Pseudomonas aeruginosa biofilms are composed of exopolysaccharides (EPS), exogenous DNA, and proteins that hold these communities together. P. aeruginosa produces lectins LecA and LecB, which possess affinities towards sugars found in matrix EPS and mediate adherence of P. aeruginosa to target host cells. Here, we demonstrate that LecB binds to Psl, a key matrix EPS, and this leads to increased retention of both cells and EPS in a growing biofilm. This interaction is predicted to occur between the lectin and the branched side chains present on Psl. Finally, we show that LecB coordinates Psl localization in the biofilm. This constitutes a unique function for LecB and identifies it as a matrix protein that contributes to biofilm structure through EPS interactions.

Pseudomonas aeruginosa forms multicellular aggregates or biofilms using both exopolysaccharides and the CdrA matrix adhesin. We showed for the first time that P. aeruginosa can use CdrA to build biofilms that do not require known matrix exopolysaccharides. It is appreciated that biofilm growth is protective against environmental assaults. However, little is known about how the interactions between individual matrix components aid in this protection. We found that interactions between CdrA and the exopolysaccharide Psl fortify the matrix by preventing CdrA proteolysis. When both components—CdrA and Psl—are part of the matrix, robust aggregates form that are tightly packed and protease resistant. These findings provide insight into how biofilms persist in protease-rich host environments.

Pseudomonas aeruginosa is an opportunistic, nosocomial bacterial pathogen that forms persistent infections due to the formation of protective communities, known as biofilms. Once the biofilm is formed, the bacteria embedded within it are recalcitrant to antimicrobial treatment and host immune defenses. Moreover, the presence of biofilms in wounds is correlated with chronic infection and delayed healing. The current standard of care for chronic wound infections typically involves physical disruption of the biofilm via debridement and subsequent antimicrobial treatment. The glycoside hydrolases PelAh and PslGh have been demonstrated in vitro to disrupt biofilm integrity through degradation of the key biofilm matrix exopolysaccharides Pel and Psl, respectively. Herein, we demonstrate that PslGh hydrolase therapy is a promising strategy for controlling P. aeruginosa wound infections. Hydrolase treatment of P. aeruginosa biofilms resulted in increased antibiotic efficacy and penetration into the biofilm. PslGh treatment of P. aeruginosa biofilms also improved innate immune activity leading to greater complement deposition, neutrophil phagocytosis, and neutrophil reactive oxygen species production. Furthermore, when P. aeruginosa-infected wounds were treated with a combination of PslGh and tobramycin, we observed an additive effect leading to greater bacterial clearance than treatments of tobramycin or PslGh alone. This study demonstrates that PelAh and PslGh have promising therapeutic potential and that PslGh may aid in the treatment of P. aeruginosa wound infections.

There is an increasing interest in studying interspecies bacterial interactions in diseases of animals and plants as it is believed that the great majority of bacteria found in nature live in complex communities. Plant pathologists have thus far mainly focused on studies involving single species or on their interactions with antagonistic competitors. A bacterial disease used as model to study multispecies interactions is the olive knot disease, caused by Pseudomonas savastanoi pv. savastanoi (Psv). Knots caused by Psv in branches and other aerial parts of the olive trees are an ideal niche not only for the pathogen but also for many other plant-associated bacterial species, mainly belonging to the genera Pantoea, Pectobacterium, Erwinia, and Curtobacterium. The non-pathogenic bacterial species Erwinia toletana, Pantoea agglomerans, and Erwinia oleae, which are frequently isolated inside the olive knots, cooperate with Psv in modulating the disease severity. Co-inoculations of these species with Psv result in bigger knots and better bacterial colonization when compared to single inoculations. Moreover, harmless bacteria co-localize with the pathogen inside the knots, indicating the formation of stable bacterial consortia that may facilitate the exchange of quorum sensing signals and metabolites. Here we discuss the possible role of bacterial communities in the establishment and development of olive knot disease, which we believe could be taking place in many other bacterial plant diseases.

Although the great majority of bacteria found in nature live in multispecies communities, microbiological studies have focused historically on single species or competition and antagonism experiments between different species. Future directions need to focus much more on microbial communities in order to better understand what is happening in the wild. We are using olive knot disease as a model to study the role and interaction of multispecies bacterial communities in disease establishment/development. In the olive knot, non-pathogenic bacterial species (e.g. Erwinia toletana) co-exist with the pathogen (Pseudomonas savastanoi pv. savastanoi); we have demonstrated cooperation among these two species via quorum sensing (QS) signal sharing. The outcome of this interaction is a more aggressive disease when co-inoculations are made compared with single inoculations. In planta experiments show that these two species co-localize in the olive knot, and this close proximity most probably facilitates exchange of QS signals and metabolites. In silico recreation of their metabolic pathways showed that they could have complementing pathways also implicating sharing of metabolites. Our microbiome studies of nine olive knot samples have shown that the olive knot community possesses great bacterial diversity; however. the presence of five genera (i.e. Pseudomonas, Pantoea, Curtobacterium, Pectobacterium and Erwinia) can be found in almost all samples.

