Characterization of the role of putative Aeromonas caviae-specific virulence factor, flgB, in virulence and...

Aeromonas caviae, Gram-negative bacteria ubiquitous in the environment, are an emerging human pathogen associated with various infectious diseases, particularly gastroenteritis. Despite recent studies demonstrating A. caviae is the most predominant Aeromonas species underlying human infection, A. caviae remains understudied, and no A. caviae-specific virulence factors associated with human disease have been identified. To identify A. caviae-specific putative virulence factors, we conducted comparative genomic analyses among clinical Aeromonas isolates (n = 431), which identified a variant of flgB, predicted to encode a polar flagellum machinery protein, as over-represented in A. caviae isolates. To examine the role of flgB in virulence and host-pathogen interactions, we generated an A. caviae flgB deletion mutant and genetic complementation constructs. Swimming motility and polar flagella assembly were abolished in the mutant and functionally rescued with genetic complementation. As it remains unknown where A. caviae infects the human gastrointestinal tract, we assessed host-pathogen interactions in HT-29 and Caco2 human intestinal cell lines, representative of the large and small intestine, respectively. Deletion of flgB significantly decreased bacterial adherence in only HT-29 cells and also decreased production of proinflammatory cytokines, IL-8, IL-13, IL-1β, and IL-6, by both cell types. Given the lack of relevant mammalian models for studying most enteric pathogens in vivo, we characterized in vivo virulence in a Galleria mellonella larval survival model, where the flgB deletion modestly attenuated virulence. Deletion of flgB altered aspects of virulence and host-pathogen interactions, and this study provides a framework for identification and characterization of A. caviae-specific putative virulence factors.IMPORTANCEAeromonas caviae is an emerging human bacterial pathogen associated with gastroenteritis, wound infections, and numerous other infectious diseases. Recent studies demonstrate that A. caviae accounts for the greatest burden of human Aeromonas infections. Despite this, A. caviae is understudied as a human pathogen. To address this gap in knowledge, this study characterizes A. caviae-specific virulence genes. We examined 431 clinical Aeromonas isolates using comparative genomics and identified and functionally characterized a putative A. caviae-specific virulence factor, flgB. Genetic deletion of flgB in A. caviae resulted in deficiencies in bacterial motility, adherence, host-cell proinflammatory cytokine production, and in vivo virulence in an invertebrate model. This work establishes the foundation for further study of additional A. caviae-specific virulence factors.