Bactericidal antibody response against P6, protein D, and OMP26 of nontypeable<i>Haemophilus influenzae</i>after acute otitis media in otitis-prone children

Khan, M. Nadeem; Kaur, Ravinder; Pichichero, Michael E.

doi:10.1111/j.1574-695x.2012.00967.x

Cited by 25 publications

(29 citation statements)

References 37 publications

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“…Why anti-P6 antibody was not increased to the same extent as the anti-PD antibody in the same cohort may due to the different immunogenicities of the two proteins (29). Khan et al (29) compared the serum IgG levels of protein D and P6 in paired acute-and convalescent-phase sera from children with acute otitis media. They found that the increased IgG titers to protein D were significantly higher than those to P6, indicating that the immunogenicity of P6, with a low molecular weight, was inferior to that of protein D (PD).…”

Section: Discussionmentioning

confidence: 99%

“…Several OMPs of NTHi have been proposed as potential vaccine antigens on the basis of their sequence conservation, immunogenicity, and demonstration of significant protection in animal models following immunization (27). Two highly conserved proteins among NTHi strains have shown significant potential as vaccine candidates: P6 and protein D (13,14,22,(28)(29)(30). It was reported in a chinchilla model that immunization with P6 provides protection against AOM due to NTHi (31), and intranasal immunization with P6 has been shown to confer antigen-specific mucosal immunity and enhance mucosal clearance of NTHi (32).…”

Section: Discussionmentioning

confidence: 99%

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Serum Concentrations of Antibodies against Outer Membrane Protein P6, Protein D, and T- and B-Cell Combined Antigenic Epitopes of Nontypeable Haemophilus influenzae in Children and Adults of Different Ages

Chen

Shang

et al. 2016

Clin Vaccine Immunol

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e Nontypeable Haemophilus influenzae (NTHi) is one of the most common etiologies of acute otitis media, rhinosinusitis, and pneumonia. Outer membrane proteins (OMPs) are the main focus in new vaccine development against NTHi, as the H. influenzae type b (Hib) vaccine does not cover noncapsulated NTHi. The OMPs P6 and protein D are the most promising candidate antigens for an NTHi vaccine, and low antibody levels against them in serum may be correlated with infection caused by NTHi. In the current study, we measured the antibody titers against P6, protein D, and their T-and B-cell combined peptide epitopes in healthy individuals of different ages. We found that children <1 month old had the lowest antibody levels against NTHi P6, protein D, and their T-and B-cell combined antigenic epitopes. Antibody titers increased at ages 1 to 6 months, peaked at 7 months to 3 years, and remained high at 4 to 6 years. The antibody titers started to decrease after 6 years and were the lowest in the 21-to 30-year group. The geometric mean titers (GMTs) of T-and B-cell combined antigenic epitopes in P6 and protein D were positively correlated with those of the protein antigens. Among 12 peptides tested, P6-61, P6-123, and protein D-167 epitopes were better recognized than others in human serum. These findings might contribute to the development of an effective serotypeindependent vaccine for H. influenzae. Haemophilus influenzae is one of the normal inhabitants of the human nasopharynx and is responsible for pneumonia, acute otitis media (AOM), and acute rhinosinusitis (1-3). The presence or absence of a polysaccharide capsule segregates this bacterial species into two well-defined groups: one group of encapsulated strains and another group of noncapsulated strains, commonly referred to as nontypeable H. influenzae (NTHi) (3). Common infections caused by NTHi include otitis media in children and lower airway infections of chronic obstructive pulmonary disease in adults (4, 5). Vaccines composed of polysaccharide capsule conjugated to protein carriers have virtually eliminated infections caused by encapsulated H. influenzae type b, including meningitis and other systemic infections, in regions where the vaccines are widely administered (6, 7). However, these conjugate vaccines have no effect on infections caused by NTHi, and in regions with H. influenzae type b vaccination programs, nontypeable strains are now the most common cause of noninvasive H. influenzae infection, so that the development of the vaccine against NTHi is an urgent and challenging task (8-10).Since NTHi organisms are noncapsulated bacteria, the outer membrane proteins (OMPs) are the main targets for vaccine designers. Several research groups have identified conserved surface proteins and tested them as putative vaccines, and the conserved NTHi antigens with demonstrated preclinical protective capacity have been identified, among which P6 and protein D are the most widely studied (11)(12)(13)(14).Experimental data derived from humans and animal models indicate t...

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Serum Concentrations of Antibodies against Outer Membrane Protein P6, Protein D, and T- and B-Cell Combined Antigenic Epitopes of Nontypeable Haemophilus influenzae in Children and Adults of Different Ages

Chen

Shang

et al. 2016

Clin Vaccine Immunol

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“…The reciprocal bactericidal titers were compared with unadsorbed sera to determine the bactericidal activity mediated by anti-P6 antibodies. To ensure the selective depletion of only anti-P6 antibodies, polystyrene beads were adsorbed with protein D (less bactericidal) or OMP26 proteins (nonbactericidal) as a control; the depletion of anti-OMP26 antibodies did not affect the levels of either protein D-or P6-specific antibodies in the serum or vice versa (24). OMP26-depleted sera were also used (as a control) to perform the bactericidal assay; bactericidal titers before and after OMP26 adsorption were the same (data not shown), confirming that (i) P6 adsorption was specific and (ii) the reduction in the bactericidal activity as a result of the depletion was attributed to anti-P6 antibodies.…”

