title: Brucella abortus INTA2, a novel strain 19 (Delta)bp26::luc (Delta)bmp18 double mutant lacking drug resistance markers.
abstract: Brucella abortus INTA2, a novel mutant strain, was constructed by inactivation of two B. abortus S19 genes: bp26 and bmp18, with the objective of obtaining a mutant strain that could be compatible with a diagnostic test and have less residual virulence than strain 19. The double mutant was constructed by replacing a large section of the bp26 coding region with the luciferase (luc) coding gene, resulting in mutant strain B. abortus M1luc, followed by partial deletion of bmp18 coding sequence. Both genes were inactivated by allelic replacement assisted by sacB counter-selection. Luciferase expression was evaluated and confirmed that it is a valid marker in the construction of mutant strains. When B. abortus INTA2 was inoculated in BALB/c mice, significantly fewer colony forming units (CFUs) were recovered from mice spleens during initial phase of infection. No splenomegaly was observed in strain INTA2-immunized mice at any time suggesting that strain INTA2 has lost some residual virulence of the parental strain. Nevertheless, similar protection levels against virulent challenge were observed in mice immunized with strains INTA2 or S19. Although strain INTA2 would still induce O-side antibodies, it does not express BP26. This would allow differentiation of INTA2-vaccinated animals from animals infected with field strains by measuring anti-BP26 antibodies, either by an agglutination test or ELISA using BP26 as antigen. Altogether these results indicate that B. abortus INTA2 might be a promising vaccine strain against brucellosis.
authors.affiliation: Instituto de Biotecnología, Centro de Investigaciones en Ciencias Veterinarias y Agronómicas, P.O. Box 77, 1708 Morón, Buenos Aires, Argentina.
authors.forename: Eleonora
authors.initials: E
authors.lastname: Campos
authors.forename: Silvio L
authors.initials: SL
authors.lastname: Cravero
authors.forename: Laura
authors.initials: L
authors.lastname: Delgui
authors.forename: Inés
authors.initials: I
authors.lastname: Mora
authors.forename: Nancy
authors.initials: N
authors.lastname: Kahn
authors.forename: Alicia I
authors.initials: AI
authors.lastname: Arese
authors.forename: Osvaldo L
authors.initials: OL
authors.lastname: Rossetti
mesh_terms: D000818:Animals; D001428:Bacterial Vaccines / Q000276:immunology / Q000592:standards; D002003:Brucella abortus / Q000235:genetics* / Q000276:immunology / Q000472:pathogenicity*; D002006:Brucellosis / Q000175:diagnosis / Q000276:immunology / Q000517:prevention & control; D024881:Drug Resistance, Bacterial / Q000235:genetics; D005260:Female; D020868:Gene Silencing; D005819:Genetic Markers; D007114:Immunization; D008156:Luciferases / Q000032:analysis / Q000235:genetics; D008565:Membrane Proteins / Q000235:genetics*; D051379:Mice; D008807:Mice, Inbred BALB C; D016296:Mutagenesis; D010957:Plasmids / Q000235:genetics; D014774:Virulence
chemical_list: C106040:BP26 protein, Brucella; D001428:Bacterial Vaccines; D005819:Genetic Markers; D008565:Membrane Proteins; D008156:Luciferases