Cultural methods for the production of heat-stable enterotoxin by porcine strains of Escherichia coli and its detection by the infant mouse test.

Casamino acids-yeast extract medium (CAY) and a tryptone-yeast extract medium (TY-1) were evaluated in testing for production of heat-stable enterotoxin (ST) by porcine strains of Escherichia coli using the infant mouse assay. More strains were ST-positive when grown in CAY medium than in TY-1 medium. Questionably or indeterminately ST positive strains were investigated in detail to determine whether or not they were weak ST producers. Growth in four different media and in different batches of CAY medium, inactivation of culture supernatant fluids at a lower temperature, addition of mitomycin C to growing cultures and preparation of periplasm-cytoplasm fractions of bacteria by sonication, all failed to yield ST positive samples. ST value limits (i.e. ratios of intestinal weight to remaining body weight of challenged mice), which clearly differentiated positive or negative strains for ST production, were set for CAY medium. A minimal salts-amino acids medium (MSA) was devised. Both in shake flask and fermenter cultures MSA medium gave better ST yields than CAY and a previously described defined medium.