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Capsular polysaccharides (CPSs) are important antigenic targets against bacterial infections. As T-independent antigens, however, CPSs elicit short-lived immune responses in adults and are poorly immunogenic in young children. Coupling CPS with protein carriers enhances anti-CPS responses and generates long-lasting immune memory. However, the mechanisms whereby carrier proteins accomplish this are not fully understood. Here, we dissect different mechanisms whereby carrier proteins enhance anti-CPS immunity. We show how coupling CPS with protein carriers modifies the interaction of CPS with antigen-presenting cells, enables a dual-activation mechanism for CPS-specific B cells via interaction with CPS- or carrier-specific T helper cells, and potentiates the recall of anti-CPS responses by engaging memory T helper cells during subsequent vaccination or bacterial exposure. Our findings provide new insights into the immunological basis of carrier-mediated anti-CPS immunity and may help in the design of more effective CPS-based vaccines. IMPORTANCE Polysaccharide capsules, the outermost shells of many bacterial pathogens, play a role in pathogenesis and protect bacteria against the immune system. Generating antipolysaccharide antibodies by vaccination has provided effective protection against infectious diseases caused by encapsulated bacteria. However, most pure polysaccharide preparations are poorly immunogenic, particularly in young children. To circumvent this problem, vaccines have been developed using polysaccharides associated with protein carriers. The precise mechanism whereby protein carriers enhance the immunogenicity of the polysaccharide remains unclear. The significance of our research is in elucidating the different roles played by carriers in facilitating polysaccharide processing and presentation, priming polysaccharide-specific B cells, and potentiating recall antipolysaccharide responses. Overall, our work provides new insights into the immunological basis of carrier-mediated antipolysaccharide immunity and may help in the design of more effective polysaccharide-based vaccines.

Lactobacillus strains can synthesize capsular polysaccharides (CPS), which are important substances in the dairy industry-they exhibit many important technological as well as health-promoting properties. Technological advancements have made it possible to detect bacterial capsules using costly and labor-intensive methods, such as serological reactions, molecular genetic techniques, and electron microscopy. Light microscopy, which is the method of interest in this paper, is one of the most widely accessible and cheapest techniques. CPS may be observed under a light microscope after staining bacterial cells and the background with a basic die and an acidic die, respectively (negative-positive staining), with the capsules remaining transparent. The literature offers many polysaccharide staining methods, but due to the considerable structural diversity of CPS and possible dye-capsule interactions, a suitable staining technique should be carefully selected for each strain. The current study showed that not all methods adequately reveal Lactobacillus CPS, with the most effective ones being those proposed by Hiss and Maneval.

Haemophilus influenzae type b (Hib) is a human pathogen that causes severe infections such as pneumonia, sepsis and meningitis. Vaccines for Hib infections are based on its capsular polysaccharide conjugated to a protein. This conjugated Hib antigen is included as one of the components of polyvalent vaccines and accounts for more than 50% of the total cost of the formulations. The instability of the polysaccharide is responsible for the high cost of the vaccine. In this study, the factors affecting the spontaneous degradation of the polysaccharide from Hib were evaluated based on the decrease in its molecular mass, as measured by size-exclusion chromatography. Temperature and pH were found to be the most significant variables, and the results showed that the conditions of bacterial cell growth (37 °C and pH 7.5) are favourable for depolymerization. An increase in the concentration of sodium ions up to 200 mM intensified the effect of pH, allowing higher rates of depolymerization at lower pH values, whereas the presence of magnesium ions showed no effects., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

Abundant autologous proteins, like serum albumin, should be immunologically inert. However, individuals with no apparent predisposition to autoimmune disease can develop immune responses to autologous therapeutic proteins. Protein aggregation is a potential major trigger of these responses. Adsorption of proteins to particles provides macromolecular size and may generate structural changes in the protein, resembling aggregation. Using aldehyde/sulfate latex beads coated with murine serum albumin (MSA), we found that BALB/c mice mounted MSA-specific IgG responses that were dependent on CD4(+) T cells. IgGs were specific for MSA adsorbed to solid surfaces and noncross-reactive with human, bovine, or pig albumins. T cells induced in response to MSA augmented the primary and induced boosted secondary IgG and IgM responses specific for the T cell-independent antigen, capsular polysaccharide of Streptococcus pneumoniae type 14 (PPS14), when the latter was attached to the same bead. Similar to the anti-MSA IgG response, the boosted PPS14-specific IgG secondary response was CD4(+) T-cell dependent, displayed a typical carrier effect, and was enhanced by, but did not require, Toll-like receptor stimulation. These results provide a potential mechanism for the induction of responses to autoantigens unable to induce specific T-cell responses, and provide new insights into polysaccharide-specific immunity., (Published 2014. This article is a U.S. Government work and is in the public domain in the USA.)

Streptococcus agalactiae (Group B Streptococci, GBS) constitutes a risk factor for infections of the newborns born by colonized mothers. The adherence of GBS to epithelial cells has been proved to be an important factor in the colonization of mucus membranes of both human rectum and vagina. The objective of the study was to assess the adhesion of the selected GBS strains to the human colon adenocarcinoma cell line (HT-29) and human epidermoid vulvo-vaginal cells (A-431) in relation to the capsular polysaccharides and alpha-like protein genes. GBS strains from the human sources belonging to Ia, Ib, II, III and V serotypes possessing different surface alpha-like protein genes such as the alp 2, alp 3, bca, epsilon and rib in the conventional adherence assay were examined. The adherence of GBS strains to the HT-29 cell line was considerably higher than to the A-431 cell line. For GBS serotype Ia and III, a significant difference between the adhesion to the HT-29 and A-431 cell lines was presented. The adhesion of GBS strains to the HT-29 cell line depended on alpha-like protein genes. The most adhesive ones were the GBS strains containing the rib and alp 2 genes. The adherence of GBS strains to the A-431 cell line depended on both their serotype and alpha-like protein genes. Serotype III adhered to the A-431 cells most tightly, particularly the strains containing the rib and alp 2 genes. GBS strains containing the rib gene adhered to the HT-29 and A-431 cell lines more firmly than GBS strains containing other alpha-like protein genes.

Background: Combined diphtheria-tetanus-whole-cell pertussis (DTPw) vaccines remain the cornerstone of many childhood vaccination programs. Larger DTPw-based combination vaccines facilitate high coverage and protection against other childhood pathogens, such as hepatitis B (HBV) and Haemophilus influenzae type b (Hib). Such vaccines have been available since the mid-1990s (Tritanrix-HBV and Tritanrix-HBV/Hib; GlaxoSmithKline [GSK] Biologicals), demonstrating excellent immunogenicity in various schedules. In order to address growing demand for DTPw-based vaccines, GSK Biologicals have developed the Zilbrix range using a new DTPw antigen source and containing reduced quantities of polyribosylribitol phosphate [PRP]., Objective: This article presents clinical trial results for both DTPw-based vaccines., Methods: GSK Biologicals provided a comprehensive data package, which was supplemented with a Medline literature search., Conclusion: New antigen sources and reduced PRP will ensure the continued supply of DTPw-based combination vaccines for vital global mass vaccination campaigns.

