Traditional treatment of infectious diseases is based on compounds that kill or inhibit growth of bacteria. A major concern with this approach is the frequent development of resistance to antibiotics. The discovery of communication systems (quorum sensing systems) regulating bacterial virulence has afforded a novel opportunity to control infectious bacteria without interfering with growth. Compounds that can override communication signals have been found in the marine environment. Using Pseudomonas aeruginosa PAO1 as an example of an opportunistic human pathogen, we show that a synthetic derivate of natural furanone compounds can act as a potent antagonist of bacterial quorum sensing. We employed GeneChip^® microarray technology to identify furanone target genes and to map the quorum sensing regulon. The transcriptome analysis showed that the furanone drug specifically targeted quorum sensing systems and inhibited virulence factor expression. Application of the drug to P.aeruginosa biofilms increased bacterial susceptibility to tobramycin and SDS. In a mouse pulmonary infection model, the drug inhibited quorum sensing of the infecting bacteria and promoted their clearance by the mouse immune response.

Novel molecular tools have been constructed which allow for in situ detection of N-acyl homoserine lactone (AHL)-mediated quorum sensing in Pseudomonas aeruginosa biofilms. The reporter responds to AHL activation of LasR by expression of an unstable version of the green-fluorescent protein (Gfp). Gfp-based reporter technology has been applied for non-destructive, single-cell-level detection of quorum sensing in laboratory-based P. aeruginosa biofilms. It is reported that a synthetic halogenated furanone compound, which is a derivative of the secondary metabolites produced by the Australian macroalga Delisea pulchra, is capable of interfering with AHL-mediated quorum sensing in P. aeruginosa. It is demonstrated that the furanone compound specifically represses expression of a PlasB-gfp reporter fusion without affecting growth or protein synthesis. In addition, it reduces the production of important virulence factors, indicating a general effect on target genes of the las quorum sensing circuit. The furanone was applied to P. aeruginosa biofilms established in biofilm flow chambers. The Gfp-based analysis reveals that the compound penetrates microcolonies and blocks cell signalling and quorum sensing in most biofilm cells. The compound did not affect initial attachment to the abiotic substratum. It does, however, affect the architecture of the biofilm and enhances the process of bacterial detachment, leading to a loss of bacterial biomass from the substratum.

Leaf-litter decomposition is a central process in carbon cycling; however, our knowledge about the microbial regulation of this process is still scarce. Metaproteomics allows us to link the abundance and activity of enzymes during nutrient cycling to their phylogenetic origin based on proteins, the ‘active building blocks’ in the system. Moreover, we employed metaproteomics to investigate the influence of environmental factors and nutrients on the decomposer structure and function during beech litter decomposition. Litter was collected at forest sites in Austria with different litter nutrient content. Proteins were analyzed by 1-D-SDS-PAGE followed by liquid-chromatography and tandem mass-spectrometry. Mass spectra were assigned to phylogenetic and functional groups by a newly developed bioinformatics workflow, assignments being validated by complementary approaches. We provide evidence that the litter nutrient content and the stoichiometry of C:N:P affect the decomposer community structure and activity. Fungi were found to be the main producers of extracellular hydrolytic enzymes, with no bacterial hydrolases being detected by our metaproteomics approach. Detailed investigation of microbial succession suggests that it is influenced by litter nutrient content. Microbial activity was stimulated at higher litter nutrient contents via a higher abundance and activity of extracellular enzymes.

Killing Pseudomonas: Gram-negative Pseudomonas bacteria are opportunistic pathogens, and drug-resistant strains present a serious health problem. Srinivas et al. (p. 1010 ) synthesized a family of peptidomimetic antibiotics that is active only against Pseudomonas . These antibiotics do not lyse the cell membrane, but instead target an essential outer membrane protein, LptD, which plays a role in the assembly of lipopolysaccharide in the outer cell membrane. Activity in a mouse infection model suggests that the antibiotics might have therapeutic potential. In addition, LptD is widely distributed in gram-negative bacteria and so its validation as a target has the potential to drive development of antibiotics with a broader spectrum of activity against gram-negative pathogens.

Evidence shows that bacteria contribute actively to the decomposition of cellulose and hemicellulose in forest soil; however, their role in this process is still unclear. Here we performed the screening and identification of bacteria showing potential cellulolytic activity from litter and organic soil of a temperate oak forest. The genomes of three cellulolytic isolates previously described as abundant in this ecosystem were sequenced and their proteomes were characterized during the growth on plant biomass and on microcrystalline cellulose. Pedobacter and Mucilaginibacter showed complex enzymatic systems containing highly diverse carbohydrate-active enzymes for the degradation of cellulose and hemicellulose, which were functionally redundant for endoglucanases, β-glucosidases, endoxylanases, β-xylosidases, mannosidases and carbohydrate-binding modules. Luteibacter did not express any glycosyl hydrolases traditionally recognized as cellulases. Instead, cellulose decomposition was likely performed by an expressed GH23 family protein containing a cellulose-binding domain. Interestingly, the presence of plant lignocellulose as well as crystalline cellulose both trigger the production of a wide set of hydrolytic proteins including cellulases, hemicellulases and other glycosyl hydrolases. Our findings highlight the extensive and unexplored structural diversity of enzymatic systems in cellulolytic soil bacteria and indicate the roles of multiple abundant bacterial taxa in the decomposition of cellulose and other plant polysaccharides.

Burkholderia cepacia and Pseudomonas aeruginosa often co-exist as mixed biofilms in the lungs of patients suffering from cystic fibrosis (CF). Here, the isolation of random mini-Tn5 insertion mutants of B. cepacia H111 defective in biofilm formation on an abiotic surface is reported. It is demonstrated that one of these mutants no longer produces N-acylhomoserine lactones (AHLs) due to an inactivation of the cepR gene. cepR and the cepI AHL synthase gene together constitute the cep quorum-sensing system of B. cepacia. By using a gene replacement method, two defined mutants, H111-I and H111-R, were constructed in which cepI and cepR, respectively, had been inactivated. These mutants were used to demonstrate that biofilm formation by B. cepacia H111 requires a functional cep quorum-sensing system. A detailed quantitative analysis of the biofilm structures formed by wild-type and mutant strains suggested that the quorum-sensing system is not involved in the regulation of initial cell attachment, but rather controls the maturation of the biofilm. Furthermore, it is shown that B. cepacia is capable of swarming motility, a form of surface translocation utilized by various bacteria to rapidly colonize appropriate substrata. Evidence is provided that swarming motility of B. cepacia is quorum-sensing-regulated, possibly through the control of biosurfactant production. Complementation of the cepR mutant H111-R with different biosurfactants restored swarming motility while biofilm formation was not significantly increased. This result suggests that swarming motility per se is not essential for biofilm formation on abiotic surfaces.