Abstract
The tad operons encode the machinery required for adhesive Flp (fimbrial low-molecular-weight protein) pili biogenesis. Vibrio vulnificus, an opportunistic pathogen, harbors three distinct tad loci. Among them, only tad1 locus was highly upregulated in in vivo growing bacteria compared to in vitro culture condition. To understand the pathogenic roles of the three tad loci during infection, we constructed single, double and triple tad loci deletion mutants. Interestingly, only the Δtad123 triple mutant cells exhibited significantly decreased lethality in mice. Ultrastructural observations revealed short, thin filamentous projections disappeared on the Δtad123 mutant cells. Since the pilin was paradoxically non-immunogenic, a V5 tag was fused to Flp to visualize the pilin protein by using immunogold EM and immunofluorescence microscopy. The Δtad123 mutant cells showed attenuated host cell adhesion, decreased biofilm formation, delayed RtxA1 exotoxin secretion and subsequently impaired translocation across the intestinal epithelium compared to wild type, which could be partially complemented with each wild type operon. The Δtad123 mutant was susceptible to complement-mediated bacteriolysis, predominantly via the alternative pathway, suggesting stealth hiding role of the Tad pili. Complement depletion by treating with anti-C5 antibody rescued the viable count of Δtad123 in infected mouse bloodstream to the level comparable to wild type strain. Taken together, all three tad loci cooperate to confer successful invasion of V. vulnificus into deeper tissue and evasion from host defense mechanisms, ultimately resulting in septicemia.
Highlights
Vibrio vulnificus is an opportunistic Gram-negative marine pathogen that causes fatal septicemia and necrotizing wound infections in susceptible individuals with underlying hepatic diseases and other immunocompromised conditions
We show an important bacterial factor that should be used to adhere to human cells and avoid from host immune system
To understand the roles of the three Tad operons in the pathogenesis, we deleted each of those three gene loci
Summary
Vibrio vulnificus is an opportunistic Gram-negative marine pathogen that causes fatal septicemia and necrotizing wound infections in susceptible individuals with underlying hepatic diseases and other immunocompromised conditions. V. vulnificus is halophilic and found worldwide in warm coastal and brackish waters in association with shellfish such as oysters and other sea animals. In humans, this pathogen frequently causes rapidly progressing fatal sepsis with a mortality rate of greater than 50% within a few days post-infection after eating raw seafood and contamination of preexisting wounds [1,2,3,4]. To establish successful infections in vivo, V. vulnificus must manage spatiotemporally coordinated changes in the expression levels of various virulence genes. To understand the genome-wide gene expression changes in V. vulnificus after infection, we recently performed a transcriptomic analysis of cells grown in vivo using a rat peritoneal infection model. The Tad proteins have been reported to be essential for adherence, biofilm formation, colonization, and pathogenesis in a number of genera and are considered to be instrumental in the colonization of diverse environmental niches [6, 7, 10,11,12]
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