One of five strains of Vibrio cholerae non-O1 isolated from environmental sources caused fluid accumulation in an initial rabbit ileal loop (RIL) test. The four strains that caused little or no accumulation of fluid gave a positive response after one-to-three consecutive passages through RILs. The amount of fluid produced increased after each passage. Filtrates of cultures of all five environmental isolates caused fluid accumulation similar to that produced by live cells. The enterotoxin showed a precipitin band with new cholera antitoxin and was neutralised completely by new cholera antitoxin diluted 1 in 32, indicating its close immunobiological relationship to the new cholera toxin. The present study indicates that V. cholerae non-O1 strains produce an enterotoxin that is similar to the new cholera toxin.