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The (p)ppGpp synthetase RelA contributes to stress adaptation and virulence in Enterococcus faecalis V583

Vianney Pichereau^1,

¹ Laboratoire Microbiologie de l'Environnement, EA956 – USC INRA 2017 – IFR146 ICORE, Université de Caen, 14032 Caen Cedex, France
² Schepens Eye Research Institute, Department of Ophthalmology, Harvard Medical School, Boston, MA, USA

Guanosine penta- and tetraphosphate [(p)ppGpp] are two unusual nucleotides implied in the bacterial stringent response. In many pathogenic bacteria, mutants unable to synthesize these molecules lose their virulence. In Gram-positive bacteria such as Enterococcus faecalis, the synthesis and degradation of (p)ppGpp mainly depend on the activity of a bifunctional enzyme, encoded by the relA gene. By analysing relA and relQ (which encodes a protein harbouring a ppGpp synthetase activity) deletion mutants, we showed that RelA is by far the main system leading to (p)ppGpp production under our experimental conditions, and during the development of a stringent response induced by mupirocin. We also constructed a mutant (relAsp) in which a small part of the relA gene (about 0.7 kbp) encoding the carboxy-terminal domain of the RelA protein was deleted. Both relA mutants were more resistant than the wild-type strain to 0.3 % bile salts, 25 % ethanol and acid (pH 2.3) challenges. Interestingly, the relAsp mutant grew better than the two other strains in the presence of 1 mM H₂O₂, but did not display increased tolerance when subjected to lethal doses of H₂O₂ (45 mM). By contrast, the relA mutant was highly sensitive to 45 mM H₂O₂ and displayed reduced growth in a medium containing 1 M NaCl. The two mutants also displayed contrasting virulence phenotypes towards larvae of the Greater Wax Moth infection model Galleria mellonella. Indeed, although the relA mutant did not display any phenotype, the relAsp mutant was more virulent than the wild-type strain. This virulent phenotype should stem from its increased ability to proliferate under oxidative environments.

Correspondence
Vianney Pichereau
vianney.pichereau{at}univ-brest.fr

Present address: Laboratoire des Sciences de l'Environnement Marin, UMR CNRS 6539, Institut Universitaire Européen de la Mer, Technopole Brest Iroise, 29280 Plouzané, France.