Limits...
The vaccine potential of Bordetella pertussis biofilm-derived membrane proteins.

de Gouw D, Serra DO, de Jonge MI, Hermans PW, Wessels HJ, Zomer A, Yantorno OM, Diavatopoulos DA, Mooi FR - Emerg Microbes Infect (2014)

Bottom Line: As proof of concept, mice were vaccinated with recombinantly produced BipA.Immunization significantly reduced colonization of the lungs and antibodies to BipA were found to efficiently opsonize bacteria.Together, these data suggest that biofilm proteins and in particular BipA may be of interest for inclusion into future pertussis vaccines.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Pediatric Infectious Diseases, Department of Pediatrics, Radboud University Medical Center , Nijmegen 6500 HB, The Netherlands ; Laboratory of Medical Immunology, Department of Laboratory Medicine, Radboud University Medical Center , Nijmegen 6500 HB, The Netherlands.

ABSTRACT
Pertussis is an infectious respiratory disease of humans caused by the gram-negative pathogen Bordetella pertussis. The use of acellular pertussis vaccines (aPs) which induce immunity of relative short duration and the emergence of vaccine-adapted strains are thought to have contributed to the recent resurgence of pertussis in industrialized countries despite high vaccination coverage. Current pertussis vaccines consist of antigens derived from planktonic bacterial cultures. However, recent studies have shown that biofilm formation represents an important aspect of B. pertussis infection, and antigens expressed during this stage may therefore be potential targets for vaccination. Here we provide evidence that vaccination of mice with B. pertussis biofilm-derived membrane proteins protects against infection. Subsequent proteomic analysis of the protein content of biofilm and planktonic cultures yielded 11 proteins which were ≥three-fold more abundant in biofilms, of which Bordetella intermediate protein A (BipA) was the most abundant, surface-exposed protein. As proof of concept, mice were vaccinated with recombinantly produced BipA. Immunization significantly reduced colonization of the lungs and antibodies to BipA were found to efficiently opsonize bacteria. Finally, we confirmed that bipA is expressed during respiratory tract infection of mice, and that anti-BipA antibodies are present in the serum of convalescent whooping cough patients. Together, these data suggest that biofilm proteins and in particular BipA may be of interest for inclusion into future pertussis vaccines.

No MeSH data available.


Related in: MedlinePlus

Bacterial opsonization by BipA-specific antibodies. (A) Post-immune mouse sera (day 28) were used to determine the amount of total IgG specific for rBipA using an ELISA. Each symbol represents one mouse and the geometrical mean is represented by a horizontal line. Dashed lines indicate the lower limit of detection (93 ng/mL). *P<0.05; one-tailed Wilcoxon signed-rank test. The binding of serum IgG (B) and nasal lavage IgG (C) and IgA (D) antibodies from vaccinated mice to Bvg+ or Bvg− grown B. pertussis B1917 was determined by flow cytometry. Bars represent the geometric mean±95% CI of six individual mice. **P<0.005, ***P<0.0005 relative to PBS group; Kruskal–Wallis test followed by a Dunns post-hoc test (α=5%). (E) To determine the distribution of IgG subtypes, the amount of pertussis-specific IgG1, IgG2a and IgG2b binding to Bvg+ grown whole B1917 cells was assessed. Each symbol represents one mouse and the geometrical mean is represented by a line. Dashed lines indicate the lower limit of detection (23, 4 and 27 ng/mL for IgG1, IgG2a and IgG2b, respectively). For IgG1, a one-tailed Wilcoxon signed-rank test was performed because IgG1 levels of the PBS mice were below the detection limit. IgG2a and IgG2b levels were statistically compared to the PBS mice using a two-tailed Mann–Whitney U test. *P<0.05, **P<0.005. CI, confidence interval; FI, fluorescence index.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4150286&req=5

