Tetrodotoxin-producing bacteria from the blue-ringed octopus Octopus maculosus

Abstract

Several live specimens of the blue-ringed octopus Octopus maculosus were collected from the Philippines in November 1985, and from Japan in February 1986, and the distribution of toxicity, along with toxin composition, in the posterior salivary gland and other soft parts were examined. Tetrodotoxin (TTX: 1400 mouse units g-1) was detected in the posterior salivary gland of a Japanese specimen, while not only the salivary gland but other soft parts were toxic in the Philippine specimens. The Philippine specimens contained TTX and anhydrotetrodotoxin, the Japanese specimen TTX, 4-epitetrodotoxin, and an unknown toxin. The posterior salivary gland, intestine and other parts were excised from the Philippine specimens and examined for bacterial flora. Twenty-two dominant strains were isolated and cultured in a 2xORI medium (Ocean Research Institute, Simidu and Tsukamoto 1985) at 20°C for 20 to 48 h. Cells were harvested by centrifugation, and disrupted by ultrasonication. The toxins were partially purified from the cell lyzate by ultrafiltration and Bio-Gel P-2 column-chromatography. Instrumental analyses disclosed that 16 of the 22 strains produced TTX and/or related substances. Six strains which clearly exhibited TTX productivity were identified as Alteromonas (2 strains), Bacillus (2), Pseudomonas (1) and Vibrio (1), based on biochemical and biological characteristics. Of these, one strain each of Bacillus and Pseudomonas produced TTX at a level detectable by the mouse assay.