Objective: To study the effectiveness of the Haemophilus influenzae type b (Hib) vaccination program in Uganda., Methods: Case-control study of Hib vaccine effectiveness against Hib meningitis. Cases were children hospitalized with Hib meningitis confirmed by culture and/or latex agglutination. Cases were identified retrospectively from July 2002 to July 2004, and prospectively from July 2004 to July 2005. Each case-patient was matched by age to three neighbourhood and three hospital controls; all children were eligible to receive Hib vaccine through the routine schedule. Vaccine effectiveness was evaluated by conditional logistic regression, controlling for confounding variables., Results: We enrolled 41 cases; their median age was 6 months. Only six (15%) cases, all HIV-negative, had received >/=2 doses of Hib vaccine, compared with 64% of neighbourhood controls and 70% of hospital controls. Controlling for maternal education, the only variable which remained in the multivariable model, vaccine effectiveness for two or three doses vs. no dose was 99% [95% confidence intervals (CI) 92-100%] and 96% (95% CI 80-100%) when cases were compared with neighbourhood and hospital controls, respectively., Conclusion: In Uganda, Hib vaccine was highly effective in the context of the routine immunization schedule. Sustained routine use of Hib vaccine will contribute to the prevention of childhood morbidity and mortality.

Haemophilus influenzae type b (Hib) disease is estimated to cause 3 million cases of meningitis and severe pneumonia and approximately 386,000 deaths worldwide per year in children aged <5 years. Safe and effective Hib conjugate vaccines have been widely used in industrialized countries for nearly 20 years. However, primarily because of financial constraints and lack of awareness among both public health officials and the public regarding Hib disease burden and benefits of the vaccine, use of these vaccines has been low in developing countries, where most Hib disease and deaths occur. In 2000, the GAVI Alliance (formerly known as the Global Alliance for Vaccines and Immunizations) began providing financial support for Hib vaccine in 72 countries that had a gross national income of < or =$1,000 (USD) per capita. Despite this support, before 2005, adoption of Hib vaccine by these countries remained low. In response, in June 2005, the GAVI Alliance established the Hib Initiative to accelerate evidence-informed decision making regarding use of Hib vaccine in GAVI-eligible countries. During 2004-2007, the number of GAVI-eligible countries using Hib vaccine or approved to use the vaccine increased from 13 (18%) to 47 (65%).

Microbial exposure is necessary for the development of normal immune function, which has driven the idea of using probiotics for treatment and prevention of immune-mediated diseases in infancy and childhood. Mounting evidence indicates that probiotics have immunomodulatory effects. However, the mechanisms are still poorly understood. Specific antibody response is a valuable proxy for immune system maturation status in infancy. We aimed at determining the impact of Lactobacillus F19 (LF19) during weaning on infections and IgG antibody responses to routine vaccines. In a double-blind, placebo-controlled randomized intervention trial, infants were fed cereals with (n = 89) or without LF19 (n = 90) from 4 to 13 months of age. Infants were immunized with DTaP (diphtheria and tetanus toxoid and acellular pertussis), polio and Hib-conjugate vaccines at (3), 5(1/2) and 12 months of age. We assessed the number of days with infections, antibiotic prescriptions and antibody concentrations to Hib capsular polysaccharide (HibPS), diphtheria toxin (D) and tetanus toxoid (T) before and after the second and third doses. Days with infectious symptoms did not differ between the groups. Days with antibiotic prescriptions were fewer in the LF19 group (p = 0.044). LF19 enhanced anti-D concentrations when adjusting for breastfeeding duration and colonization with LF19 (p = 0.024). There was an interaction of the intervention and colonization with LF19 on anti-T concentrations during the course of vaccination (p = 0.035). The anti-HibPS concentrations were higher after the first and second dose of Hib vaccine in infants breastfed <6 months compared with those breastfed > or =6 months (p < 0.05), with no effect by LF19. In conclusion, feeding LF19 did not prevent infections, but increased the capacity to raise immune responses to protein antigens, with more pronounced effects in infants breastfed <6 months.

Background: Despite the excellent immunogenicity of Haemophilus influenzae type b (Hib) conjugate vaccines, breakthrough cases of Hib disease still affect a small proportion of vaccinated children in the United Kingdom. We performed a retrospective study to compare the avidity of antibody directed against the Hib polysaccharide capsule (PRP) in children who experienced Hib vaccine failure in the United Kingdom among 3 historical cohorts and with age-matched healthy control subjects., Methods: Serum samples from vaccinated children with invasive Hib disease were collected beginning in 1992 as part of enhanced surveillance for Hib disease following vaccine introduction. A total of 251 children who experienced Hib vaccine failure were identified from 3 historical cohorts (1992-1995, 1996-1999, and 2000-2003). The anti-PRP antibody concentration and avidity from healthy age-matched control subjects was obtained for the 3 contemporary time points (1995, 1999, and 2002). Serum anti-PRP antibody concentration was measured in each of the samples using a standard Hib ELISA, and antibody avidity was determined using thiocyanate elution., Results: Within the first 60 days after disease onset, there was no change in the anti-PRP antibody avidity, and there was no statistically significant difference in the geometric mean Hib antibody avidity over the 3 study periods. However, the children who experienced Hib vaccine failure had significantly lower Hib antibody avidity than did healthy control subjects, despite a marked antibody response following infection., Conclusions: Children who experience Hib disease despite vaccination appear to have a defect in immunological priming, leading to a qualitative difference in Hib-specific memory B cells. Low anti-PRP antibody avidity decreases the functional activity of anti-PRP antibody in the sera of these children experiencing vaccine failure, leading to disease susceptibility.

Despite the demonstration of effectiveness and public health impact of Hib conjugate vaccines, the majority of infants from poorest countries do not yet have access to this safe and effective preventive intervention. This paper provides a descriptive analysis of the main characteristics of countries that have included Hib vaccine in their national immunization programmes before 2006. It highlights the importance of regularly analyzing the process of decision-making involved in implementation of public health interventions, in order to learn from cumulative experience and expedite the introduction of future interventions.

Recombinant human betacellulin binds strongly to heparin, requiring of the order of 0.8 M NaCl for its elution from a heparin affinity matrix. This is in complete contrast to the prototypic member of its cytokine superfamily, epidermal growth factor, which fails to bind to the column at physiological pH and strength. We used a well-established heparin binding ELISA to demonstrate that fucoidan and a highly sulfated variant of heparan sulfate compete strongly for heparin binding. Low sulfated heparan sulfates and also chondroitin sulfates are weaker competitors. Moreover, although competitive activity is reduced by selective desulfation, residual binding to extensively desulfated heparin remains. Even carboxyl reduction followed by extensive desulfation does not completely remove activity. We further demonstrate that both hyaluronic acid and the E. coli capsular polysaccharide K5, both of which are unsulfated polysaccharides with unbranched chains of alternating N-acetylglucosamine linked beta(1-4) to glucuronic acid, are also capable of a limited degree of competition with heparin. Heparin protects betacellulin from proteolysis by LysC, but K5 polysaccharide does not. Betacellulin possesses a prominent cluster of basic residues, which is likely to constitute a binding site for sulfated polysaccharides, but the binding of nonsulfated polysaccharides may take place at a different site.

