e252 13th International Congress on Infectious Diseases Abstracts, Poster Presentations 43.002 Isolation, Cloning, Expression and Immunoactivity of Periplasmic Binding Protein, FepB I. Rasooli 1,∗ , M. Alipour 2 , S.L. Mousavi Gargari 1 1 Shahed University, Tehran, Iran (Islamic Republic of) 2 Islamic Azad university, Babol, Iran (Islamic Republic of) Background: Iron, an essential element for bacterial life. Native FepB functions in the periplasm to facilitate the transfer of ferric enterobactin through the two- protein inner membrane channel FepDG into cytoplasm by FepC catalyzed ATP hydrolysis. These data support the idea that production of antibody against FepB may contribute to pro- tection against gram negative bacteria. The present study was designed to identify, produce and immunoactivate the periplasmic binding protein, FepB from Escherichia coli O157:H7 Methods: The gene coding for ferric enterobactin biding protein from E. coli O157:H7 was amplified. This gene was cloned and expressed as C-terminal His(6)-tagged protein. Results: The SDS-PAGE analysis of the total protein revealed only two distinct bands, with molecular masses of 31 kDa and 34 kDa. The Ni-NTA chromatography purified FepB and the osmotically shocked periplasmic fraction of IPTG induced cells showed only a single band of 31 kDa. Polyclonal mouse antibody was raised against the recombinant protein during 4 weeks after immunization. Western blot analysis of the recombinant FepB with mouse antiserum revealed a single band of 31 kDa. Polyclonal antibody raised against the recombinant protein reacted with bacterial FepB. Conclusion: Identification and purification of FepB helped reveal its appropriate molecular mass. Reaction of the recombinant FepB antiserum with bacterial FepB finds its immunoactive contribution to protection against gram neg- ative bacteria harbouring the FepB protein. The successful production of antibody by periplasmic product of FepB gene can find a room for further research aimed at broad spec- trum vaccines production. doi:10.1016/j.ijid.2008.05.682 43.003 Immunogenicity of Recombinant Omp28 from Brucella Melitensis in Mice P. Kaushik ∗ , P. Chaudhury, G. Shukla, D.K. Singh IVRI, Bareilly, India Recombinant Omp28 (r-Omp28) from Brucella meliten- sis produced in Escherichia coli was previously identified as group 3 proteins. In this study, we evaluated the immunogenicity of r-Omp28 in mice. This r-Omp28 was injected intramuscularly in the mice with and without CpG oligodeoxynucleotides (ODN). The booster was given on 3rd week with the same antigenic preparation. Specific anti- bodies to purified r-Omp28 were detected in mice sera by western blotting and indirect ELISA (iELISA). In addi- tion, isotype (IgG1 and IgG2a) specific antibodies were also measured by iELISA. Cellular immune response was mea- sured by lymphocyte proliferation assay (MTT assay) after in vitro stimulation of spleenocytes by r-Omp28. Immu- nization with r-Omp28 induced a vigorous immunoglobulin G (IgG) response, with higher IgG1 than IgG2a. Whereas, immunization of mice with rOmp28 + CpG induced IgG2a dominated response, suggesting the induction of a T helper 1 (Th1) response by CpG ODN. Spleenocyte from immunized mice showed significant proliferative response (3 week post booster), but it was higher (P < 0.05) in r-Omp28 + CpG than r-Omp28 immunized mice. Our results indicate that r-Omp28 is a good immunogen, capable of inducing both humoral and cellular immune response. The humoral response was biased towards Th1 type when it was co-administered with CpG ODN. doi:10.1016/j.ijid.2008.05.683 43.004 Safety and Efficacy of ML29 Reassortant Lassa Fever Vac- cine in Non-Human Primates I.S. Lukashevich 1,∗ , J.C. Zapata 1 , M. Goicochea 1 , R. Car- rion Jr. 2 , J.L. Patterson 2 , K. Brasky 2 , K. Mansfield 3 , M.S. Salvato 1 1 Institute of Human Virology, University of Maryland School of Medicine, Baltimore, MD, USA 2 Southwest Foundation for Biomedical Research, San Anto- nio, TX, USA 3 New England Primate Research Center, Harvard Medical School, Southborough, MA, USA Lassa virus (LASV) is the cause of widespread human infection in West Africa that occasionally develops into Lassa Fever (LF) and is responsible for the deaths of thousands of people each year. The ‘‘at risk’’ LASV sero-negative pop- ulation in the region may be as high as 200 million. In the general population the case fatality rate is 1-2% and can be as high as 15—30% among hospitalized patients. The sizeable disease burden, numerous imported cases worldwide, and possibility that LASV can be used as an agent of biological warfare make a strong case for vaccine development. Reassortant technology was used to make ML29 virus encoding the major antigens (NP and GPC) of LASV and the RNA polymerase and Z protein of Mopeia virus (MOPV), a naturally attenuated virus genetically related to LASV. Plaque purification and restricted numbers of cell culture passages resulted in eighteen mutations distinguishing the ML29 genome from the parental LASV and MOPV strains. These mutations additionally contributed to the attenuated phenotype. In the current study we evaluated safety, immunogenic- ity, and efficacy of the ML29 reassortant vaccine using a recently-established human LF model in common mar- mosets, Callithrix jacchus. We have shown that a single injection of ML29 induced low, transient viremia and ML29 replicated at low or moderate levels in target tissues depending on dose of the vaccination. Exposure of human PBMC with ML29 and MOPV has been used as an additional safety test. The gene expression patterns of ML29-exposed cells and control, mock-exposed cells, clustered together and maintained a significant distance from the MOPV group. A single ML29 vaccination induced specific and non-specific immune responses and completely protected monkeys in challenge experiments against fatal LF by induction of ster-