Essential role of the VirB machinery in the maturation of the Brucella abortus-containing vacuole

Abstract

SummaryIn epithelial cells, the intracellular pathogen Brucella abortus escapes from the endocytic pathway, exploits the autophagic machinery of the host cell and establishes a unique replication niche in the endoplasmic reticulum. The molecular mechanisms underlying these processes are still poorly understood. Recently, a B. abortus type IV-related secretion system encoded by the virB operon has been described as being involved in the intracellular trafficking of the bacteria. In this study, we have analysed the intracellular pathway of B. abortus virB10 mutant strains by confocal microscopy. We demonstrate that a functional virB operon is essential for the biogenesis of the Brucella-containing vacuole. Polar mutation preventing the transcription of virB10 and downstream sequences did not allow Brucella to bypass the endocytic pathway. Consequently, polar mutant-containing vacuoles fused with lysosomes in which bacteria underwent a degradation process. In contrast, virB10 non-polar mutants were capable of avoiding interactions with the endocytic pathway but, diverging to wild-type Brucella, were unable to reach the endoplasmic reticulum to establish their intracellular replication niche and seemed to be recycled to the cell surface. Based on the two particular phenotypes described in this work, a model of maturation of the Brucella-containing vacuole is proposed.

The two-component system BvrR/BvrS essential for Brucella abortus virulence regulates the expression of outer membrane proteins with counterparts in members of the Rhizobiaceae

“…However, because BvrR͞BvrS mutants have properties indicative of overall outer membrane alteration, it cannot be excluded that deficient internalization relates to this rather than to the Omp3 deficiency by itself. It also possible that an outer membrane structural stability is necessary for the correct assembly and functioning of the VirB membrane components known to be involved in the control of intracellular trafficking (34). Although these hypotheses and observations concern the role of Omp3 in intracellular survival, they do not exclude the contribution of other outer membrane components, such as LPS, in virulence (1), or their regulation by BvrR͞BvrS.…”

“…mAbs to the following Omps were used (17)(18)(19): Omp10 (A68͞07G11͞C10), Omp16 (A68͞08C03͞ G03), Omp19 (A76͞05C10͞A08, A76͞18B02͞D06), Omp25 (A18͞13D02͞F05, A19͞12B10͞F04, A59͞05F01͞C09, A70͞ 06B05͞A07, A76͞02C12͞C11, A68͞04B10͞F05, A68͞07D11͞ B03, A68͞28G06͞C07), Omp2b (Omp36) (A63͞08D08͞C07, A63͞05A07͞A08, A63͞03H02͞B01, A63͞11E05͞D11, A63͞ 13G02͞C04, A63͞04D11͞G01, A68͞25G05͞A05), and Omp1 (Omp89) (A53͞10B02͞A01). Anti-Omp25 mAbs A18͞13D02͞ F05, A19͞12B10͞F04, and A59͞05F01͞C09 recognize a linear epitope close to the amino-terminal section (amino acids [21][22][23][24][25][26][27][28][29][30][31][32][33][34][35][36][37][38][39][40]. A mAb against the C͞Y epitope was conjugated with peroxidase (20), and was used for LPS detection.…”

Zinc Uptake System (znuA Locus) of Brucella abortus is Essential for Intracellular Survival and Virulence in Mice

“…Intracellular survival and replication are key virulence features of Brucella because mutants defective for these attributes are also avirulent [10,14]. A key aspect of the virulence of Brucella successfully bypasses the bactericidal effects of phagocytes, and their virulence and chronic infections are thought to be due to their ability to avoid the killing mechanisms within host cells [5,20].…”

Desenvolvimento e avaliação de uma cepa knockout de Brucella abortus obtida pela deleção do gene virB10

“…e fazem parte do sistema de secreção do tipo IV que, aliado ao LPS, é o principal fator de virulência (Ding et al 2003, Lapaque et al 2005. Para que ocorra a sobrevivência intracelular e a multiplicação de B. suis, B. abortus e B. melitensis foi demonstrada a essencialidade de genes virB (Boschiroli et al 2002, Wu et al 2006), a qual está envolvida no amadurecimento dos vacúolos parasitóforos, possibilitando sua interação estável com o retículo endoplasmático rugoso e o escape da via de degradação, gerada pela fusão com lisossomos (Comerci et al 2001, Celli & Gorvel 2004. Mutantes em diferentes genes virB já foram atenuados, sendo a virulência restabelecida com plasmídeo contendo toda a região virB (O'Collaghan et al 1999).…”