Limits...
Reducing Campylobacter jejuni colonization of poultry via vaccination.

Neal-McKinney JM, Samuelson DR, Eucker TP, Nissen MS, Crespo R, Konkel ME - PLoS ONE (2014)

Bottom Line: Seven days following challenge, chickens were necropsied and cecal contents were serially diluted and plated to determine the number of C. jejuni per gram of material.The sera from the chickens were also analyzed to determine the concentration and specificity of antibodies reactive against the C. jejuni SECPs.Vaccination of chickens with the CadF, FlaA, and FlpA peptides resulted in a reduction in the number of C. jejuni in the ceca compared to the non-vaccinated C. jejuni-challenged group.

View Article: PubMed Central - PubMed

Affiliation: School of Molecular Biosciences, College of Veterinary Medicine, Washington State University, Pullman, Washington, United States of America.

ABSTRACT
Campylobacter jejuni is a leading bacterial cause of human gastrointestinal disease worldwide. While C. jejuni is a commensal organism in chickens, case-studies have demonstrated a link between infection with C. jejuni and the consumption of foods that have been cross-contaminated with raw or undercooked poultry. We hypothesized that vaccination of chickens with C. jejuni surface-exposed colonization proteins (SECPs) would reduce the ability of C. jejuni to colonize chickens, thereby reducing the contamination of poultry products at the retail level and potentially providing a safer food product for consumers. To test our hypothesis, we injected chickens with recombinant C. jejuni peptides from CadF, FlaA, FlpA, CmeC, and a CadF-FlaA-FlpA fusion protein. Seven days following challenge, chickens were necropsied and cecal contents were serially diluted and plated to determine the number of C. jejuni per gram of material. The sera from the chickens were also analyzed to determine the concentration and specificity of antibodies reactive against the C. jejuni SECPs. Vaccination of chickens with the CadF, FlaA, and FlpA peptides resulted in a reduction in the number of C. jejuni in the ceca compared to the non-vaccinated C. jejuni-challenged group. The greatest reduction in C. jejuni colonization was observed in chickens injected with the FlaA, FlpA, or CadF-FlaA-FlpA fusion proteins. Vaccination of chickens with different SECPs resulted in the production of C. jejuni-specific IgY antibodies. In summary, we show that the vaccination of poultry with individual C. jejuni SECPs or a combination of SECPs provides protection of chickens from C. jejuni colonization.

Show MeSH

Related in: MedlinePlus

Domains of the C. jejuni proteins targeted for vaccination.The residues of CadF, FlaA, FlpA, and CmeC used for vaccination of chicks are shown above the diagram for each peptide. The portions used for the 6X His-tagged peptides are shown in light gray, the GST-tagged 90 mer fragments are shown in dark-gray, and the 30 mer peptides used to create the trifecta peptide are shown in black. The signal peptides (CadF, FlpA, and CmeC) or sequences required for Type 3 secretion (FlaA) were excluded from the 6X His-tagged peptides, and are shown in white. The arrows under the diagrams for CadF and FlpA indicate residues determined to be required for adherence to fibronectin. The box under the FlaA diagram represents the area chosen to create a synthetic FlaA 30 mer peptide based on the consensus sequence shown below. This region falls within the borders of the D2 and D3 domains of FlaA, which are predicted to be surface-exposed in the native protein. The 90 mer region chosen for CmeC falls within a hydrophilic region of the protein that is predicted to be surface-exposed.
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4256221&req=5

pone-0114254-g001: Domains of the C. jejuni proteins targeted for vaccination.The residues of CadF, FlaA, FlpA, and CmeC used for vaccination of chicks are shown above the diagram for each peptide. The portions used for the 6X His-tagged peptides are shown in light gray, the GST-tagged 90 mer fragments are shown in dark-gray, and the 30 mer peptides used to create the trifecta peptide are shown in black. The signal peptides (CadF, FlpA, and CmeC) or sequences required for Type 3 secretion (FlaA) were excluded from the 6X His-tagged peptides, and are shown in white. The arrows under the diagrams for CadF and FlpA indicate residues determined to be required for adherence to fibronectin. The box under the FlaA diagram represents the area chosen to create a synthetic FlaA 30 mer peptide based on the consensus sequence shown below. This region falls within the borders of the D2 and D3 domains of FlaA, which are predicted to be surface-exposed in the native protein. The 90 mer region chosen for CmeC falls within a hydrophilic region of the protein that is predicted to be surface-exposed.

Mentions: The rationale for this experimental design was to narrow the site of interest within a C. jejuni SECP to 30 residues and to test if a combination of fragments from different proteins would generate enhanced protection (i.e., a reduction in C. jejuni colonization) versus injection with an individual C. jejuni SECP. We defined the ‘site of interest’ for the CadF, FlaA, and FlpA proteins to be the site most likely to be surface exposed, immunogenic, and protective (i.e., required for protein function). Our strategy also involved generating both GST-tagged 90 mers and His-tagged full-length proteins (minus the signal peptide) for CadF, FlaA, FlpA, and CmeC (Figure 1). The trifecta (CadF-FlaA-FlpA) peptide, composed of three 30 mers, was generated only as a GST-tagged protein. The rationale for using two distinct peptides is to generate an antibody response against a specific region of the protein using the first injection with the GST-90 mers. The booster injection then used a full-length version of the peptide to enhance the immune response without including the 26 kDa GST tag, resulting in specificity against the 90 mer fragment (and not GST). The 90 mer regions used were chosen based on putative surface exposure (i.e., hydrophilicity predictions), conservation amongst C. jejuni strains, and the presence of sequences required for protein function.


