Type VI Secretion System and Its Effectors PdpC, PdpD, and OpiA Contribute to; Francisella; Virulence in Galleria mellonella Larvae

Francisella tularensis causes the deadly zoonotic disease tularemia in humans and is able to infect a broad range of organisms including arthropods, which are thought to play a major role in; Francisella; transmission. However, while mammalian; in vitro; and; in vivo; infection models are widely used to investigate; Francisella; pathogenicity, a detailed characterization of the major; Francisella; virulence factor, a noncanonical type VI secretion system (T6SS), in an arthropod; in vivo; infection model is missing. Here, we use Galleria mellonella larvae to analyze the role of the; Francisella; T6SS and its corresponding effectors in F. tularensis subsp.; novicida; virulence. We report that G. mellonella larvae killing depends on the functional T6SS and infectious dose. In contrast to other mammalian; in vivo; infection models, even one of the T6SS effectors PdpC, PdpD, or OpiA is sufficient to kill G. mellonella larvae, while sheath recycling by ClpB is dispensable. We further demonstrate that treatment by polyethylene glycol (PEG) activates; Francisella; T6SS in liquid culture and that this is independent of the response regulator PmrA. PEG-activated IglC secretion is dependent on T6SS structural component PdpB but independent of putative effectors PdpC, PdpD, AnmK, OpiB; 1; , OpiB; 2; , and OpiB; 3; . The results of larvae infection and secretion assay suggest that AnmK, a putative T6SS component with unknown function, interferes with OpiA-mediated toxicity but not with general T6SS activity. We establish that the easy-to-use G. mellonella larvae infection model provides new insights into the function of T6SS and pathogenesis of; Francisella;