Pseudomonas aeruginosa is a ubiquitous environmental bacterium capable of causing a variety of life-threatening human infections. The genetic basis for preferential infection of certain immunocompromised patients or individuals with cystic fibrosis by P. aeruginosa is not understood. To establish whether variation in the genomic repertoire of P. aeruginosa strains can be associated with a particular type of infection, we used a whole-genome DNA microarray to determine the genome content of 18 strains isolated from the most common human infections and environmental sources. A remarkable conservation of genes including those encoding nearly all known virulence factors was observed. Phylogenetic analysis of strain-specific genes revealed no correlation between genome content and infection type. Clusters of strainspecific genes in the P. aeruginosa genome, termed variable segments, appear to be preferential sites for the integration of novel genetic material. A specialized cloning vector was developed for capture and analysis of these genomic segments. With this capture system a site associated with the strain-specific ExoU cytotoxin-encoding gene was interrogated and an 80-kb genomic island carrying exoU was identified. These studies demonstrate that P. aeruginosa strains possess a highly conserved genome that encodes genes important for survival in numerous environments and allows it to cause a variety of human infections. The acquisition of novel genetic material, such as the exoU genomic island, through horizontal gene transfer may enhance colonization and survival in different host environments.

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Species identification within the genus Mycobacteriumand subsequent antibiotic susceptibility testing still rely on time-consuming, culture-based methods. Despite the recent development of DNA probes, which greatly reduce assay time, there is a need for a single platform assay capable of answering the multitude of diagnostic questions associated with this genus. We describe the use of a DNA probe array based on two sequence databases: one for the species identification of mycobacteria (82 unique 16S rRNA sequences corresponding to 54 phenotypical species) and the other for detectingMycobacterium tuberculosis rifampin resistance (rpoB alleles). Species identification or rifampin resistance was determined by hybridizing fluorescently labeled, amplified genetic material generated from bacterial colonies to the array. Seventy mycobacterial isolates from 27 different species and 15 rifampin-resistant M. tuberculosis strains were tested. A total of 26 of 27 species were correctly identified as well as all of the rpoB mutants. This parallel testing format opens new perspectives in terms of patient management for bacterial diseases by allowing a number of genetic tests to be simultaneously run.

The araC gene encodes a positive regulatory protein required for L-arabinose utilization in Escherichia coli. Transcription from the araC promoter has been shown to be under positive control by cAMP receptor protein and under negative control by its protein product (autoregulation). This work describes the identification of the region of the araC promoter that interacts with the cAMP receptor protein to mediate catabolite repression. A 3-base-pair deletion centered 60 base pairs from the transcriptional initiation site results in a mutant araC promoter that, in the absence of araC protein, reduces transcriptional activity when compared with the wildtype promoter and is unresponsive to various concentrations of intracellular cAMP in vivo. The same deletion results in a lowered affinity of the araC promoter for cAMP receptor protein in vitro. However, this lowered affinity for the mutant araC promoter does not result in substantial reduction of intracellular araC protein because autoregulation of the araC gene dominates catabolite repression. The 3-base-pair deletion in the cAMP receptor protein binding site of the araC promoter does not affect catabolite repression of the adjacent araBAD operon. The implications of these results on current models for expression of the araBAD operon and the araC gene are discussed.L-Arabinose utilization in the bacterium Escherichia coli B/r requires the activation of three unlinked genetic loci by a single regulatory gene, araC (1, 2). The araBAD operon encodes three enzymes that are responsible for the initial catabolism of L-arabinose; the araE and araF genes encode proteins that are responsible for the transport of L-arabinose into the bacterium. In the presence of L-arabinose, araC protein is an activator of transcription for the araBAD operon and the araE and araF genes (1, 2). In the presence or absence of L-arabinose, araC protein is a repressor of its own synthesis (3, 4). In addition to regulation by its own protein product, araC gene expression is under positive control of the cAMP receptor protein (CRP) (3).The regulatory region for the araC gene is adjacent to the regulatory region for the araBAD operon. Their respective promoters are transcribed in opposite directions (5) and their transcriptional initiation sites are separated by 147 base pairs (bp) (6). The nucleotide sequence of this region of DNA has been determined (7,8) and it will be referred to as the ara regulatory region. Binding sites for regulatory proteins have been identified between the two transcriptional initiation sites (refs. 4 and 9; see Fig. 1). The localization of the regulatory protein binding sites in the promoter region has been the basis for several proposed models of the regulation of the araBAD operon and the araC gene (4, 9, 10). The portions of these models that are relevant to this work are the following. The DNase I protection studies form the basis for the most recent models for araBAD and araC expression (4, 9). Although consistent with physiological data, these models have...