Wspólnotowy Serwis Informacyjny Badan i Rozwoju - CORDIS


RECOMBINANT BCG Streszczenie raportu

Project ID: ICA4-CT-2000-30032
Źródło dofinansowania: FP5-INCO 2
Kraj: Argentina

immunogenicity and protective efficacy of rBCG-babesia antigen strains

We demonstrated the capability of BCG expressing Rap-1 to stimulate an specific immune response against the B.bovis antigen in mice. The specific anti RAP-1 cellular immune response was analyzed measuring gIFN produced by antigen-stimulated splenocytes and the humoral immune response was followed up by Western blot. The groups of mice that received a prime/booster schema showed better specific immnue response. Moreover and in accordance to stability data the highest gIFN mesurements were observed in animals vaccinated with the most stable rBCG(pMip12:rap1) and boostered with rRap-1. The results are in accordance to previous publications showing a specific T limphocyte priming by BCG and its usefulnes as part of a combined vaccine strategy in order to induce long lasting humoral an cellular inmune response.

As a second step we perfomed a bovine inoculation assay of rBCG to test for protection against Babesia bovis in cattle as there is no experimental laboratory model for bovine babesiosis. Male Holstein calves of 200kg were used for evaluation of the rBCG. Five groups of 5 calves were employed for the vaccination trial.

Two groups were injected with rBCG-Rap, the three other were the control groups injected with BCG, Babesia bovis R1A (B. bovis attenuated strain) and non vaccinated animals respectively. A biopsy of the subscapular lymphonode of eight animals was performed the day after inoculation for identification of BCG by PCR and in vitro culture of the sample. Although all of the samples rendered negative results for the in vitro culture, 40% were PCR positive using specific primers for Mycobacteria .

Antibody response against rRap-1 in sera of immunized cattle were quentified by enzyme- lynked immunosorbent assays (ELISA) using rRap-1. No anti rRap-1 antibodies were detected in any of the groups until 22 days after challenge. At day 36 after challenge a slight incresase of antibody titer was detected for the BCG innoculated group and even higher in the rBCG- pMIP12-RAP-1 innoculated group.

Whether or not it is an specifici response will be confirmed by Immunoflurescence assay and Western blotting. At day 45 after innoculation production of bovine g-IFN in complete blood sample was detected using the commercial available kit. No specific stimulation was detected at that time point in any of the tested groups. Although no protection was obtained a slight improvement in clinical parameters post challenge in the 3 innoculated groups were observed in comparison with the control group. However most likely reflects a nonspecific effect associated with BCG.

This study represents the first application of rBCG technology to the development of candidate vaccines for the control of a veterinary protozoan pathogen.

The results obtained so far provide encouragement for continued research into the development of rBCG as a convenient and practical means through which to control bovine infections with B. bovis.

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