Related Experiment Video
Updated: Jun 3, 2026
Directed Protein Packaging within Outer Membrane Vesicles from Escherichia coli: Design, Production and Purification
Published on: November 16, 2016
Cloning, expression and protective capacity of 37 kDa outer membrane protein gene (ompH) of Pasteurella multocida
H Y Tan1, N H Nagoor, S D Sekaran
1Department of Medical Microbiology, Faculty of Medicine, University of Malaya, 50603 Kuala Lumpur, Malaysia.
Abstract:
The major outer membrane protein (OmpH) of 4 local Malaysian strains of Pasteurella multocida serotype B:2 were characterized in comparison to ATCC strains. Three major peptide bands of MW 26, 32 and 37 kDa were characterized using SDSPAGE. Two of these fragments, the 32 kDa and 37 kDa were observed to be more reactive with a mouse polyclonal antiserum in all of the local isolates as well as the ATCC strains in a Western blot. However, the 32 kDa fragment was found to cross react with other Gram negative bacteria. Therefore, the 37 kDa OmpH was selected as vaccine candidate. The 37 kDa ompH gene of the isolated strain 1710 was cloned into an Escherichia coli expression vector to produce large amounts of recombinant OmpH (rOmpH). The 37 kDa ompH gene of strain 1710 was sequenced. In comparison to a reference strain X-73 of the ompH of P. multocida, 39bp was found deleted in the 37 kDa ompH gene. However, the deletion did not shift the reading frame or change the amino acid sequence. The rOmpH was used in a mice protection study. Mice immunized and challenged intraperitoneally resulted 100% protection against P. multocida whilst mice immunized subcutaneously and challenged intraperitoneally only resulted 80% protection. The rOmpH is therefore a suitable candidate for vaccination field studies. The same rOmpH was also used to develop a potential diagnostic kit in an ELISA format.
Insights
A novel outer membrane protein H (OmpH) from Pasteurella multocida was identified as a promising vaccine candidate. Recombinant OmpH demonstrated high protection rates in mice and potential for diagnostic kit development.
Area of Science:
- Veterinary Microbiology
- Bacterial Pathogenesis
- Vaccine Development
Background:
- Pasteurella multocida serotype B:2 is a significant pathogen in livestock.
- Outer membrane proteins (OMPs) are key targets for vaccine development.
- Characterization of OmpH from local Malaysian strains is crucial for effective disease control.
Purpose of the Study:
- To characterize the major outer membrane protein (OmpH) of local Malaysian Pasteurella multocida strains.
- To evaluate the immunogenic and protective potential of a recombinant OmpH (rOmpH) as a vaccine candidate.
- To explore the utility of rOmpH in developing a diagnostic kit.
Main Methods:
- Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and Western blotting were used to characterize OmpH.
- The ompH gene was cloned into an Escherichia coli expression vector for recombinant protein production.
- Mice were immunized with rOmpH and challenged to assess protection.
- Enzyme-linked immunosorbent assay (ELISA) was explored for diagnostic applications.
Main Results:
- A 37 kDa OmpH fragment showed high reactivity and was selected as a vaccine candidate.
- Recombinant OmpH (rOmpH) was successfully produced.
- Mice immunized intraperitoneally with rOmpH showed 100% protection against Pasteurella multocida challenge.
- Subcutaneous immunization resulted in 80% protection.
- The rOmpH showed potential for ELISA-based diagnostics.
Conclusions:
- The 37 kDa OmpH is a suitable candidate for Pasteurella multocida serotype B:2 vaccine development.
- Recombinant OmpH offers significant protection and warrants further field studies.
- rOmpH can be utilized for developing diagnostic tools for Pasteurella multocida infections.
