Jury

  • Prof. Régis HALLEZ (UNamur), Président
  • Prof. Jean-Yves MATROULE (UNamur), Secrétaire
  • Dr Rob VAN HOUDT (SCK CEN)
  • Dr Kristel MIJNENDONCKX (SCK CEN)
  • Prof. Liselot DEWACHTER (UCLouvain)

Abstract

The viable-but-nonculturable (VBNC) state is a type of bacterial dormancy triggered by sublethal stress, where cells remain intact but lose the ability to grow on standard media. This poses challenges for microbial monitoring and public health, as VBNC cells can evade detection and might regain virulence upon resuscitation. Copper compounds are explored as antimicrobial agents, however sublethal Cu concentrations were shown to induce the VBNC state in certain bacteria. This thesis investigates the Cu-induced VBNC state in Cupriavidus metallidurans, a metal-resistant betaproteobacterium, and examines the involvement of its Cu resistance determinants (CRDs). While resuscitation is usually mediated by external factors, we aimed to uncover intrinsic processes that enable spontaneous resuscitation, a rare phenotype lacking mechanistic understanding. Proteomic analysis revealed that expression of CRDs, among others, correlated with mitigated dormancy. Time-resolved profiling showed that VBNC cells exhibit highly dynamic proteomes: VBNC entry involved oxidative stress response, and resuscitation correlated with metabolic reconstitution and the strong induction of periplasmic CRDs. Temporal clustering corroborated the explored proteomic modifications. Through mutational studies we identified the plasmid-encoded copAB system as the minimal resuscitation factor, where integrity of the CopA methionine-rich domain proved critical. ICP-MS analysis indicated that detoxification relies on Cu sequestration rather than export. Altogether, this work uncovers key intrinsic factors and proposes a mechanistic basis for spontaneous resuscitation from the Cu-induced VBNC state in C. metallidurans. These insights refine our understanding of the VBNC state as a dynamic survival strategy and of bacterial Cu resistance.