The chronicity of Brucella abortus
infection in humans and animals depends on the organism's ability to escape host defenses by gaining entry and surviving inside the macrophage. Although no human
vaccine exists for
Brucella, vaccine development in other bacteria has been based on deletions of selective nutritional as well as regulatory systems. Our goal is to develop a
vaccine for Brucella. To further this aim, we have used a
green fluorescent protein (GFP) reporter system to identify constitutively and intracellularly induced B. abortus genes. Constitutively producing gfp clones exhibited sequence homology with genes associated with
protein synthesis and metabolism (
initiation factor-1 and
tRNA ribotransferase) and detoxification (organic hydroperoxidase resistance). Of greater interest, clones negative for constitutively produced gfp in
agar were examined by fluorescence microscopy to detect promoter activity induced within macrophages 4 and 24 h following
infection. Bacterial genes activated in macrophages 4 h postinfection appear to be involved in adapting to intracellular environmental conditions. Included in this group were genes for detoxification (lactoglyglutathione
lyase gene), repair (
formamidopyrimidine-DNA glycosylase gene), osmotic protection (K(+) transport gene), and site-specific recombination (xerD gene). A gene involved in metabolism and biosynthesis (
deoxyxylulose 5'
phosphate synthase gene) was also identified. Genes activated 24 h following
infection were biosynthesis- and metabolism-associated genes (
iron binding protein and
rhizopine catabolism). Identification of B. abortus genes that are activated following macrophage invasion provides insight into Brucella pathogenesis and thus is valuable in
vaccine design utilizing selective targeted deletions of newly identified Brucella genes.