Construction And Biological Characterization Of A Omp10-deficient Brucella Suis S2Strain

Vaccination is a good means to control brucellosis.At present, domestic andforeign brucella vaccines are various,.but due to various reasons, which haveserious defect. In China, attenuated brucella suis S2strain vaccine have beenmainly used，which have broad host range, good immune effect and low virulence.But brucella suis S2strain still exist certain virulence, which is similar to the othersmooth Brucella vaccines. Between attenuated live vaccine strain and wild virulentstrain have the lack of available for identifying effective antigens, resulting inserological test which is difficult to distinguish natural infection andvaccination.Application of attenuated live vaccines have been limited, which makebrucellosis can been not fundamentally controlled.Brucella outer membrane protein OMP10is bacterial outer membranelipoprotein,which is a immunogenic protein. At present, all known brucella speciesand biological types have the lipoprotein. OMP10which is also considered closelycontact with brucella virulence.The deletion strains were identified attenuatedbrucella virulence. In addition to cattle, brucella outer membrane protein OMP10can be applied in the animal brucellosis serological test,which is one of brucelladetectable protein.Brucella attenuated live vaccine S2reform genetic gene to overcome theabove defect in the experiment. PCR amplify OMP10gene upstream anddownstream homologous arm, which are connected to the pBluescript IISK (+)plasmid. At the same time, PCR amplify sacB gene of pEMOC2plasmid, which isconnected to the pBluescript II SK (+) plasmid, successfully constructingbrucella suis S2suicide plasmid of OMP10gene deletion strains. When the suicideplasmid is translated into S2, the suicide plasmid will be linearized plasmid in thereceptor bacteria. With the same gene homologous sequence, the suicide plasmidis inserted into the chromosome of S2receptor bacteria. Then, the suicide plasmidand omp10gene occur homologous recombination. Only the suicide plasmidwhich exchange omp10gene of the S2chromosome can copy. AMP is a negative selection marker, and sacB gene is positive selection marker. A omp10-deficientBrucella suis S2strain is successfully constructed without antibiotic resistancemarker, with reducing the resistance genes,which may lead to more biologicalsafety. The gene deletion strain and vaccine strain were used to infect mice tocompare their survival capability, Results from mice infection showed that the newstrain could survive in mice, indicating the feasibility as live attenuated vaccinestrain and infection capability decreased. The conclusion is that the gene omp10may be impact on Brucella virulence and survival capability in vivo. The studyprovide a new candidate for further tagged vaccine development.