Abh regulation of biofilm formation by Bacillus subtilis

A biofilm can be defined as a community of bacteria that live together. In a biofilm bacteria adhere to a surface and coat themselves in a protective layer of sugars and proteins. Common examples of where biofilms can be found include dental plaque, endocarditis, which is a chronic infection of a heart valve, and catheter infections. Bacteria living in biofilms are difficult to treat in humans as they are highly resistant to many antibiotics and other antimicrobial agents. It appears that bacteria living in a biofilm 'club' together and protect each other. We are interested in understanding how bacteria coordinate and come together to form a biofilm. Information about this process should help us to understand the mechanisms that bacteria use to associate together in a community. We study the process of biofilm formation using a bacterium called Bacillus subtilis. This bacterium is commonly used as a model to understand basic processes regarding gene regulation and expression in bacteria from which fundamental principles about the regulatory mechanisms used can be drawn and applied to other bacteria. In our study a mutant strain of B. subtilis that is unable to form mature biofilms that recently been identified will be utilised. This strain can attach to a surface at the beginning stages of biofilm formation but cannot develop properly into a mature biofilm structure. We plan to investigate why the mutant is defective in forming biofilms by identifying which components are missing from the mutant that are essential for biofilm formation. We will look at the behaviour of the bacteria in the biofilm community at both the single-cell and population level in order to be able to generate a full picture of the regulation during the transition process from planktonic cells to a biofilm. Understanding how biofilms are formed by B. subtilis could help us to understand how other medically important bacteria form biofilms with the long-term goal of helping to prevent or eradicate biofilm infections.

Biofilms are surface-associated aggregations of microorganisms that are embedded in a self-produced polymeric matrix. Bacillus subtilis is a Gram-positive bacterium that has been used for many years as a model for bacterial physiology. It is an ideal organism to study the molecular mechanisms employed during the differentiation process from the planktonic state into the biofilm state. Recent studies indicate that biofilm formation by many species of bacteria is subject to complex regulation. It is of interest to determine how bacteria integrate the different signals and outputs from the multiple regulators to co-ordinate and control the complex, decision-making process of biofilm formation as at a rudimentary level it can be compared to the processes involved in the formation of a multicellular organism. In addition understanding such mechanisms could have important implications for the treatment of biofilm related infections. However, in order to be able to understand the global picture of how biofilm formation is regulated, we first need to understand the contribution that each of the individual regulators makes during biofilm formation. We have identified that the transcription factor Abh is a positive regulator of biofilm formation, thereby providing the first physiological role for Abh. The abh mutant strain attaches to a surface at the initial stages of biofilm formation at a level comparable to that of the wild type strain yet when analysed by confocal microscopy fails to form a wild type mature biofilm structure. Abh is part of a family of proteins that contains more than 30 known or potential regulators that are found in species including the pathogenic bacteria Bacillus, Listeria, Staphylococcus and Streptococcus. This proposal aims to identify at the population and single-cell level how Abh regulates biofilm formation using a combination of fluorescent reporter protein fusions, live cell imaging techniques, flow cytometry, and mutant analysis.