A bacterium S184 isolated from the liver of puffer fish was found to produce tetrodotoxin (TTX). The toxin was purified from the cultures by activated charcoal column and C18 cartridge column after being cultured at 28 °C in broth medium. The results of toxicity mouse bioassay, liquid chromatography/fluorescent detection (LC/FLD), liquid chromatography/mass spectrometry (LC/MS), liquid chromatography/tandem mass spectrometry (LC/MS/MS) suggested that the strain could produce TTX during cultivation. The strain was identified asBacillus horikoshii according to its physiological-biochemical characteristics and phylogenetic analysis based on 16S rDNA sequences.