Polysaccharide-protein Conjugate Vaccines Research Articles

Humoral and cellular immune responses induced by serogroup W135 meningococcal conjugate and polysaccharide vaccines

Investigating the mechanisms by which W135 meningococcal conjugate (PSW135-TT) activates adaptive immune responses in mice can provide a comprehensive understanding of the immune mechanisms of bacterial polysaccharide conjugate vaccines. We compared B-cell and T-cell immune responses immunized with W135 meningococcal capsular polysaccharides (PSW135), tetanus toxoid (TT) and PSW135-TT in mice. The results showed that PSW135-TT could induce higher PSW135-specific and TT-specific IgG antibodies with a significant enhancement after two doses. All serum antibodies immunized with PSW135- TT had strong bactericidal activity, whereas none of the serum antibodies immunized with PSW135 had bactericidal activity. Besides, IgM and IgG antibodies immunized with PSW135-TT after two doses were positively correlated with the titer of bactericidal antibodies. We also found Th cells favored Th2 humoral immune responses in PSW135-TT, PSW135, and TT-immunized mice, especially peripheral blood lymphocytes. Furthermore, PSW135-TT and TT could effectively activate bone marrow derived dendritic cells (BMDCs) and promote BMDCs to highly express major histocompatibility complex Ⅱ (MHCⅡ), CD86 and CD40 molecules in mice, whereas PSW135 couldn’t. These data verified the typical characteristics of PSW135-TT and TT as T cell dependent antigen (TD-Ag) and PSW135 as T cell independent antigen (TI-Ag), which will be very helpful for further exploration of the immune mechanism of polysaccharide-protein conjugate vaccines and improvement of the quality of bacterial polysaccharide conjugate vaccines in future.

Streptococcus agalactiae serotype Ia capsular polysaccharide production from a Brazilian strain

The pathogen S. agalactiae, categorized as Lancefield group B streptococci (GBS), is a Gram-positive bacterium, encapsulated, anaerobic facultative bacillus, which occurs in pairs or short chains. It was responsible for more than 319,000 cases of infections in neonates. Early and late streptococci, in the course of an illness or medical condition, are infections that have high mortality and morbidity rates, with an incidence of 2.0 to 5.0 per 1000 live births in North America, Europe, and Australia. The infection of newborns usually happens through transmission from infected mothers during and after childbirth, with GBS being among the critical bacterial infections in the Brazilian epidemiological picture. S. agalactiae strains isolated from different regions of Brazil, indicate a great diversity among the isolated serotypes but with a higher incidence of serotype Ia. Publications report that, in pregnant women in Brazil, the prevalence of serotype Ia has been constant between 2002 and 2018, followed by serotypes II and IV. Serotypes Ib, III, and V also appear, but with some fluctuations in different regions of the country. The pathogen which causes invasive disease are usually surrounded by a polysaccharide capsule and is its major virulence factor, being that the key antigen in protective response triggered by polysaccharides and, more recently, protein-polysaccharide conjugate vaccines. The development of antibiotic resistance by these microorganisms to different antibiotics has been the guideline in leading therapeutic options against these infections including the use of prophylactic vaccines in pregnant women, immunosuppressed or older people. Such prophylactic alternatives can be constructed from capsular polysaccharides.Based on the experience in the production and purification of Neisseria meningitidis polysaccharides for vaccines, Bio-Manguinhos developed a process for the production, purification and characterization of S. agalactiae Ia polysaccharides. In the present study, the capsular polysaccharide from the strain isolated in clinical material of a public hospital, in Rio de Janeiro – Brazil, was obtained from simple batch cultures, with purification process achieving about 20 mg g−1 in yield for cell dry weight (CDW) and ca. of 100 % of purity. Hence, to prove that the CPS of GBS Ia can be consistently produced, purified, and characterized by NMR, HRGC-MS, and SEC-MALS – from such strain – to be used in vaccine productions against S. agalactiae.