Bacteria are social creatures that are able to interact and coordinate behaviors with each other in a multitude of ways. The study of such group behaviors in microbes was coined “sociomicrobiology” in 2005. Two such group behaviors in bacteria are quorum sensing (QS) and biofilm formation. At a very basic level, QS is the ability to sense bacterial density via cell-to-cell signaling using self-produced signals called autoinducers, and biofilms are aggregates of cells that are attached to one another via a self-produced, extracellular matrix. Since cells in biofilm aggregates are in close proximity, biofilms represent an ecologically relevant environment for QS. While QS is known to affect biofilm formation in both Gram-negative and Gram-positive species, in this review, we will focus exclusively on Gram-negative bacteria, with an emphasis on Pseudomonas aeruginosa. We will begin by describing QS systems in P. aeruginosa and how they affect P. aeruginosa biofilm formation. We then expand our review to other Gram-negative bacteria and conclude with interesting questions with regard to the effect of biofilms on QS.

Resolution of bacterial infections is often hampered by both resistance to conventional antibiotic therapy and hiding of bacterial cells inside biofilms, warranting the development of innovative therapeutic strategies. Here, we report the efficacy of blue laser light in eradicating Pseudomonas aeruginosa cells, grown in planktonic state, agar plates and mature biofilms, both in vitro and in vivo, with minimal toxicity to mammalian cells and tissues. Results obtained using knock-out mutants point to oxidative stress as a relevant mechanism by which blue laser light exerts its anti-microbial effect. Finally, the therapeutic potential is confirmed in a mouse model of skin wound infection. Collectively, these data set blue laser phototherapy as an innovative approach to inhibit bacterial growth and biofilm formation, and thus as a realistic treatment option for superinfected wounds.

Members of the genus Burkholderia are known to be adept at biofilm formation, which presumably assists in the survival of these organisms in the environment and the host. Biofilm formation has been linked to quorum sensing (QS) in several bacterial species. In this study, we characterized Burkholderia thailandensis biofilm development under flow conditions and sought to determine whether QS contributes to this process. B. thailandensis biofilm formation exhibited an unusual pattern: the cells formed small aggregates and then proceeded to produce mature biofilms characterized by "dome" structures filled with biofilm matrix material. We showed that this process was dependent on QS. B. thailandensis has three acyl-homoserine lactone (AHL) QS systems (QS-1, QS-2, and QS-3). An AHL-negative strain produced biofilms consisting of cell aggregates but lacking the matrix-filled dome structures. This phenotype was rescued via exogenous addition of the three AHL signals. Of the three B. thailandensis QS systems, we show that QS-1 is required for proper biofilm development, since a btaR1 mutant, which is defective in QS-1 regulation, forms biofilms without these dome structures. Furthermore, our data show that the wild-type biofilm biomass, as well as the material inside the domes, stains with a fucose-binding lectin. The btaR1 mutant biofilms, however, are negative for fucose staining. This suggests that the QS-1 system regulates the production of a fucose-containing exopolysaccharide in wild-type biofilms. Finally, we present data showing that QS ability during biofilm development produces a biofilm that is resistant to dispersion under stress conditions. IMPORTANCEThe saprophyte Burkholderia thailandensis is a close relative of the pathogenic bacterium Burkholderia pseudomallei, the causative agent of melioidosis, which is contracted from its environmental reservoir. Since most bacteria in the environment reside in biofilms, B. thailandensis is an ideal model organism for investigating questions in Burkholderia physiology. In this study, we characterized B. thailandensis biofilm development and sought to determine if quorum sensing (QS) contributes to this process. Our work shows that B. thailandensis produces biofilms with unusual dome structures under flow conditions. Our findings suggest that these dome structures are filled with a QS-regulated, fucose-containing exopolysaccharide that may be involved in the resilience of B. thailandensis biofilms against changes in the nutritional environment. In the environment, many bacteria reside in biofilms. Biofilm growth helps protect the resident bacteria from environmental stresses, such as desiccation, nutrient limitation, and predation (1). Biofilm formation is a coordinated process among community members and can differ from species to species. One key feature of biofilm communities is that they produce an extracellular matrix that serves to hold the community together. This matrix is usually composed of a mixture of extracellular DNA, exopolysaccharides, proteins...