Reducing Campylobacter jejuni colonization of poultry via vaccination.

Neal-McKinney JM, Samuelson DR, Eucker TP, Nissen MS, Crespo R, Konkel ME - PLoS ONE (2014)

Domains of the C. jejuni proteins targeted for vaccination.The residues of CadF, FlaA, FlpA, and CmeC used for vaccination of chicks are shown above the diagram for each peptide. The portions used for the 6X His-tagged peptides are shown in light gray, the GST-tagged 90 mer fragments are shown in dark-gray, and the 30 mer peptides used to create the trifecta peptide are shown in black. The signal peptides (CadF, FlpA, and CmeC) or sequences required for Type 3 secretion (FlaA) were excluded from the 6X His-tagged peptides, and are shown in white. The arrows under the diagrams for CadF and FlpA indicate residues determined to be required for adherence to fibronectin. The box under the FlaA diagram represents the area chosen to create a synthetic FlaA 30 mer peptide based on the consensus sequence shown below. This region falls within the borders of the D2 and D3 domains of FlaA, which are predicted to be surface-exposed in the native protein. The 90 mer region chosen for CmeC falls within a hydrophilic region of the protein that is predicted to be surface-exposed.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4256221&req=5

pone-0114254-g001: Domains of the C. jejuni proteins targeted for vaccination.The residues of CadF, FlaA, FlpA, and CmeC used for vaccination of chicks are shown above the diagram for each peptide. The portions used for the 6X His-tagged peptides are shown in light gray, the GST-tagged 90 mer fragments are shown in dark-gray, and the 30 mer peptides used to create the trifecta peptide are shown in black. The signal peptides (CadF, FlpA, and CmeC) or sequences required for Type 3 secretion (FlaA) were excluded from the 6X His-tagged peptides, and are shown in white. The arrows under the diagrams for CadF and FlpA indicate residues determined to be required for adherence to fibronectin. The box under the FlaA diagram represents the area chosen to create a synthetic FlaA 30 mer peptide based on the consensus sequence shown below. This region falls within the borders of the D2 and D3 domains of FlaA, which are predicted to be surface-exposed in the native protein. The 90 mer region chosen for CmeC falls within a hydrophilic region of the protein that is predicted to be surface-exposed.
Mentions: The rationale for this experimental design was to narrow the site of interest within a C. jejuni SECP to 30 residues and to test if a combination of fragments from different proteins would generate enhanced protection (i.e., a reduction in C. jejuni colonization) versus injection with an individual C. jejuni SECP. We defined the ‘site of interest’ for the CadF, FlaA, and FlpA proteins to be the site most likely to be surface exposed, immunogenic, and protective (i.e., required for protein function). Our strategy also involved generating both GST-tagged 90 mers and His-tagged full-length proteins (minus the signal peptide) for CadF, FlaA, FlpA, and CmeC (Figure 1). The trifecta (CadF-FlaA-FlpA) peptide, composed of three 30 mers, was generated only as a GST-tagged protein. The rationale for using two distinct peptides is to generate an antibody response against a specific region of the protein using the first injection with the GST-90 mers. The booster injection then used a full-length version of the peptide to enhance the immune response without including the 26 kDa GST tag, resulting in specificity against the 90 mer fragment (and not GST). The 90 mer regions used were chosen based on putative surface exposure (i.e., hydrophilicity predictions), conservation amongst C. jejuni strains, and the presence of sequences required for protein function.

Bottom Line: Seven days following challenge, chickens were necropsied and cecal contents were serially diluted and plated to determine the number of C. jejuni per gram of material.The sera from the chickens were also analyzed to determine the concentration and specificity of antibodies reactive against the C. jejuni SECPs.Vaccination of chickens with the CadF, FlaA, and FlpA peptides resulted in a reduction in the number of C. jejuni in the ceca compared to the non-vaccinated C. jejuni-challenged group.

View Article: PubMed Central - PubMed

Affiliation: School of Molecular Biosciences, College of Veterinary Medicine, Washington State University, Pullman, Washington, United States of America.

ABSTRACT
Campylobacter jejuni is a leading bacterial cause of human gastrointestinal disease worldwide. While C. jejuni is a commensal organism in chickens, case-studies have demonstrated a link between infection with C. jejuni and the consumption of foods that have been cross-contaminated with raw or undercooked poultry. We hypothesized that vaccination of chickens with C. jejuni surface-exposed colonization proteins (SECPs) would reduce the ability of C. jejuni to colonize chickens, thereby reducing the contamination of poultry products at the retail level and potentially providing a safer food product for consumers. To test our hypothesis, we injected chickens with recombinant C. jejuni peptides from CadF, FlaA, FlpA, CmeC, and a CadF-FlaA-FlpA fusion protein. Seven days following challenge, chickens were necropsied and cecal contents were serially diluted and plated to determine the number of C. jejuni per gram of material. The sera from the chickens were also analyzed to determine the concentration and specificity of antibodies reactive against the C. jejuni SECPs. Vaccination of chickens with the CadF, FlaA, and FlpA peptides resulted in a reduction in the number of C. jejuni in the ceca compared to the non-vaccinated C. jejuni-challenged group. The greatest reduction in C. jejuni colonization was observed in chickens injected with the FlaA, FlpA, or CadF-FlaA-FlpA fusion proteins. Vaccination of chickens with different SECPs resulted in the production of C. jejuni-specific IgY antibodies. In summary, we show that the vaccination of poultry with individual C. jejuni SECPs or a combination of SECPs provides protection of chickens from C. jejuni colonization.

Show MeSH
Related in: MedlinePlus