Section: Methodsmentioning

confidence: 99%

Dual Orientation of the Outer Membrane Lipoprotein P6 of Nontypeable Haemophilus influenzae

Michel

Snyder

Schmidt

et al. 2013

Journal of Bacteriology

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The majority of outer membrane (OM) lipoproteins in Gram-negative bacteria are tethered to the membrane via an attached lipid moiety and oriented facing in toward the periplasmic space; a few lipoproteins have been shown to be surface exposed. The outer membrane lipoprotein P6 from the Gram-negative pathogenic bacterium nontypeable Haemophilus influenzae (NTHi) is surface exposed and a leading vaccine candidate for prevention of NTHi infections. However, we recently found that P6 is not a transmembrane protein as previously thought (L. V. Michel, B. Kalmeta, M. McCreary, J. Snyder, P. Craig, M. E. Pichichero, Vaccine 29:1624 -1627, 2011). Here we pursued studies to show that P6 has a dual orientation, existing infrequently as surface exposed and predominantly as internally oriented toward the periplasmic space. Flow cytometry using three monoclonal antibodies with specificity for P6 showed surface staining of whole NTHi cells. Confocal microscopy imaging confirmed that antibodies targeted surface-exposed P6 of intact NTHi cells and not internal P6 in membrane-compromised or dead cells. Western blots of two wild-type NTHi strains and a mutant NTHi strain that does not express P6 showed that P6 antibodies do not detect a promiscuous epitope on NTHi. Depletion of targets to nonlipidated P6 significantly decreased bactericidal activity of human serum. Protease digestion of surface-exposed P6 demonstrated that P6 is predominantly internally localized in a manner similar to its homologue Pal in Escherichia coli. We conclude that P6 of NTHi is likely inserted into the OM in two distinct orientations, with the predominant orientation facing in toward the periplasm.T he outer membrane (OM) of Gram-negative bacteria is asymmetrically structured with lipopolysaccharides in its outer leaflet and phospholipids in its inner leaflet (1). The OM is also comprised of numerous lipoproteins that are typically tethered to the inner leaflet of the OM (via their N-terminally attached lipid moieties) and oriented toward the periplasmic space of the cell (1-4). However, a subset of lipoproteins, including the outer membrane protein P6 from nontypeable Haemophilus influenzae (NTHi), are surface exposed (5-8).Since its discovery in the mid-1980s, P6 has been a leading vaccine candidate for prevention of NTHi infections in humans (acute otitis media, sinusitis, acute exacerbations of chronic bronchitis, and pneumonia). P6 is a strong vaccine candidate because it is immunogenic in children and adults, it is surface exposed, and it is highly conserved among pathogenic strains (5,(8)(9)(10)(11)(12)(13)(14)(15)(16).Previous work demonstrated noncovalent binding of P6 to the peptidoglycan layer of the cell (17-19). Therefore, P6 was thought to be a transmembrane protein, able to access both intracellular and extracellular molecules by physically spanning the OM. In 2011, we demonstrated that P6 could not be a transmembrane protein based on structural and computational studies (20) utilizing the nuclear magnetic resonance (NMR) solution structu...

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“…We hypothesized and showed that the propensity to recurrent AOM could be attributed to poor adaptive immune responses following infection by the dominant otopathogens Streptococcus pneumoniae and Haemophilus influenzae . Specifically we found low or absent antibody and cellular responses to vaccine candidate antigens PhtD, PhtE, Ply and LytB but less so to PcpA of Streptococcus pneumoniae [24, 27] and to protein D and OMP26 but less so to P6 of Haemophilus influenzae [28, 29]. Also, the children exhibited poor antigen-specific memory T-cell responses to Streptococcus pneumoniae and Haemophilus influenzae antigens, although they responded normally to Staphylococcal enterotoxin B, suggesting the primary immune defect might involve multiple factors such as poor antigen presenting cell (APC) function, altered innate responses or lower toll-like receptor expression [22, 23, 26, 32, 33].…”

Section: Introductionmentioning

confidence: 99%

Challenges in vaccination of neonates, infants and young children

Pichichero

2014

Vaccine

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All neonates, infants and young children receive multiple priming doses and booster vaccinations in the 1st and 2nd year of life to prevent infections by viral and bacterial pathogens. Despite high vaccine compliance, outbreaks of vaccine-preventable infections are occurring worldwide. These data strongly argue for an improved understanding of the immune responses of neonates, infants and young children to vaccine antigens and further study of the exploitable mechanisms to achieve more robust and prolonged immunity with fewer primary and booster vaccinations in the pediatric population. This review will focus on our recent work involving infant and young child immunity following routine recommended vaccinations. The discussion will address vaccine responses with respect to four areas: (1) systemic antibody responses, (2) memory B-cell generation, (3) CD4 T-cell responses, and (4) APC function.

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Bactericidal antibody response against P6, protein D, and OMP26 of nontypeableHaemophilus influenzaeafter acute otitis media in otitis-prone children

Cited by 25 publications

References 37 publications

Serum Concentrations of Antibodies against Outer Membrane Protein P6, Protein D, and T- and B-Cell Combined Antigenic Epitopes of Nontypeable Haemophilus influenzae in Children and Adults of Different Ages

Serum Concentrations of Antibodies against Outer Membrane Protein P6, Protein D, and T- and B-Cell Combined Antigenic Epitopes of Nontypeable Haemophilus influenzae in Children and Adults of Different Ages

Dual Orientation of the Outer Membrane Lipoprotein P6 of Nontypeable Haemophilus influenzae

Challenges in vaccination of neonates, infants and young children

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