fig5: Bacterial opsonization by BipA-specific antibodies. (A) Post-immune mouse sera (day 28) were used to determine the amount of total IgG specific for rBipA using an ELISA. Each symbol represents one mouse and the geometrical mean is represented by a horizontal line. Dashed lines indicate the lower limit of detection (93 ng/mL). *P<0.05; one-tailed Wilcoxon signed-rank test. The binding of serum IgG (B) and nasal lavage IgG (C) and IgA (D) antibodies from vaccinated mice to Bvg+ or Bvg− grown B. pertussis B1917 was determined by flow cytometry. Bars represent the geometric mean±95% CI of six individual mice. **P<0.005, ***P<0.0005 relative to PBS group; Kruskal–Wallis test followed by a Dunns post-hoc test (α=5%). (E) To determine the distribution of IgG subtypes, the amount of pertussis-specific IgG1, IgG2a and IgG2b binding to Bvg+ grown whole B1917 cells was assessed. Each symbol represents one mouse and the geometrical mean is represented by a line. Dashed lines indicate the lower limit of detection (23, 4 and 27 ng/mL for IgG1, IgG2a and IgG2b, respectively). For IgG1, a one-tailed Wilcoxon signed-rank test was performed because IgG1 levels of the PBS mice were below the detection limit. IgG2a and IgG2b levels were statistically compared to the PBS mice using a two-tailed Mann–Whitney U test. *P<0.05, **P<0.005. CI, confidence interval; FI, fluorescence index.

Mentions: To determine the role of antibody-mediated protection by rBipA, an ELISA was performed using sera from vaccinated mice. Figure 5A shows that vaccination with rBipA induced high levels of rBipA-specific IgG. Similarly, rBipA-specific antibodies were also detected in serum from biofilm-vaccinated mice, while these were below the detection limit in serum derived from mice vaccinated with planktonic culture-derived membrane proteins (Figure 5A). Next, the ability of serum and NL-derived rBipA-specific antibodies to bind its native epitopes on the surface of Bvg+ and Bvg−B. pertussis was assessed by flow cytometry. We found significant IgG-mediated opsonization of Bvg+B. pertussis using serum, but not NL, from rBipA-vaccinated mice (Figure 5B). For aP-vaccinated mice, strong opsonization of Bvg+B. pertussis was observed with serum IgG and NL IgG and IgA (Figures 5B–5D). Both aP- and rBipA-induced antibodies showed significantly reduced binding to Bvg− bacteria, which confirms the low expression of these antigens by Bvg−-phase bacteria. These data show that rBipA vaccination-induced antibodies recognize native BipA epitopes. Since IgG binding alone does not necessarily reflect the quality of the subsequent antibody response,42 the subtype distribution of the opsonizing IgG molecules was also determined using a whole-cell ELISA.31 Analysis of the IgG1, IgG2a and IgG2b response showed that rBipA and aP antigen-specific antibodies were predominantly of the phagocytosis-mediating IgG1 subtype43,44 (Figure 5E).


The vaccine potential of Bordetella pertussis biofilm-derived membrane proteins.

de Gouw D, Serra DO, de Jonge MI, Hermans PW, Wessels HJ, Zomer A, Yantorno OM, Diavatopoulos DA, Mooi FR - Emerg Microbes Infect (2014)

Bacterial opsonization by BipA-specific antibodies. (A) Post-immune mouse sera (day 28) were used to determine the amount of total IgG specific for rBipA using an ELISA. Each symbol represents one mouse and the geometrical mean is represented by a horizontal line. Dashed lines indicate the lower limit of detection (93 ng/mL). *P<0.05; one-tailed Wilcoxon signed-rank test. The binding of serum IgG (B) and nasal lavage IgG (C) and IgA (D) antibodies from vaccinated mice to Bvg+ or Bvg− grown B. pertussis B1917 was determined by flow cytometry. Bars represent the geometric mean±95% CI of six individual mice. **P<0.005, ***P<0.0005 relative to PBS group; Kruskal–Wallis test followed by a Dunns post-hoc test (α=5%). (E) To determine the distribution of IgG subtypes, the amount of pertussis-specific IgG1, IgG2a and IgG2b binding to Bvg+ grown whole B1917 cells was assessed. Each symbol represents one mouse and the geometrical mean is represented by a line. Dashed lines indicate the lower limit of detection (23, 4 and 27 ng/mL for IgG1, IgG2a and IgG2b, respectively). For IgG1, a one-tailed Wilcoxon signed-rank test was performed because IgG1 levels of the PBS mice were below the detection limit. IgG2a and IgG2b levels were statistically compared to the PBS mice using a two-tailed Mann–Whitney U test. *P<0.05, **P<0.005. CI, confidence interval; FI, fluorescence index.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4150286&req=5

