Stress induced virulence of Staphylococcus aureus and Listeria monocytogenes

Abstract: Popular Abstract in English The food we consume is hardly, if ever, sterile. From fresh produce to processed products, foods harbor one or more types of microorganisms. The presence of some ‘beneficial’ microorganisms is desirable in food production, because they contribute to the characteristics of the final product, as, for example, in the production of wine, beer or cheese. In other cases, microorganisms can be ‘spoilage’ or ‘pathogenic’, and have negative effects on food quality and safety because they cause spoilage and foodborne diseases respectively. Foodborne diseases constitute global health and economic issues and every sector relevant to food production and public health care has at some time focused on developing ways to control the dissemination of pathogenic bacteria in food. The measures employed include preservation through the application of physical (heating, freezing, drying) and chemical (low pH, preservatives) treatments (stresses) during food production in order to eliminate any pathogens present. Although these treatments have been proven effective at delivering safe and high quality products to consumers, bacteria have found ways to adapt and survive their application. The evolution of foodborne pathogens in response to preservation treatments has brought new challenges for food safety. In recent decades, new pathogenic strains have emerged, exhibiting enhanced virulence and resistance to preservation treatments. Therefore, for effective control of their dissemination in food, it is necessary to understand the stress response mechanisms bacteria employ, and the events which lead to the development of enhanced virulence and resistance. In this way, hazard analysis before and during food production will be improved and the risk of disease more efficiently controlled and reduced. The work presented in this thesis concentrates on understanding the virulence relevant to foodborne diseases of the pathogenic bacteria Staphylococcus aureus and Listeria monocytogenes. The primary interest was the impact of adverse food-related environmental conditions on gene expression. The first part of the thesis discusses S. aureus and the disease it causes through the formation of enterotoxins, staphylococcal food poisoning (SFP), with a particular focus on the production of enterotoxin A (SEA). In the second part, the adaptation and survival of L. monocytogenes in acidic and osmotic environments is investigated, and related to the expression of key stress and virulence genes. Staphylococcal food poisoning is a common foodborne disease. It is caused by consumption of food containing pre-formed enterotoxins, without live bacterial cells necessarily being present. Therefore the challenge in controlling SFP is to prevent and control the production of enterotoxins in food. Common preservation methods that are effective in the elimination of bacterial cells, have no impact on the staphylococcal enterotoxins, as they are stable against thermal treatment, acidity and proteolysis. Enterotoxin A has been reported as the cause of 80% of SFP outbreaks worldwide. A key characteristic of SEA is that it is encoded by a gene (sea) carried on Siphoviridae temperate bacteriophages. These phages are able to enhance the virulence of the host bacterial cell by altering its life cycle and expressing virulence factors such the sea gene when conditions in the environment diverge from optimal. The work presented in this thesis explored the relationship between the phage life cycle, expression of the sea gene, and production of SEA. The impact of food composition and preservatives such as salt and weak acids was assessed on the phage and the SEA produced, to highlight food processing treatments that could potentially increase the risk of SFP through the formation of high levels of SEA. In summary, it was found that the life cycle of the sea-carrying phages has a critical role in the expression of the gene and the levels of SEA produced by the S. aureus strains, and it is further affected by the conditions of the cell environment and food preservation treatments. The second part of the thesis studies another major foodborne pathogen, L. monocytogenes, which is responsible for the infectious disease, listeriosis. The pathogen is robust and can survive in harsh environments and at refrigeration temperatures, and thus constitutes a high risk for food safety. It can also adapt to adverse environments and develop resistance, which increases its persistence in food and in the processing environment. The focus of this work was the response of the pathogen to osmotic and acid stress, and how the intensity and sequence of these stresses can affect its capability for survival and growth in a new environment. The genetic events behind the phenotypic responses of the pathogen were also investigated, in order to potentially establish a correlation between them that could be used to improve control of L. monocytogenes in food. The knowledge obtained during this thesis work could be used to improve existing predictive models in the assessment of hazards during food production, and contribute to the design of effective food safety strategies.