Haemophilus influenzae b conjugate vaccines (Hib) are almost entirely evaluated by physico-chemical methods to ensure the consistency of manufacture of batches. As different assays are employed for the quantification of Hib capsular polysaccharide PRP (polyribosyl ribitol phosphate; 5-D-ribitol-(1-->1)-beta-D-ribose-3-phosphate) in final formulations and bulk components, there was deemed a need for an International Standard of Hib PRP polysaccharide to be made available. Ten laboratories from 8 different countries participated in a collaborative study to determine the PRP content and assess the suitability of a candidate International Standard PRP preparation (02/208). The results illustrate that a reduction in between-laboratory variability could be achieved by use of a common reference preparation and data analysis showed no significant differences in the values obtained by the different assays: ribose, phosphorus, and high performance anion exchange chromatography-pulsed amperometric detection (HPAEC-PAD), suggesting the suitability of the proposed reference for use across these assays for quantification of PRP content in Hib vaccines. On the basis of the results of this study, the First International Standard for PRP, NIBSC Code 02/208, has been established by the Expert Committee of Biological Standards of the World Health Organisation, with a content of 4.933+/-0.267mg/ampoule, as determined by the ribose assays carried out by 7 of the participating laboratories.

Rwanda introduced Haemophilus influenzae type b (Hib) conjugate vaccine in January 2002 and simultaneously implemented pediatric bacterial meningitis surveillance at a major referral hospital in the capital Kigali. We reviewed clinical and laboratory information collected during January 2002 to June 2006. Due to a variety of laboratory limitations, only eight confirmed Hib cases were identified, all before 2004. However, the proportion of cerebrospinal fluid with purulence decreased from 26.0% during 2002, to 15.9% during 2003, 9.7% during 2004 and 8.4% in 2005 (p<0.001). Vaccine effectiveness of two or three doses of Hib vaccine against purulent meningitis was 52% (95% confidence interval, 5-75%). In an African setting with few resources and in which few confirmed Hib meningitis cases were identified, Hib vaccine impact nevertheless could be demonstrated against the outcome of purulent meningitis and was found to be high.

Escherichia coli K1 is the leading cause of human neonatal sepsis and meningitis and is important in other clinical syndromes of both humans and domestic animals; in this strain the polysialic acid capsule (K1 antigen) functions by inhibiting innate immunity. Recent discovery of the phase-variable capsular O acetylation mechanism indicated that the O-acetyltransferase gene, neuO, is carried on a putative K1-specific prophage designated CUS-3 (E. L. Deszo, S. M. Steenbergen, D. I. Freedberg, and E. R. Vimr, Proc. Natl. Acad. Sci. USA 102:5564-5569, 2005). Here we describe the isolation and characterization of a CUS-3 derivative (CUS-3a), demonstrating its morphology, lysogenization of a sensitive host, and the distribution of CUS-3 among a collection of 111 different K1 strains. The 40,207-bp CUS-3 genome was annotated from the strain RS218 genomic DNA sequence, indicating that most of the 63 phage open reading frames have their closest homologues in one of seven different lambdoid phages. Translational fusion of a reporter lacZ fragment to the hypervariable poly-Psi domain facilitated measurement of phase variation frequencies, indicating no significant differences between switch rates or effects on rates of the methyl-directed mismatch repair system. PCR analysis of poly-Psi domain length indicated preferential loss or gain of single 5'-AAGACTC-3' nucleotide repeats. Analysis of a K1 strain previously reported as "locked on" indicated a poly-Psi region with the least number of heptad repeats compatible with in-frame neuO expression. The combined results establish CUS-3 as an active mobile contingency locus in E. coli K1, indicating its capacity to mediate population-wide capsule variation.

Following the reduction in efficacy of Hib-TT vaccines in the primary immunization schedule observed in the UK between 1999 and 2003, batches of vaccine manufactured by two different companies were retrospectively examined by the National Institute for Biological Standards and Control. The study evaluated 41 batches of the Hib-TT vaccines manufactured between 1994 and 2003, assaying potency (total PRP saccharide content), integrity (% free saccharide), consistency (molecular sizing), and immunogenicity, as well as reviewing data previously obtained at the time of release. The study indicated the stability of the lyophilized final fill vaccines to extend well past their assigned shelf-lives, and found no trends in the endotoxin content, total saccharide or % free saccharide content. A trend towards slightly larger conjugates was observed over time in Hib-TT A, evidenced in both the manufacturer's data obtained at the time that samples were submitted for testing and in data obtained from the retrospective analysis. The study confirmed that that there had been no significant change in the quality of the Hib vaccines that could possibly account for the change reported in their protective efficacy in the UK. The study also demonstrated the value of independent testing of vaccines from the time of licensure and in the ongoing monitoring and re-examination of selected batches, as necessary, to assure their continuing quality, safety and consistency.

Objective: There are conflicting reports as to whether breastfed infants respond with higher antibody levels to conjugate Haemophilus influenzae type b (Hib) vaccine compared with formula-fed infants. These observations prompted us to investigate the effect of feeding method on the antibody concentration to Hib polyribosylribitol (PRP) both prior to and 3 months after the primary course of immunisation with Hib (PRP-OMP)., Methods: We measured plasma concentrations of IgG antibody to Hib PRP by enzyme-linked immunosorbent assay in blood samples from a total of 272 breastfed and formula-fed infants prior to immunisation (7 weeks of age, n = 82 and n = 148, respectively) and again 3 months after completion of the primary course of immunisation with Hib PRP-OMP (7 months of age, n = 88 and n = 132, respectively)., Results: Breastfeeding was associated with lower plasma antibody titres at both times (P < 0.01, T-test) with 49% of breastfed infants having anti-PRP concentrations below 1.0 microg/mL at age 7 months. There was no reported invasive Hib disease in this cohort of infants, and nationally the effectiveness of the Hib vaccination programme remains high., Conclusions: These data suggest that breastfeeding may be associated with immunomodulation of infant Hib immunisation responses with this immunisation regime. Further research is needed to determine whether differences in antibody concentration described here are primarily determined by factors directly attributed to breastfeeding or whether other environmental factors may play a significant role.

Invasive Hib disease, which remains a major cause of childhood mortality and morbidity in most of the developing world, was eliminated in The Gambia by 2002 following the introduction of conjugate Hib vaccine in 1997. Formal disease surveillance was stopped in 2002 but five cases (including three of meningitis) were detected non-systematically between July 2005 and April 2006. This equates to an incidence of 3 per 100,000 annually for meningitis, a likely underestimate. The age distribution of cases (median 15 months, range 0-36 months) was older than previously seen and there were examples of apparent vaccine failure, but the cause for this re-emergence is not clear. No evidence was found of the emergence of a hypervirulent strain. The re-establishment of continuing surveillance is required to answer the questions raised by this report, and is particularly important in settings like The Gambia, where a booster dose is not given, to determine long-term effects of national immunisation with Hib vaccine.