A novel approach to determine residual aldehyde content in conjugated polysaccharides with 3-methyl-2-benzothiazolinonehydrazon (MBTH) by colorimetric method, automation, and HP-SEC

Polysaccharide activation by periodate oxidation to form aldehydes, followed by conjugation with proteins via reductive amination is a general procedure to make polysaccharide-protein conjugate vaccines. Controlling the level of residue aldehyde content after conjugation is critical to ensure vaccine product stability. Herein, to support conjugation process optimization, we developed a 3-methyl-2-benzothiazolone hydrazone (MBTH) chemistry-based colorimetric method, which can measure low-level residual aldehyde polysaccharide-protein conjugate in high throughput 96-well plate format. This mechanism of detection works in two steps. First, MBTH reacts with the aldehyde group to form azine. Then the excess MBTH was oxidized by ferric ammonium sulfate in sulfamic acid to form a reactive cation, which reacts further with azine to form formazan, a characteristic blue chromophore at 610 nm, for colorimetric detection. The method performance, including detection limit, linearity range, matrix effect, kinetics, and color stability was systematically studied. For samples with severe matrix interference, a supplemental size exclusion chromatography (SEC) method was also developed as an alternative method.

Prognoses of patients undergoing hemodialysis administered 23-valent pneumococcal polysaccharide versus 13-valent pneumococcal protein conjugate vaccines

IntroductionSequential vaccination with the 13-valent pneumococcal protein conjugate vaccine (PCV13) and 23-valent pneumococcal polysaccharide vaccine (PPSV23) is recommended for patients undergoing hemodialysis; however, evidence for the efficacy of these pneumococcal vaccines for patients undergoing hemodialysis is limited to a single dose. We aimed to evaluate the prognosis of patients undergoing hemodialysis who received vaccination with PPSV23 alone versus sequential vaccination with PCV13 and PPSV23. MethodsPatients undergoing hemodialysis who were vaccinated with PPSV23 alone (PPSV23 group) or PCV13 followed by PPSV23 (PCV13+PPSV23 group) between 2014 and 2016 were included; the observation period was three years from the first injection. Patients who underwent hemodialysis between 2011 and 2012 were included as controls. After propensity score matching using age, sex, dialysis vintage, diabetes history, pneumonia history, and serum albumin and creatinine levels, survival analysis was performed. ResultsThe study included 89, 71, and 319 patients in the PPSV23, PCV13+PPSV23, and control groups, respectively. After propensity score matching, the PPSV23 and control group 1 (79 patients each) and the PCV13+PPSV23 and control group 2 (61 patients each) were compared. Significant differences were observed in the survival rate between the PPSV23 group and control group 1 (p = 0.005) but not between the PCV13+PPSV23 group and control group 2. Pneumonia-related mortality in the two vaccinated groups did not differ significantly during the observation period. ConclusionsPatients who received PPSV23 had a favorable prognosis; however, no positive effect was demonstrated in the PCV13+PPSV23 group.

Individual polysaccharide quantification in polyvalent pneumococcal conjugate vaccine using rate nephelometry

The multivalent pneumococcal conjugate vaccine (PCV) contains purified polysaccharides of different serotypes conjugated to a carrier protein. Testing the final formulated product for individual serotype polysaccharide content is critical in vaccine quality control which requires an assay specific to each serotype polysaccharide present in the formulated product. Antibodies specific to the serotypes specific polysaccharide were used in rate nephelometry assay for quantifying individual serotype polysaccharides in the formulated vaccine. Generally, native polysaccharide (NP) have been used as reference standard. However, the polysaccharide antigen in the vaccine product is in the conjugate form (CRM197 linked) and hence using NP as a reference standard may not be suitable. Activated quenched polysaccharide (AQP) as a reference standard in rate nephelometry would be more appropriate. The epitope structure of AQP closely represents the polysaccharide-protein conjugate drug product (DP) after trypsin digestion. Hence, AQP was evaluated as a novel reference standard for the accurate and precise determination of individual polysaccharides in the multivalent DP. Rate nephelometry assay using AQP could be used for DP release and stability for monitoring time-dependent changes in the product and establishing the shelf life. A similar strategy could be applied to test and release monovalent or multivalent polysaccharide-protein conjugate vaccines (Meningococcal, Haemophilus influenza Type B, Typhoidal, and non-typhoidal salmonella).

Early-onset group B streptococcal disease in African countries and maternal vaccination strategies.

