MARTX Toxin in the Zoonotic Serovar of Vibrio vulnificus Triggers an Early Cytokine Storm in Mice

Abstract

<i>Vibrio vulnificus</i> biotype 2-serovar E is a zoonotic clonal complex that can cause death by sepsis in humans and fish. Unlike other biotypes, Bt2 produces a unique type of MARTX_Vv (Multifunctional-Autoprocessive-Repeats-in-Toxin; RtxA1₃), which is encoded by a gene duplicated in the pVvBt2 plasmid and chromosome II. In this work, we analyzed the activity of this toxin and its role in human sepsis by performing <i>in vitro, ex vivo</i>, and <i>in vivo</i> assays. First, we demonstrated that the ACD domain, present exclusively in this toxin variant, effectively has an actin-cross-linking activity. Second, we determined that the whole toxin caused death of human endotheliocytes and monocytes by lysis and apoptosis, respectively. Finally, we tested the hypothesis that RtxA1₃ contributes to human death caused by this zoonotic serovar by triggering an early cytokine storm in blood. To this end, we used a Bt2-SerE strain (R99) together with its <i>rtxA1</i>_<i>3</i> deficient mutant, and a Bt1 strain (YJ016) producing RtxA1₁ (the most studied MARTX_Vv) together with its <i>rtxA1</i>_<i>1</i> deficient mutant, as controls. Our results showed that RtxA1₃ was essential for virulence, as R99ΔΔ<i>rtxA1</i>_<i>3</i> was completely avirulent in our murine model of infection, and that R99, but not strain YJ016, induced an early, strong and dysregulated immune response involving the up-regulation of a high number of genes. This dysregulated immune response was directly linked to RtxA1₃. Based on these results and those obtained <i>ex vivo</i> (human blood), we propose a model of infection for the zoonotic serovar of <i>V. vulnificus</i>, in which RtxA1₃ would act as a sepsis-inducing toxin.