Objective: To evaluate the immunogenicity of the Hepatitis B and Haemophilus influenzae type b components and the overall safety and reactogenicity of the DTPw-HBV/Hib vaccine when given as primary vaccination to Indian infants., Design and Methods: At 3 centers in India, 225 healthy infants (who had received HBV at birth) received three doses of DTPw-HBV/Hib vaccine at 6, 10 and 14 weeks of age. Serum anti-HBs and anti-PRP antibody levels were measured prior to vaccination and one month post dose 3. Solicited local and general symptoms reported during the 4-day follow-up period and unsolicited adverse event reported during the 30-day follow-up period after each dose were recorded. Serious adverse events were recorded throughout the study., Results: A total of 219 subjects completed the study. 2.7% and 11.5% of all administered doses led to redness and swelling >20 mm, respectively; only 3.6% of doses were followed by severe pain (cried when limb was moved, spontaneously painful) within 4 days after vaccination. Fever exceeding 39.5C was recorded following only one dose in one subject. The percentage of doses followed by severe solicited general symptoms (symptoms that prevented normal activity) did not exceed 0.8%. Two SAEs were reported, neither of which were considered as related to vaccination. One month post-dose 3, all subjects had seroprotective antiPRP antibody concentrations (> or =0.15 microgram/mL) and 98.6% had concentrations > or =1 microgram/mL; 99% were seropositive for antiHBs (concentrations > or = 3 mIU/mL) and 99% were seroprotected (concentrations > or = 10 mIU/mL)., Conclusion: The combination DTPw-HBV/Hib vaccine is immunogenic (for the antigens tested), safe and well tolerated in Indian infants.

This study presents detailed analyses of total and specific serum antibody levels among 26 and 24 adult volunteers before vaccination and after the third dose of the meningococcal serogroup B outer membrane vesicle (OMV) vaccines MeNZB and MenBvac, respectively, in a clinical trial in New Zealand (V. Thornton, D. Lennon, K. Rasanathan, J. O'Hallahan, P. Oster, J. Stewart, S. Tilman, I. Aaberge, B. Feiring, H. Nokleby, E. Rosenqvist, K. White, S. Reid, K. Mulholland, M. J. Wakefield, and D. Martin, Vaccine 24:1395-1400, 2006). With the homologous vaccine strains as targets, both vaccines induced significant increases in serum bactericidal and opsonophagocytic activities and in the levels of immunoglobulin G (IgG) to OMV antigens in an enzyme-linked immunosorbent assay (ELISA) and to live meningococci by flow cytometry. They also induced high levels of activity against the heterologous strains, particularly in terms of opsonophagocytic activity and IgG binding to live bacteria. The antibody levels with the homologous and heterologous strains in the four assays showed high and significant positive correlations. Specific IgG binding to 10 major OMV antigens in each vaccine was measured by scanning of immunoblots; ELISAs for two antigens, lipopolysaccharide and Neisseria surface protein A (NspA), were also performed. Both vaccines elicited significant increases in IgG binding to all homologous and heterologous OMV antigens except NspA. The total IgG band intensity on the blots correlated significantly with the IgG levels determined by the OMV ELISA and flow cytometry. In conclusion, the results of the various immunological assays showed that both OMV vaccines gave rise to high levels of specific and cross-reacting antibodies.

Heparin-like polysaccharides possess the capacity to inhibit cancer cell proliferation, angiogenesis, heparanase-mediated cancer cell invasion, and cancer cell adhesion to vascular endothelia via adhesion receptors, such as selectins. The clinical applicability of the antitumor effect of such polysaccharides, however, is compromised by their anticoagulant activity. We have compared the potential of chemically O-sulfated and N,O-sulfated bacterial polysaccharide (capsular polysaccharide from E. COLI K5 [K5PS]) species to inhibit metastasis of mouse B16-BL6 melanoma cells and human MDA-MB-231 breast cancer cells in two in vivo models. We demonstrate that in both settings, O-sulfated K5PS was a potent inhibitor of metastasis. Reducing the molecular weight of the polysaccharide, however, resulted in lower antimetastatic capacity. Furthermore, we show that O-sulfated K5PS efficiently inhibited the invasion of B16-BL6 cells through Matrigel and also inhibited the in vitro activity of heparanase. Moreover, treatment with O-sulfated K5PS lowered the ability of B16-BL6 cells to adhere to endothelial cells, intercellular adhesion molecule-1, and P-selectin, but not to E-selectin. Importantly, O-sulfated K5PSs were largely devoid of anticoagulant activity. These findings indicate that O-sulfated K5PS polysaccharide should be considered as a potential antimetastatic agent.

Shewanella strains have previously been studied with regard to their cell surface ultrastructure and LPS composition. They have now been further characterized with respect to their surface physicochemistry and ability to adhere to haematite. The surfaces of the Shewanella strains were found to be electronegative and hydrophilic, and these properties could be correlated with LPS composition or the presence of capsular polysaccharides. Strains expressing rough LPS with no capsule were more hydrophobic and electronegative than those possessing smooth LPS or capsules. By combining different approaches, such as contact-angle measurement, hydrophilic/hydrophobic chromatography, microelectrophoresis, adhesion assays and calculation of interaction energies, it was shown that electrostatic interactions predominate over hydrophobic interactions at the cell-iron oxide interface. Bacterial adhesion to haematite was significantly reduced in strains expressing smooth LPS or a capsule. These findings remained true for Shewanella strains grown under either aerobic or anaerobic conditions, although the surfaces of anaerobic cells appeared to be less electronegative and more hydrophilic than those of aerobic cells.

Vitamin A supplementation reduces child mortality and severe morbidity in less developed countries, and the Expanded Program on Immunization (EPI) offers an ideal opportunity to deliver supplements in developing countries. High-dose vitamin A supplementation has been shown to have no effect on the immunogenicity of oral polio vaccine, tetanus toxoid, pertussis, or on measles vaccine given at 9 mo, but a negative effect on measles vaccine administered at 6 mo and a potentiating effect on diphtheria vaccine. Its effect on the antibody response to hepatitis B and Haemophilus influenzae type b antigens has not yet been established. To assess these effects, the present trial was carried out in the Offinso district of Ghana; 1077 infants were enrolled shortly after birth and randomized either to receive or not to receive 15 mg retinol equivalent with vitamin A together with the pentavalent "diphtheria-polio-tetanus-Haemophilus influenzae b-hepatitis B" vaccine at 6, 10, and 14 wk of age. All mothers received a postpartum supplement of 120 mg retinol equivalent vitamin A as per national policy. Blood samples were taken from infants at 6 and 18 wk of age. The results are based on 888 infants (82.4%) who completed the trial. The vitamin A supplementation did not affect the immune response to Haemophilus influenzae type b, but there was a significant improvement in the immune response to hepatitis B vaccine (93.9 vs. 90.2%, P = 0.04). However, given the high percentage of infants with seroprotection in the control group, it is doubtful that inclusion of vitamin A in the EPI would be justified on these grounds alone.

Neisseria meningitidis serogroup B (MenB) is a leading cause of sepsis and meningitis in children. No vaccine is available for the prevention of these infections because the group B capsular polysaccharide (CP) (MenB CP) is unable to stimulate an immune response, due to its similarity with human polysialic acid. Because the MenB CP bears both human cross-reactive and non-cross-reactive determinants, we developed immunogenic peptide mimics of the latter epitopes. Peptides were selected from phage display libraries for their ability to bind to a protective anti-MenB CP mAb. One of these peptides (designated 9M) induced marked elevations in serum bactericidal activity, but not polysialic acid cross-reacting Abs, after gene priming followed by carrier-conjugate boosting. Moreover, the occurrence of bacteremia was prevented in infant rats by administration of immune sera before MenB challenge. 9M is a promising lead candidate for the development of an effective and affordable anti-MenB vaccine.