Invasive group B streptococcal (GBS) disease is the commonest perinatally-acquired bacterial infection in newborns; the burden is higher in African countries where intrapartum antibiotic prophylaxis strategies are not feasible. In sub-Saharan Africa, almost one in four newborns with GBS early-onset disease will demise, and one in ten survivors have moderate or severe neurodevelopmental impairment. A maternal GBS vaccine to prevent invasive GBS disease in infancy is a pragmatic and cost-effective preventative strategy for Africa. Hexavalent polysaccharide protein conjugate and Alpha family surface protein vaccines are undergoing phase II clinical trials. Vaccine licensure may be facilitated by demonstrating safety and immunological correlates/thresholds suggestive of protection against invasive GBS disease. This will then be followed by phase IV effectiveness studies to assess the burden of GBS vaccine preventable disease, including the effect on all-cause neonatal infections, neonatal deaths and stillbirths.

Impact of aluminum adjuvants on the stability of pneumococcal polysaccharide-protein conjugate vaccines

Development of a vaccine drug product requires formulation optimization to ensure that the vaccine’s effectiveness is preserved upon storage throughout the shelf-life of the product. Although aluminum adjuvants have been widely used in vaccine formulations to safely and effectively potentiate an immune response, careful attention must be directed towards ensuring that the type of aluminum adjuvant does not impact the stability of the antigenic composition. PCV15 is a polysaccharide-protein conjugate vaccine comprising the pneumococcal polysaccharide (PnPs) serotypes (1, 3, 4, 5, 6A, 6B, 7F, 9V, 14, 18C, 19A, 19F, 22F, 23F and 33F), each individually conjugated to the protein carrier CRM197. PCV15 was formulated with either amorphous aluminum hydroxyphosphate sulfate adjuvant (AAHS) or aluminum phosphate adjuvant (AP) and examined for both stability and immunogenicity. Using a collection of methods to evaluate vaccine stability, it was discovered that certain PCV15 serotypes (e.g., 6A, 19A, 19F) formulated with AAHS resulted in a reduction of immunogenicity in vivo and a reduction in recoverable dose as tested by an in vitro potency assay. The same polysaccharide-protein conjugates formulated with AP were stable regarding all measures tested. Moreover, the reduction in potency of certain serotypes correlated with chemical degradation of the polysaccharide antigen caused by the aluminum adjuvant as measured by reducing polyacrylamide gel electrophoresis (SDS-PAGE), High-Pressure Size Exclusion Chromatography coupled with UV detection (HPSEC-UV) and ELISA immunoassay. This study suggests a formulation, which includes AAHS, may negatively impact the stability of a pneumococcal polysaccharide-protein conjugate vaccine that contains phosphodiester groups. This decrease in stability would likely result in a decrease in the “active” concentration of antigen dose, and herein, it is shown that such instability directly compromised vaccine immunogenicity in an animal model. The results presented in this study help to explain critical degradation mechanisms of pneumococcal polysaccharide-protein conjugate vaccines.

Strategies to Prevent Early and Late-Onset Group B Streptococcal Infection via Interventions in Pregnancy

Group B Streptococcus is a Gram-positive bacterium that typically colonizes 10-30% of pregnant women, causing chorioamnionitis, preterm birth, and stillbirth, as well as neonatal sepsis and meningitis with early-onset disease (EOD) or late-onset disease (LOD) due to ascending infection or transmission during delivery. While there are some differences between EOD and LOD in terms of route of transmission, risk factors, and serotypes, the only preventive approach currently is maternal intrapartum antibiotic prophylaxis (IAP) which will not be able to fully address the burden of the disease since this has no impact on LOD. Probiotics and immunization in pregnancy may be more effective than IAP for both EOD and LOD. There is mixed evidence of probiotic effects on the prevention of GBS colonization, and the data from completed and ongoing clinical trials investigating different GBS vaccines are promising. Current vaccine candidates target bacterial proteins or the polysaccharide capsule and include trivalent, tetravalent, and hexavalent protein-polysaccharide conjugate vaccines. Some challenges in developing novel GBS vaccines include the lack of a correlate of protection, the potential for serotype switching, a need to understand interactions with other vaccines, and optimal timing of administration in pregnancy to maximize protection for both term and preterm infants.

