Nitric Oxide-Mediated Intracellular Growth Restriction of Pathogenic Rhodococcus equi Can Be Prevented by Iron

Abstract

Rhodococcus equi is an intracellular pathogen which causes pneumonia in young horses and in immunocompromised humans. R. equi arrests phagosome maturation in macrophages at a prephagolysosome stage and grows inside a privileged compartment. Here, we show that, in murine macrophages activated with gamma interferon and lipopolysaccharide, R. equi does not multiply but stays viable for at least 24 h. Whereas infection control of other intracellular pathogens by activated macrophages is executed by enhanced phagosome acidification or phagolysosome formation, by autophagy or by the interferoninducible GTPase Irgm1, none of these mechanisms seems to control R. equi infection. Growth control by macrophage activation is fully mimicked by treatment of resting macrophages with nitric oxide donors, and inhibition of bacterial multiplication by either activation or nitric oxide donors is annihilated by cotreatment of infected macrophages with ferrous sulfate. Transcriptional analysis of the R. equi ironregulated gene iupT demonstrates that intracellular R. equi encounters iron stress in activated, but not in resting, macrophages and that this stress is relieved by extracellular addition of ferrous sulfate. Our results suggest that nitric oxide is central to the restriction of bacterial access to iron in activated macrophages.

Virulence‐associated protein A fromRhodococcus equiis an intercompartmental pH‐neutralising virulence factor

“…2 hr of infection. Phagosomes containing strain 103− steadily acidify to a pH of 5.0 (von Bargen, Wohlmann, Taylor, Utermöhlen, & Haas, 2011). We now show that the pH of phagosomes containing strain ΔvapA decreases quickly to~5.8 and does not show any pH rebound as with phagosomes containing strain 103+ ( Figure 5d).…”

“…Multiplication of virulent strain 103+ was not affected by addition of rVapA ( Figure S7A) indicating that virulent R. equi produce saturating amounts of VapA. It also argues that the extra rVapA does not interfere with multiplication of virulent R. equi, for example, by damaging or activating macrophages which, in turn, could kill R. equi (von Bargen et al, 2011). Of note, the vacuoles, which contained avirulent strain 103− in rVapA-fed cells at 24 hr of infection were packed with vesicular cargo ( Figure S7B) and thus also revealed the typical complex RCV pattern (compare Figures S6B and S7C).…”

“…Similarly as above ("Determination of lysosome pH"), phagosome pH was quantified by ratiometric fluorometry as before (von Bargen et al, 2011). To quantify phagosome colocalization with LT microscopically, J774E cells were infected as above with R. equi as above.…”

Investigation of infectivity of neonates and adults from different rat strains to Toxoplasma gondii Prugniaud shows both variation which correlates with iNOS and Arginase-1 activity and increased susceptibility of neonates to infection

“…However, the true identity of Toxo1 has not been confirmed. A large number of reports have demonstrated that nitric oxide (NO) is a major effector molecule for macrophage-mediated cytotoxicity in mouse macrophages and is a key anti-pathogen factor used by the infected host to control progression of intracellular pathogens including Toxoplasma (Adams et al, 1990;Davis et al, 2007;EI Kasmi et al, 2008;James, 1995;Von Bargen et al, 2011). Recent evidence indicates that the high expression of inducible nitric oxide synthase (iNOS), which is also located on chromosome 10, and low expression of Arg-1 in the macrophages of rats are strongly linked to this resistance when T. gondii RH strain was used to infect the cells (Li et al, 2012;Zhao et al, 2013).…”

Mycobacteria and the Intraphagosomal Environment: Take It With a Pinch of Salt(s)!

“…Interestingly, the authors determined that macrophage activation results in Fe 2+ depletion that can be compensated by external supplementation in the medium. Restoring R. equi access to Fe 2+ fully reversed the growth‐inhibitory effects of RNI .…”