The Utilization of Amino Acids and Related Compounds by Clostridium tetani

Abstract

Studies on the metabolism of Clostridium tetani have been confined to the general changes during growth in various media. Woods and Clifton, 1937, and Barker, 1937, reported that Clostridium tetanomorphum, an anaerobe apparently closely related to C. tetani, obtains its energy by the fermentation of certain amino acids, while Clostridium sporogenes (Stickland, 1934-35) and C. botulinum (Clifton, 1940) utilize amino acids primarily by means of the "Stickland reac-tion", a coupled oxidation-reduction between pairs of different amino acids. These studies on the utilization of amino acids and of possible intermediate products by the pathogenic anaerobes (and closely related forms) have been extended to include the metabolic activities of C. tetani. The methods employed were essentially those previously described (Clifton, 1940). The strain of C. tetani (E-35), obtained from Dr. Ivan Hall's collection (No. 130A), is an active toxin producer. RESULTS The organisms were cultivated in broth containing 1 per cent glucose and 0.1 per cent yeast extract (Difco), growth appearing to be more rapid in the presence of glucose altlough no evidence was obtained indicating that glucose was utilized. After 7 days at 37°C. the main products were alcohol, volatile and non-volatile acids, ammonia, carbon dioxide and small amounts of hydrogen. Du-claux distillations of the bichromate oxidation products of the alcohol suggested a mixture of ethyl and butyl alcohols. The actual formation of butyl alcohol and the ratio of the alcohols were not determined. Acetic and butyric acids were found in a ratio of approximately 2 to 1. The non-volatile, ether-soluble acid appeared to be approximately 3 parts of lactic to 1 part of an acid tentatively identified as malic. Carbon dioxide and hydrogen production by washed suspensions of 20-hour cultures of C. tetani in the presence of the more common amino acids, glucose, glycerol, and certain non-amino acids was determined at 37°C. by the Warburg technic. The contents of the Warburg vessels were also analyzed for ammonia when amino acids were employed as the substrate. The amino acids attacked were fermented directly, pairs of amino acids not being required as with C. sporo-genes or C. botulinum. The amounts of hydrogen produced were in most instances not markedly greater than those produced by the control suspensions. Hydrogen production varied to some extent with different suspensions and sub-strates but never exceeded 2-5 per cent of the total gas produced. The rate of