TlyC, a conserved hemolysin in Rickettsia, contributes to spotted fever pathogenesis in mice

Rickettsia circulates between mammalian hosts and hematophagous arthropod vectors by exploiting their intracellular environment. With advances in rickettsial genetic tools, recent studies have identified novel molecular mechanisms involved in Rickettsia-host-vector interactions. However, a significant knowledge gap exists in understanding how Rickettsia modulates its virulence functions to survive in two drastically different environments (mammalian hosts vs arthropod vectors). Bacterial hemolysins play a crucial role in neutralizing innate immune functions through pore-forming activities and direct interactions with cognate receptors. Prior work suggested that typhus group Rickettsia (e.g., R. typhi and R. prowazekii), but not spotted fever group Rickettsia (e.g., R. conorii and R. parkeri), can induce hemolytic activities upon direct interactions with red blood cells. Here, we demonstrate that typhus and spotted fever groups exhibit comparable hemolytic activities. Furthermore, by characterizing an R. conorii transposon insertional variant (HK27), we document that TlyC, a factor conserved in Rickettsia, is responsible for pH-, temperature-, and host species-dependent hemolytic activities. Our biochemical and genetic studies confirmed that the first 10 amino acids are critical in facilitating hemolytic activities without affecting TlyC localization to the outer membrane. Compared to wild-type R. conorii, the HK27 variant showed reduced intracellular survival in primary endothelial cells and attenuated virulence in mice. These findings suggest a functional role for a conserved hemolysin in rickettsial pathogenesis.