The Cytolethal Distending Toxin Produced by Nontyphoidal Salmonella Serotypes Javiana, Montevideo, Oranienburg, and Mississippi Induces DNA Damage in a Manner Similar to That of Serotype Typhi

Select nontyphoidal Salmonella enterica (NTS) serotypes were recently found to encode the Salmonella cytolethal distending toxin (S-CDT), an important virulence factor for serotype Typhi, the causative agent of typhoid fever. Using a PCR-based assay, we determined that among 21 NTS serotypes causing the majority of food-borne salmonellosis cases in the United States, genes encoding S-CDT are conserved in isolates representing serotypes Javiana, Montevideo, and Oranienburg but that among serotype Mississippi isolates, the presence of S-CDT-encoding genes is clade associated. HeLa cells infected with representative strains of these S-CDT-positive serotypes had a significantly higher proportion of cells arrested in the G(2)/M phase than HeLa cells infected with representative strains of S-CDT-negative serotypes Typhimurium, Newport, and Enteritidis. The G(2)/M cell cycle arrest was dependent on CdtB, the active subunit of S-CDT, as infection with isogenic Delta cdtB mutants abolished their ability to induce a G(2)/M cell cycle arrest. Infection with S-CDT-encoding serotypes was significantly associated with activation of the host cell's DNA damage response (DDR), a signaling cascade that is important for detecting and repairing damaged DNA. HeLa cell populations infected with S-CDT-positive serotypes had a significantly higher proportion of cells with DDR protein 53BP1 and gamma H2AX foci than cells infected with either S-CDT-negative serotypes or isogenic Delta cdtB strains. Intoxication with S-CDT occurred via autocrine and paracrine pathways, as uninfected HeLa cells among populations of infected cells also had an activated DDR. Overall, we show that S-CDT plays a significant role in the cellular outcome of infection with NTS serotypes. IMPORTANCE The recent discovery that multiple serotypes encode S-CDT, which was previously established as an important virulence factor for serotype Typhi, suggested that this toxin may also contribute to the outcome of infection with nontyphoidal serotypes. In this study, we demonstrate that at a cellular level, S-CDT significantly alters the outcome of infection by inducing DNA damage which is associated with a cell cycle arrest and activation of the host cell's DDR. Importantly, these results contribute valuable information for assessing the public health implications of S-CDT in infections with NTS serotypes. Our data suggest that infection with Salmonella strains that encode S-CDT has the potential to result in DNA damage, which may contribute to long-term sequelae.