title: The invA gene of Brucella melitensis is involved in intracellular invasion and is required to establish infection in a mouse model.
abstract: Some of the mechanisms underlying the invasion and intracellular survival of B. melitensis are still unknown, including the role of a subfamily of NUDIX enzymes, which have been described in other bacterial species as invasins and are present in Brucella spp. We have generated a mutation in the coding gene of one of these proteins, the invA gene (BMEI0215) of B. melitensis strain 133, to understand its role in virulence. HeLa cell invasion results showed that mutant strain survival was decreased 5-fold compared with that of the parental strain at 2 h pi (P<0.001). In a goat macrophage infection assay, mutant strain replication was 8-fold less than in the parental strain at 24 h pi (P<0.001); yet, at 48 h pi, no significant differences in intracellular replication were observed. Additionally, colocalization of the invA mutant with calregulin was significantly lower at 24 h pi compared with that of the parental strain. Furthermore, the mutant strain exhibited a low level of colocalization with cathepsin D, which was similar to the parental strain colocalization at 24 h pi. In vivo infection results demonstrated that spleen colonization was significantly lower with the mutant than with the parental strain. The immune response, measured in terms of antibody switching and IFN-γ transcription, was similar for Rev1 and infection with the mutant, although it was lower than the immune response elicited by the parental strain. Consequently, these results indicate that the invA gene is important during invasion but not for intracellular replication. Additionally, mutation of the invA gene results in in vivo attenuation.
authors.affiliation: Department of Microbiology and Immunology; College of Veterinary Medicine; National Autonomous University of Mexico; Mexico DF, Mexico.
authors.forename: Jorge
authors.initials: J
authors.lastname: Alva-Pérez
authors.affiliation: Department of Microbiology and Immunology; College of Veterinary Medicine; National Autonomous University of Mexico; Mexico DF, Mexico.
authors.forename: Beatriz
authors.initials: B
authors.lastname: Arellano-Reynoso
authors.affiliation: Department of Ecology of Pathogen Agents; General Hospital "Dr. Manuel Gea González"; National Health Department; Mexico DF, Mexico.
authors.forename: Rigoberto
authors.initials: R
authors.lastname: Hernández-Castro
authors.affiliation: Department of Microbiology and Immunology; College of Veterinary Medicine; National Autonomous University of Mexico; Mexico DF, Mexico.
authors.forename: Francisco
authors.initials: F
authors.lastname: Suárez-Güemes
mesh_terms: D000818:Animals; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D017347:Brucella melitensis / Q000201:enzymology* / Q000235:genetics / Q000472:pathogenicity; D002006:Brucellosis / Q000382:microbiology*; D005260:Female; D006801:Humans; D051379:Mice; D008807:Mice, Inbred BALB C; D014774:Virulence
chemical_list: D001426:Bacterial Proteins; C075518:invA protein, Bacteria
keywords: Brucella melitensis; NUDIX enzyme gene; intracellular replication; invasion; strain attenuation