Protection of chickens against fowl cholera by supernatant proteins of Pasteurella multocida cultured in an iron-restricted medium

Pasteurella multocida (P. multocida), a causative agent of fowl cholera, is an important pathogen in the poultry industry. In the present study, we found that the inactivated vaccine of P. multocida grown in an iron-restricted medium provided better protection than that grown in normal medium. Thus, we adopted a comparative proteomics approach, by using two-dimensional gel electrophoresis (2-DE), coupled with matrix-assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF/TOF MS), to profile the supernatant proteins associated with P. multocida under both conditions. Eleven upregulated proteins were identified, including aspartate ammonia-lyase (AspA), diacylglycerol kinase (DgK), 30S ribosomal protein S6 (RpsF), and eight outer membrane proteins (OMPs). To further characterize the three novel supernatant proteins identified under iron-restricted conditions, the AspA, DgK and RpsF proteins were expressed and purified, and used as immunogens to vaccinate chickens. The results showed that AspA, DgK and RpsF proteins induced 80.0%, 66.7%, and 80.0% immunity, respectively. These data indicate that the three novel proteins identified in the supernatant of the culture media might play important roles in the survival of bacteria under iron-restricted conditions, and thus protect chickens against P. multocida. These findings also suggest that the proteins identified can be used as subunit vaccines.