Analytical technology development to monitor the stability of Polysaccharide-Protein conjugate vaccines

The covalent attachment of a bacterial-derived capsular polysaccharide to protein is of critical importance in transforming the polysaccharide from an antigen with limited immunogenicity in infants and older adults to an antigen that can prevent potentially fatal disease. For a polysaccharide-protein conjugate vaccine (PCV) candidate to be successful, it must be sufficiently stable. Chemical breakage of carbohydrate bonds in the polysaccharide may result in the reduction of “conjugate dose” and could negatively impact immunogenicity and the ability of the vaccine to prime for memory responses. Therefore, development of analytical tools to monitor the integrity of a polysaccharide-protein conjugate (glycoconjugate) vaccine is of practical significance. In this work, reducing SDS-PAGE, Intrinsic Protein Fluorescence Spectroscopy (IPFS), Differential Scanning Fluorimetry (DSF) were evaluated methods to study the impact of time, temperature, and formulation composition on the stability of a glycoconjugate vaccine prepared by multisite coupling of polysaccharide to a carrier protein. In addition, an automated capillary Western system was also evaluated to study the impact of storage on glycoconjugate vaccine stability. Two streptococcus pneumoniae polysaccharide-protein conjugates (serotype 3 and serotype 19A) were chosen to examine their physicochemical stability when formulated as a single antigen vaccine. While all methods require only a small amount of test article and can test multiple samples per assay run, automated capillary Western has the additional advantage of being highly sensitive even at low concentrations in complex vaccine formulations that contain aluminum adjuvant and multiple antigens. Results suggest that automated capillary Western is stability-indicating and may be an effective analytical technology tool for the formulation development of a multivalent glycoconjugate vaccine.

A Mouse Immunogenicity Model for the Evaluation of Meningococcal Conjugate Vaccines.

The identification of an appropriate animal model for use in the development of meningococcal vaccines has been a challenge as humans are the only natural host for Neisseria meningitidis. Small animal models have been developed and are widely used to study the efficacy or immunogenicity of vaccine formulations generated against various diseases. Here, we describe the development and optimization of a mouse model for assessing the immunogenicity of candidate tetravalent meningococcal polysaccharide (MenACYW-TT) protein conjugate vaccines. Three inbred (BALB/c [H-2d], C3H/HeN [H-2k], or C57BL/6 [H-2b]) and one outbred (ICR [H-2g7]) mouse strains were assessed using serial two-fold dose dilutions (from 2 µg to 0.03125 µg per dose of polysaccharide for each serogroup) of candidate meningococcal conjugate vaccines. Groups of 10 mice received two doses of the candidate vaccine 14 days apart with serum samples obtained 14 days after the last dose for the evaluation of serogroup-specific anti-polysaccharide IgG by ELISA and bactericidal antibody by serum bactericidal assay (SBA). C3H/HeN and ICR mice had a more dose-dependent antibody response to all four serogroups than BALB/c and C57Bl/6 mice. In general, ICR mice had the greatest antibody dose-response range (both anti-polysaccharide IgG and bactericidal antibodies) to all four serogroups and were chosen as the model of choice. The 0.25 µg per serogroup dose was chosen as optimal since this was in the dynamic range of the serogroup-specific dose-response curves in most of the mouse strains evaluated. We demonstrate that the optimized mouse immunogenicity model is sufficiently sensitive to differentiate between conjugated polysaccharides, against unconjugated free polysaccharides and, to degradation of the vaccine formulations. Following optimization, this optimized mouse immunogenicity model has been used to assess the impact of different conjugation chemistries on immunogenicity, and to screen and stratify various candidate meningococcal conjugate vaccines to identify those with the most desirable profile to progress to clinical trials.

M174 HAEMOPHILUS INFLUENZAE TYPE B MENINGITIS – A CASE OF VACCINE FAILURE

Haemophilus influenzae is a pleomorphic gram-negative bacteria commonly found within the respiratory tract. Haemophilus influenzae serotype b (Hib) is a typeable (encapsulated) strain known to cause invasive disease. Hib polysaccharide-protein conjugate vaccines stimulate a highly immunogenic T-cell dependent response which has dramatically decreased the incidence of invasive disease. Here we present a case of Hib meningitis resulting in sensorineural hearing loss in an immunized infant, presumed to be secondary to vaccine failure.