Background: Necrotizing fasciitis caused by Klebsiella pneumoniae is rare, with 11 documented cases in the literature, all occurring in Asia and the Middle East. These reports have become more frequent in recent years in association with the emergence of the highly virulent K1 capsular serotype of Klebsiella. We report a case of Klebsiella necrotizing fasciitis and con-current septic arthritis in the Western hemisphere., Methods: Case report and review of the literature., Results: Our patient, a 50-year-old Cambodian man with advanced hepatitis B-associated cirrhosis, who had last visited an endemic area six months prior to presentation, was hospitalized after several days of left knee and thigh pain. He was hypotensive, tachycardic, and bacteremic with K. pneumoniae. A computed tomography scan showed gas in the tissues of the thigh and left knee joint. Despite aggressive surgical debridement and antibiotic therapy, the patient died on day three. Serotyping of blood, wound, and synovial cultures revealed K1-type K. pneumoniae., Conclusions: Although K. pneumoniae is common, monomicrobial necrotizing fasciitis and septic arthritis caused by this pathogen has not been reported previously outside Asia, where it is rare. The appearance of this infection in the Western hemisphere may reflect geographic spread of the aggressive K1 phenotype. Physicians treating patients with symptoms and signs of necrotizing fasciitis or septic arthritis, particularly in the setting of underlying chronic illness or recent travel to Asia, should consider K. pneumoniae as a potential cause and treat accordingly.

The safety of DTaP-HepB-IPV vaccine coadministered with PCV and Hib was compared with separate administration of DTaP, HepB, IPV, Hib, and PCV at 2, 4, and 6 months of age. Healthy 2-month-old infants (N=1008) were randomized to the two groups. Following dose 1, there was no significant difference between the groups in the incidence of fever >101.3 degrees F. After each dose, the incidence of any fever (> or =100.4 degrees F) was significantly higher in the Combination Vaccine Group. The rate of fever >103.1 degrees F was < or =1.4% in both groups after any of the doses. Medical advice visits for fever were infrequent in both groups (< or =1.2%). DTaP-HepB-IPV was safe and well tolerated when coadministered with PCV and Hib.

Capsular serotypes, magA, and rmpA have been documented in high prevalence for Klebsiella pneumoniae liver abscess. To investigate the regional difference and the correlation of capsular serotype, magA, and rmpA with virulence, 73 isolates were collected in Singapore and Taiwan. Capsular serotypes were determined by countercurrent immunoelectrophoresis, the presence of magA and rmpA was determined by PCR, and virulence was determined by phagocytosis and mouse inoculation. Isolates from Singapore were similar to those from Taiwan in genomic heterogeneity, prevalence of serotype, and the presence of magA and rmpA. The most common serotype was K1 (34/73; 46.6%), followed by K2 (15/73; 20.5%). magA was restricted to serotype K1. All K1 or K2 isolates and 66.7% (16/24) of isolates that were neither serotype K1 nor serotype K2 (non-K1/K2) carried rmpA. Serotype K1 or K2 isolates demonstrated significantly more phagocytic resistance and virulence than did rmpA-positive and -negative groups of non-K1/K2 isolates. In the non-K1/K2 group, the virulence profiles of rmpA-positive strains from Taiwan and Singapore were different by phagocytosis assay and in the mouse model, indicating that factors other than rmpA contributed to virulence. The characteristics of K. pneumoniae liver abscess in Singapore and Taiwan are similar. Capsular serotype K1 or K2 plays a more important role than magA and rmpA in determining virulence in K. pneumoniae liver abscess.

When protein-polysaccharide conjugated vaccines were first developed for the immunization of small children against meningitis caused by infection with Haemophilus influenzae type b (Hib), the vaccine preparations varied in immunogenicity. Testing for immunogenicity was time-consuming and alternative analytical procedures for determining vaccine quality were unsatisfactory. For example, due to the very high molecular weight of the vaccine particles, immunogens could only be physically characterized as a fraction in the void volume of Sepharose gel filtration. In search of better analytical methods, a computer-assisted electrophoretic technique for analyzing such vaccines was developed in the period from 1983 to 1995. This new approach made it possible to analyze highly negatively charged particles as large as or larger than intact viruses. 2-D gel patterns were generated that varied depending on the conditions of the particular vaccine preparation and were therefore characteristic of each vaccine sample. Thus, vaccine particle populations with a continuous size variation over a wide range (polydisperse) could be characterized according to size and free mobility (related to particle surface net charge density). These advances are reviewed in this article, since the developed methods are still a promising tool for vaccine quality control and for predicting immunogen effectiveness in the production of vaccines. The technique is potentially beneficial for Hib immunogens and other high-molecular-mass vaccines. Additional biomedical applications for this nondenaturing electrophoretic technique are briefly discussed and detailed information about computational and mathematical procedures and theoretical aspects is provided in the Appendices.

The separation and characterization of oligosaccharides obtained by hyaluronidase [E.C. 3.2.1.35] digestion of Escherichia coli K4 polysaccharide using online high-performance liquid chromatography/electrospray ionization mass spectrometry (HPLC/ESI-MS) are presented. Complete identification and structural information for oligosaccharides containing 2-24 monomers (from 2- to 24-mers) were obtained. In particular, smaller K4 species, from 2-mers to 4-mers, exhibited mainly [M-H](-1) anions, whereas the 6- to 8-mers existed predominantly at the charge state of -2. The K4 oligomers from 10-mers to 14-mers were mainly represented by [M-3H](-3) anions while species from 16- to 20-mers were characterized by a charge state of -4. K4 oligosaccharides from 22- to 24-mers existed as [M-4H](-4) and [M-5H](-5) anions and, for this latter species, ions having a charge state of -6 appeared. For smaller K4 species, in particular from 6-mers to 10-mers, ESI-MS revealed anions related to the loss of one monosaccharide unit from the oligomers due to apparent collisional activation and ion source fragmentation. However, no odd-numbered anions were produced for K4 2/4-mer species or for oligosaccharides greater than 12-mers, while for K4 species 8/10-mer, ESI-MS revealed odd-numbered anions generally in low relative abundance making the interpretation of the spectra easier. The ESI-MS spectra of oligosaccharides separated by online HPLC were applied to the evaluation of the K4 polymerization process, confirming that the addition of fructose units is not critical for chain elongation as variously fructosylated oligomer species were detected directly on the K4 carbohydrate backbone., (Copyright (c) 2007 John Wiley & Sons, Ltd.)

Biosynthetic, semisynthetic, and synthetic analogues of heparins are currently developed as substitute antithrombotic agents for heparin. Sulfaminoheparosan (SAH) represents a bacterial polysaccharide (K5)-derived antithrombotic polymer from which pharmacologically active products with varying molecular weights (5-25 kDa) can be derived. These agents have been shown to exhibit pharmacologic effects comparable to heparins. The objective of this investigation is to determine the relative neutralization profile of various SAH derivatives, also called as bioheparins, by protamine sulfate. Four SAH fractions with varying molecular weights (20, 9, 7, and 6 kDa), a low molecular weight heparin (LMWH), tinzaparin, and unfractionated heparin (UFH) were supplemented to normal human pool plasma over a concentration range of 6.2 to 100 microg/mL. A fixed amount of protamine sulfate at 25 microg/mL (final concentration) was supplemented to determine the neutralization profile by performing tests such as prothrombin time (PT), activated partial thromboplastin time (APTT), Heptest, prothrombin-induced clotting time (PiCT), and amidolytic anti-Xa and anti-IIa assays. Protamine sulfate produced varying degrees of neutralization of all bioheparin fractions in the clotting assays. In the amidolytic anti-IIa assay relatively stronger inhibition was noted for all agents than inhibition of FXa. On a cumulative basis the neutralization profile of SAHs was comparable with heparins. These results suggest that the anticoagulant activities of SAH derivatives can be antagonized by protamine sulfate. These studies warrant further in vivo investigation to validate the relative neutralization profile of sulfaminoheparosans.

