It was already reported that even if Dharmendra antigen produced by chemical treatment of leprom was intracutaneously injected into healthy guinea pigs 8 times at an interval of two weeks, the size of reaction was only less than 5mm, and the injection of Dharmendra antigen produced no immunity. Accordingly, the investigation was made as to what effect was produced by the frequent injection of Mitsuda antigen without chemical treatment.
In the animals to which Mitsuda antigen was intracutaneously injected 6 times at an interval of 2 or 4 weeks, the size of reaction caused by Mitsuda antigen increased from 4mm to 15mm, and the magnitude of the tuberculin reaction and the reaction due to Dharmendra antigen as examined in the 8th and 14th weeks was increased as in the results of the reaction caused by Mitsuda antigen. In consequence, it can be said that the intracutaneous reaction caused by Mitsuda antigen produces immunity against leprosy and tuberculosis.
For the investigation on the cause of these phenomenon, therefore, comparison of each skin reaction was made between the reaction in animals sensitized by heat-killed BCG suspension with the human cutaneous tissue and the reaction in animals sensi- tized by heat-killed BCG suspension only, and the size of reaction due to Dharmendra antigen or tuberculin reaction in the foremer animals was larger than that in the latter animals. This fact proves that the tissue and its component acts as adjuvant, sub- stantiating the mode of a sensitization by the intracutaneous injection of Mitsuda antigen. However, it seems that the injection of Mitsuda antigen into human beings should be carried out with attention paid to the fact that the intracutaneous injection of human cutaneous tissue makes the animals allergic.
Moreover, the size of the intracutaneous reaction in animals sensitized by the heat-killed BCG suspension with Mitsuda antigen is larger than that in animals sensi- tized by only either Mitsuda antigen or the heat-killed BCG with tissue suspension, and then, a mixture of Mitsuda antigen and heat-killed BCG suspension is considered to be effective for the prevention of leprosy.

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Though treatment of leprosy has become easy by the introduction of the sulfone drugs and other anti-leprosy agents, cases difficult to treat because of various types of lepra reactions or drug-resistance are still encountered today.
The effect of the Toyoda vaccine, a chloroform extract of BCG in combination with other agents, was examined in cases of this type. The results are here presented.
It has been reported that the Toyoda vaccine is effective in tubercular lym-phadenitis of the cervical lymphnodes and in pulmonary tuberculosis. It has also been tried in leprosy and there have been no undesirable side effects. Of the cases in which the vaccine has been tried, a permanent effect has been noted in many cases of ENL and progressive lepra reaction and it has become possible to administer other forms of anti-leprosy agents by this in some cases while clinical improvement has been noted in several difficult to treat cases.
The use of the vaccine in combination with other agents still required further investigation but it is believed that it will become a valuable aid in the treatment of leprosy.
The finding that a non-specific vaccine such as this plays a role in the improve-ment of clinical symptomes is interesting from the standpoint of the host-parasite relationship in leprosy.

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It was already reported that the repeated injection of Mitsuda antigen produced the accelerated phenomenon on its reactivity, as seen in tuberculin reaction. However, Mitsuda antigen may have the immunogenicity, because this antigen contains the tissue and its component in addition to bacillary body. The immunogenicity of Mitsuda antigen was already reported in animal examination. Accordingly, this phenomenon was investigated in leprosy patients.
The reactions caused by Mitsuda antigen and tissue suspension were compared between the patients who had been subjected to the reaction by Mitsuda antigen and the patients who had been subjected chiefly to the reaction by Dharmendra antigen. In consequence, the sizes of reactions caused by both antigens were larger in the foremer patients than in the latter patients.
This fact should show that Mitsuda antigen had the immunogenicity and particularly the skin-tissue contained in this antigen related with this immunity. Therefore, when Mitsuda antigen is used for the intracutaneous examination in human body, unless this immunogenicity is taken into consideration as shown in the results of animal experiment, it may be the danger of erroneously reading the modified reaction against Mitsuda antigen.

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In order to prepare the standard lepromin from the material with low bacterial contents, it was necessary to remove the majority of tissue components from lepromin without any influence upon its potency. This urgent problem was solved by the follow-ing experiment. The pooled nodules from relapsed patients were divided into two parts, one of which was used for the preparation of the standard lepromin (Ms) by the usual method, and the other part of the material was treated with a mixture of trypsin and bile salt so that the majority of tissue components could be digested, and the partially purified bacillus suspension (Mp) was prepared through centrifugation and washing. In spite of such treatment as above, no increased morphological change of bacilli in Mp was observed, comparing with that of Ms, by the microscopic as well as electron-microscopic examinations.
As the results of comparative tests with leprosy patients, the size of early reaction with Mp in non-lepromatous cases was larger than that of Ms, but the late reactions did not show any significant difference between Mp and Ms. On the other hand, the late reactions with Mp in lepromatous cases showed a gradual decrease from 15 days to 42 days after injection, while no marked change in the period was observed in case of Ms. This difference was considered to be caused by the sensitization due to tissue components in Ms, but not to be caused by the lowered potency of Mp due to the purification procedures.

