One of the hallmarks of the Gram-negative bacterium Pseudomonas aeruginosa is its ability to thrive in diverse environments that includes humans with a variety of debilitating diseases or immune deficiencies. Here we report the complete sequence and comparative analysis of the genomes of two representative P. aeruginosa strains isolated from cystic fibrosis (CF) patients whose genetic disorder predisposes them to infections by this pathogen. The comparison of the genomes of the two CF strains with those of other P. aeruginosa presents a picture of a mosaic genome, consisting of a conserved core component, interrupted in each strain by combinations of specific blocks of genes. These strain-specific segments of the genome are found in limited chromosomal locations, referred to as regions of genomic plasticity. The ability of P. aeruginosa to shape its genomic composition to favor survival in the widest range of environmental reservoirs, with corresponding enhancement of its metabolic capacity is supported by the identification of a genomic island in one of the sequenced CF isolates, encoding enzymes capable of degrading terpenoids produced by trees. This work suggests that niche adaptation is a major evolutionary force influencing the composition of bacterial genomes. Unlike genome reduction seen in host-adapted bacterial pathogens, the genetic capacity of P. aeruginosa is determined by the ability of individual strains to acquire or discard genomic segments, giving rise to strains with customized genomic repertoires. Consequently, this organism can survive in a wide range of environmental reservoirs that can serve as sources of the infecting organisms.

Pseudomonas aeruginosa is a metabolically versatile bacterium that is found in a wide range of biotic and abiotic habitats. It is a major human opportunistic pathogen causing numerous acute and chronic infections. The critical traits contributing to the pathogenic potential of P. aeruginosa are the production of a myriad of virulence factors, formation of biofilms and antibiotic resistance. Expression of these traits is under stringent regulation, and it responds to largely unidentified environmental signals. This review is focused on providing a global picture of virulence gene regulation in P. aeruginosa. In addition to key regulatory pathways that control the transition from acute to chronic infection phenotypes, some regulators have been identified that modulate multiple virulence mechanisms. Despite of a propensity for chaotic behaviour, no chaotic motifs were readily observed in the P. aeruginosa virulence regulatory network. Having a ‘birds-eye’ view of the regulatory cascades provides the forum opportunities to pose questions, formulate hypotheses and evaluate theories in elucidating P. aeruginosa pathogenesis. Understanding the mechanisms involved in making P. aeruginosa a successful pathogen is essential in helping devise control strategies.

The leading cause of mortality in patients with cystic fibrosis (CF) is respiratory failure due in large part to chronic lung infection with Pseudomonas aeruginosa strains that undergo mucoid conversion, display a biofilm mode of growth in vivo and resist the infiltration of polymorphonuclear leukocytes (PMNs), which release free oxygen radicals such as H202. The mucoid phenotype among the strains infecting CF patients indicates overproduction of a linear polysaccharide called alginate. T o mimic the inflammatory environment o f the CF lung, P. aeruginosa PAO1, a typical non-mucoid strain, was grown in a biofilm. This was treated with low levels of H202, as if released by the PMNs, and the formation of mucoid variants was observed. These mucoid variants had mutations in mucA, which encodes an anti-o factor; this leads t o the deregulation of an alternative c factor (022, AlgT or AlgU) required for expression of the alginate biosynthetic operon. All of the mucoid variants tested showed the same mutation, the mucA22 allele, a common allele seen in CF isolates. The mucoid mucA22 variants, when compared to the smooth parent strain PAO1, (i) produced 2-6-fold higher levels of alginate, (ii) exhibited no detectable differences in growth rate, (iii) showed an unaltered LPS profile, (iv) were -72% reduced in the amount of inducible-P-lactamase and (v) secreted little or no LasA protease and only showed 44% elastase activity. A characteristic -54 kDa protein associated with alginate overproducing strains was identified as AlgE (Alg76) by N-terminal sequence analysis. Thus, the common phenotype of the mucoid variants, which included a genetically engineered mucA22 mutant, suggested that the only mutation incurred as a result of H202 treatment was in mucA. When a P. aeruginosa biofilm was repeatedly exposed to activated PMNs in vitro, mucoid variants were also observed, mimicking in vivo observations. Thus, PMNs and their oxygen by-products may cause P. aeruginosa t o undergo the typical adaptation to the intractable mu-coid form in the CF lung. These findings indicate that gene activation in bacteria by toxic oxygen radicals, similar to that found in plants and mammalian cells, may serve as a defence mechanism for the bacteria. This suggests that mucoid conversion is a response to oxygen radical exposure and that this response is a mechanism of defence by the bacteria. This is the f i r s t report to show that PMNs and their oxygen radicals can cause this phenotypic and genotypic change which is so typical of the intractable form o f P. aeruginosa in the CF lung. These findings may provide a basis for the development of anti-oxidant and anti-inflammatory therapy for the early stages of infection in CF patients.

