Attempts at Growth of M. leprae in mice

Abstract

The technical procedures of experiment were explained briefly in Table 1. Bacterial suspension prepared from lepromas was injected into mice once or several times at weekly intervals by subcutaneous or intravenous route, or by both routes. Animals. were killed at various intervals 2-16 months after injection. At necropsy, lesions were sought by gross inspection. Portions of various organs were removed and ground in mortar to make the homogenates. Smears made from the homogenate were stained by Ziehl-Neelsen's method and examined microscopically. The homogenate treated with one percent sodium hydroxide solution and then inoculated onto the egg yolk medium (for M. lepraemurium; also for M. leprae (?)) and Ogawa 1% egg medium (for cultivable mycobacreria). The tubes were incubated at 37°C for over 3 months. The details of single inoculation experiments and multiple inoculation experiments were shown in Table 2 and 3.
Ten experiments containing four with single inoculation and the other six with multiple inoculation were carried out. But one experiment, Expt. (4), exhibited a probable contamination and its results will be described in the following paper separately.
In nine experiments, gross findings were all negative. Cultivation trials showed a few, smooth, and buff colonies, supposedly atypical mycobacseria, from two specimens only, but no colonies of mycobacteria, especially suspected of M. leprae, have been isolated.
On the other hand, microscopic examination revealed the presence of acid-fast bacilli in the tissues of various organs. Among the two experiments, Expts. (3)-1 & -2, showed remaykable microscopscal findings were summasized in Table 6. As shown, in the subcutaneous experiment Expt. (3)-1, acid-fast bacilli found in the injection site and superficial lymph nodes, and none of the tissue of viscera. In the intravenous experiment, Expt. (3)-2, acid-fast bacilli were detected in the spleen, liver and lungs, but smaller in number, comparing with those of the injection site just mentioned. No bacilli were found in the superficial lymph nodes. In both of the experiments the acidfast bacilli had a tendency to decrease in number steadily. In where numbers of bacilli were present, globi were often seen, but these were usually small in size and loose in arrangement. And, indeed, it was uncertain whether the bacilli had multiplied within the tissue or not. The microscopic findings obtained in all the experiments were summarized in Table 7.
As the materials of leproma used differed from experiment to experiment, it was impossible to compare directly the values of percentage for smear-positive specimens. As a whole, however, it seemed fairly justified in concluding that the microscopic findings were superior in the multiple inoculation than in the single inoculation. This fact was accorded with the observations reported by previous workers.
ACKNOWLEDGMENT
This work was supported in part by grants from U. S.-Japan Cooperative Medical Science Program.