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Psychological investigations were conducted in Aisei-en National Leprosarium. The findings are summarized as follows:I) Questionnaire applied to 300 cases showed the following facts :
(1) Leprosy patients reveal psychological trouble connected with the course ofleprosy.
(2) They are tired with various conflicts about the actual life in the sanatorium.
(3) Though the long life in the sanatorium, their mind and body are very closely related and interact on each other, therefore we must always lead them medically including mental hygiene.
II) Interviews with 72 cases showed that there are usually three stages through which the patients go after being diagnosed as leprosy patients:
(1) The period of mental blow; the time of diagnosis is usually an extremely trying period filled with anxiety and emotion.
(2) At this stage their feelings are hidden under a cover, so to speak, this being a defense mechanism.
(3) Struggle with the emptiness of their lives and with consequent despair.

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Nishimura et al.1)2) reported that acid-fast bacilli were present in the skin tissue of apparently healthy mice without giving to any symptom. We have also detected abundant acid-fast bacilli in the skin of healthy guinea pig, rabbit and monkey, which could not grow on a medium. The present paper described the results of our attempts to detect acid-fast bacilli in human skin tissues of non-leprosy cases. Some characterizations of the organism collected from one case are also presented. Collection Methods of Acid fast Bacilli from Specimens
Since it was difficult to detect bacilli in the specimens smeared from the skin section, we employed two methods for collection of the bacilli modifying the methods devised by Kar et al.4) and Dharmendra et al.5) According to Kar's method a small amount of 2 per cent of acetic acid was added to 0.1 to 0.5g of the minced skin tissue in order to make the tissue loose. The acid-fast bacilli were extracted with chloroform from the loosened tissue according to the Dharmendra's method. After addition of 20ml of chloroform, the mixture was homogenized for a minute in a Waring blender. Homogenizing with chloroform was repeated five times and the chloroform layers were pooled. Chloroform was evaporated to dryness. The residue was washed with ethyl ether, which was removed by centrifugation at 2, 000 rpm for 2 minutes. The sediment was suspended in a drop of distilled water, one drop put on a slide glass, after dryness the smear was stained with Ziehl-Neelsen stain. For another method of bacilli collection, 5g of skin tissue were minced and divided into five equal portions. To each portion were added 5ml of 2 per cent of acetic acid to minimize the contamination of tissue components and the mixture was homogenized in a Waring blender. The homogenate was stirred well with about 80ml of chloroform and the chloroform layer was transferred into an Erlenmyer flask. The chloroform layers from five portions were pooled and left standing overnight in a refrigerator. Since tissue particles contaminated in chloroform extracts attached by this procedure on the wall of flask, the clear chloroform extracts were obtained. The extracts were treated as mentioned above. Summary and Discussion
1) Attempts were made to isolate acid-fast bacilli from the specimens of ap-parently healthy skin of the human body, which were taken out from non-leprosy patients in the cases of surgical operations or in autopsies. The isolation was performed by the acetic acid-chloroform-ether collection method. The acid-fast bacilli were detected in 80 out of total 105 specimens tested. The detection was irrespective of age. The abundant bacilli were detected in 9 per cent of the positive cases. In the most abundant case, 5*106 bacilli were observed in 5g of the specimen.
2) In a specimen the aggregated bacilli were observed within the Subpapillary layer of the skin, though no sign of inflammation such as infiltration of the inflammatory cells was observed.
3) The specimens were emulsified and inoculated on neutral or acidic egg slants and incubated at both 33°C and 37°C. So far no acid-fast bacilli have grown on the slants.
4) Lepromin test was performed with the antigen prepared either from the isolated acid-fast bacilli or from the human leprosy bacilli. Both antigens gave negative results to the carriers of the isolated acid-fast bacilli and to the lepromatous patients. The tuberculoid leprosy patients reacted to the human leprosy lepromin, but not to the antigen prepared from the isolated acid-fast bacilli.
Figueredo et al.8) reported that acid-fast bacilli were detected in 31 out of 48 individuals who were apparently healthy but had some contacts with leprosy patients. Although he has not characterized the acid-fast bacilli, they suggested that those positive individuals would have been the healthy carriers of leprosy bacilli.

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Since the discovery of anti-leprotic agents such as Sulfone, long acting Sulfamine, Antibiotics and Thiourea etc., other new drugs have been studied in respect to theiranti-leprotic activity.
The thiourea family drugs have been applied in leprosy by many authors, all reporting good results. The chemical structural formula of thiourea and thiosemicar-bazide are exactly like each other. There-fore, the author has expected good effect to treatment of leprosy by thiosemicarbazide drugs. On this idea, the author has synthesed new some of compounds as follow.DOT OH NH CS NHN=CH N CH3CH3 Methoxy-DOT CH3O NH CS NHN=CH N CH3CH3 Ethoxy-DOT C2H5O NH CS NHN=CH N CH3CH3 These compounds have proved antibacterial action (H37RV) in 70, 000 100, 000 times diluted solutions, and these toxicity was very little in experiment.
Accordingly, the author believes these compounds can be used to application forclinic of leprosy or tuberculosis.

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