Genotoxic stress stimulates eDNA release via explosive cell lysis and thereby promotes streamer formation of Burkholderia cenocepacia H111 cultured in a microfluidic device.

DNA is a component of biofilms, but the triggers of DNA release during biofilm formation and how DNA contributes to biofilm development are poorly investigated. One key mechanism involved in DNA release is explosive cell lysis, which is a consequence of prophage induction. In this article, the role of explosive cell lysis in biofilm formation was investigated in the opportunistic human pathogen Burkholderia cenocepacia H111 (H111). Biofilm streamers, flow-suspended biofilm filaments, were used as a biofilm model in this study, as DNA is an essential component of their matrix. H111 contains three prophages on chromosome 1 of its genome, and the involvement of each prophage in causing explosive cell lysis of the host and subsequent DNA and membrane vesicle (MV) release, as well as their contribution to streamer formation, were studied in the presence and absence of genotoxic stress. The results show that two of the three prophages of H111 encode functional lytic prophages that can be induced by genotoxic stress and their activation causes DNA and MVs release by explosive cell lysis. Furthermore, it is shown that the released DNA enables the strain to develop biofilm streamers, and streamer formation can be enhanced by genotoxic stress. Overall, this study demonstrates the involvement of prophages in streamer formation and uncovers an often-overlooked problem with the use of antibiotics that trigger the bacterial SOS response for the treatment of bacterial infections.

Beyond the Wall: Exopolysaccharides in the Biofilm Lifestyle of Pathogenic and Beneficial Plant-Associated Pseudomonas.

The formation of biofilms results from a multicellular mode of growth, in which bacteria remain enwrapped by an extracellular matrix of their own production. Many different bacteria form biofilms, but among the most studied species are those that belong to the Pseudomonas genus due to the metabolic versatility, ubiquity, and ecological significance of members of this group of microorganisms. Within the Pseudomonas genus, biofilm studies have mainly focused on the opportunistic human pathogen Pseudomonas aeruginosa due to its clinical importance. The extracellular matrix of P. aeruginosa is mainly composed of exopolysaccharides, which have been shown to be important for the biofilm architecture and pathogenic features of this bacterium. Notably, some of the exopolysaccharides recurrently used by P. aeruginosa during biofilm formation, such as the alginate and polysaccharide synthesis loci (Psl) polysaccharides, are also used by pathogenic and beneficial plant-associated Pseudomonas during their interaction with plants. Interestingly, their functions are multifaceted and seem to be highly dependent on the bacterial lifestyle and genetic context of production. This paper reviews the functions and significance of the exopolysaccharides produced by plant-associated Pseudomonas, particularly the alginate, Psl, and cellulose polysaccharides, focusing on their equivalents produced in P. aeruginosa within the context of pathogenic and beneficial interactions.

Role of extracellular matrix components in the formation of biofilms and their contribution to the biocontrol activity of Pseudomonas chlororaphis PCL1606.

Pseudomonas chlororaphis PCL1606 (PcPCL1606) displays plant-colonizing features and exhibits antagonistic traits against soil-borne phytopathogenic fungi. Biofilm formation could be relevant for the PcPCL1606 lifestyle, and in this study the role of some putative extracellular matrix components (EMC; Fap-like fibre, alginate and Psl-like polysaccharides) in the biofilm architecture and biocontrol activity of this bacterium were determined. EMC such as the Fap-like fibre and alginate polysaccharide play secondary roles in biofilm formation in PcPCL1606, because they are not fundamental to its biofilm architecture in flow cell chamber, but synergistically they have shown to favour bacterial competition during biofilm formation. Conversely, studies on Psl-like polysaccharide have revealed that it may contain mannose, and that it is strongly involved in the PcPCL1606 biofilm architecture and niche competition. Furthermore, the Fap-like fibre and Psl-like exopolysaccharide play roles in early surface attachment and contribute to biocontrol activity against the white root rot disease caused by Rosellinia necatrix in avocado plants. These results constitute the first report regarding the study of the extracellular matrix of the PcPCL1606 strain and highlight the importance of a putative Fap-like fibre and Psl-like exopolysaccharide produced by PcPCL1606 in the biofilm formation process and interactions with the host plant root.

A Large Tn7-like Transposon Confers Hyper-Resistance to Copper in Pseudomonas syringae pv. syringae.

