Study Of Twitching Motility And Surface Sensing Mechanisms Of Pseudomonas Aeruginosa

Most of nosocomial infection is related the formation of biofilm, which makes the study and prevention of biofilms to become an important, health-related and mul-tidisciplinary research direction. It is well established that the biofilms formation is programmable and developmental process, in which the typical life cycle of a biofilm includes five stages as below; namely,1) planktonic bacteria attach to a surface; 2) bac-teria adapt to the surface so that cells irreversibly attach to the surface; 3) bacteria start to form micocolonies on the surface; 4) micocolonies start to differentiate to form a mature biofilm; and 5) planktonic bacteria release form the mature biofilm. Deeply understand-ing each stage of biofilm formation is predicted to provide a theoretical guidance on the design, research and development of a new generation anti-bacteria and anti-biofilms materials. In this context, here, we systematically investigate two topics that have been known to greatly impact to the biofilm formation during the stage of 1) to 3); theese topics include:1) how bacteria use their twitching motilities to adapt to a surface?; 2) what is the external signal that trigger the phenotype switching of a planktonic cell to a surface-adapted cell?.Topic 1:The structure of bacterial biofilms depends on environmental conditions, such as availability of nutrients, during biofilm formation. In turn, variations in biofilm struc-ture in part reflect differences in bacterial motility during early biofilm formation. Pseu-domonas aeruginosa deprived of nutrients remain dispersed on a surface, whereas cells supplemented with additional nutrients cluster and form microcolonies. At the single-cell scale, how bacteria modify their motility to favor distinct life-cycle outcomes re-mains poorly understood. High-throughput algorithms were used to track thousands of P. aeruginosa moving using TFP on surfaces in varying nutrient conditions and hence identify four distinct motility types. A minimal stochastic model were used to repro-duces the TFP-driven motility types. Here, we report that P. aeruginosa cells differently deploy type-IV pili (TFP) to alter the distribution of motility types under different nu-trient conditions. Bacteria preferentially crawl unidirectionally under nutrient-limited conditions but preferentially stall under nutrient-supplemented conditions. These motil-ity types correlate with subcellular localization of FimX, a protein required for TFP as-sembly and implicated in environmental response. The subcellular distribution of FimX is asymmetric for unidirectional crawling, consistent with TFP assembled primarily at the leading pole, whereas for non-translational types FimX expression is symmetric or non-existent. These results are consistent with a minimal stochastic model that repro- duces the motility types from the subcellular average concentration and asymmetry of FimX. These findings reveal that P. aeruginosa deploy TFP symmetrically or asymmet-rically to modulate motility behaviors in different nutrient conditions and thereby form biofilms only where nutrients are sufficient, which greatly enhances their competitive capacity in diverse environments.Topic 2:The answer of the question that what is the external signal that trigger the pheno-type switching of a planktonic cell to a surface-adapted cell remains open. To unlock the puzzle, understanding the molecular mechanism by which bacteria sense or recognize a surface is the key step. In the last two decades, several putative mechanisms have been proposed, including that bacteria can mechanically sense a solid surface by us-ing their surface appendages or some secreted outer-membrane proteins. All of which, however, remains to be proved. By contrast, it is well established that a second mes-senger, called c-di-GMP, dominate the phenotype switching of a planktonic cell to a surface-adapted cell; i.e., a planktonic cell relative a surface-adapted cell usually pos-sesses a much lower intracellular concentration of c-di-GMP. Therefore, here, we in situ monitor the concentration of c-di-GMP in single cell scale by using a fluorescent molecular probe. We find that the after initial attachments, intracellular concentration of c-di-GMP undergoes a plateau-increase-plateau process, and this transition highly correlates to the surface density of cell. Furthermore, we exclude some factors that may cue the increase of c-di-GMP concentration, including that turn on of quorum sensing signaling, physical contact of bacteria, limited of nutrients supplement and metabolism waste accumulation. Finally, we propose that this transition may cause by the oxygen limitation, and two possible mechanisms including fluorescent protein maturation and oxygen induced genetic regulation were discussed.