Background: This phase II study evaluated the immunogenicity and reactogenicity of primary vaccination with a novel Hib-MenC conjugate vaccine (GlaxoSmithKline [GSK] Biologicals) coadministered with DTPa-HBV-IPV (GSK Biologicals) at 2, 4 and 6 months., Methods: Healthy infants were randomized to receive Hib-MenC coadministered with DTPa-HBV-IPV (N = 117) or MenC-CRM (Wyeth) coadministered with DTPa-HBV-IPV/Hib (GSK Biologicals; N = 120) at 2, 4 and 6 months. Antibody concentrations were measured before vaccination and after doses 2 and 3. Solicited local and general symptoms, unsolicited symptoms and serious adverse events (SAEs) were recorded., Results: All subjects in the Hib-MenC group had seroprotective titers of anti-PRP antibodies (>or=0.15 microg/mL) and SBA-MenC titers (>or=1:8) 1 month after the third dose. These responses were noninferior to those seen in the control group, in which a 99.1% seroprotection rate was observed for both Hib and MenC. At that time, anti-PRP and SBA-MenC GMTs were significantly higher in the Hib-MenC group (12.8 microg/mL and 2467.1 microg/mL, respectively) than in the control group (3.8 microg/mL and 1833.7 microg/mL). High seroprotection rates were already observed after the second dose of Hib-MenC; 96.4% and 100% of subjects were seroprotected to Hib and MenC, respectively. Immune responses to coadministered antigens were unimpaired; seroprotection/vaccine response rates >or=96.5% were recorded postdose 3 in the Hib-MenC group. No differences in reactogenicity were seen between the 2 study groups., Conclusions: Coadministration of a Hib-MenC conjugate vaccine with DTPa-HBV-IPV is well tolerated and immunogenic, and does not impair the immune response to any of the coadministered antigens.

Multiplex-PCR assay for identification of Klebsiella pneumoniae isolates carrying gene clusters for biosynthesis of capsular polysaccharide (CPS) types K1 and K2 was developed. Genes wzc and orf10 of the cps cluster were applied as K1 and K2 specific markers respectively. The assay specificity was confirmed using 147 isolates of Klebsiella spp. including 77 K-antigen reference strains. The multiplex-PCR assay was found simple and cost-effective tool for identification of K. pneumoniae clinical isolates of K1 and K2 geno-serotypes.

Background: Prematurity may be a risk factor for Haemophilus influenzae type b vaccine failure. This article evaluates the Haemophilus influenzae type b immunogenicity of a hexavalent diphtheria-tetanus-acellular pertussis-hepatitis B-inactivated poliovirus/Haemophilus influenzae type b vaccine in preterm infants (< 37 weeks' gestation)., Methods: This was an open-label, parallel group study. Preterm (N = 94) and term infants (N = 92) received 3 doses of a diphtheria-tetanus-acellular pertussis-hepatitis B-inactivated poliovirus/Haemophilus influenzae type b vaccine at 2, 4, and 6 months with a booster dose at 18 to 20 months. Antipolyribosyl ribitol phosphate antibody concentrations were determined in serum samples taken before and 1 month after primary and booster vaccination., Results: Postprimary seroprotection rates (antipolyribosyl ribitol phosphate > or = 0.15 microg/mL) were lower in preterm than in term infants (92.5% vs 97.8%), with antipolyribosyl ribitol phosphate geometric mean concentrations of 2.241 vs 4.247 microg/mL. A progressive reduction in immune response to the Haemophilus influenzae type b antigen was observed with decreasing length of gestation and decreasing birth weight when cutoff > or = 1 microg/mL was considered. Prebooster seroprotection rates and antipolyribosyl ribitol phosphate geometric mean concentrations were low in both groups (antipolyribosyl ribitol phosphate > or = 1.0 microg/mL in 10.7% of preterm and 28.4% of term infants). A vigorous response to booster vaccination was seen in both groups, with no differences in postbooster seroprotection rates or antipolyribosyl ribitol phosphate geometric mean concentrations between the 2 groups (antipolyribosyl ribitol phosphate > or = 1.0 microg/mL in 100% of preterm and 98.5% of term infants)., Conclusions: Primary vaccination with a hexavalent diphtheria-tetanus-acellular pertussis-hepatitis B-inactivated poliovirus/Haemophilus influenzae type b vaccine at 2, 4, and 6 months with a booster dose at 18 to 20 months elicits a satisfactory antipolyribosyl ribitol phosphate response in preterm infants compared with term controls. Immunologic response decreased with decreased gestational age and birth weight.

A conjugate meningococcal vaccine against serogroup B/C consisting of capsular PS (polysaccharide) from serogroup C conjugated to OMV (outer membrane vesicle) from serogroup B would be a very useful vaccine in regions where there is a prevalence of both serogroups, for example in Brazil. For this purpose, the conjugation method that uses ADHy (adipic acid dihydrazide) as spacer and a carbodi-imide derivative, EDAC [1-ethyl-3-(3-dimethylaminopropyl)carbodi-imide], as catalyser was optimized looking for synthesis yield and maintenance of the antigenicity of both components. The best synthesis conditions preserving the vaccine immunogenicity resulted in a final yield of approx. 17%. Immunogenicity of the vaccine was highest when 10% of the sialic acid residues of the PS were occupied by the ADHy spacer. Sterilization of the conjugate by filtration through a 0.22-microm-pore-size membrane resulted in a low recovery of protein and PS (approximately 50%), although the vaccine immunogenicity was maintained. Using gamma irradiation on freeze-dried sample, it was possible to maintain the integrity of OMV structure and, consequently, its ability to induce bactericidal antibodies.

Heparan sulfate plays a wide range of physiological and pathological roles. Heparan sulfate consists of glucosamine and glucuronic/iduronic acid repeating disaccharides with various sulfations. Synthesis of structurally defined heparan sulfate oligosaccharides remains a challenge. Access to nonsulfated and unepimerized heparan sulfate backbone structures represents an essential step toward de novo enzymatic synthesis of heparan sulfate. The nonsulfated, unepimerized backbone heparan sulfate is similar to the capsular polysaccharide from Escherichia coli strain K5. The biosynthesis of this capsular polysaccharide involves in N-acetylglucosaminyltransferase (KfiA) and d-glucuronyltransferase (KfiC). In this study, we report the characterization of purified KfiA. KfiA was expressed in a C-terminal six-His fusion protein in BL21 star cells coexpressing chaperone proteins GroEL and GroES. The recombinant KfiA was purified to homogeneity with a Ni-agarose column. The binding affinities of various UDP-sugars for KfiA were determined using isothermal calorimetry titration, indicating that both the N-acetyl group and sugar type may be essential for donor substrates to bind KfiA. Kinetic analysis of KfiA toward different sizes of oligosaccharide revealed that KfiA is less sensitive to the size of the acceptor substrates. The results from this study open a new approach for the synthesis of the heparan sulfate backbone.