fig5: Bacterial opsonization by BipA-specific antibodies. (A) Post-immune mouse sera (day 28) were used to determine the amount of total IgG specific for rBipA using an ELISA. Each symbol represents one mouse and the geometrical mean is represented by a horizontal line. Dashed lines indicate the lower limit of detection (93 ng/mL). *P<0.05; one-tailed Wilcoxon signed-rank test. The binding of serum IgG (B) and nasal lavage IgG (C) and IgA (D) antibodies from vaccinated mice to Bvg+ or Bvg− grown B. pertussis B1917 was determined by flow cytometry. Bars represent the geometric mean±95% CI of six individual mice. **P<0.005, ***P<0.0005 relative to PBS group; Kruskal–Wallis test followed by a Dunns post-hoc test (α=5%). (E) To determine the distribution of IgG subtypes, the amount of pertussis-specific IgG1, IgG2a and IgG2b binding to Bvg+ grown whole B1917 cells was assessed. Each symbol represents one mouse and the geometrical mean is represented by a line. Dashed lines indicate the lower limit of detection (23, 4 and 27 ng/mL for IgG1, IgG2a and IgG2b, respectively). For IgG1, a one-tailed Wilcoxon signed-rank test was performed because IgG1 levels of the PBS mice were below the detection limit. IgG2a and IgG2b levels were statistically compared to the PBS mice using a two-tailed Mann–Whitney U test. *P<0.05, **P<0.005. CI, confidence interval; FI, fluorescence index.
Mentions: To determine the role of antibody-mediated protection by rBipA, an ELISA was performed using sera from vaccinated mice. Figure 5A shows that vaccination with rBipA induced high levels of rBipA-specific IgG. Similarly, rBipA-specific antibodies were also detected in serum from biofilm-vaccinated mice, while these were below the detection limit in serum derived from mice vaccinated with planktonic culture-derived membrane proteins (Figure 5A). Next, the ability of serum and NL-derived rBipA-specific antibodies to bind its native epitopes on the surface of Bvg+ and Bvg−B. pertussis was assessed by flow cytometry. We found significant IgG-mediated opsonization of Bvg+B. pertussis using serum, but not NL, from rBipA-vaccinated mice (Figure 5B). For aP-vaccinated mice, strong opsonization of Bvg+B. pertussis was observed with serum IgG and NL IgG and IgA (Figures 5B–5D). Both aP- and rBipA-induced antibodies showed significantly reduced binding to Bvg− bacteria, which confirms the low expression of these antigens by Bvg−-phase bacteria. These data show that rBipA vaccination-induced antibodies recognize native BipA epitopes. Since IgG binding alone does not necessarily reflect the quality of the subsequent antibody response,42 the subtype distribution of the opsonizing IgG molecules was also determined using a whole-cell ELISA.31 Analysis of the IgG1, IgG2a and IgG2b response showed that rBipA and aP antigen-specific antibodies were predominantly of the phagocytosis-mediating IgG1 subtype43,44 (Figure 5E).

Bottom Line: As proof of concept, mice were vaccinated with recombinantly produced BipA.Immunization significantly reduced colonization of the lungs and antibodies to BipA were found to efficiently opsonize bacteria.Together, these data suggest that biofilm proteins and in particular BipA may be of interest for inclusion into future pertussis vaccines.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Pediatric Infectious Diseases, Department of Pediatrics, Radboud University Medical Center , Nijmegen 6500 HB, The Netherlands ; Laboratory of Medical Immunology, Department of Laboratory Medicine, Radboud University Medical Center , Nijmegen 6500 HB, The Netherlands.

ABSTRACT
Pertussis is an infectious respiratory disease of humans caused by the gram-negative pathogen Bordetella pertussis. The use of acellular pertussis vaccines (aPs) which induce immunity of relative short duration and the emergence of vaccine-adapted strains are thought to have contributed to the recent resurgence of pertussis in industrialized countries despite high vaccination coverage. Current pertussis vaccines consist of antigens derived from planktonic bacterial cultures. However, recent studies have shown that biofilm formation represents an important aspect of B. pertussis infection, and antigens expressed during this stage may therefore be potential targets for vaccination. Here we provide evidence that vaccination of mice with B. pertussis biofilm-derived membrane proteins protects against infection. Subsequent proteomic analysis of the protein content of biofilm and planktonic cultures yielded 11 proteins which were ≥three-fold more abundant in biofilms, of which Bordetella intermediate protein A (BipA) was the most abundant, surface-exposed protein. As proof of concept, mice were vaccinated with recombinantly produced BipA. Immunization significantly reduced colonization of the lungs and antibodies to BipA were found to efficiently opsonize bacteria. Finally, we confirmed that bipA is expressed during respiratory tract infection of mice, and that anti-BipA antibodies are present in the serum of convalescent whooping cough patients. Together, these data suggest that biofilm proteins and in particular BipA may be of interest for inclusion into future pertussis vaccines.

No MeSH data available.


Related in: MedlinePlus