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Distinct roles of reactive nitrogen and oxygen species to control infection with the facultative intracellular bacterium Francisella tularensis.
Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Clinical Bacteriology. Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Infectious Diseases.
Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Infectious Diseases.
Umeå University, Faculty of Medicine, Department of Clinical Microbiology.
Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Infectious Diseases.
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2004 (English)In: Infection and Immunity, ISSN 0019-9567, E-ISSN 1098-5522, Vol. 72, no 12, 7172-7182 p.Article in journal (Refereed) Published
Abstract [en]

Reactive nitrogen species (RNS) and reactive oxygen species (ROS) are important mediators of the bactericidal host response. We investigated the contribution of these two mediators to the control of infection with the facultative intracellular bacterium Francisella tularensis. When intradermally infected with the live vaccine strain F. tularensis LVS, mice deficient in production of RNS (iNOS(-/-) mice) or in production of ROS by the phagocyte oxidase (p47(phox-/-) mice) showed compromised resistance to infection. The 50% lethal dose (LD(50)) for iNOS(-/-) mice was <20 CFU, and the LD(50) for p47(phox-/-) mice was 4,400 CFU, compared to an LD(50) of >500,000 CFU for wild-type mice. The iNOS(-/-) mice survived for 26.4 +/- 1.8 days, and the p47(phox-/-) mice survived for 10.1 +/- 1.3 days. During the course of infection, the serum levels of gamma interferon (IFN-gamma) and interleukin-6 were higher in iNOS(-/-) and p47(phox-/-) mice than in wild-type mice. Histological examination of livers of iNOS(-/-) mice revealed severe liver pathology. Splenocytes obtained 5 weeks after primary infection from antibiotic-treated iNOS(-/-) mice showed an in vitro recall response that was similar in magnitude and greater secretion of IFN-gamma compared to cells obtained from wild-type mice. In summary, mice lacking expression of RNS or ROS showed extreme susceptibility to infection with F. tularensis LVS. The roles of RNS and ROS seemed to be distinct since mice deficient in production of ROS showed dissemination of infection and died during the early phase of infection, whereas RNS deficiency led to severe liver pathology and a contracted course of infection.

Place, publisher, year, edition, pages
2004. Vol. 72, no 12, 7172-7182 p.
Keyword [en]
Animals, Colony Count; Microbial, Disease Susceptibility, Interferon Type II/blood, Liver/microbiology/pathology, Lymphocyte Activation, Mice, Mice; Inbred C57BL, Nitric Oxide Synthase/physiology, Nitric Oxide Synthase Type II, Reactive Nitrogen Species/*physiology, Reactive Oxygen Species/*metabolism, Skin/microbiology, Superoxides/metabolism, Tularemia/*immunology
URN: urn:nbn:se:umu:diva-7172DOI: 10.1128/IAI.72.12.7172-7182.2004PubMedID: 15557642OAI: diva2:146843
Available from: 2008-01-04 Created: 2008-01-04 Last updated: 2011-04-15Bibliographically approved
In thesis
1. Reactive oxygen and nitrogen in host defence against Francisella tularensis
Open this publication in new window or tab >>Reactive oxygen and nitrogen in host defence against Francisella tularensis
2005 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Francisella tularensis, the causative agent of tularemia, is a potent human and animal pathogen. Initially upon infection of the host, intramacrophage proliferation of F. tularensis occurs but after activation of the acquired host immunity, the phagocytes become activated to kill the bacterium. In my thesis, I focused on mechanisms utilized by F. tularensis to survive intracellularly and on host mechanisms responsible for macrophage-mediated killing and control of infection.

The F. tularensis-specific protein IglC has been previously shown to be essential to the intramacrophage proliferation and virulence of the bacterium in mice. By electron microscopy of macrophages infected with either the live vaccine strain of F. tularensis or an isogenic mutant, denoted ∆iglC, expression of IglC was found to be necessary for the bacterium to escape from the phagosome. IFN-g-activated macrophages significantly inhibited the escape of the live vaccine strain of F. tularensis from the phagosome.

iNOS and phox generate NO and O2-, respectively. These molecules and their reaction products possess both bactericidal and immunoregulatory properties. We investigated the capability of IFN-g-activated peritoneal exudate cells from gene deficient iNOS-/- or p47phox-/- mice to control an intracellular F. tularensis LVS infection. iNOS was found to contribute significantly to the IFN-g induced killing, while phox contributed only to a minor extent. Unexpectedly, bacteria were eradicated even in the absence of both a functional phox and an active iNOS. The eradication was found to depend on ONOO-, the reaction product of NO and O2-, because addition of a decomposition catalyst of ONOO- completely inhibited the killing.

Studies on iNOS-/- or p47phox-/- mice infected with F. tularensis LVS showed phox to be important during the first days of infection, a stage when iNOS seemed dispensable. Eventually, iNOS-/- mice died of the infection, suggesting a role of iNOS later in the course of infection. iNOS-/- mice exhibited elevated IFN-g serum levels and severe liver damage suggesting that the outcome of infection was at least in part related to an uncontrolled immune response.

Several pathogenic bacteria express Cu,Zn-SOD, which in combination with other enzymes detoxifies reactive oxygen species produced by the host. A deletion mutant of F. tularensis LVS lacking the gene encoding Cu,Zn-SOD was attenuated at least 100-fold compared to LVS in mice. In peritoneal exudate cells from mice, Cu,Zn-SOD was found to be required for effective intramacrophage proliferation and, in mice, important for bacterial replication at the very early phase of infection.

In summary, the most conspicuous findings were a capability of IFN-g activated macrophages to retain F. tularensis LVS in the phagosome, an essential role of ONOO- in intracellular killing of F. tularensis, and an importance of Cu,Zn-SOD to the virulence of F. tularensis LVS.

Place, publisher, year, edition, pages
Umeå: Klinisk mikrobiologi, 2005. 55 p.
Umeå University medical dissertations, ISSN 0346-6612 ; 954
Cell and molecular biology, Francisella tularensis, inducible nitric oxide synthase, phagocyte oxidase, macrophages, Cell- och molekylärbiologi
National Category
Biochemistry and Molecular Biology
Research subject
Clinical Bacteriology
urn:nbn:se:umu:diva-474 (URN)91-7305-851-3 (ISBN)
Public defence
2005-04-15, E04, 6E, Norrlands Universitetssjukhus i Umeå, Umeå, 09:00 (English)
Available from: 2005-03-15 Created: 2005-03-15 Last updated: 2009-11-19Bibliographically approved

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Lindgren, HelenaStenmark, StephanTärnvik, ArneSjöstedt, Anders
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