Objective: To analyse trends in reported invasive Haemophilus influenzae disease in South Africa within the first five years of introduction of conjugate Haemophilus influenzae type b (Hib) vaccine in the routine child immunization schedule., Methods: We used national laboratory-based surveillance data to identify cases of invasive H. influenzae disease between July 1999 and June 2004, and submitted isolates for serotyping and antimicrobial susceptibility testing., Findings: The absolute number of Hib cases (reported to the national surveillance system) among children below one year of age decreased by 65%, from 55 cases in 1999-2000 to 19 cases in 2003-04. Enhanced surveillance initiated in 2003, identified human immunodeficiency virus (HIV)-infection and incomplete vaccination as contributing factors for Hib transmission. The total number of laboratory-confirmed cases of H. influenzae remained unchanged because non-type b disease was being increasingly reported to the surveillance system concomitant with system enhancements. Children with non-typable disease were more likely to be HIV-positive (32 of 34, 94%) than children with Hib disease (10 of 14, 71%), P = 0.051. Recent Hib isolates were more likely to be multidrug resistant (2% in 1999-2000 versus 19% in 2003-04, P = 0.001)., Conclusion: Data from a newly established national laboratory-based surveillance system showed a decrease in Hib disease burden among South African children following conjugate vaccine introduction and identified cases of non-typable disease associated with HIV infection.

Objective: Conjugate vaccines are the best public health tools available for preventing most invasive diseases caused by Haemophilus influenzae type b (Hib), but the high cost of the vaccines has so far kept them from being introduced worldwide. The objective of this study was to estimate the cost-effectiveness of introducing Hib conjugate vaccines for the prevention of meningitis and pneumonia among children under 2 years of age in Colombia., Methods: We estimated the direct and indirect costs of managing in-hospital pneumonia and meningitis cases. In addition, following the recommendations of the World Health Organization, we assessed the cost-effectiveness of Hib vaccination programs. We also estimated the costs for preventing Hib cases, and the cost per year of life saved in two hypothetical situations: (1) with vaccination against Hib (with 90% coverage) and (2) without vaccination., Results: The average in-hospital treatment costs were 611.50 US$ (95% confidence interval (95% CI) = 532.2 to 690.8 US$) per case of pneumonia and 848.9 US$ (95% CI = 716.8 to 981.0 US$) per case of meningitis. The average cost per Hib case prevented was 316.7 US$ (95% CI = 294.2 to 339.2 US$). In terms of cost-effectiveness, the cost would be 2.38 US$ per year of life saved for vaccination, versus 3.81 US$ per year of life saved without vaccination., Conclusion: Having an adequate Hib vaccination program in Colombia could prevent around 25,000 cases of invasive disease per year, representing a cost savings of at least 15 million US$ annually. Furthermore, the program could prevent some 700 deaths per year and save 44,054 years of life per year.

Recent efforts toward developing vaccines against group B streptococci (GBS) have focused on increasing the immunogenicity of GBS polysaccharides by conjugation to carrier proteins. However, partial depolymerization of GBS polysaccharides for the production of vaccines is a difficult task because of their acid-labile, antigenically critical sialic acids. Here we report a method for the partial depolymerization of type II and III polysaccharides by mild deaminative cleavage to antigenic fragments with reducing-terminal 2,5-anhydro-d-mannose residues. Through the free aldehydes of their newly formed end groups, the fragments were conjugated to tetanus toxoid by reductive amination. The resulting conjugates stimulated the production in animals of high-titer type II- and III-specific antibodies which induced opsonophagocytic killing of type II and III strains of group B streptococci. For the type II conjugates, immunogenicity increased as oligosaccharide size decreased, whereas for type III conjugates, the size of the oligosaccharides did not significantly influence immunogenicity. When oligosaccharides of defined size were conjugated through sialic acid residues, the resulting cross-linkages were shown to affect immunogenicity. When oligosaccharides were conjugated through terminal aldehyde groups generated by deamination, modification of the exocyclic chain of sialic acid did not influence immunogenicity.

Background: Combined vaccines containing diphtheria-tetanus-pertussis whole-cell (DTPw), Haemophilus influenzae type b (Hib), and hepatitis-B vaccines are essential for the continuing success of vaccination programs in developing nations. This randomized, dose-ranging study assessed the immunogenicity and reactogenicity of primary and booster vaccination with pentavalent DTPw-HBV/Hib vaccines containing 10, 5 or 2.5 microg of polyribosylribitol phosphate (PRP) conjugated to tetanus toxoid (trials Hib-052/064)., Methods: Six hundred eighty infants were randomized to receive one of 5 vaccine combinations at 6, 10, and 14 weeks of age. Of these, 351 received the same vaccine at 15-24 months of age. The immune response was evaluated on blood samples collected 1 month after the 3-dose primary course and before and 6 weeks after the booster dose. Reactogenicity was assessed during a 4-day period after each vaccine dose using diary cards., Results: After primary vaccination, all subjects had seroprotective anti-PRP antibody concentrations (> or = 0.15 microg/mL) and > 95% had concentrations > or = 1.0 microg/mL, irrespective of the PRP dose administered. Anti-PRP antibody avidity after primary vaccination and antibody persistence until the second year of life were similar among groups. The booster dose induced marked increases in anti-PRP antibody GMCs and antibody avidity, indicative of effective priming and the presence of immune memory. All vaccination regimens elicited good immune responses and comparable antibody persistence to the other vaccine antigens, with significant increases in all antibody concentrations observed after boosting. All vaccination regimens were safe, with similar overall reactogenicity profiles., Conclusion: Hib conjugate vaccines containing reduced amounts of PRP can be effectively combined with the licensed DTPw-HBV vaccine to provide protection against 5 major childhood pathogens in a single injection.

Objective: Before 1991, Alaska Native children experienced one of the highest rates of invasive Haemophilus influenzae type b disease. H influenzae type b vaccine has led to a near-elimination of invasive H influenzae type b disease in the United States. We describe challenges encountered in controlling H influenzae type b disease in Alaska and update the current status of H influenzae disease and carriage in Alaska as lessons to other populations., Patients and Methods: We reviewed data from statewide H influenzae disease surveillance conducted during 1980-2004. Vaccine coverage data were based on audits from tribal facilities and the National Immunization Survey. H influenzae type b colonization data were based on 6 carriage studies., Results: After universal infant vaccination in 1991, H influenzae type b disease among Alaska Native and non-Native children < 5 years of age decreased by 94% and 96%, respectively. After a 1996 change in H influenzae type b vaccine from polyribosylribitol phosphate-outer membrane protein conjugate vaccine to H influenzae type b oligosaccharide-CRM197 vaccine, the incidence of H influenzae type b disease increased in rural Alaska Natives from 19.8 to 91.1 cases per 100000 per year < 5 years of age. During 2001-2004, with use of polyribosylribitol phosphate-outer membrane protein conjugate vaccine, the rate of H influenzae type b disease in Alaska Native and non-Native children aged < 5 years decreased to 5.4 and 0 per 100000 per year, respectively. In postvaccine studies, H influenzae type b carriage has decreased in Alaska Native children < 5 years of age., Conclusions: H influenzae type b vaccination has resulted in a dramatic decrease in invasive H influenzae type b disease in Alaska; however, despite high rates of H influenzae type b vaccine coverage, H influenzae type b disease rates among rural Alaska Native children < 5 years of age remain higher than the rates among non-Native Alaska and other US children. Equity in disease rates may not be achieved in indigenous populations with the current vaccines unless other environmental and household factors contributing to disease transmission are addressed.