Copper resistance mechanisms provide an important adaptive advantage to plant pathogenic bacteria under exposure to copper treatments. Copper resistance determinants have been described in Pseudomonas syringae pv. syringae (Pss) strains isolated from mango intimately associated with 62 kb plasmids belonging to the pPT23A family (PFP). It has been previously described that the indiscriminate use of copper-based compounds promotes the selection of copper resistant bacterial strains and constitutes a selective pressure in the evolution of copper resistance determinants. Hence, we have explored in this study the copper resistance evolution and the distribution of specific genetic determinants in two different Pss mango populations isolated from the same geographical regions, mainly from southern Spain with an average of 20 years of difference. The total content of plasmids, in particular the 62 kb plasmids, and the number of copper resistant Pss strains were maintained at similar levels over the time. Interestingly, the phylogenetic analysis indicated the presence of a phylogenetic subgroup (PSG) in the Pss mango phylotype, mostly composed of the recent Pss population analyzed in this study that was strongly associated with a hyper-resistant phenotype to copper. Genome sequencing of two selected Pss strains from this PSG revealed the presence of a large Tn7-like transposon of chromosomal location, which harbored putative copper and arsenic resistance genes (COARS Tn7-like). Transformation of the copper sensitive Pss UMAF0158 strain with some putative copper resistance genes and RT-qPCR experiments brought into light the role of COARS Tn7-like transposon in the hyper-resistant phenotype to copper in Pss.IMPORTANCECopper compounds have traditionally been used as standard bactericides in agriculture in the past few decades. However, the extensive use of copper has fostered the evolution of bacterial copper resistance mechanisms. Pseudomonas syringae is a plant pathogenic bacterium used worldwide as a model to study plant-pathogen interactions. The adaption of P. syringae to plant surface environment is the most important step prior to an infection. In this scenario, copper resistance mechanisms could play a key role in improving its epiphytic survival. In this work, a novel Tn7-like transposon of chromosomal location was detected in P. syringae pv. syringae strains isolated from mango. This transposon conferred the highest resistance to copper sulfate described to date for this bacterial phytopathogen. Understanding in depth the copper resistance mechanisms and their evolution are important steps to the agricultural industry to get a better improvement of disease management strategies.

Biological role of EPS from Pseudomonas syringae pv. syringae UMAF0158 extracellular matrix, focusing on a Psl-like polysaccharide.

Pseudomonas syringae is a phytopathogenic model bacterium that is used worldwide to study plant-bacteria interactions and biofilm formation in association with a plant host. Within this species, the syringae pathovar is the most studied due to its wide host range, affecting both, woody and herbaceous plants. In particular, Pseudomonas syringae pv. syringae (Pss) has been previously described as the causal agent of bacterial apical necrosis on mango trees. Pss exhibits major epiphytic traits and virulence factors that improve its epiphytic survival and pathogenicity in mango trees. The cellulose exopolysaccharide has been described as a key component in the development of the biofilm lifestyle of the P. syringae pv. syringae UMAF0158 strain (PssUMAF0158). PssUMAF0158 contains two additional genomic regions that putatively encode for exopolysaccharides such as alginate and a Psl-like polysaccharide. To date, the Psl polysaccharide has only been studied in Pseudomonas aeruginosa, in which it plays an important role during biofilm development. However, its function in plant-associated bacteria is still unknown. To understand how these exopolysaccharides contribute to the biofilm matrix of PssUMAF0158, knockout mutants of genes encoding these putative exopolysaccharides were constructed. Flow-cell chamber experiments revealed that cellulose and the Psl-like polysaccharide constitute a basic scaffold for biofilm architecture in this bacterium. Curiously, the Psl-like polysaccharide of PssUMAF0158 plays a role in virulence similar to what has been described for cellulose. Finally, the impaired swarming motility of the Psl-like exopolysaccharide mutant suggests that this exopolysaccharide may play a role in the motility of PssUMAF0158 over the mango plant surface.

The Compound 2-Hexyl, 5-Propyl Resorcinol Has a Key Role in Biofilm Formation by the Biocontrol Rhizobacterium Pseudomonas chlororaphis PCL1606.

The production of the compound 2-hexyl-5-propyl resorcinol (HPR) by the biocontrol rhizobacterium Pseudomonas chlororaphis PCL1606 (PcPCL1606) is crucial for fungal antagonism and biocontrol activity that protects plants against the phytopathogenic fungus Rosellinia necatrix. The production of HPR is also involved in avocado root colonization during the biocontrol process. This pleiotrophic response prompted us to study the potential role of HPR production in biofilm formation. The swimming motility of PcPLL1606 is enhanced by the disruption of HPR production. Mutants impaired in HPR production, revealed that adhesion, colony morphology, and typical air-liquid interphase pellicles were all dependent on HPR production. The role of HPR production in biofilm architecture was also analyzed in flow chamber experiments. These experiments revealed that the HPR mutant cells had less tight unions than those producing HPR, suggesting an involvement of HPR in the production of the biofilm matrix.