Characterization of the Role of Transcriptional Regulator of AraC/XylS Family in Tularemia Pathogenesis

The Tier 1 Select Agent, Francisella tularensis causes an acute and fatal disease known as tularemia. Many studies have devoted enormous efforts to understand how F. tularensis avoids host defense mechanisms, replicates within an extremely secure immune system, and eventually causes the deadly disease tularemia. The extremely high virulence of Francisella depends on its ability to manipulate gene expression according to the surrounding environment. This process requires the involvement of unique transcriptional regulators. Francisella possesses very few transcriptional regulators, and a majority of them characterized to-date have been shown to regulate genes involved in virulence and cellular functions. The role of a transcriptional regulator of F. tularensis belonging to the AraC/XylS in gene regulation and virulence remains uncharacterized to-date. This study characterized the role of AraC in gene regulation, intramacrophage survival, and virulence of F. tularensis. In specific aim 1, we generated a deletion mutant (DaraC) of FTL_0689 gene encoding AraC of F. tularensis Live Vaccine Strain (LVS), and its transcomplemented strain (DaraC+paraC). Characterization of the DaraC mutant demonstrated that AraC does not regulate genes involved in arabinose utilization. Genomic organization of the araC gene suggested that it may have a role in the regulation of a unique multidrug efflux pump (MEP) located downstream of it. Our results revealed that the phenotype of the DaraC mutant mirrors that of the emrA1 and the silC mutants, the components of the MEP. However, these phenotypic similarities are not due to the direct regulation of MEP genes by AraC. Further characterization revealed that AraC is involved in providing resistance against oxidative stress. In specific aim 2, we investigated the role of AraC as a global transcriptional regulator in F. tularensis LVS strain. We studied gene expression profiles of the wild type F. tularensis LVS and the DaraC mutant under normal and oxidative stress conditions. The results revealed that AraC serves as a transcriptional regulator only when the bacteria are exposed to oxidative stress conditions. AraC is involved in the regulation of virulence genes encoded on Francisella Pathogenicity Island, stress response genes, energy production, genes encoding enzymes in the tricarboxylic acid (TCA) cycle, metabolism and regulatory proteins indicating its role as a global regulator. The differential expression of these genes also impaired the ability of the DaraC mutant to survive in macrophages and attenuated its virulence in mice. Collectively, this study identified and characterized a novel transcription factor, AraC, required for adaptation of Francisella to oxidative stress conditions encountered outside or within a host

An AraC/XylS Family Transcriptional Regulator Modulates the Oxidative Stress Response of Francisella Tularensis

Francisella tularensis is a Gram-negative bacterium that causes a fatal human disease known as tularemia. The Centers for Disease Control and Prevention have classified F. tularensis as a category A tier 1 select agent. The virulence mechanisms of Francisella are not entirely understood. Francisella possesses very few transcription regulators, and most of these regulate the expression of genes involved in intracellular survival and virulence. The F. tularensis genome sequence analysis reveals an AraC (FTL_0689) transcriptional regulator homologous to the AraC/XylS family of transcriptional regulators. In Gram-negative bacteria, AraC activates genes required for l-arabinose utilization and catabolism. The role of the FTL_0689 regulator in F. tularensis is not known. In this study, we characterized the role of FTL_0689 in the gene regulation of F. tularensis and investigated its contribution to intracellular survival and virulence. The results demonstrate that FTL_0689 in Francisella is not required for l-arabinose utilization. Instead, FTL_0689 specifically regulates the expression of the oxidative and global stress response, virulence, metabolism, and other key pathways genes required by Francisella when exposed to oxidative stress. The FTL_0689 mutant is attenuated for intramacrophage growth and virulence in mice. Based on the deletion mutant phenotype, FTL_0689 was termed osrR (oxidative stress response regulator). Altogether, this study elucidates the role of the osrR transcriptional regulator in tularemia pathogenesis. IMPORTANCE The virulence mechanisms of category A select agent Francisella tularensis, the causative agent of a fatal human disease known as tularemia, remain largely undefined. The present study investigated the role of a transcriptional regulator and its overall contribution to the oxidative stress resistance of F. tularensis. The results provide an insight into a novel gene regulatory mechanism, especially when Francisella is exposed to oxidative stress conditions. Understanding such Francisella- specific regulatory mechanisms will help identify potential targets for developing effective therapies and vaccines to prevent tularemia

On the Use of Radar and Optical Satellite Imagery for the Monitoring of Flood Hazards on Heritage Sites in Southern Sinai, Egypt

This study focuses on the use of radar and optical satellite imagery for flood hazard mapping and monitoring around the archaeological sites of the Wadi Baba area, situated at Sinai (Egypt) and well known for its heritage treasures belonging to diverse historical periods and civilizations from the Pharaonic, Nabateans, Christian, and Islamic eras. Although this area is located in an arid to semi-arid climatic region, it is intermittently flooded due to torrential rainstorms. To assess the amount of rainfall expected and its impacts on heritage sites, satellite Sentinel-1 (C-Band) and Tropical Rainfall Monitoring Mission (TRMM) data were jointly used with measurements from meteorological stations and the Digital Elevation Model (DEM) from Shuttle Radar Topography Mission (SRTM). Envi5.1, ArcGIS 10.4.1, Snap 6.0, and the GEE platform were used to process optical and radar data, which were further analysed using the ArcHydro model. In this study, the TRMM accumulated rainfall data acquired on 17 January 2010, Sentinel-1 radar images between 2017 and 2019, and Sentinel-1 data captured from 1 to 30 March 2020 processed by GEE platform were chosen to assess the effects of flood events on the archaeological sites in the study area. The results indicated that the study area is exposed to flood risk that significantly threatens these heritage sites. Based on that, mitigation strategies were devised and recommended to mitigate the flood hazard impact around the archaeological areas

Role of Peroxiredoxin of the AhpC/TSA Family in Antioxidant Defense Mechanisms of Francisella tularensis

Francisella tularensis is a Gram-negative, facultative intracellular pathogen and the causative agent of a lethal human disease known as tularemia. Due to its extremely high virulence and potential to be used as a bioterror agent, F. tularensis is classified by the CDC as a Category A Select Agent. As an intracellular pathogen, F. tularensis during its intracellular residence encounters a number of oxidative and nitrosative stresses. The roles of the primary antioxidant enzymes SodB, SodC and KatG in oxidative stress resistance and virulence of F. tularensis live vaccine strain (LVS) have been characterized in previous studies. However, very fragmentary information is available regarding the role of peroxiredoxin of the AhpC/TSA family (annotated as AhpC) of F. tularensis SchuS4; whereas the role of AhpC of F. tularensis LVS in tularemia pathogenesis is not known. This study was undertaken to exhaustively investigate the role of AhpC in oxidative stress resistance of F. tularensis LVS and SchuS4. We report that AhpC of F. tularensis LVS confers resistance against a wide range of reactive oxygen and nitrogen species, and serves as a virulence factor. In highly virulent F. tularensis SchuS4 strain, AhpC serves as a key antioxidant enzyme and contributes to its robust oxidative and nitrosative stress resistance, and intramacrophage survival. We also demonstrate that there is functional redundancy among primary antioxidant enzymes AhpC, SodC, and KatG of F. tularensis SchuS4. Collectively, this study highlights the differences in antioxidant defense mechanisms of F. tularensis LVS and SchuS4