Acid-fast bacilli were detected in 13 (27%) of 49 skin samples in surgical operation under the procedures of collection of bacilli by centrifuging the filtrate of tissue homogenate through adsorbent cotton. Ten specimens (20%) contained cultivable organisms, including M. simiae (9 specimens) and M. gordonae (one specimen). The other 3 specimens did not contain any cultivable organism, although microscopic observation revealed the presence of acid-fast bacilli.
Eight (17%) of 48 raw umbilical codes of babies received Cesarian operation were positive for acid-fast bacilli in the smear preparation. Six (13%) were positive in cultivation and the organisms were identified as M. simiae (4 cases), M. scrofulaceum (1 case) and M. aviumcomplex (1 case). The remaining two specimens were negative in cultivable bacteria in spite of obvious presence of acid-fast bacilli. In the case of frozen umbilical codes, 9 specimens (16%) were positive in acid-fast bacilli, only 3 cases of which were positive in cultivable organisms, including M. gordonae (2 cases) and M. scrofulaceum (one case). M. simiae was not detected in cultivation of frozen materials.
The purpose of this experiment was to isolate the microscopically detectable but uncultivable acid-fast bacilli, using experimental infection system induced in nude mouse. However, two experiments separately performed failed to achieve this purpose, because of contamination of the cultivable acid-fast bacilli among mice or death of the organisms during storage.

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When leprosy bacilli grown in nude mouse foot pad were used for culture experiments, cultivable acid-fast bacillus was sometimes isolated as a contaminant. Whenever bacilli were inoculated to nude mice, the same leprosy bacilli were killed by autoclaving and were inoculated in to foot pads of 5 nude mice for examination of this cause of the contamination. Acid-fast bacillus was cultivated on 3% Ogawa egg medium at 33°C from homogenates of foot pads of mude mice infected with M. leprae after one year and a while of infection. Foot pad of nude mouse injected with leprosy bacilli was cut off, ground in mortar and passed through sterile absorbent cotton and the filtrate was centrifuged at 10, 000 rpm for 30 minutes. The sediment was inoculated on 3% Ogawa egg medium after treating with a small amount of sterile 1 N sodium hydroxide.
Acid-fast bacilli were isolated from 3 out of 41 mice inoculoted with heat killed bacilli. The isolated acid-fast bacillus did not be observed in the same experimental group inocudated with live bacilli, positive cases were scattered in another groups. Four out of 16 tubes were positive for acid-fast bacilli in mice infected with Kurume-naha and 5 out of 7 tubes in the Amami-KM infected mouse group. The two negative tubes were discarded due to contamination. Kurume-Oki strain which has yellow colonial morphology was isolated from one out of 6 culture tubes. Strains Kurume-naha and Amami-KM have the same characteristics as follows: slow grower with pale yellow smooth colonial morphology, strongly positive for niacin production and urease; positive for nicotinamidase, pyradinamidase and 68°C catalase; no growth at 45°C, negative for nitrate reduction, hydrolysis of Tween 80, diamine oxidase, heat stable acid-phosphatase and arylsulphatase; resistant to streptomycin, isoniazid, rif ampicin and B 663. Two isolates were identified as Mycobacterium simiae from these characteristics. Characteristics of a Kurume-Oki isolate was as follows: slow grower with yellow smooth colonial morphology, positive for urease, 68°C catalase, hydrolysis of Tween 80 and arylsulfatase; no growth at 45°C, negative for niacin production, nicotinamidase, pyradinamidase, nitrate reduction, daimine oxidase and heat stable acid-phosphatase; resistant to streptomycin, isoniazid, rifampicin and B. 663. This bacillus was identified as Mycobacterium gordonae from these characteristics.
The isolated bacilli from nude muose inoculoted with Kurume-naha strain were infected to mouse but no pathogenic behavior was seen one year after infection, but a nodule was produced at the site of infection of nude mice foot pads. On the other hand, Kurume-oki was nonpathogenic for mice.
The isolation of M. simiae and M. gordonae from healthy human skin has been reported. These organisms may be infected latently in healthy humans. Since nude mice were sometime contaminated with such mycobacteria, we have to keep in mind this fact when carrying out the cultivation of M. leprae from nude mouse leproma and moreover take care to use for inoculation material. More than 50 nude mice should not be inoculated at the same time with M. leprae of the same source.

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Histopathological findings of intrafascicular-edema found in n. ulnaris running at forearm, palmside of hand and fingers of rhesus monkey 8664 and african green monkey 8175 were studied by semithin section method. These two monkeys were inoculated with leprosy bacilli from lepromata of mangabey monkey A022. The mangabey monkey A022 had been experimental leprosy after inoculum of the leprosy bacilli from lepromata of the mangabey monkey A015-natural infection.
Period between date of inoculum and sacrifice of the rhesus monkey 8664 was about 2 years, and the african green monkey 8175 was about 5 years.

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For the purpose of understanding subclinical infection with Mycobacterium leprae among the inhabitants in an endemic area, 3, 547 schoolchildren and 1, 487 adults in several regions of Okinawa were surveyed by using clinical examinations and immunological tests, i. e. the fluorescent leprosy antibody absorption (FLA-ABS) test and the lepromin test using Dharmendra's antigen, during a period from 1978 to 1984. The enlargement of peripheral nerves, especially that of lateral or bilateral auricular and/or ulnar nerves, without loss of sensation was found in 8.4% of the schoolchildren and in 9% of the adults. A frequency of these signs and symptoms was significantly higher in male than in female and tended to increase with age. The percentage of positive reactions in FLA-ABS tests for schoolchildren and adults was 21.8 and 22.5, respectively, the values suggesting a minimum frequency of subclinical infection with M. leprae among these inhabitants. The percentage of positive FLA-ABS tests was significantly higher in those with neural signs and symptoms than in those without. Such a correlation was not found in the lepromin test. Ill-defined depigmentation of the skin without loss of sensation was found in a few percentage of both schoolchildren and adults. However, this sings and symptoms did not correlate with the FLA-ABS test nor with the lepromin test. A history of tuberculin test and BCG vaccination in the schoolchildren seemed to influence the lepromin-reactivity. Among 770 schoolchildren tested with both FLA-ABS and lepromin, 70 (9.1%) were the FLA-ABS positive but lepromin non-positive responders. Neural signs and symptoms were found in 31 of these children

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