The Staphylococcus aureus response to bactericidal fatty acids and its role in pathogenicity

The bacterium Staphylococcus aureus is sometimes called a superbug because it causes a large amount of disease in mostly hospitalised patients and can often resist the drugs used to treat it. In the UK, the government estimates that at least 5,000 people die every year from S. aureus infections caught in hospital. Because the bacteria can quickly become resistant to the drugs or antibiotics that are used to treat it, we must develop new ones and this requires that we understand more about how the bacterium causes disease. In this study we aim to examine how S. aureus responds to and survives the killing capacity of linoleic acid which is a chemical called a fatty acid. It is toxic to the bacteria and is believed to be important for controlling the growth of bacteria in an abscess. An abscess is a structure that forms around the bacteria in the body if the immune cells that attempt to consume them are not successful. These immune cells are targeted to the abscess but are not always successful because of the dead cells and debris resulting from unsuccessful clearance of the bacteria, and this prevents them getting at the bacteria in the core of the abscess. The bacteria would rapidly increase in number if their growth was not limited and one factor that limits their growth is linoleic acid. S. aureus attempts to survive damage from linoleic acid by producing an enzyme to convert the toxic linoleic acid into a less toxic fatty acid. The bacterium also produces a large amount of slime around it that might protect it from being removed by immune cells. One of the less toxic fatty acids, oleic acid, also stimulates the cells to produce large amounts of slime. The killing effect of the fatty acid on the cell is thought to be due to it making the membrane around the cell more fluid. This will destroy the ability of the bacterium to regulate processes essential for life so it responds by producing a pigment that restores fluidity of the membrane back to normal. The bacteria have a large lattice structure around them called a cell wall that protects them and maintains their shape and structure. Linoleic acid interferes with the production of this structure and this might also account for toxicity. It is not understood how the bacterium controls its responses to survive damaging levels of linoleic acid. We have found there are more components important for survival than those that are already known. In the study that is proposed, we aim to identify the changes in the S. aureus proteins that result from exposure to linoleic acid. This is important to determine the extent and different types of processes that are affected in the cell. To complement this study of proteins we will also study a large bank of uncharacterised mutants and determine which of these have a defect in survival after exposure to linoleic acid. Mutant S. aureus that are defective in survival will tell us about the processes that must occur for the cell to adapt and respond to its toxic effect. The different mutants obtained and some that have already been identified will be thoroughly characterised to find out more about the effect that the defect has on the cell. This will be done using specific tests for altered behaviour that will provide information about the survival strategy towards linoleic acid . To determine how several identified components actually contribute to abscess formation by S. aureus, a mouse model of infection will be used to compare normal and mutant strains to look for differences in severity of infection. In this way we will learn more about the features that make S. aureus such a harmful pathogen.

A common feature of many S. aureus infections is the development of an abscess, which envelops the bacteria in a vascularised compartment. Phagocytes attempt to resolve the infection within the abscess and the presence of elevated levels of bactericidal long-chain free fatty acids (LC-FFAs) limits bacterial growth. The proposed research will characterise the response of S. aureus to linoleic acid, a bactericidal LC-FFA found in abscesses, by identifying differential protein synthesis using proteomic and mass spectrometry methods. Using transposon mutagenesis, mutants will be identified with reduced fatty acid resistance to add to several key components identified already. The phenotype of mutants will be thoroughly characterised with respect to the nature of the survival defects and their influence on identified changes in the cell biology of S. aureus exposed to linoleic acid. The contribution of several thoroughly characterised key components to pathogenicity will be achieved using an established model of infection.