Infant serotype specific anti-capsular immunoglobulin G antibody and risk of invasive group B Streptococcal disease

Past studies have mainly investigated the association of serotype-specific capsular IgG in the mother and risk reduction of invasive Group B Streptococcus (GBS) in their young infants. The efficiency of transplacental transfer of IgG could be affected by multiple maternal factors. Hence, investigation of infant serum GBS anti-capsular IgG and risk reduction for invasive GBS disease may be more robust and generalizable. In a matched case-control study, infant serum serotype-specific capsular polysaccharide Ia and III IgG concentrations were analyzed in infants with invasive GBS cases and healthy controls born to women with recto-vaginal colonization by the homotypic serotype. Using Bayesian modeling, an antibody concentration of 2.5 µg/mL and 1 µg/mL predicted a 90% reduced risk of invasive disease for serotype Ia and III, respectively. These data contribute to the possible licensure of a GBS polysaccharide-protein conjugate vaccine, targeted at pregnant women, based on serological correlates of protection against invasive GBS disease.

Saccharide dosage content of meningococcal polysaccharide conjugate vaccines determined using WHO International Standards for serogroup A, C, W, Y and X polysaccharides

Potency of meningococcal polysaccharide-protein conjugate vaccines relies on the polysaccharide content to prevent meningitis. NIBSC, as the official national control laboratory in UK, analysed ten different mono- and multi-meningococcal conjugate vaccines, using established International Standards for meningococcal serogroups A, C, W, Y and X, by resorcinol or HPAEC-PAD assay. Most saccharide contents were within ±20% of their claimed content for licensure with taking different O-acetylation levels into consideration, with only MenC content in two vaccines below (by 60% and 54%) the labelled value, however, previous study showed different dosage was not necessarily correlated to the immunogenicity of those vaccines. This study demonstrated the use of International Standards to quantify saccharide content in polysaccharide-based vaccines with different percentage of O-acetylation. These International Standards are suitable to serve as either quantitative standard or calibrator of in-house standards, with supplied stability data.

An affordable pneumococcal conjugate vaccine after 20 years

An affordable pneumococcal conjugate vaccine after 20 years

Association of Group B Streptococcus (GBS) Serum Serotype-Specific Anticapsular Immunoglobulin G Concentration and Risk Reduction for Invasive GBS Disease in South African Infants: An Observational Birth-Cohort, Matched Case-Control Study.

Licensure of a group B Streptococcus (GBS) polysaccharide-protein conjugate vaccine for protecting infants against invasive GBS disease (IGbsD) will likely need to be based on demonstrating vaccine safety in pregnant women, and benchmarking immunogenicity against a serological threshold associated with risk reduction of IGbsD. We investigated the association between naturally derived GBS serotype Ia and III IgG and risk reduction of IGbsD in infants ≤90 days of age. In a matched case-control study, IGbsD cases were identified from a cohort of 38 233 mother-newborn dyads. Mothers colonized vaginally with serotype Ia or III at birth and their healthy infants were eligible as matched controls. GBS serotype-specific anticapsular immunoglobulin G (IgG) was measured on maternal and cord blood/infant sera by multiplex Luminex assay, and the IgG threshold associated with 90% risk reduction of IGbsD was derived by estimating absolute disease risk. In infants born at ≥34 weeks' gestational age, cord-blood IgG geometric mean concentrations (GMCs) were lower in cases than controls for serotypes Ia (0.05 vs 0.50 µg/mL; P = .004) and III (0.20 vs 0.38 µg/mL; P = .078). Cord-blood IgG concentrations ≥1.04 and ≥1.53 µg/mL were associated with 90% risk reduction of serotype Ia and III IGbsD, respectively. The maternal sera IgG threshold associated with 90% risk reduction was ≥2.31 µg/mL and ≥3.41 µg/mL for serotypes Ia and III, respectively. The threshold associated with a reduced risk for serotype Ia and III IGbsD identified on infant sera supports the case for licensure of a GBS polysaccharide-protein conjugate vaccine based on an immunogenicity evaluation benchmarked against the defined thresholds. NCT02215226.

Activation of Soluble Polysaccharides with 1-Cyano-4-Dimethylaminopyridine Tetrafluoroborate (CDAP) for Use in Protein-Polysaccharide Conjugate Vaccines and Immunological Reagents. III Optimization of CDAP Activation.

