II. THE BEHAVIOR OF TETANUS SPORES INJECTED SUBCUTANE OUSLY INTO GUINEA PIGS AND WHITE MICE. Since the subcutaneous injection of guinea pigs and white mice with vaccine virus constitutes one of the methods of testing the virus for contamination, a thorough knowledge of the resistance and susceptibility of these animals to tetanus is therefore highly desirable. THE EFFECT IN GUINEA PIGS OF THE INJECTION OF QUININE OR STAPHYLOCOCCI AT THE SITE OF INJECTION OF TETANUS SPORES. Tetanus spores in pure culture and free from tetanus toxin, when injected subcutaneously into a guinea pig, do not give rise to tetanus. The animal remains well. (See Table No. 9, guinea pigs 5, 6, 11, and 12.) The spores lie dormant as an inert body at the point of injection awaiting either destruction by the phagocytes or an awakening into activity and toxin production by the irritation of some foreign body, particularly quinine or staphylococci. If quinine or staphylococci be combined with the spores before injection and both are injected at the same time into the same site, tetanus and death of the animal follow promptly and regularly within 3 to 6 days. (See Table No. 9, guinea pigs 1, 2, 3, 4, 7, 8, 9, and 10.) TABLE NO. 9.-Activation in guinea pigs of tetanus spores by quinine or staphylococcus. If a length of time be allowed to elapse after the injection of the spores, before injecting quinine or staphylococci into the site of the spore injection, tetanus will or will not follow in a guinea pig depending upon whether the interval has been sufficient to allow the devour ing process of the phagocytes to effect only a partial or a complete destruction of the spores. An effort was made to determine how long tetanus spores will be harbored by a guinea pig in its subcutaneous tissues before they become destroyed by the phagocytes. Quinine or staphylococci injected at the site of the spore injection was depended upon to activate any spores which had escaped phagocytosis. Forty-five tests in guinea pigs were made in which various spaces of time elapsed between the subcutaneous injection of tetanus spores and the injection of quinine or staphylococci into the site of the spore injection. (See Table No. 10, guinea pigs Nos. 1 to 25; see Table No. 13, guinea pigs Nos. 1 to 15, except 7; guinea pigs 23, 24, 25, 32, 33, and 34; see summary, Table No. 12.) The intervals between the two injections varied from 9 to 120 days and the result was indicated either as a success in producing tetanus or as a failure to do so. TABLE NO. 10.—Effect of lapse of time between injection of tetanus spores and injection of activator, both at same site, in guinea pigs. An interval of 9 days was followed by 1 success and 1 failure. An interval of 14 days was followed by 1 success and 1 failure. An interval of 16 days was followed by 3 successes and 1 failure. An interval of 30 days was followed by 6 successes and 10 failures. An interval between 30 and 60 days was followed by 7 failures. An interval between 60 and 90 days was followed by 2 successes and 11 failures. An interval between 90 and 120 days was followed by 1 failure. The shorter the interval, the more constant is the activating power of quinine or staphylococcus. The longer the interval, the more complete is the destruction of spores by the phagocytes. When the injections were simultaneous, 8 guinea pigs all contracted tetanus; when the interval was between 9 and 30 days, 11 contracted tetanus and 13 did not; when the interval was between 30 and 90 days, 2 contracted tetanus and 18 did not; when the interval was 120 days, 1 did not contract tetanus-only 1, however, was tested. While a simultaneous injection of tetanus spores and quinine or staphylococci is certain to produce tetanus in a guinea pig, an interval of 9 to 30 days between the injections gives the phagocytes and spores about an equal chance, and half of the animals will contract tetanus and half will not; any lengthening of the interval beyond 30 days very rapidly shifts the advantage to the phagocytes and no tetanus follows. The data indicates that after an interval of 30 days, tetanus spores are practically all destroyed in the subcutaneous tissue of a guinea pig, and the chances of any spores remaining dormant and susceptible of activation longer than 30 days are very remote. THE EFFECT IN WHITE MICE OF THE INJECTION OF QUININE OR STAPHYLOCOCCI AT THE SITE OF INJECTION OF TETANUS SPORES. White mice injected subcutaneously on the back with a pure culture of tetanus spores free from tetanus toxin do not contract tetanus in the absence of a suitable activator of the spores. (See Table No. 11, Mice 1 to 20.) |