In the previous report (Tsukamura et al.: Kekkaku, 41: 401, 1966), the present writers stated that pathogenic scotochromogens isolated in Japan showed fairly uniform characters and were to be recognized as a species. These pathogenic scotochromogens were shown to be identical as Mycobacterium scrofulaceum (Tsukamura et al.: Kekkaku, 42: 219, 1967). On the other hand, slowly growing scotochromogens occurring in the soil showed similar characters with the pathogenic scotochromogens and were considered to belong to the same species (Tsukamura et al. : Kekkaku, 42: 15, 1967). These results suggest a hypothesis that the scotochromogens occurring in the soil or water (M. scrofulaceum) enter the human body and, if there exist some strains with high virulence among them, they cause an infection in the human, being influenced by the condition of the host. If this hypothesis is correct, it would be expected that sporadic isolates of the scotochromogens occurring in sputa are M. scrofulaceum and their virulence is weaker than that of the pathogenic scotochromogens (multiple isolates). The present paper dealt with the characters and virulence of the sporadic isolatescotochromogens.

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There are some different opinions on the problem concerning the reduction of activity of antituberculous drugs added into the egg media. According to the “Handbook of Tuberculosis Laboratory Method” issued by Veterans Administration, the amount of streptomycin (SM) and viomycin (VM) customarily increased three times and 25% respectively of the required concentration to compensate for the loss of drug activity on inspissation in egg-containing media. And the “guide to the examination of tubercle bacilli” in Japan describes the increased concentration of antituberculous drugs in the drug sensitivity test, such as, two-fold for SM, ten fold for kanamycin (KM) and 1.25 fold for VM. The report on the worldwide research of drug sensitivity test techniques performed by I. U. A. T. committee revealed that a considerable number of laboratories in the world have taken this matter into consideration. On the other hand, a part of investigators deny this phenomenon.
In this paper, we attempt to c larify the reasonable concentrations of drugs which are added. into egg media for drug sensitivity test of tubercle bacilli.
Experiment I: Minimal inhibitory concentrations o f antituberculous drugs in both the serum liquid medium and the egg solid medium.
SM, VM, KM, CPM, CS, PAS, TB₁, INH, TH and EB were tested under the serious control repeatedly, and the results are shown in Fig.1. In this figure, the length of each bar means the growth grade in each medium and strain (H₃₇Rv-standard strain, Yamamoto-fresh isolated strain, BCG). Of these drugs, SM, VM, KM, CPM and TH show remarkable differences between the drug-activities in the serum liquid medium and the egg solid medium.
Experiment II: Activity of drugs in water extracts of drug-containing media.
The activity of drugs in the supernatants of centrifuged homoginates o f drug-containing media was measured by the vertical diffusion method. As shown in Fig.2, a∼j, each drug has the characteristic lines according to the media used. The similar results as in Experiment I were obtained in this experiment, and the grade of reduction of drugs in the egg medium is different depending on the concentration of drugs. The equations which show the correlation between the drug-concentration added into egg media and the activity expressed on media were led from these lines. These equations are considered to be very useful for a practical purpose in the drug sensitivity test.

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In a previous experiment niacin test on the cultures of tubercle bacilli grown on Ogawa egg slant with an airtight rubber stopper revealed that approximately 28% of the test strains were negative. Such a high incidence of the negative strains was considered to be due to an insufficient amount of the growth used. The objective of this paper is, firstly, to confirm the above assumption using a large number of strains and, secondly, to answer the question whether or not the resistance to the primary antituberculous drugs brings about the negative reaction in the bacilli.
472 strains isolated from treated or non-treated patients were submitted to the routine drugsusceptibility test. Each of the control cultures was transferred by loop onto four Ogawa egg slants which were then divided into equal two; the first two tubes were sealed with an airtight rubber stopper while the other with a cork stopper. After 4 weeks of incubation at 37°C, each one tube exhibiting more abundant growth was directly subjected to the niacin test according to BrCN-benzidine method. The results were summarized in Table 1. It is seen that the niacin positive rate in the series with cork stopper was 98.7% while the rate of the series with airtight rubber stopper was 89.4%. Moreover, the difference in degree of the positive reaction was observed between the two series. Namely, most of the cultures with cork stopper, 93.6%, revealed strongly or moderately positive reaction, whereas in the series with rubber stopper about 80% of the cultures were found to be only weakly or doubtfully positive. Tables 1 and 2 show that the amont of growth as well as the number of colonies developed was superior in the series of cork stopper. It will be suggested that in order to perform direct niacin test the culture tube must be sealed with such a stopper or cap allowing free-entry of air.