Quorum sensing (QS) is a key regulator of virulence and biofilm formation in Pseudomonas aeruginosa and other medically relevant bacteria. Aqueous extracts of six plants, Conocarpus erectus, Chamaesyce hypericifolia, Callistemon viminalis, Bucida buceras, Tetrazygia bicolor, and Quercus virginiana, were examined in this study for their effects on P. aeruginosa virulence factors and the QS system. C. erectus, B. buceras, and C. viminalis caused a significant inhibition of LasA protease, LasB elastase, pyoverdin production, and biofilm formation. Additionally, each plant presented a distinct effect profile on the las and rhl QS genes and their respective signaling molecules, suggesting that different mechanisms are responsible for efficacy. Extracts of all plants caused the inhibition of QS genes and QS-controlled factors, with marginal effects on bacterial growth, suggesting that the quorum-quenching mechanisms are unrelated to static or cidal effects.Pneumonia due to microbial infections is a major cause of morbidity and mortality in immunocompromised patients. Pseudomonas aeruginosa hails as the leading pathogen among patients with cystic fibrosis, diffused panbronchitis, and chronic obstructive pulmonary disease (16,29,37). In addition, P. aeruginosa remains one of the major causes of nosocomial infections (10). The success of this organism is largely due to the production of a myriad of virulence factors (including LasA protease, LasB elastase, pyoverdin, pyocyanin, and alginate) and its ability to form intractable biofilms (38).Expression of many of the virulence factors in P. aeruginosa is controlled by a quorum-sensing (QS) system (59), an intercellular communication scheme in which bacteria are able to detect the population density (via signaling molecules and receptors) and control gene expression accordingly (55). P. aeruginosa elaborates two main sets of QS systems: lasI-lasR and rhlI-rhlR (55). LasI and RhlI are synthetases that manufacture the autoinducer signaling molecules N-(3-oxododecanoyl)-L-homoserine lactone (OdDHL) and N-butanoyl-L-homoserine lactone (BHL), respectively. These molecules diffuse out into the environment, and when they reach a putative threshold concentration, they activate the receptors lasR and rhlR. These receptors, in turn, coordinate the regulation of pathogenicity. A third signal, the Pseudomonas quinolone signal, also plays an integral role in the QS system (50). This secondary metabolite of P. aeruginosa is incorporated into the QS hierarchy in times of cell stress (43). This intricate communication system of P. aeruginosa is mirrored in many gramnegative pathogenic bacteria, where it coordinates the regulation of virulence, including motility, biofilm formation, and toxin production (18,19,21,48,54).The misuse and abuse of antibiotics in pharmacotherapy have led to the development of widespread resistance in the target organism. The failure of existing antibiotics to control infection makes it crucial to find alternatives to currently available drugs. Since pathogenici...

SUMMARYThis review focuses on the era of antibiosis that led to a better understanding of bacterial morphology, in particlar the cell wall component peptidoglycan. This is an effort to take readers on a tour de force from the concept of antibiosis, to the serepidity of antibiotics, evolution of betalactam development, and the molecular biology of antibiotic resistance. These areas of research have culminated in a deeper understanding of microbiology, particularly in the area of bacterial cell wall synthesis and recycling. In spite of this knowledge, which has enabled design of new even more effective therapeutics to combat bacterial infection and has provided new research tools, antibiotic resistance remains a worldwide health care problem.