The Dot/Icm Effector SdhA Is Necessary for Virulence of Legionella pneumophila in Galleria mellonella and A/J Mice

Abstract

bLegionella pneumophila is an intracellular bacterium that resides within amoebae and macrophages in a specialized compartment termed the Legionella-containing vacuole (LCV). As well as providing an intracellular niche for replication, the LCV helps to prevent the release of bacterial components into the cytoplasm. Recognition of these components as danger signals by the host activates immune responses leading to clearance of the bacterium. Here, we examined the role of two important virulence factors of L. pneumophila, the potent danger signal flagellin and the translocated Dot/Icm type IVB secretion system effector SdhA, which is crucial to maintain LCV integrity, in the Galleria mellonella infection model. We demonstrate that flagellin expression does not contribute to virulence, replication, or induction of clearance mechanisms. Conversely, SdhA expression is important for virulence. We found that in the absence of SdhA, the LCV in hemocytes showed signs of instability and leakage. Furthermore, in contrast to wild-type L. pneumophila, a ⌬sdhA mutant caused a transient depletion of hemocytes and reduced mortality. Analysis of the ⌬sdhA mutant in the A/J mouse model also showed a significant replication defect. Together, our data underline the crucial importance of SdhA in infection across different model organisms.

Use of <em>Galleria mellonella</em> as a Model Organism to Study <em>Legionella pneumophila</em> Infection

“…As L. pneumophila is expected to lyse host cells at the end of its replication cycle, measuring hemocyte depletion may also serve as an indirect measurement of bacterial replication. Hemocyte depletion has previously been correlated with insect mortality in infection 3,52 , although recent results suggest that this picture is more complex than first belived 37 . Recently, it has been shown that starvation of larvae leads to increased susceptibility to infection through a suppression of immune responses 53 .…”

“…In addition, use of this model will allow the identification and further characterization of novel virulence factors including secreted and translocated effector proteins. Recently, it has been shown that phospholipase C activity of L. pneumophila has a role in G. mellonella virulence 60 and that the Dot/Icm effector protein SdhA is required for virulence 37 . In addition, we have recently demonstrated that there is a correlation between the phenotypes observed in G. mellonella and in the A/J mouse strain 37 .…”

Legionella pneumophila Effector LpdA Is a Palmitoylated Phospholipase D Virulence Factor

“…All mouse procedures were approved by the University of Melbourne Animal Ethics Committee and carried out as described previously (41). Briefly, groups of mice were inoculated separately with ϳ2.5 ϫ 10 6 CFU of each L. pneumophila 130b strain under investigation.…”

Creating a customized intracellular niche: subversion of host cell signaling byLegionellatype IV secretion system effectors

“…This triggers a type I interferon immune response and rapid inflammasome-mediated caspase-1 activation and proinflammatory cell death, called pyroptosis, depriving Legionella of its intracellular niche before it can efficiently replicate (Monroe et al 2009;Ge et al 2012). Deletion of SdhA results in a growth defect of L. pneumophila in the Galleria mellonella infection model, in the lungs of mice, in murine bone-marrowderived macrophages and, to a lesser extent, human U937 macrophage-like cells, and in amoebae (Laguna et al 2006;Harding et al 2013b).…”