Capstone Title

An Examination of Tetrodotoxin Origin in Cultured and Wild Puffer Fishes

Defense Date


Document Type


Degree Name

M.S. Marine Biology

First Advisor

Donald McCorquodale

Second Advisor

Richard E. Spieler


Several members of the puffer fish family Tetraodontidae, are known to contain tetrodotoxin (TTX), a potent toxin that acts on the peripheral and central nervous systems when consumed. Although it is well known that "fugu", or prepared puffer fish, can sometimes prove fatal if eaten, it still remains a delicacy in Japan. Initially, it was believed that the origin of the toxin was endogenous, or produced directly by the fish. However, research conducted by Mari Yotsu in 1986 isolated the TTX producing bacteria, Pseudomonas sp., from the skin of the puffer fish, Fugu poecilonotus. In 1987, Simidu et al. (1987), found what they believed to be a symbiotic relationship between TTX producing bacteria and host puffer fishes. In that same year, a study was published by Noguchi et al. (1987), that found that the bacteria Vibrio alginolyticus was closely involved in the toxification of aquatic animals containing TTX. It was also discovered that the bacteria Pseudomonas sp had been implicated in the toxification of several marine organisms known to produce TTX (Noguchi et al., 1987). Further research conducted in 1989 by Takashi Matsui (1989), also demonstrated that the source of TTX in the puffer fish Takifugu niphobles was exogenous and produced by the bacteria Shewanella putrefaciens. In another study, AbdAllah Aly found that when flatworms containing TTX producing bacteria were fed to nontoxic puffers, the puffers bioconcentrated the bacteria and were soon after found to contain TTX toxins within their tissue (Aly,1991).

This idea that TTX originated from an exogenous source was widely accepted unti11995, when Kendo Matsumura detected peaks and fragment ions in the polypeptone and yeast extracts used in the medium materials resulting in a false positive. This finding would require the authors of previous studies to reexamine the authenticity of their results (Matsumura, 1995). However, in a report published in 1995, Kodama eta/. discovered that when the nontoxic puffer fish T. rubripes were exposed to TTX producing bacteria extracted from the toxic puffer T. porphyreus, there was no observable negative reaction. In addition to this finding, Kodama et al. (1995), found that the nontoxic puffer T. rubripes was able to bioconcentrate these foreign bacteria and accumulate their TTX byproduct (Kodama et al., 1995).

Once again it appeared that the origin of TTX was in fact exogenous until another study conducted in 1998 by Matsumura found that the toxin was bioconcentrating in puffer fish embryos of Fugu niphobles which had been reared in a controlled environment known to have no TTX producing bacteria present (Matsumura, 1998). However, following this study, in 1999 Myoung-Ja Lee was able to identify TTX producing Vibrio strains in the puffer fish Fugu vermicularis radiatus, once again bolstering the argument that the origin of the TTX toxin may be exogenous. At the present time, no consensus has been reached as to which origin of TTX is correct, but based on the supporting evidence, most scholars appear to be leaning towards the exogenous stance.

Tetrodotoxin is a neurotoxin, blocking the movement of sodium ions across neural membranes by attaching to a sodium channel receptor and capping the sodium channel. As a result, TTX blocks nerve impulses and causes paralysis of voluntary muscles. Consequently, this interferes with the muscles of the chest wall and diaphragm, causing respiratory failure. Tetrodotoxin is 10,000 times more lethal than cyanide, as one milligram of TTX can kill a person. At the present time no antidote exists. The amount of toxicity is species dependant and certain organs in the puffer fish may contain higher concentrations of the toxin than others. The most frequently associated organs include the gastro-intestinal tract, liver, roe, ovaries, muscle, and skin.

While several members of the family Tetraodontidae contain TIX, this toxin is not exclusive to this particular family of fish. Other marine organisms such as the Australian blue-ringed octopus (Hapaloclaena macu/osa), boxfish (Ostracion spp.), starfish (Astropecten scoparius), xanthid crab (Eriphia spp.), and horshoe crab (Carcinoscorpius rotund;cauda), also employ TTX, as do many other organisms, as a means of defense and J or subduing prey. The number of organisms found to contain TTX continues to grow. It is theorized that these hosts provide the bacteria with optimal living conditions and in return, the bacteria provide the hosts with a potent form of defense and I or enhanced predatory mechanism. It is well known that the Australian blue-ringed octopus uses specialized salivary glands which contain TTX producing bacteria to subdue it's prey (Johnson, 2002). There is much speculation that TTX containing members of the puffer fish family also exhibit this symbiotic relationship as recent findings have determined that several species of puffer fish have unique tetrodotoxin secreting glands in their skin (Kodama, 1986).

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