Probing the role of commensal intestinal fungi during Salmonella pathogenesis

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PROJECT SUMMARY Despite modern sanitation and hygiene, gastrointestinal infections continue to be a significant health problem in the United States and represent a tremendous economic burden in terms of loss of work. One of the crucial steps of Salmonella pathogenicity is the establishment of colonization in the gut. While the host eventually clears the infection with Salmonella, initial host responses benefit the growth of Salmonella. Similarly, also some commensal bacteria promote Salmonella intestinal colonization. These resident bacteria liberate for example otherwise inaccessible carbon sources. However, previous studies focused exclusively on the role of commensal bacteria during Salmonella pathogenesis and did not address the role of another important group of microorganisms that resides in the gut: commensal intestinal fungi. In recent years, this under-studied component of the microbiota has been implicated in a variety of diseases including Inflammatory Bowel Disease. However, the role of commensal fungi during pathogenesis of gut pathogens like Salmonella is unknown. This is despite the fact that Salmonella encodes genes that make an interaction with commensal gut fungi highly likely. In this regard, Salmonella produces three receptors for fungal and bacterial siderophores (so-called xenosiderophores). Siderophores are small molecules that are secreted by fungi and bacteria for iron acquisition in low iron environments like the inflamed gut. Our preliminary data show that expression of two fungal siderophore receptors confers a growth advantage to Salmonella in conventionally raised but not in germ-free mice. Furthermore, chitinases produced by Salmonella could weaken the chitin-containing fungal cell wall to enable Salmonella to acquire fungal nutrients including intracellularly stored siderophores. We therefore hypothesize that Salmonella acquires xenosiderophores and accesses fungal nutrients with the help of chitinases to increase colonization in the gut. We plan to investigate in Aim 1 how fungal xenosiderophores promote Salmonella colonization in the gastrointestinal tract by depletion or addition of fungi. We will sequence the fungal microbiota to identify siderophore-producing fungi. We will also determine the growth promoting effect of the third xenosiderophore receptor, which binds a bacterial siderophore. In Aim 2, we propose to study how members of the microbiota compete with Salmonella for xenosiderophores. For this, we will reduce the complexity of the microbiota. Germ-free mice allow for specific colonization with producers and competitors for fungal siderophores. We will test if common fungal and bacterial commensals that do not produce siderophores like Candida, Saccharomyces, and Bacteroides compete with Salmonella for xenosiderophores. In Aim 3, we will elucidate if Salmonella chitinases enhance Salmonella pathogenicity. We will test two different functions of chitinases: weakening the fungal cell wall to access fungal nutrients including intracellularly stored siderophores and degradation of mucin or glycans on epithelial cells. We expect that our approaches will enhance our understanding of the role of commensal fungi during infection with Salmonella. In a broader context, we expect our work will help to design treatments, i.e. effective competitors to intestinal pathogens.