Members of the Enterobacteriaceae family produce a diffusible signal factor that modulates enteric pathogen virulence

Diffusible signal factors (DSFs) are a gammaproteobacterial family of quorum-sensing signals that mediate the transition between sessile and planktonic lifestyles. In addition to coordinating behavior within producing species, DSFs can be sensed by surrounding organisms, enabling interspecies "eavesdropping." Several enteric pathogens that do not produce DSFs use these environmental signals to modulate their virulence, allowing them to conserve energy when expression of pathogenesis genes is not advantageous. Although DSFs have been detected in the gut, the microbial origins of these signals remain poorly defined. Here, we show that the ability to produce DSFs is more widespread among Gammaproteobacteria than previously appreciated. Using phylogenetic analysis, we identified more than 500 homologs of the DSF synthase RpfF that group into three distinct taxa. Our analysis further revealed evidence of horizontal gene transfer between Burkholderiales, a known reservoir of DSF production, and Enterobacterales, an order enriched in gut-associated bacteria. We found that Enterobacterales members Cronobacter sakazakii and Enterobacter cloacae produce the DSF cis-2-tetradecenoic acid in an rpfF-dependent manner. Furthermore, we demonstrated cis-2-tetradecenoic acid to be a potent repressor of virulence in the enteric pathogens Salmonella enterica, Shigella flexneri, and Vibrio cholerae. Finally, we have shown that in addition to its role in interspecies signaling, endogenous DSF production reduces biofilm formation and suppresses swarming in E. cloacae, while also promoting swimming motility, consistent with a shift to a free-living lifestyle. Together, these findings expand the known diversity of DSF signaling and identify gut-associated bacteria as a source of interspecies cues that shape pathogen behavior.

Many bacterial pathogens rely on tightly regulated gene expression to cause disease, making virulence pathways attractive targets for new therapies. Diffusible signal factors (DSFs) are communication molecules that can suppress the virulence of several enteric pathogens, but their sources in the gut have not been well defined. Here, we identify gut-associated bacteria that produce the DSF molecule cis-2-tetradecenoic acid, capable of strongly repressing virulence in multiple enteric pathogens. Additionally, our work substantially expands the distribution of bacteria with the genetic capacity for DSF production, broadening the pool of candidate signal sources and paving the way for future discoveries in microbial communication.