Enumeration of viable tubercle bacilli (BCG) by the roll-tube method.

A technique for counting viable BCG particles by the roll-tube method is described.The bacterial suspension is diluted in a 5% solution of bovine albumin (Armour, fraction V) and five drops (0.1 ml) of the appropriate dilution are delivered into each of a number of bottles containing 0.9 ml of Dubos basic medium with 2% agar; the contents of the bottles are maintained in a liquid state at 45 degrees C before inoculation. The inoculation bottles are sealed with rubber stoppers and rolled. The colonies are counted after three to five weeks' incubation at 37 degrees C.Data from a series of counts indicate conformity with a Poisson distribution.The incorporation of oleic acid in the medium is not advisable because it results in low counts and an abnormal dispersion of the numbers obtained with parallel bottles, although the colonies are considerably larger than they are when oleic acid is excluded.