CDAP (1-cyano-4-dimethylaminopyridine tetrafluoroborate) is employed in the synthesis of conjugate vaccines as a cyanylating reagent. In the published method, which used pH 9 activation at 20 °C (Vaccine, 14:190, 1996), the rapid reaction made the process difficult to control. Here, we describe optimizing CDAP activation using dextran as a model polysaccharide. CDAP stability and reactivity were determined as a function of time, pH and temperature. While the rate of dextran activation was slower at lower pH and temperature, it was balanced by the increased stability of CDAP, which left more reagent available for reaction. Whereas maximal activation took less than 2.5 min at pH 9 and 20 °C, it took 10–15 min at 0 °C. At pH 7 and 0 °C, the optimal time increased to >3 h to achieve a high level of activation. Many buffers interfered with CDAP activation, but DMAP could be used to preadjust the pH of polysaccharide solutions so that the pH only needed to be maintained. We found that the stability of the activated dextran was relatively independent of pH over the range of pH 1–9, with the level of activation decreased by 40–60% over 2 h. The use of low temperature and a less basic pH, with an optimum reaction time, requires less CDAP, improving activation levels while making the process more reliable and easier to scale up.

Meningococcal vaccines and protein-energy undernutrition in children in the African meningitis belt

BackgroundVaccines to prevent meningococcal meningitis in the African meningitis belt include PsACWY, a polysaccharide-only vaccine; and PsA-TT, a polysaccharide-protein conjugate vaccine. Protein-energy undernutrition, a condition where children do not receive enough macro- or micronutrients, is related to increased risk of infectious diseases and poor immune function. Reduced immune function could affect vaccine immunogenicity. We investigated connections between protein-energy undernutrition and vaccine immunogenicity and antibody waning to PsACWY and PsA-TT in children in the African meningitis belt. MethodsThis is a secondary analysis of data collected as part of four clinical trials testing the safety and efficacy of PsA-TT in children in Mali, Ghana, and Senegal. We identified whether anthropometric growth indices (low height-for-age, weight-for-height, or weight-for-age Z-score categories) were related to reduced vaccine-elicited antibody (measured with rabbit complement) from pre- to 1 month post-vaccination, in linear regression models. We also identified whether these growth indices were related to increased waning for vaccine-elicited antibody over time, in linear regression models. ResultsA total of 697 children were included in our analysis, of which 350 (50.2%) were female; the mean (SD) age was 1.0 (1.1) years, and 578 (83.0%) received PsA-TT. In linear regression models, no consistent statistical relationship was seen between pre-vaccination anthropometric Z-score categories and vaccine immunogenicity, or decline in antibody over time, for either vaccine, although children with low weight-for-height had a greater decline in antibody from 1 to 6 months post-vaccination. ConclusionsOur analysis did not find protein-energy undernutrition to be associated with immunogenicity or waning of PsACWY- or PsA-TT-elicited antibody in children living in the African meningitis belt. Future studies should consider measuring antibody titers at additional time points post-vaccination, and for longer periods of time, to determine if the rate of antibody waning over a period of several years is associated with protein-energy undernutrition.

Quantitation of novel pentavalent meningococcal polysaccharide conjugate vaccine (Men A-TT, Men C-CRM, Men Y-CRM, Men W-CRM, Men X-TT) using sandwich ELISA

Meningococcal vaccines have been developed over the years, to control outbreaks of meningitis. There are 12 immunologically distinct serogroups of which, A, B, C, W, Y and X are predominant and invasive in nature. Meningococcal vaccines can be sorted out as, polysaccharide vaccines, polysaccharide protein conjugate vaccines or protein based (independent of polysaccharides) vaccines. Stringent quality control analysis as per the regulatory guidelines is a requirement for any vaccine manufacturer for commercial release of vaccines. Quantitation becomes challenging when it comes to analysis of multivalent vaccines. We describe the quantitation of novel pentavalent meningococcal polysaccharide conjugate vaccine manufactured at Serum Institute of India Pvt Ltd (SIIPL). Briefly, sandwich ELISA assay was developed to quantitate five different serogroups (Men A-TT, Men C-CRM, Men Y-CRM, Men W-CRM, Men X-TT) in pentavalent meningococcal polysaccharide conjugate vaccine containing two different carrier proteins (TT and CRM). ELISA reported herein showed significant reproducibility and repeatability over a range of developed standard curve with acceptable coefficient of variation (<15%) for both intra and inter assay. Furthermore analysis showed that, polysaccharide conjugate (PC) contents were within the accepted range (±30%) of the stated label claim. The immunoassay was verified for its sensitivity, accuracy and precision. Based on the relevant experimental data; it is proposed that, reported sandwich ELISA is well suited for quantitation of individual polysaccharide conjugate from pentavalent formulation. Furthermore, the ELISA was explored for forced degradation and polysaccharide spiking studies. This leads to open up an area of research for sandwich ELISA as a tool to assess the integrity of vaccine as well as for lot to lot consistency monitoring.

