Lack of OxyR and KatG Results in Extreme Susceptibility of Francisella tularensis LVS to Oxidative Stress and Marked Attenuation In vivo

Abstract

Francisella tularensis is an intracellular bacterium and as such is expected to encounter a continuous attack by reactive oxygen species (ROS) in its intracellular habitat and efficiently coping with oxidative stress is therefore essential for its survival. The oxidative stress response system of F. tularensis is complex and includes multiple antioxidant enzymes and pathways, including the transcriptional regulator OxyR and the H2O2-decomposing enzyme catalase, encoded by katG. The latter is regulated by OxyR. A deletion of either of these genes, however, does not severely compromise the virulence of F. tularensis and we hypothesized that if the bacterium would be deficient of both catalase and OxyR, then the oxidative defense and virulence of F. tularensis would become severely hampered. To test this hypothesis, we generated a double deletion mutant, ΔoxyR/ΔkatG, of F. tularensis LVS and compared its phenotype to the parental LVS strain and the corresponding single deletion mutants. In accordance with the hypothesis, ΔoxyR/ΔkatG was distinctly more susceptible than ΔoxyR and ΔkatG to H2O2, ONOO−, and O2-, moreover, it hardly grew in mouse-derived BMDM or in mice, whereas ΔkatG and ΔoxyR grew as well as F. tularensis LVS in BMDM and exhibited only slight attenuation in mice. Altogether, the results demonstrate the importance of catalase and OxyR for a robust oxidative stress defense system and that they act cooperatively. The lack of both functions render F. tularensis severely crippled to handle oxidative stress and also much attenuated for intracellular growth and virulence.

Neutrophils to the ROScue: Mechanisms of NADPH Oxidase Activation and Bacterial Resistance

“…Nonetheless, several studies have demonstrated a role for ROS-detoxifying and/or repair enzymes during mammalian bacterial infection models, suggesting that the role of these factors in protecting against host-derived ROS must not be fully discounted (De Groote et al, 1997 ; Roggenkamp et al, 1997 ; Harris et al, 2003 ; Brenot et al, 2004 ; Hebrard et al, 2009 ; Aussel et al, 2011 ; Green et al, 2016 ; Honn et al, 2017 ). Therefore, although in vitro models do not fully replicate the oxidative environment bacterial pathogens encounter in the host, the concepts outlined above provide us with a framework to begin to understand the mechanisms by which bacterial pathogens detoxify mammalian sources of ROS or other host factors triggered by ROS and repair damage to cellular targets.…”

“…Interestingly, while suppression of oxidative burst by some pathogens is advantageous for mammalian infections, many of these organisms, including S. typhimurium, Yersinia sp, GAS, F. tularensis , and H. pylori , also require mechanisms to detoxify ROS or repair their damage in animal models of infection (De Groote et al, 1997 ; Roggenkamp et al, 1997 ; Harris et al, 2003 ; Brenot et al, 2004 ; Hebrard et al, 2009 ; Green et al, 2016 ; Honn et al, 2017 ). These data suggest that suppression of NADPH oxidase activity by secreted effector proteins and toxins is not sufficient to defend these bacterial pathogens against oxidative attack and that these factors must work in concert with cellular processes critical for detoxification and repair to prevent restriction by host-derived ROS.…”

Live Attenuated Tularemia Vaccines for Protection Against Respiratory Challenge With Virulent F. tularensis subsp. tularensis

“…Several oxidant mutants of LVS have been reported (Lenco et al, 2005 ; Bakshi et al, 2006 , 2008 ; Buchan et al, 2009 ; Melillo et al, 2009 ; Honn et al, 2012 , 2017 ; Ma et al, 2014 ; Suresh et al, 2015 ; Saha et al, 2017 ) among which LVS mutants with a point mutation in sodB (FTL_1791), encoding an iron superoxide dismutase ( sodB Ft ), with a transposon insertion in a putative gene for the EmrA1 (FTL_0687) secretion protein ( emrA1 ), or with a deletion in dsbA , encoding a disulfide oxidoreductases protein family homolog (ΔdsbA/FSC200), have been tested in mice for their efficacy against respiratory challenge with SCHU S4 (Bakshi et al, 2006 , 2008 ; Straskova et al, 2015 ; Suresh et al, 2015 ). Both sodB Ft and emrA1 LVS mutants are sensitive to oxidant stress and are more attenuated than the parental LVS in mice; C57BL/6 and BALB/c mice infected i.n.…”

Control of Francisella tularensis Virulence at Gene Level: Network of Transcription Factors

“…It is able to bind directly to the promoter of ahpC and katG [ 49 , 50 ]. Mainly, KatG and OxyR work together and constitute an important system for defense against oxidative stress [ 79 ].…”