Studies of the formation of the streptococcus toxin

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In a paper read before the American Society of Bacteriologists in December, 1924, Huntoon reported observations on the progressive formation of skin-reacting toxin in the Berkefeld filtrate from a 44 hr. culture of hemolytic streptococcus (scarlet fever). He suggested that this formation of toxin in the absence of organisms was probably due to the action of an enzyme (protease), secreted by the bacteria, in the filtrate. The following experiments were conducted to determine something of the nature of this enzyme action. The organism used was a strain of hemolytic streptococcus isolated from a scarlet fever patient and obtained from the Board of Health. Qualitative tests on filtrates obtained by passing a 36 hr. culture (bacto-veal-horse-plasma broth) through a Berkefeld filter, and on extracts of the bacterial bodies (obtained as described below), failed to show the presence of active protease, using as substrate either fibrin or a solution of proteose (prepared from bacto-peptone). Mass cultures were made by growing the streptococcus on whole blood bacto-veal agar. The growth was carefully removed and dried over concentrated sulphuric acid. Three tenths gm. of dried bacteria was well ground and suspended in a M/15 Na, HP04, KH, PO, mixture having a pH of 7.2. This was shaken at intervals during 12 hours, and filtered through Berkefeld. This filtrate was designated “bacterial extract.” Cutaneous tests were made by intradermal injections, using the Dick teichnique, in the arm of an adult male subject who was skin-sensitive to the Dick toxin. After many repeated injections in this subject, it seemed that some change occurred in the skin which made the readings of the tests difficult to interpret. The later experiments, therefore, were done on infant subjects who gave positive (4 plus) reactions to the Dick test, and negative controls.