N-acylhomoserine lactone (AHL)-based quorum sensing (QS) is important for the regulation of proteobacterial virulence determinants. Thus, the inhibition of AHL synthases offers non-antibiotics-based therapeutic potentials against QS-mediated bacterial infections. In this work, functional AHL synthases of Pseudomonas aeruginosa LasI and RhlI were heterologously expressed in an AHL-negative Escherichia coli followed by assessments on their AHLs production using AHL biosensors and high resolution liquid chromatography–mass spectrometry (LCMS). These AHL-producing E. coli served as tools for screening AHL synthase inhibitors. Based on a campaign of screening synthetic molecules and natural products using our approach, three strongest inhibitors namely are salicylic acid, tannic acid and trans-cinnamaldehyde have been identified. LCMS analysis further confirmed tannic acid and trans-cinnemaldehyde efficiently inhibited AHL production by RhlI. We further demonstrated the application of trans-cinnemaldehyde inhibiting Rhl QS system regulated pyocyanin production in P. aeruginosa up to 42.06%. Molecular docking analysis suggested that trans-cinnemaldehyde binds to the LasI and EsaI with known structures mainly interacting with their substrate binding sites. Our data suggested a new class of QS-inhibiting agents from natural products targeting AHL synthase and provided a potential approach for facilitating the discovery of anti-QS signal synthesis as basis of novel anti-infective approach.

TheYersinia enterocoliticaLuxI homologue YenI directs the synthesis ofN-3-(oxohexanoyl)homoserine lactone (3-oxo-C6-HSL) andN-hexanoylhomoserine lactone (C6-HSL). In aY. enterocolitica yenImutant, swimming motility is temporally delayed while swarming motility is abolished. Since both swimming and swarming are flagellum dependent, we purified the flagellin protein from the parent andyenImutant. Electrophoresis revealed that in contrast to the parent strain, theyenImutant grown for 17 h at 26°C lacked the 45-kDa flagellin protein FleB. Reverse transcription-PCR indicated that while mutation ofyenIhad no effect onyenR,flhDC(the motility master regulator) orfliA(the flagellar sigma factor) expression,fleB(the flagellin structural gene) was down-regulated. Since 3-oxo-C6-HSL and C6-HSL did not restore swimming or swarming in theyenImutant, we reexamined theN-acylhomoserine lactone (AHL) profile ofY. enterocolitica. Using AHL biosensors and mass spectrometry, we identified three additional AHLs synthesized via YenI:N-(3-oxodecanoyl)homoserine lactone,N-(3-oxododecanoyl)homoserine lactone (3-oxo-C12-HSL), andN-(3-oxotetradecanoyl)homoserine lactone. However, none of the long-chain AHLs either alone or in combination with the short-chain AHLs restored swarming or swimming in theyenImutant. By investigating the transport of radiolabeled 3-oxo-C12-HSL and by introducing an AHL biosensor into theyenImutant we demonstrate that the inability of exogenous AHLs to restore motility to theyenImutant is not related to a lack of AHL uptake. However, both AHL synthesis and motility were restored by complementation of theyenImutant with a plasmid-borne copy ofyenI.

Aggregating and forming biofilms on biotic or abiotic surfaces are ubiquitous bacterial behaviors under various conditions. In clinical settings, persistent presence of biofilms increases the risks of healthcare-associated infections and imposes huge healthcare and economic burdens. Bacteria within biofilms are protected from external damage and attacks from the host immune system and can exchange genomic information including antibiotic-resistance genes. Dispersed bacterial cells from attached biofilms on medical devices or host tissues may also serve as the origin of further infections. Understanding how bacteria develop biofilms is pertinent to tackle biofilm-associated infections and transmission. Biofilms have been suggested as a continuum of growth modes for adapting to different environments, initiating from bacterial cells sensing their attachment to a surface and then switching cellular physiological status for mature biofilm development. It is crucial to understand bacterial gene regulatory networks and decision-making processes for biofilm formation upon initial surface attachment. Pseudomonas aeruginosa is one of the model microorganisms for studying bacterial population behaviors. Several hypotheses and studies have suggested that extracellular macromolecules and appendages play important roles in bacterial responses to the surface attachment. Here, I review recent studies on potential molecular mechanisms and signal transduction pathways for P. aeruginosa surface sensing.

Bdellovibrio bacteriovorus is a Gram-negative bacterium that is a pathogen of other Gram-negative bacteria, including many bacteria which are pathogens of humans, animals and plants. As such Bdellovibrio has potential as a biocontrol agent, or living antibiotic. B. bacteriovorus HD100 has a large genome and it is not yet known which of it encodes the molecular machinery and genetic control of predatory processes. We have tried to fill this knowledge-gap using mixtures of predator and prey mRNAs to monitor changes in Bdellovibrio gene expression at a timepoint of early-stage prey infection and prey killing in comparison to control cultures of predator and prey alone and also in comparison to Bdellovibrio growing axenically (in a prey-or host independent “HI” manner) on artificial media containing peptone and tryptone. From this we have highlighted genes of the early predatosome with predicted roles in prey killing and digestion and have gained insights into possible regulatory mechanisms as Bdellovibrio enter and establish within the prey bdelloplast. Approximately seven percent of all Bdellovibrio genes were significantly up-regulated at 30 minutes of infection- but not in HI growth- implicating the role of these genes in prey digestion. Five percent were down-regulated significantly, implicating their role in free-swimming, attack-phase physiology. This study gives the first post- genomic insight into the predatory process and reveals some of the important genes that Bdellovibrio expresses inside the prey bacterium during the initial attack.

A new class of bacteria‐attachment‐resistant materials is discovered using a multi‐generation polymer microarray methodology that reduces bacterial attachment by up to 99.3% compared with a leading commercially available silver hydrogel anti‐bacterial material. The coverage of three bacterial species, Pseudomonas aeruginosa, Staphylococcus aureus, and uropathogenic Escherichia coli is assessed.