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ABSTRACTVibrio choleraeis a gram-negative bacterium that is the causative agent of cholera, a severe diarrheal illness. The two biotypes of V. choleraeO1 capable of causing cholera, classical and El Tor, require different in vitro growth conditions for induction of virulence gene expression. Growth under the inducing conditions or infection of a host initiates a complex regulatory cascade that results in production of ToxT, a regulatory protein that directly activates transcription of the genes encoding cholera toxin (CT), toxin-coregulated pilus (TCP), and other virulence genes. Previous studies have shown that sodium bicarbonate induces CT expression in the V. choleraeEl Tor biotype. However, the mechanism for bicarbonate-mediated CT induction has not been defined. In this study, we demonstrate that bicarbonate stimulates virulence gene expression by enhancing ToxT activity. Both the classical and El Tor biotypes produce inactive ToxT protein when they are cultured statically in the absence of bicarbonate. Addition of bicarbonate to the culture medium does not affect ToxT production but causes a significant increase in CT and TCP expression in both biotypes. Ethoxyzolamide, a potent carbonic anhydrase inhibitor, inhibits bicarbonate-mediated virulence induction, suggesting that conversion of CO2into bicarbonate by carbonic anhydrase plays a role in virulence induction. Thus, bicarbonate is the first positive effector for ToxT activity to be identified. Given that bicarbonate is present at high concentration in the upper small intestine where V. choleraecolonizes, bicarbonate is likely an important chemical stimulus that V. choleraesenses and that induces virulence during the natural course of infection.

Zebrafish (Danio rerio) is an attractive model organism to use for an array of scientific studies, including host-microbe interactions. Zebrafish contain a core (i.e., consistently detected) intestinal microbiome consisting primarily of Proteobacteria.

Zebrafish (Danio rerio) are an attractive model organism for a variety of scientific studies, including host-microbe interactions. The organism is particularly useful for the study of aquatic microbes that can colonize vertebrate hosts, including Vibrio cholerae, an intestinal pathogen. V. choleraemust colonize the intestine of an exposed host for pathogenicity to occur.

The Vibrio choleraeO1 serogroup is responsible for pandemic cholera and is divided into the classical and El Tor biotypes. Classical V. choleraeproduces acid when using glucose as a carbon source, whereas El Tor V. choleraeproduces the neutral product acetoin when using glucose as a carbon source.