Studies on the effects of D-Amino Acids on Brucella abortus.

Broth (Albimi) containing 2 per cent "M" peptone, 0.1 per cent glucose, 0.2 per cent yeast autolysate, 0.5 per cent NaCl, and 0.01 per cent sodium bisulfite was the medium used. Four and one-half ml of medium were added to each tube, sterilized for 15 min at 15 lb pressure, and the tubes of media, after inoculation, were incubated at 37 C in air or under varying amounts of carbon dioxide. Three strains of B. abortus were used: a strain which required added carbon dioxide, strain no. 1335; a relatively avirulent strain no. 19; and a highly virulent strain no. 2308. The latter two strains had no added carbon dioxide requirement. The inoculum was prepared by growing the desired strain in 10 ml of "Albimi" broth in air or under 10 per cent carbon dioxide at 37 C for 48 hours. After centrifuging the cells and resuspending them in sterile physiological saline, 0.1 ml of the saline suspension containing approximately 3 X 108 was used as inoculum for each tube of medium. The desired amount of carbon dioxide was obtained by displacing the air with carbon dioxide in an anaerobic jar in which the inoculated tubes of broth were placed. The essential amino acids (Rose, 1937) were made up separately in 0.1 M solutions, sterilized in the autoclave at 15 lb pressure for 15 min, and added aseptically to the broth in amounts to give the desired concentrations. Growth was measured turbidimetrically with an Evelyn colorimeter using the 660 m,u filter. Five ml of water were added to the heat-killed broth cultures before they were read in the colorimeter. The colorimeter scale was set at 100 per cent transmission with the broth diluted with 5 ml of water. To determine the degree of inhibition of B. abortus by the amino acids, the control broth cultures were diluted with the diluted broth medium to 75, 50, 25, and 10 per cent of maximum growth and read in the Evelyn colorimeter. From these readings, a turbidity curve was plotted which was used as a standard of reference. The readings of the growth in the broth tubes containing the amino acids were then used to determine from this curve the amount of inhibition attributable to the amino acids.