Zebrafish as a Natural Host Model for Vibrio cholerae Colonization and Transmission

Abstract

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. Vibrio cholerae, the cause of the severe human diarrheal disease cholera, is also a ubiquitous inhabitant of coastal regions around the globe. As is the case for all species within the Vibrio genus, V. cholerae is an aquatic bacterium that may be found both freely swimming and in association with various forms of aquatic flora and fauna (1-5). The environmental lifestyle and reservoirs of V. cholerae have only in recent years become the subject of vigorous research and remain poorly understood.Over 200 V. cholerae serogroups have been identified from environmental sampling. However, only the O1 and O139 serogroups are capable of causing cholera. The O1 serogroup is further subdivided into two biotypes, classical and El Tor (6). Classical biotype V. cholerae is thought to have caused the first six of the seven known cholera pandemics beginning in 1817 and produces a more severe form of cholera. El Tor V. cholerae is responsible for the seventh pandemic, which began in 1961 and continues to the present day. El Tor strains are thought to be better suited for environmental survival, although the reasons for this are not clear. However, classical biotype strains are currently very difficult, if not impossible, to isolate from the environment, suggesting that El Tor strains have fully occupied the V. cholerae environmental niche. O139 serogroup strains, which caused large cholera outbreaks in the 1990s, have been shown to be derived from El Tor strains (7). In recent years some hybrid strains that closely resemble El Tor strains but also contain genetic material from classical strains have been isolated from cholera patients (8-10).To become a human pathogen, V. cholerae must be ingested in contaminated water or seafood. After ingestion, V. cholerae senses numerous signals resulting in production of virulence factors that permit colonization of the human intestine and ultimately cause the diarrhea that will transmit V. cholerae back into the environment...

Quantifying Vibrio cholerae Enterotoxicity in a Zebrafish Infection Model

“…Previous work in our laboratory confirmed this finding in zebrafish infected with V. cholerae. We found that in time course experiments, zebrafish infected with classical strain O395 remained highly colonized only for about 24 h postinfection and were cleared by 72 h postinfection (18). However, as we have also demonstrated in this study, zebrafish infected with the El Tor strain remained robustly colonized for at least 144 h postinfection, or 6 days.…”

“…Fish were observed to have diarrhea even when infected with ΔtoxT strains of V. cholerae that do not produce cholera toxin, which suggests that there are other factors responsible for inducing diarrhea in infected fish (18). Accessory toxins are the most likely cause of this noncholera diarrhea.…”

“…Our laboratory has developed the zebrafish as a natural host model for the study of V. cholerae infection (18). During colonization experiments with zebrafish, it was observed that within several hours of infection (ϳ 6 to 8 h), fish had what appeared to be diarrhea.…”

Vibrio cholerae Colonization of Soft-Shelled Turtles

“…Compared with those using mammalian animal models, studies with soft-shelled turtles provide a new method for evaluating V. cholerae colonization on aquatic animal surfaces; animals can be continuously observed without euthanization. In addition, zooplankton, fish, shellfish, chironomid egg masses, waterfowl, and crustaceans are environmental hosts of V. cholerae (6,(38)(39)(40)(41)(42)(43), and fish and shellfish are proven vectors of V. cholerae (44). In studies of colonization in aquatic organisms, soft-shelled turtles can be anesthetized and sampled out of water for relatively long periods without the risk of hydropenia and death, due to their amphibious nature, making them a better model than zebrafish and other fish or shrimp models.…”

Engineering Microbial Physiology with Synthetic Polymers: Cationic Polymers Induce Biofilm Formation inVibrio choleraeand Downregulate the Expression of Virulence Genes

“…Zebrafish (Danio rerio) have been established as an aquatic host which can be colonised and infected by V. cholerae in a concentration dependent manner, and infection eventually leads to mortality. [46][47][48] Zebrafish are a suitable natural host model for V. cholerae colonisation and transmission as their gastrointestinal development and physiology closely mimics that of mammalian organisms. 49 Additionally, ease of propagation and live imaging made them a good choice of host for our in vivo studies, in particular at this early .…”