Dataset Information

Increased Resistance to Intradermal Francisella tularensis LVS Infection by Inactivation of the Sts Phosphatases.

ABSTRACT: The Suppressor of TCR signaling proteins (Sts-1 and Sts-2) are two homologous phosphatases that negatively regulate signaling pathways in a number of hematopoietic lineages, including T lymphocytes. Mice lacking Sts expression are characterized by enhanced T cell responses. Additionally, a recent study demonstrated that Sts^-/- mice are profoundly resistant to systemic infection by Candida albicans, with resistance characterized by enhanced survival, more rapid fungal clearance in key peripheral organs, and an altered inflammatory response. To investigate the role of Sts in the primary host response to infection by a bacterial pathogen, we evaluated the response of Sts^-/- mice to infection by a Gram-negative bacterial pathogen. Francisella tularensis is a facultative bacterial pathogen that replicates intracellularly within a variety of cell types and is the causative agent of tularemia. Francisella infections are characterized by a delayed immune response, followed by an intense inflammatory reaction that causes widespread tissue damage and septic shock. Herein, we demonstrate that mice lacking Sts expression are significantly resistant to infection by the live vaccine strain (LVS) of F. tularensis Resistance is characterized by reduced lethality following high-dose intradermal infection, an altered cytokine response in the spleen, and enhanced bacterial clearance in multiple peripheral organs. Sts^-/- bone marrow-derived monocytes and neutrophils, infected with F. tularensis LVS ex vivo, display enhanced restriction of intracellular bacteria. These observations suggest the Sts proteins play an important regulatory role in the host response to bacterial infection, and they underscore a role for Sts in regulating functionally relevant immune response pathways.

SUBMITTER: Parashar K

PROVIDER: S-EPMC5563576 | biostudies-literature | 2017 Sep

REPOSITORIES: biostudies-literature

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Increased Resistance to Intradermal Francisella tularensis LVS Infection by Inactivation of the Sts Phosphatases.

Parashar Kaustubh K Kopping Erik E Frank David D Sampath Vinaya V Thanassi David G DG Carpino Nick N

Infection and immunity 20170818 9

The Suppressor of TCR signaling proteins (Sts-1 and Sts-2) are two homologous phosphatases that negatively regulate signaling pathways in a number of hematopoietic lineages, including T lymphocytes. Mice lacking Sts expression are characterized by enhanced T cell responses. Additionally, a recent study demonstrated that Sts-/- mice are profoundly resistant to systemic infection by Candida albicans, with resistance characterized by enhanced survival, ...[more]

PMID: 28630061

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Project description:Francisella tularensis, the causative agent of tularemia, is capable of causing disease in a multitude of mammals and remains a formidable human pathogen due to a high morbidity, low infectious dose, lack of a FDA approved vaccine, and ease of aerosolization. For these reasons, there is concern over the use of F. tularensis as a biological weapon, and, therefore, it has been classified as a Tier 1 select agent. Fluoroquinolones and aminoglycosides often serve as the first line of defense for treatment of tularemia. However, high levels of resistance to these antibiotics has been observed in gram-negative bacteria in recent years, and naturally derived resistant Francisella strains have been described in the literature. The acquisition of antibiotic resistance, either natural or engineered, presents a challenge for the development of medical countermeasures. In this study, we generated a surrogate panel of antibiotic resistant F. novicida and Live Vaccine Strain (LVS) by selection in the presence of antibiotics and characterized their growth, biofilm capacity, and fitness. These experiments were carried out in an effort to (1) assess the fitness of resistant strains; and (2) identify new targets to investigate for the development of vaccines or therapeutics. All strains exhibited a high level of resistance to either ciprofloxacin or streptomycin, a fluoroquinolone and aminoglycoside, respectively. Whole genome sequencing of this panel revealed both on-pathway and off-pathway mutations, with more mutations arising in LVS. For F. novicida, we observed decreased biofilm formation for all ciprofloxacin resistant strains compared to wild-type, while streptomycin resistant isolates were unaffected in biofilm capacity. The fitness of representative antibiotic resistant strains was assessed in vitro in murine macrophage-like cell lines, and also in vivo in a murine model of pneumonic infection. These experiments revealed that mutations obtained by these methods led to nearly all ciprofloxacin resistant Francisella strains tested being completely attenuated while mild attenuation was observed in streptomycin resistant strains. This study is one of the few to examine the link between acquired antibiotic resistance and fitness in Francisella spp., as well as enable the discovery of new targets for medical countermeasure development.

Project description:BackgroundThe NF-κB activating kinases, IKKα and IKKβ, are key regulators of inflammation and immunity in response to infection by a variety of pathogens. Both IKKα and IKKβ have been reported to modulate either pro- or anti- inflammatory programs, which may be specific to the infectious organism or the target tissue. Here, we analyzed the requirements for the IKKs in myeloid cells in vivo in response to Francisella tularensis Live Vaccine Strain (Ft. LVS) infection.Methods and principal findingsIn contrast to prior reports in which conditional deletion of IKKβ in the myeloid lineage promoted survival and conferred resistance to an in vivo group B streptococcus infection, we show that mice with a comparable conditional deletion (IKKβ cKO) succumb more rapidly to lethal Ft. LVS infection and are unable to control bacterial growth at sublethal doses. Flow cytometry analysis of hepatic non-parenchymal cells from infected mice reveals that IKKβ inhibits M1 classical macrophage activation two days post infection, which has the collateral effect of suppressing IFN-γ(+) CD8(+) T cells. Despite this early enhanced inflammation, IKKβ cKO mice are unable to control infection; and this coincides with a shift toward M2a polarized macrophages. In comparison, we find that myeloid IKKα is dispensable for survival and bacterial control. However, both IKKα and IKKβ have effects on hepatic granuloma development. IKKα cKO mice develop fewer, but well-contained granulomas that accumulate excess necrotic cells after 9 days of infection; while IKKβ cKO mice develop numerous micro-granulomas that are less well contained.ConclusionsTaken together our findings reveal that unlike IKKα, IKKβ has multiple, contrasting roles in this bacterial infection model by acting in an anti-inflammatory capacity at early times towards sublethal Ft. LVS infection; but in spite of this, macrophage IKKβ is also a critical effector for host survival and efficient pathogen clearance.

Dataset Information

Increased Resistance to Intradermal Francisella tularensis LVS Infection by Inactivation of the Sts Phosphatases.

Publications

Increased Resistance to Intradermal Francisella tularensis LVS Infection by Inactivation of the Sts Phosphatases.

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