quantifying-vibrio-cholerae-colonization-diarrhea-adult-zebrafish

Quantifying <em>Vibrio cholerae</em> Colonization and Diarrhea in the Adult Zebrafish Model

Vibrio cholerae is best known as the infectious agent that causes the human disease cholera. Outside the human host, V. cholerae primarily exists in the ...

Wayne State University School of Medicine

The human diarrheal disease cholera is caused by the aquatic bacterium Vibrio cholerae. V. cholerae in the environment is associated with several varieties of aquatic life, including insect egg masses, shellfish, and vertebrate fish. Here we describe a novel animal model for V. cholerae, the zebrafish. Pandemic V. cholerae strains specifically colonize the zebrafish intestinal tract after exposure in water with no manipulation of the animal required. Colonization occurs in close contact with the intestinal epithelium and mimics colonization observed in mammals. Zebrafish that are colonized by V. cholerae transmit the bacteria to naive fish, which then become colonized. Striking differences in colonization between V. cholerae classical and El Tor biotypes were apparent. The zebrafish natural habitat in Asia heavily overlaps areas where cholera is endemic, suggesting that zebrafish and V. cholerae evolved in close contact with each other. Thus, the zebrafish provides a natural host model for the study of V. cholerae colonization, transmission, and environmental survival. 

Zebrafish as a natural host model for Vibrio cholerae colonization and transmission.

Vibrio cholerae is autochthonous to various aquatic niches and is the etiological agent of the life-threatening diarrheal disease cholera. The persistence of V. cholerae in natural habitats is a crucial factor in the epidemiology of cholera. In contrast to the well-studied V. cholerae-chitin connection, scarce information is available about the factors employed by the bacteria for the interaction with collagens. Collagens might serve as biologically relevant substrates, because they are the most abundant protein constituents of metazoan tissues and V. cholerae has been identified in association with invertebrate and vertebrate marine animals, as well as in a benthic zone of the ocean where organic matter, including collagens, accumulates. Here, we describe the characterization of the V. cholerae putative collagenase, VchC, encoded by open reading frame VC1650 and belonging to the subfamily M9A peptidases. Our studies demonstrate that VchC is an extracellular collagenase degrading native type I collagen of fish and mammalian origin. Alteration of the predicted catalytic residues coordinating zinc ions completely abolished the protein enzymatic activity but did not affect the translocation of the protease by the type II secretion pathway into the extracellular milieu. We also show that the protease undergoes a maturation process with the aid of a secreted factor(s). Finally, we propose that V. cholerae is a collagenovorous bacterium, as it is able to utilize collagen as a sole nutrient source. This study initiates new lines of investigations aiming to uncover the structural and functional components of the V. cholerae collagen utilization program. 

A metalloprotease secreted by the type II secretion system links Vibrio cholerae with collagen.

 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.  is an aquatic bacterium that inhabits coastal and estuarine areas, and 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  intestinal colonization, pathogenesis, and transmission.As early as six hours after exposure to , 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  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  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. Identification of the accessory toxins that cause diarrhea in zebrafish can help us understand more about the role of fish in the wild as aquatic reservoirs for  It is plausible that accessory toxins can act to prolong colonization and subsequent shedding of  back into the environment, thus perpetuating and facilitating transmission during an outbreak. It is also possible that accessory toxins help to maintain low levels of intestinal colonization in fish, giving  an advantage when environmental conditions are not optimal for survival in the water. Studies such as this one are critical because fish could be an overlooked source of cholera transmission in the environment.

Kristie Mitchell

Department of Microbiology,
Immunology,
and Biochemistry,
Department of Microbiology, Immunology, and Biochemistry

Wayne State University School of Medicine

Quantifying Vibrio cholerae Colonization and Diarrhea in the Adult Zebrafish Model

Kristie Mitchell

.css-o250i3{font-size:18px;font-family:Open Sans,sans-serif;line-height:21.6px;}Department of Microbiology,Immunology,and Biochemistry,Department of Microbiology, Immunology, and Biochemistry

Wayne State University School of Medicine

Quantifying Vibrio cholerae Colonization and Diarrhea in the Adult Zebrafish Model

Department of Microbiology,
Immunology,
and Biochemistry,
Department of Microbiology, Immunology, and Biochemistry