Quantifying Vibrio cholerae enterotoxicity in a zebrafish infection model

Abstract

Vibrio cholerae is the etiological agent of cholera, an acute intestinal infection in humans characterized by voluminous watery diarrhea. Cholera is spread through ingestion of contaminated food or water, primarily in developing countries that lack the proper infrastructure for proper water and sewage treatment. Vibrio cholerae is an aquatic bacterium that inhabits coastal and estuarine areas, and it is known to have several environmental reservoirs, including fish. Our laboratory has recently described the use of the zebrafish as a new animal model for the study of V. cholerae intestinal colonization, pathogenesis, and transmission. As early as 6 h after exposure to V. cholerae, zebrafish develop diarrhea. Prior work in our laboratory has shown that this is not due to the action of cholera toxin. We hypothesize that accessory toxins produced by V. cholerae are the cause of diarrhea in infected zebrafish. In order to assess the effects of accessory toxins in the zebrafish, it was necessary to develop a method of quantifying diarrheal volume as a measure of pathogenesis. Here, we have adapted cell density, protein, and mucin assays, along with enumeration of V. cholerae in the zebrafish intestinal tract and in the infection water, to achieve this goal. Combined, these assays should help us determine which toxins have the greatest diarrheagenic effect in fish and, consequently, which toxins may play a role in environmental transmission.