Effect of HIV-exposure and timing of antiretroviral treatment initiation in children living with HIV on antibody persistence and memory responses to Haemophilus influenzae type b and pneumococcal polysaccharide-protein conjugate vaccines

BackgroundWe investigated the effect of in utero HIV-exposure, timing of antiretroviral treatment (ART) initiation, and ART interruption on memory responses and persistence of immunity induced by pneumococcal (PCV) and Haemophilus influenzae type b (HibCV) polysaccharide-protein conjugate vaccines. MethodsChildren were enrolled (6–12 weeks of age), and vaccinated with a three-dose primary series of 7-valent PCV (PCV7) and HibCV at 6, 10 and 14 weeks of age. Study groups included infants infected with HIV perinatally with CD4+ ≥ 25% initiating ART following immunological or clinical deterioration (ART-Def), or immediately upon enrolment followed by interruption at 40 (ART-Immed/40w) or 96 weeks (ART-Immed/96w); and HIV-uninfected infants with (HEU), and without HIV (HIV-unexpsoed) exposure in utero. Within each group, children were randomized to receive either a booster dose of PCV7 or HibCV at 15 months of age. PCV serotype-specific and polyribosyl ribitol phosphate (PRP) IgG were measured pre-boost, two-weeks post-boost and at two-years of age. Opsonophagocytic activity (OPA) to serotypes 9V, 19F and 23F was measured post-booster dose. ResultsPersistence of IgG to PCV vaccine–serotypes and anti-PRP was similar in all groups of children living with HIV (CLWH) compared to HIV-unexposed children. Anamnestic responses to PCV and HibCV were also similar in all three groups of CLWH compared to HIV-unexposed children. CLWH, however, tended to have lower functional antibody (OPA) titers than HIV-unexposed children after the PCV booster dose for some serotypes. Immunity to PCV and HibCV was similar between the ART-Immed/40w and ART-Immed-96w groups. There were no differences in IgG kinetics between HEU and HIV-unexposed children. ConclusionsA three dose primary series, with or without PCV or HibCV booster doses in CLWH initiated on ART during infancy, would likely be similarly effective in preventing invasive bacterial disease as in HIV-unexposed children.

Designing ecologically optimized pneumococcal vaccines using population genomics.

Streptococcus pneumoniae (the pneumococcus) is a common nasopharyngeal commensal that can cause invasive pneumococcal disease (IPD). Each component of current protein-polysaccharide conjugate vaccines (PCVs) generally induces immunity specific to one of the approximately 100 pneumococcal serotypes, and typically eliminates it from carriage and IPD through herd immunity. Overall carriage rates remain stable owing to replacement by non-PCV serotypes. Consequently, the net change in IPD incidence is determined by the relative invasiveness of the pre- and post-PCV-carried pneumococcal populations. In the present study, we identified PCVs expected to minimize the post-vaccine IPD burden by applying Bayesian optimization to an ecological model of serotype replacement that integrated epidemiological and genomic data. We compared optimal formulations for reducing infant-only or population-wide IPD, and identified potential benefits to including non-conserved pneumococcal carrier proteins. Vaccines were also devised to minimize IPD resistant to antibiotic treatment, despite the ecological model assuming that resistance levels in the carried population would be preserved. We found that expanding infant-administered PCV valency is likely to result in diminishing returns, and that complementary pairs of infant- and adult-administered vaccines could be a superior strategy. PCV performance was highly dependent on the circulating pneumococcal population, further highlighting the advantages of a diversity of anti-pneumococcal vaccination strategies.