Identification of new virulence factors in Francisella tularensis

Sammanfattning: Francisella tularensis, the causative agent of tularemia, is a highly virulent bacterium with an infection dose of less than ten bacteria. The ability of a pathogen to cause infection relies on different virulence mechanisms, but in Francisella tularensis relatively few virulence factors are known. Two F. tularensis subspecies are virulent in humans; the highly virulent subspecies tularensis, also referred to as type A, and the less virulent subspecies holarctica, also called type B. The aim of this thesis has been to improve the knowledge regarding the ability of Francisella to cause disease, with the emphasis on surface located and membrane associated proteins and structures. In addition I have also investigated how virulence is regulated by studying the role of the small RNA chaperone, Hfq.The genome of Francisella appears to encode few regulatory genes. In my work I found that Hfq has an important role in regulation of virulence associated genes in Francisella. Similar to what has been found in other pathogens, Hfq functions in negative regulation, and this is the first time a negative regulation has been described for genes in the Francisella pathogenicity island. Another protein with a key role in virulence is a homologue to a disulphide oxidoreductase, DsbA, which was identified as an outer membrane lipoprotein in Francisella. A dsbA mutant was found to be severely attenuated for virulence and also induced protection against wild-type infections, thus making it a candidate for exploration as a new live vaccine. Additional genes with homology to known virulence determinants include a type IV pilin system. The pilin homologue, PilA, was identified to be required for full virulence in both type A and type B strains. In addition, genes involved in pili assembly and secretion, pilC and pilQ, were also found to be virulence associated in the type A strain.In summary, dsbA, hfq and type IV pili associated genes were indentified to be virulence determinants in F. tularensis. DsbA is a potential target for drug development and a dsbA mutant a candidate for a new live vaccine strain. Furthermore the identification of Hfq as a novel regulatory factor opens new insights into the virulence regulatory network in Francisella.