We assessed the safety and immunogenicity of a fully liquid, DTPw-HepB-Hib combination vaccine (Quinvaxem) in comparison with separately administered DTPw-Hib and hepatitis B vaccines. Infants participating in this open-label, randomized, phase II study received a primary vaccination course using a 2-3-4 month schedule. Blood samples were taken immediately prior to the first and one month after the third vaccination. Adverse events were assessed over a 7-day post-vaccination period using subject diaries. After completion of the primary vaccination course, 94.7% [95% CI: 89.8-97.7%] of infants receiving the combination vaccine achieved protective anti-HBs antibody titers (> or = 10 mIU/mL) with a mean 39-fold increase in GMTs in comparison with 99.3% [95% CI: 96.3-100%] seroprotection and a mean 29-fold GMT increase in the comparator group. Diphtheria, tetanus and Haemophilus influenzae type b (Hib) seroprotection rates and pertussis seroconversion rates were also similar between the two groups. There was no statistically significant difference in GMTs for diphtheria between the two groups, but significant differences were shown for tetanus, Hib, and pertussis with higher GMTs for each antigen observed in the comparator group. The combination vaccine was well tolerated, with fever (body temperature > or = 38 degrees C) being the most frequently reported adverse event in both the DTPw-HepB-Hib (12.5% [95% CI: 7.7-18.8%]) and comparator (12.6% [95% CI: 7.7-19.0%]) groups. This study demonstrated that the fully liquid DTPw-HepB-Hib combination vaccine has safety and immunogenicity profiles similar to the DTPw-Hib and hepatitis B vaccines when administered separately.

Objective: To identify evidence of the impact of Haemophilus influenzae type b (Hib) conjugate vaccine on the epidemiology of invasive Hib disease., Sources of Data: This review was based on a search of MEDLINE, LILACS, technical reports, national and international guidelines (publications from 1991 to 2005). The keywords Haemophilus influenzae type b, immunization, impact, and effectiveness, alone or in combination, were used to retrieve the articles. Studies published before 1991 and cited in the references of the studies reviewed were analyzed for useful information., Summary of the Findings: Introduction of the Hib conjugate vaccine produced great decline in the incidence of invasive Hib disease in childhood in countries where this vaccine was introduced into the routine immunization schedule. Nevertheless, the resurgence of invasive Hib disease in some regions has challenged several researchers to identify the reasons for this epidemiological pattern, as well as the measures to be implemented in order to avoid such a phenomenon., Conclusions: The use of Hib conjugate vaccine on a population scale has been greatly effective; nonetheless, changes in the vaccination scheme seem to be necessary to keep invasive Hib disease under control.

Sulfaminoheparosans (alternatively known as bioheparins) represent sulfated derivatives obtained from the K5 capsular polysaccharide of Escherichia coli. Previous studies have shown that these agents are structurally comparable to heparins and capable of exerting anticoagulant and antiprotease effects like heparins. Furthermore they are also able to release tissue factor pathway inhibitor (TFPI). Tissue factor (TF) plays a vital role in the pathogenesis of thrombotic and cardiovascular disorders. Anticoagulants such as heparins and bioheparins inhibit this thrombogenic mediator and thereby downregulate the activation of prothrombin and factor X. This study was carried out to determine the effects of several bioheparin fractions and heparins on TF-mediated platelet activation and their direct effect on platelets using human whole blood flow cytometry. Four different sulfaminoheparosan fractions with mean molecular weights of 20, 9, 7, and 6 kDa were tested for their inhibitory effects on platelet activation at two different concentrations (100 and 10 microg/mL). Unfractionated heparin and a low-molecular-weight heparin, tinzaparin, were also tested under the same experimental conditions for comparative modulatory responses. Fresh whole blood from healthy female and male volunteers (n = 5) was mixed with each of these agents and incubated with TF (diluted thromboplastin C) to activate platelets. Platelets were labeled with the antibodies CD61 FITC (GP IIIa) and CD62 PE (P-selectin). The data were analyzed in terms of percent platelet aggregation and platelet P-selectin expression. At 100 microg/mL, all of these agents strongly and significantly inhibited (approximately 40%) the platelet activation induced by TF in comparison to saline control. The inhibitory effects of each of these agents were slightly weaker (approximately 24% inhibition) at 10 microg/mL. The inhibitory effects of these agents on P-selectin expression correspond to their effects on platelet aggregation. At 100 microg/mL all the agents produced greater than 80% inhibition of P-selectin expression whereas at 10 microg/mL, the inhibition is greater than 70% except for bio-20 kDa, which produced less than 50% of inhibition. No molecular weight dependence was observed with bioheparin fractions in terms of inhibitory effects on platelet aggregation or P-selectin expression. None of the bioheparins and heparins exhibited any direct effects on platelets. These observations suggest that both the bioheparins and heparins are capable of inhibiting TF-mediated activation of platelets. Thus the therapeutic effects of bioheparins in the TF-mediated pathogenesis of platelet activation may be similar to those of heparins.

Background: The introduction of Hemophilus influenzae type b (Hib) conjugate vaccine as part of the routine childhood vaccination schedule in Canada has resulted in a dramatic reduction in the cases of Hib meningitis. We describe the epidemiology and outcome of bacterial meningitis in Canadian children six years after the introduction of Hib conjugate vaccine and prior to the introduction of the conjugate Streptococcus pneumoniae vaccine., Methods: A retrospective chart review from January 1998 to December 1999 of children with meningitis identified at eight Canadian tertiary care children's hospitals belonging to the PICNIC network., Results: Bacterial meningitis was documented in 104 (11%) of 970 children presenting with meningitis. The most common isolated organisms were: Streptococcus pneumoniae (54%), group B streptococci (13%), and Neisseria meningitidis (11%). The mean age was 2.2 +/- 3.5 yr. Forty seven percent of the children required admission to Intensive Care Unit (ICU), and 19% required artificial ventilation. Sequelae were documented among 32 children (31%) prior to discharge and there were 6 (5.6%) deaths attributable to meningitis and sepsis., Conclusions: Bacterial meningitis is an important cause of morbidity in Canadian children with S. pneumoniae replacing H. influenzae as the leading potentially vaccine preventable cause. Despite proper initiation of antimicrobial therapy, meningitis results in great morbidity and mortality in children in Canada.

Paediatric cases of epiglottitis declined markedly in England following the introduction of safe effective immunization against Haemophilus influenzae type b (Hib). With the recently described resurgence in Hib infections, a corresponding rise in the number of presentations of clinical epiglottitis in children was observed, although numbers were still well below those reported prior to vaccine availability. This was seen both in microbiology reports and hospital admissions data for England. In keeping with the more diverse aetiology of epiglottitis in adults, Hib vaccination had minimal impact on hospital presentations with upper airway infections in those aged 15 years and over, which showed an overall increasing trend over 10 years. The need for a high index of suspicion to allow early diagnosis of this life-threatening clinical presentation is reinforced.