Σύγκριση πρωτεώματος ευαίσθητων και ανθεκτικών, σε αντιβιοτικά επιλογής, στελεχών Βρουκέλλας και γονιδίων - στόχων τους

Abstract

MethodologyIn the present study we created resistant strains of Brucella abortus were created in vitro for the antibiotics doxycycline, rifampicin and streptomycin. Possible instances of cross-resistance to other antibiotics was also tested. Genes known to be involved in these antibiotics resistance were amplified and analysed. The proteomics study was performed using a gel-free shotgun comparative proteomics approach. The proteomics study results were analysed using bioinformatics methods with online software.After completion of the experiments all resistant strains were destroyed.Results - DiscussionSpecific primers were designed for the amplification of the target genes of this study. The genes examined by PCR-Sequencing are rpsL, rpoB, bepC θαη 16SrRNA. There was only one mutation detected in the rpoB gene.The rifampicin resistant strain had an MIC of 128κg/ml, the doxycycline resistant revealed an MIC of 2,4κg/ml and the MIC for the streptomycin resistant strain was 4.096κg/ml.Cross-resistance testing showed that rifampicin resistance elevates the MIC levels of trimethoprim/sulfamethoxazole. Doxycycline resistance affects almost all other groups of antibiotics and streptomycin resistance affected mostly the MIC of rifampicin.In brucella doxycycline resistance is related to the blocking of the drug entry within the cell and the restriction of all of the cells metabolic activitiesResistance in rifampicin is not only a result of mutations in the rpoB gene. Reduced susceptibility in this antibiotic is the end result of many altered metabolic processes.The comparison of the differentially expressed proteins of the two datasets revealed 10 common proteins that alter their expression levels in both instances of resistance (Doxycycline, Rifampicin).4Finally a further 180 proteins were added to the list of the experimentally detected proteins and 109 proteins have been described as worthy to investigate for possible use in diagnosis, vaccines and/or as drug targets