in patients with relapsed low-grade lymphoma treated with four infusions of R. In patients treated with R-chemotherapy or high-dose therapy after R-purged autologous graft, the B-cell recovery is postponed further. The efficacy of active immunization against various bacterial and viral infections of R- treated patients has been debated as total level of immunoglobulin (Ig) M has been shown to decrease during prolonged treatment with R and humoral T cell-independent response might be defect due to lack of B-cells. Aim:To evaluate antibody-response to two vaccines after two timepoints of vaccination. Methods:An open randomized study of patients with B-cells lymphoma vaccinated six or twelve month after last treatment with R (Mabthera® Hoffman-LaRoche, Basel, Switzerland) with or without chemotherapy. Pneumococcal polysaccharide vaccine, Pneumo23® (Sanofi Pasteur MSD), assessed T-cell–independent antibody response and conjugated H. influenza type b-vaccine, Act-Hib® (Sanofi Pasteur MSD), evaluated combined B-cell/T-cell pathway. The patients randomized to vaccination at six months and without response, were revaccinated after further six months. Blood samples were collected immediately before vaccination and four weeks after. Serum was separated, frozen to −20 degrees C, sent to the Division of Microbiology, Statens Seruminstitut, Copenhagen, Denmark for analyses. A modified enzyme-linked assay for measuring type-specific anti-pneumococcal capsular polysaccharide antibodies for 6 pneumococcal serotypes (1, 4, 7F, 14, 18C, 19F) was used and a geometrical mean value for the total titer of antibody was calculated. The definition of protective response was determined to >39 arbitrary units/ml. Total Ig antibodies to the capsular polysaccharide of Haemophilus influenzae type b (HbPs) was measured by ELISA. An anti-HbPs antibody titer of >1 μg/ml was recorded as a protective response after immunization. Blood samples for flow cytometry analysis, including CD19+ and CD20+ B-cell counts, were drawn at the time of vaccination and four weeks after. Results:Nine patients, age 56–73 years, six females and three males, were randomized to vaccination; four at six months and five at twelve months after last rituximab treatment. Five patients with follicular lymphoma (FL) grade 2 and one with marginalzone lymphoma (MZL) had received single R, one of these in combination with alfa-interferon in a clinical phase II trial. One patient with mantle cell lymphoma (MCL) and one with follicular centroblastic lymphoma were treated with R-CHOP-21 × 8 prior to vaccination. One female patient with MZL randomized to vaccination at twelve months, dropped out because of progression of disease. Vaccination with Pneumo23® resulted in a protective response of pneumococcal antibodies in only one out of eight (12,5 %) patients. This responding patient had MZL treated with R 375 mg weekly x4 during 2 courses, and was randomized to vaccination 6 months after last R and also showed a response to Act-Hib vaccine. Further four patients had an immunization response to act Hib vaccine, one with FL randomized to vaccination six months after R and three after twelve months, two with FL and one with MCL. One patient was revaccinated after twelve months and had no response neither with Pneumo23® or Act-Hib® vaccine. The percentage CD19+ and CD20 + B-cells in the lymphocyte gate (by flow cytometry) was low in all patients at the time of vaccination and four weeks after, but showed a slight increase with time (0,04, 0,26, 0,42, 0,54 % and 0,24, 0,6, 0,6, 1,26, 1,0 % at six and twelve months after R, respectively). Conclusion:Conjugate vaccine is more potent to give immune response than polysaccharide vaccine after R-therapy in patients with B cell lymphoma. Despite deficient recovery of B-cells cells, five of eight (62,5 %) patients responded to the conjugate vaccine Act-Hib®. The longstanding effect on the immune system of antibody therapy has to be further studied. Disclosures:No relevant conflicts of interest to declare.

Kinetics of the Natural, Humoral Immune Response toSalmonella entericaSerovar Typhi in Kathmandu, Nepal

Typhoid fever is a major public health problem in developing countries, conservatively estimated to occur in 17 million cases and be responsible for 200,000 deaths annually. We investigated the acquisition of natural immunity to Salmonella enterica serovar Typhi in a region where typhoid is endemic by testing sera from an age-stratified sample of 210 healthy participants in Kathmandu, Nepal, for bactericidal activity toward S. Typhi and for anti-Vi capsular polysaccharide antibodies. Bactericidal titers in children were significantly lower than those in newborns and adults (P < 0.0001). Anti-S. Typhi bactericidal geometric mean titers were age dependent, increasing 10-fold during childhood. Anti-Vi polysaccharide antibody geometric mean concentrations were also lower in children than in adults. Data presented here indicate the possibility of a relationship between low levels of bactericidal activity toward S. Typhi in serum and susceptibility to disease, as observed for other polysaccharide-encapsulated bacteria. Bactericidal antibody may be a marker of protective immunity against S. Typhi.