Identification of Francisella tularensis Live Vaccine Strain CuZn Superoxide Dismutase as Critical for Resistance to Extracellularly Generated Reactive Oxygen Species

Center for Immunology and Microbial Disease, MC 151, Albany Medical College, 47 New Scotland Ave., Albany, NY 12208, USA.
Journal of bacteriology (Impact Factor: 2.81). 09/2009; 191(20):6447-56. DOI: 10.1128/JB.00534-09
Source: PubMed


Francisella tularensis is an intracellular pathogen whose survival is in part dependent on its ability to resist the microbicidal activity of host-generated reactive oxygen species (ROS) and reactive nitrogen species (RNS). In numerous bacterial pathogens, CuZn-containing superoxide dismutases (SodC) are important virulence factors, localizing to the periplasm to offer protection from host-derived superoxide radicals (O(2)(-)). In the present study, mutants of F. tularensis live vaccine strain (LVS) deficient in superoxide dismutases (SODs) were used to examine their role in defense against ROS/RNS-mediated microbicidal activity of infected macrophages. An in-frame deletion F. tularensis mutant of sodC (DeltasodC) and a F. tularensis DeltasodC mutant with attenuated Fe-superoxide dismutase (sodB) gene expression (sodB DeltasodC) were constructed and evaluated for susceptibility to ROS and RNS in gamma interferon (IFN-gamma)-activated macrophages and a mouse model of respiratory tularemia. The F. tularensis DeltasodC and sodB DeltasodC mutants showed attenuated intramacrophage survival in IFN-gamma-activated macrophages compared to the wild-type F. tularensis LVS. Transcomplementing the sodC gene in the DeltasodC mutant or inhibiting the IFN-gamma-dependent production of O(2)(-) or nitric oxide (NO) enhanced intramacrophage survival of the sod mutants. The DeltasodC and sodB DeltasodC mutants were also significantly attenuated for virulence in intranasally challenged C57BL/6 mice compared to the wild-type F. tularensis LVS. As observed for macrophages, the virulence of the DeltasodC mutant was restored in ifn-gamma(-/-), inos(-/-), and phox(-/-) mice, indicating that SodC is required for resisting host-generated ROS. To conclude, this study demonstrates that SodB and SodC act to confer protection against host-derived oxidants and contribute to intramacrophage survival and virulence of F. tularensis in mice.

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    • "In contrast, katG mutants of Schu S4 were sensitive to hydrogen peroxide and reactive nitrogen species in vitro but retained lethality in mice, indicating that KatG is a sufficient but not necessary virulence factor.29 Similarly, the superoxide dismutases SodB and SodC are essential for bacterial resistance to superoxide radicals, as F. tularensis LVS mutants of either enzyme are highly attenuated in mice challenged intranasally.30,31 "
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    ABSTRACT: Francisella tularensis is an intracellular Gram-negative bacterium that causes life-threatening tularemia. Although the prevalence of natural infection is low, F. tularensis remains a tier I priority pathogen due to its extreme virulence and ease of aerosol dissemination. F. tularensis can infect a host through multiple routes, including the intradermal and respiratory routes. Respiratory infection can result from a very small inoculum (ten organisms or fewer) and is the most lethal form of infection. Following infection, F. tularensis employs strategies for immune evasion that delay the immune response, permitting systemic distribution and induction of sepsis. In this review we summarize the current knowledge of F. tularensis in an immunological context, with emphasis on the host response and bacterial evasion of that response.
    Infection and Drug Resistance 09/2014; 7:239-51. DOI:10.2147/IDR.S53700
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    • "This mechanism occurs over and above the activity of the superoxide dismutase that neutralises both reactive oxygen and nitrogen spp. [31] and is just one of many mechanisms employed by F. tularensis to manipulate and evade the host response [32]. "
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    ABSTRACT: Reactive nitrogen is critical for the clearance of Francisella tularensis infections. Here we assess the role of nitric oxide in control of intracellular infections in two murine macrophage cell lines of different provenance: the alveolar macrophage cell line, MH-S, and the widely used peritoneal macrophage cell line, J774A.1. Cells were infected with the highly virulent Schu S4 strain or with the avirulent live vaccine strain (LVS) with and without stimuli. Compared to MH-S cells, J774A.1 cells were unresponsive to stimulation and were able to control the intracellular replication of LVS bacteria, but not of Schu S4. In MH-S cells, Schu S4 demonstrated control over cellular NO production. Despite this, MH-S cells stimulated with LPS or LPS and IFN-γ were able to control intracellular Schu S4 numbers. However, only stimulation with LPS induced significant cellular NO production. Combined stimulation with LPS and IFN-γ produced a significant reduction in intracellular bacteria that occurred whether high levels of NO were produced or not, indicating that NO secretion is not the only defensive cellular mechanism operating in virulent Francisella infections. Understanding how F. tularensis interacts with host macrophages will help in the rational design of new and effective therapies.
    Research Journal of Immunology 07/2014; 2014(3):694717. DOI:10.1155/2014/694717
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    • "The importance of altering NADPH complexes is not unique to Francisella, as multiple bacteria including Helicobacter pylori and Salmonella typhimurium have been shown to alter NADPH oxidase assembly in cells (Gallois et al., 2001; Allen et al., 2005). If Francisella does encounter ROS, catalases and super oxide dismutases (SOD) enzymes are necessary for survival, as ΔSOD F. tularensis LVS have increased susceptibility to IFNγ-induced death (Melillo et al., 2009). Indeed, it has been shown that antioxidants produced by Francisella Schu S4 can dampen macrophage inflammatory responses (Melillo et al., 2010). "
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    ABSTRACT: Francisella tularensis can bypass and suppress host immune responses, even to the point of manipulating immune cell phenotypes and intercellular inflammatory networks. Strengthening these responses such that immune cells more readily identify and destroy the bacteria is likely to become a viable (and perhaps necessary) strategy for combating infections with Francisella, especially given the likelihood of antibiotic resistance in the foreseeable future. Monocytes and macrophages offer a niche wherein Francisella can invade and replicate, resulting in substantially higher bacterial load that can overcome the host. As such, understanding their responses to Francisella may uncover potential avenues of therapy that could promote a lowering of bacterial burden and clearance of infection. These response pathways include Toll-like Receptor 2 (TLR2), the caspase-1 inflammasome, Interferons, NADPH oxidase, Phosphatidylinositide 3-kinase (PI3K), and the Ras pathway. In this review we summarize the literature pertaining to the roles of these pathways during Francisella infection, with an emphasis on monocyte/macrophage responses. The therapeutic targeting of one or more such pathways may ultimately become a valuable tool for the treatment of tularemia, and several possibilities are discussed.
    Frontiers in Cellular and Infection Microbiology 02/2014; 4:18. DOI:10.3389/fcimb.2014.00018 · 3.72 Impact Factor
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