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The authors titrated precipitins and complement fixing activities to tuberculoproteins in sera from patients with pulmonary tuberculosis. Cells of human tubercle bacilli (Frankfurt stra in)were mechanically disintegrated by the equipment designed by one of the authors. A crude protein fraction extracted from them with minimum denaturation was used as the antigen. Precipitin titration was carried out by the agglutination reaction of antigen-coated collodion particles elaborated by the authors.
Complement fixation performed against the native tuberculoproteins was of high diseasespecificity. No healthy positive reactor to tuberculin had 1/20 or higher titers, while vast majority of 335 tuberculous patients revealed higher titers than 1/20 reaching 1/640 in serious cases. The serological reaction, therefore, was useful in making differential diagnosis of abnormal chest shadows, in speculating prognosis and confirming recovery of pulmonary tuberculosis.
Precipitins to tuberculoproteins were comparatively scant (1/5-1/20) in hopeless patients suffering from severe disease and in recovered patients as well as in healthy positive reactors. Precipitins were plentiful (1/40-4/160) only in improving patients whose chest shadows were diminishing steady.
It was o f interest to notice that excessive production of the precipitins to the mostly noxious bacillary element (tuberculoproteins) accompanied a favourable clinical condition, whereas low blood concentration of the antibodies occurred in patients running hopeless course. The authors postulated basing on their studies that the precipitins (antiproteins) would be reduced in the circulation of severe patients not only by the paralysis of production but also by increased union with corresponding antigen from active lesion.

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In order to clarity the results of pneumonectomy in Japan and to find out how to improve the results, the co-operative follow-up studies were carried out on the 550 cases who received pneumonectomy during 3 year period from 1961 to 1963, and were followed-up at least 8 months post-operatively in 50 sanatoria and hospitals belonging to the Tuberculosis Research Committee (Ryoken).
The rate of success (the rate of the persons who were working with negative sputum after operation among all cases) was 75%, and the post-operative mortality rate excluding the cases who died from non-tuberculous disease later than two months after operation was 7%. The factors reducing the rate of success were low pulmonary function and preoperative posit ive sputum. The result was the worst when both these factors coexisted together in a person. Therefore, in order to improve the result of pneumonectomy, efforts must be made to convert sputum negative before operation.
Moreover, as the po s t -operative deaths were most frequently caused by bleeding or pulmonary dysfunction, it is desired to keep patients under good management during and after operation.

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It is of ten seen that antituberculous agents are ineffective for cases of chronic lung tuberculosis with caseous lesions encapsulated by connective tissue. Possibility of softening and resolution of such lesions through destruction of fibrous capsule by artificial means combined with antituberculous drugs was first discussed by Hart.
In order to attain such changes in the lesion, v arious kinds of stimulation therapy have been tried. The purpose of this study is to evaluate the stimulation therapy with steroid hormone.
Thirty two patients were divided into three groups, each of which received prednisolone and/or ACTH in addition to isoniazid and PAS as described in Fig.1.
For cases showing such reactions as perifocal exudation or cavity formation during the stimulation therapy, SM or secondary antituberculous drugs (KM, 1314 Th or CS) were immediately added.
Changes in chest roentgenogram, blood chemistry, pituitary-adrenocortical function and other side effects were examined during the therapy.
On six cases of group C (see Fig.1), fur t h ermore, changes of 17-ketosteroid fractions were analysed by gaschromatography.
The results were summarized as follows:
1) No marked changes in chest roentgeno g ram was noted in group A.
2) Slight changes in chest roentgenogram was observed in group B.
3) In group C, nine out of eleven cases showed improvement in r oentgenogram, and in four cases, marked clearing of lesions was noted. It could be said that the large dose steroid and ACTH-Z therapy for 9 months is the best steroid-hormone stimulation therapy among these three methods.
4) Moon face, acne vulgaris, striae cutis and disturbance of menstruation were often observed, but they disappeared soon after the cessation of the stimulation therapy. Severe complications such as grave infection, glycosuria, hypertension and peptic ulcer were not found.
5) Cases in all the three groups suffered from pituitary-adrenocortical hypof unction due to the therapy, although this side effect was relieved soon after the withdrawal of hormone administration.
6) According to gaschromatographic analysis of urinary 17-ketosteroids, dehydroepiandrosterone fraction was significantly decreased in cases of group C compared with that of normal adults. In group C, decrease of androsterone fraction and increase of etiocholanolone fraction were also observed after ACTH loading.
Interesting finding was that e tiocholanolone fraction increased at first, and decreased later by SU-4885 administration in some cases of group C.

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