Cutaneous resistance against Francisella tularensis

Abstract: Francisella tularensis, the causative agent of tularemia, is a potent pathogen in humans and other mammals. The ulceroglandular form of the disease is the most common expression in humans with a clinical picture characterized by a skin ulcer, enlarged regional lymph nodes and fever. Despite being a preferred route of infection, the skin also affords an effective defense barrier against F. tularensis. Doses required to induce infection by intradermal inoculation are several logs higher than those needed for infection by other routes. In the present thesis, the requirements for the local and systemic host defense to intradermal infection with F. tularensis was studied in experimental mouse models. Naïve mice and mice immunized by previous infection were challenged, mostly with the live vaccine strain F. tularensis LVS but also with a clinical isolate of F. tularensis.In naïve mice, intradermal inoculation of F. tularensis LVS resulted in a rapid increase of bacterial numbers during the first few days in the skin, lymph nodes, spleen and liver, followed by a decrease and eradication of the bacteria within two weeks of inoculation. Immune mice controlled the infection at the site of infection and very few bacteria spread to internal organs. When immunohistochemical staining of skin specimens was performed during the first 3 days, naïve mice showed a weak or barely discernible local expression of TNF-?, IL-12 and IFN-?. In immune mice, the expression of all three cytokines was strongly enhanced, TNF-? and IL-12 within 24 h and IFN-? within 72 h of inoculation.To investigate the role of T cells in the defense against intradermal infection with F. tularensis LVS, naïve and immune T-cell knockout mice (e.g., ?? TCR-/-, ?? TCR-/-, ???? TCR-/-) were used. Naïve mice lacking the ?? TCR had persistently high bacterial numbers in all organs and died at 4 weeks. Mice lacking the ?? TCR, on the other hand, controlled the infection as effectively as did wild-type mice. To enable ?? TCR-/- and ???? TCR-/- mice to survive, antibiotic treatment was given from day 10 to 20 of infection. When intradermally challenged 2 weeks later, these animals were found to control a secondary infection, resulting in decreasing viable counts in skin and lymph nodes and prevention of spread to liver and spleen. The results indicated the presence of a T-cell independent mechanism of resistance and analyses of serum showed high levels of F. tularensis-specific IgM, findings suggesting a role for antibodies in the protection against cutaneous tularemia.To study the effect of F. tularensis-specific antibodies on host resistance, we adoptively transferred immune serum to B-cell-deficient mice. After receiving immune serum, both naïve and immunized mice became capable of surviving an otherwise lethal dose of F. tularensis LVS. Moreover, transfer of immune serum to wild type mice, afforded significant protection to a lethal dose of a wild-type strain of F. tularensis subsp. holarctica, as disclosed by reduced bacterial counts in spleen and liver. Finally, we studied the effect of immune serum on the local expression of proinflammatory cytokines and neutrophils in response to an intradermal injection of F. tularensis LVS. As compared to normal serum, transfer of immune serum resulted in increased expression of TNF-?, IL-12 and neutrophils. These findings afford a possible explanation for the effect of specific antibodies in the local host protection in the skin against tularemia.