The Measurement of the Response of Immunized Mice to Infection with Mycobacterium Tuberculosis Var. Hominis

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Abstract
Summary: Data have been presented which show the response of normal and vaccinated mice, in terms of time of death, to intravenously administered increasing doses of virulent human-type tubercle bacilli strain H37Rv. The deaths of normal mice infected intravenously with small doses and the deaths of vaccinated mice infected intravenously with slightly larger doses were found to be so anormally distributed that neither the mean survival time nor the median survival time could be used as the measurement of the average response. Instead, the response of such mice to the tuberculous infection was measured by recording the number of mice which survived 30 days or longer, and the rationale for this procedure is given. Only by a determination of the relationship between the number of tubercle bacilli in the challenge dose and the response of both normal and vaccinated mice can a suitable single challenge test dose of virulent tubercle bacilli be selected to measure the degree of immunity to this disease in mice. Using this method of evaluation, it was shown that neither large single doses nor small single challenge doses of the H37Ra strain of Mycobacterium tuberculosis var. hominis could be used to measure the immune state of mice, since both normal and vaccinated mice responded to such doses in a similar fashion. Only when intermediate doses (0.5 to 1.5 mg) were used was an adequate differentiation between normal and vaccinated mice obtained. Alternatively, the degree of immunity in mice may be measured by determining the 30-day lethal dose/50 (30-DLD50). This can be done by determining the number of tubercle bacilli which will kill 50% of the normal mice within 30 days, and comparing this with the number of tubercle bacilli required to kill 50% of the vaccinated animals within the same period. Application of the above principles to the problem of the measurement of the immunity produced in mice by the injection of various agents has shown that vaccination with 3 strains of BCG produced a significantly greater degree of immunity in mice than did vaccinaton with a fourth strain of BCG or with the H37Ra strain. The latter 2 strains were equivalent in their immunity-producing ability. Ultraviolet irradiation almost completely abolished the ability of tubercle bacilli to stimulate production of immunity in mice, whereas heat (126°C for 20 min) reduced the immunizing capacity of the H37Ra and BCG strains about 50%. Unrelated organisms such as Bacillus subtilis, Bacillus sphaericus, and Hemophilus pertussis did not stimulate the production of any immunity in “Strong A” strain mice. Booster doses of the H37Ra strain administered either 2 or 4 days before challenge nearly doubled the immune response of “Strong A” mice to tuberculous infection. The “Strong A” strain mice were found to develop a higher degree of acquired immunity to tuberculous infection than mice of the “Rockland” all-purpose strain.