Biomedical subjects

Michel Ramuz

Publications and source records attributed to Michel Ramuz.

8 recordsLinked to original sources

PubMed2005

Identification of a new virulence factor, BvfA, in Brucella suis.

We report the identification of BvfA (for Brucella virulence factor A), a small periplasmic protein unique to the genus Brucella, which is essential for the virulence of Brucella suis. A BvfA knockout mutant was highly attenuated both in in vitro macrophage infection assays and in vivo in the murine model of brucellosis. Fluorescence-activated cell sorting analysis with green fluorescent protein fusions showed that the expression of bvfA is induced within macrophages by phagosome acidification and coregulated with the B. suis virB operon, suggesting that it too may play a role in the establishment of the intracellular replication niche.

Animals↗

PubMed2003

What is the nature of the replicative niche of a stealthy bug named Brucella?

Brucella spp. are facultatively intracellular bacteria that persist and multiply in the macrophages of their mammalian hosts. The so-called phagosome to which they have adapted is their natural living niche. Characterization of this niche would facilitate an understanding of the true relationship between the host cell and the intracellular bacteria. This Opinion analyses and discusses the characteristic properties and genesis of this vacuole during phagocytosis as deduced from the virulence factors necessary for intracellular multiplication of the pathogen. We conclude that the replicative niche of Brucella spp.--the 'brucellosome'--differs from all other cellular organelles, and that it isolates the pathogen from certain cytoplasmic nutrients. Adaptation to the stress conditions encountered and the use of anaerobic respiration enable brucellae to replicate in the compartment they create.

Brucella↗

PubMed2003

Atypical 16S rRNA gene copies in Ochrobactrum intermedium strains reveal a large genomic rearrangement by recombination between rrn copies.

Ochrobactrum intermedium is an opportunistic human pathogen belonging to the alpha 2 subgroup of proteobacteria. The 16S rDNA sequences of nine O. intermedium isolates from a collection of clinical and environmental isolates exhibited a 46-bp insertion at position 187, which was present in only one sequence among the 82 complete or partial 16S rDNA sequences of Ochrobactrum spp. available in data banks. Reverse transcription-PCR experiments showed that the 46-bp insertion remained in the 16S rRNA. The inserted sequence folded into a stem-loop structure, which took place in and prolonged helix H184 of the 16S rRNA molecule. Helix H184 has been described as conserved in length among eubacteria, suggesting the idiosyncratic character of the 46-bp insertion. Pulsed-field gel electrophoresis experiments showed that seven of the clinical isolates carrying the 46-bp insertion belonged to the same clone. Insertion and rrn copy numbers were determined by hybridization and I-CeuI digestion. In the set of clonal isolates, the loss of two insertion copies revealed the deletion of a large genomic fragment of 150 kb, which included one rrn copy; deletion occurred during the in vivo evolution of the clone. Determination of the rrn skeleton suggested that the large genomic rearrangement occurred during events involving homologous recombination between rrn copies. The loss of insertion copies suggested a phenomenon of concerted evolution among heterogeneous rrn copies.

Alphaproteobacteria↗

PubMed2002

Type IV secretion and Brucella virulence.

The type IV secretion system, encoded by the virB region, is a key virulence factor for Brucella. The 12 genes of the region form an operon that is specifically induced by phagosome acidification in cells after phagocytosis. We speculate that the system serves to secrete unknown effector molecules, which allow Brucella to pervert the host cell endosomal pathways and to create a novel intracellular compartment in which it can replicate.

Animals↗

PubMed2002

The Brucella genome at the beginning of the post-genomic era.

The year 2002 began with the publication of the first complete genome sequence for a Brucella species, that of the two replicons of B. melitensis 16M. Hopefully in 2002, the complete genome of B. suis 1330, and, perhaps, a B. abortus strain will be published. This is the culmination of over 30 years investigation of the composition, structure, organisation and evolution of the Brucella genome. Brucella research must now adapt to the new challenges of the post-genomic era.

Brucella↗

PubMed2002

The analysis of the intramacrophagic virulome of Brucella suis deciphers the environment encountered by the pathogen inside the macrophage host cell.

The pathogen Brucella suis resides and multiplies within a phagocytic vacuole of its host cell, the macrophage. The resulting complex relationship has been investigated by the analysis of the set of genes required for virulence, which we call intramacrophagic virulome. Ten thousand two hundred and seventy-two miniTn5 mutants of B. suis constitutively expressing gfp were screened by fluorescence microscopy for lack of intracellular multiplication in human macrophages. One hundred thirty-one such mutants affected in 59 different genes could be isolated, and a function was ascribed to 53 of them. We identified genes involved in (i) global adaptation to the intracellular environment, (ii) amino acid, and (iii) nucleotide synthesis, (iv) sugar metabolism, (v) oxidoreduction, (vi) nitrogen metabolism, (vii) regulation, (viii) disulphide bond formation, and (ix) lipopolysaccharide biosynthesis. Results led to the conclusion that the replicative compartment of B. suis is poor in nutrients and characterized by low oxygen tension, and that nitrate may be used for anaerobic respiration. Intramacrophagic virulome analysis hence allowed the description of the nature of the replicative vacuole of the pathogen in the macrophage and extended our understanding of the niche in which B. suis resides. We propose calling this specific compartment "brucellosome."

Adaptation, Physiological↗

PubMed2002

The Brucella suis virB operon is induced intracellularly in macrophages.

A type IV secretion system similar to the VirB system of the phytopathogen Agrobacterium tumefaciens is essential for the intracellular survival and multiplication of the mammalian pathogen Brucella. Reverse transcriptase-PCR showed that the 12 genes encoding the Brucella suis VirB system form an operon. Semiquantitative measurements of virB mRNA levels by slot blotting showed that transcription of the virB operon, but not the flanking genes, is regulated by environmental factors in vitro. Flow cytometry used to measure green fluorescent protein expression from the virB promoter confirmed the data from slot blots. Fluorescence-activated cell sorter analysis and fluorescence microscopy showed that the virB promoter is induced in macrophages within 3 h after infection. Induction only occurred once the bacteria were inside the cells, and phagosome acidification was shown to be the major signal inducing intracellular expression. Because phagosome acidification is essential for the intracellular multiplication of Brucella, we suggest that it is the signal that triggers the secretion of unknown effector molecules. These effector molecules play a role in the remodeling of the phagosome to create the unique intracellular compartment in which Brucella replicates.

Bacterial Proteins↗

PubMed2002

[Brucella at the dawn of the third milenium: genomic organization and pathogenesis].

Bacteria of the genus Brucella, responsible for brucellosis, are pathogenic for animals and occasionally for humans. The cost of this widespread zoonotic infection is still very high for the community. Over the last few years, there have been advances in two main domains. First, the Brucella genome has been shown to be complex, with two circular chromosomes. Second, recent data on the virulence of Brucella suggest common mechanisms shared with plant pathogens and endosymbionts of the alpha-proteobacteria. Understanding virulence will have practical repercussions in the realms of vaccine development and, perhaps, development of new antibiotics. Two complete Brucella genome sequences are now available and will be a gold mine of information to guide future research.

Animals↗