Evaluation of outer membrane proteins of Pseudomonas aeruginosa as a protective agent in mice model.

The crude Outer Membrane Protein (OMP) from a strain of P. aeruginosa isolated from burn patient was purified by two different methods. One procedure involved separation of Sodium Dodecyle Sulphate (SDS) and Triton X-100, where as the other involved using lysozyme enzyme. Both methods showed very similar polypeptide pattern and the major peptide band with molecular weight of 37 KD was common in both procedures. The protein estimation of OMP extracted by lysosyme was 3 mg mL(-1) compared to 5.5 mg mL(-1) extracted by Triton-X100 method. The latter was chosen to examine for the immunogenicity study in a mice model. The efficacy of immunization with OMP and challenge with homologous strain in mice showed a very good protection compared to control mice injected with saline. The passive haemoagglutination test (PHA) in mice, injected with OMP showed increased level of antibody after the second injection and stayed constant after repeated injection. The results of this study showed that the crude OMP extracted from P. aeruginosa induced a significant protection in mice against Pseudomonas infections and could be used as a vaccine candidate.

Efficacy of immunization with outer membrane proteins for induction of pulmonary clearance of nontypeable Haemophilus influenzae in a rat respiratory model.

Three strains of nontypeable Haemophilus influenzae namely NTHi-I, NTHi-II and NTHi-III were isolated from the sputum of patients with bronchitis and identified by biochemical, serological and electron microscopy. The polypeptide patterns of isolates were compared and found to have similar sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) polypeptide patterns, although some of the bands were specific in some strains. A similar comparison was made on extracted outer membrane proteins (OMPs) on the above mentioned strains, using Triton X-100 and sodium dodecyle sulphate (SDS). It was found that the polypeptides with molecular weights of 70, 42, 33 and 27 KDa were identified as P1, P2, P4 and P5 respectively. The protein estimation of crude OMPs from the three strains were calculated, and OPM-I prepared from NTHi-I showed the highest amount of protein and was chosen for its immunogenicity in a rat respiratory model. The efficacy of immunization with OMP was determined by enhancement of pulmonary clearance of live bacteria in the rat lung. A significant protective immune response induced by OMP was observed by enhanced respiratory clearance of nontypeable H. influenzae following mucosal immunization.

Protective Immune Responses Induced in Chickens by Outer Membrane Proteins Extracted from Different Strains of Escherichia coli.

Different strains of Escherichia coli (E. coli) from human, chickens, and the common strain between human and chickens were isolated and typed with mono-specific antibody. The E. coli strains from each group of human, chicken and common between human and chicken were selected. The polypeptide patterns of selected strains were analyzed and compared with each other by Sodium dodecyle sulphate polyacrylamide gel electrophoresis (SDS - PAGE). The SDS - polyacrylamide gel electrophoresis patterns between the strains were very similar, although the densities of some of the peptide bands were different, and some were missing in some strains. A similar comparison was made on extracted outer membrane proteins (OMP) using Triton X-100 and sodium dodecyle sulphate on the above-mentioned strains. The polypeptide patterns between the strains 078 (chicken strain), 06 (human strain) and O2 (the common strain between human and chicken) were also very similar and two major bands with molecular weights of 44 KD and 25 KD were very distinctive, and seen between all the strains.The passive haemoagglutination tests (PHA) in chickens injected OMP from the common strain (O2), showed increased level of antibody after the second injection which remained constant after repeated injections.In the challenge study, with LD100 from homologous strain of O2, a significant protection was observed in the groups injected the OMP (extracted from O2 strain), compared with controls injected saline.The results of this study indicate that purified E. coli outer membrane protein, can induce a significant protection immunity against colibacillus diseases in chickens, and probably in immunocompromised hosts with repeated urinary infections.