An attempt to prepare lipopolysacchride from a halophilic bacterium No.101 was made by extracting the bacterial cells with hot phenol-water using the Westphal's method. Further purification was carried out by freezing the phenol extract at -20°C for 48 hours with successful removal of the nucleic acid moiety and some other inert contaminants. The purified preparation thus obtained showed high pyrogenic activity for rabbits (0.01μg per kg) and potent precipitation activity, its titer being 10μg/ml as measured on capillary precipitation test. On Ouchterlony's agar diffusion gave no more than one sharp line of precipitation against homologous bacterial cells antiserum, indicating an evidence for immunological homogeneity. Chemical analysis of the preparation was follows: N. 2.8%, P. 3.2%, amino-sugar 5.0%, reducing-sugar 16.0%, chloroform-methanol extractable material 57.3%.
The ecological survey and comparative investigations of the pathogenicity of V. parahaemolyticus derived from many sources were made. Results obtained are as follows: 1) The detection rate of V. parahaemolyticus subgroup 1 was calculated as 6% from the sea-water, sea-mud and plankton samples tested and as 34% from the marine products specimen in fish's shops. 2) Most of the marine source strains showed positive results by the De-test, but a few showed questionable consequences, while all the strain isolated from patients always showed positive results. 3) It is considered that the marine source organisms classified in subgroup 1 of V. Parahaemolyticus are also essentially pathogenic. 4) The strain classified as V. anguillarum, which resembling to those of V. parahaemolyticus subgroup 1, always showed negative results by the De-test and the pathogenicity of these organisms against mice were considerably weak.
An ecological investigation of alimentary flora of man was made by eight selective media and six specimens recovered from stomack, duodenum, gall bladder, gall ducts, jejumun, transverse colon of 112 patients who were undergoing laparotomy for either stomack cancer, duodenal and stomack ulcer, or gall ducts disease of them. Similarities and dissimilarities of obtained results proposed existence of four different ecological sites, stomack, upper intestine, transverse colon and gall ducts in the alimentary track. Such flora speciality of a different site, however, was much influenced by pathological changes thereon. Influences from another pathological site upon flora of a healthy site were variable. Existence of the glycolytic layer, which was found by Onisi et al in the human mounth was also proved among alimentary flora with some exceptions; Enterobacteria associated with this layer, population of Candidae did not run parallel with Lactobacilli, and fusobacterium count slightly related to that of Staphylococci, a member of the layer.
Susceptibility to sodium salicylate (0, 0.5 and 1.0mg/ml) on Sauton agar medium (3% agar) has been suggested to be useful for a rough differentiation of rapidly, growing mycobacteria, when used in combination with the test of temperature response. Growth of nonphotochromogenic mycobacteria on medium containing 1mg/ml salicylate seemed to suggest that the test organism belongs to one of M. fortuitum, M. parafortuitum (n. sp.), M. smegmatis, or M. phlei. M. fortuitum blackened markedly the medium containing salicylate.
Even if serodiagnosis of syphilis with FTA test, using fluorescent antibody to human γ-globulin has the advantages of being convenient, specific and highly sensitive, still it remains the problem of so-called biological false positive reaction in a debate. In order to investigate in detail the specificity of this test we tried first to study the distribution of antibodies, especially antibodies of 7S γ type in syphilitic sera in each stage. Two kinds of the antiserum were used as anti-human γ-globulin. In order to get purified 7S γ-globulin 800mg of the same globulin was separated by chromatography on DEAE-cellulose as follows: 200ml of normal human serum was salted out with 33% saturated (NH4)2SO4 three times, whereafter it was transfered to a column (4.0×25.0cm) of DEAE-cellulose equilibrated with 0.01M phosphate buffer, pH7.5, containing 0.015M NaCl and eluted with the same buffer. Elution with this buffer gave a pure γ2-globulin peak as shown by cellulose acetate electrophoresis, Ouchterlony agar diffusion method and immunoelectrophoresis using LKB's immunophor. Then, anti-7S γ-globulin was prepared in rabbits following a course of immunization which included intramuscular injections of alum precipitated antigen (purified 7S γ-globulin preparation) and intravenous injections of 1% aqueous solution of the antigen and then γ-globulin fraction of thus raised antiserum was conjugated with fluorescein isothiocyanate (FITC). Its F/P ratio was estimated as 1.32×10-3. Immunoelectrophoresis with normal human serum showed a strong 7S γ line only. We used FTA tests with this FITC-labeled anti-7S γ-globulin and with FITC-labeled anti-γ-globulin, in which the latter γ-globulin was prepared by routine ammonium sulfate precipitation and thus it contained both globulins of 7S γ and 19S γ types. Comparative studies were undertaken to investigate the transition of antibodies of 7S γ and 19S γ types in the syphilitic sera by both methods. From our results it was found that 7S γ class antibodies appeared always in the sera of primary stage (2 cases), secondary stage (5 cases), tertiary stage (6 cases) and congenital syphilis (4 cases), but in contrast to this 19S γ class antibody was found only in the sera of primary syphilis. These findings seem likely to show the evidence that in the very early stages of syphilis macroglobulin (γ1M-globulin) antibodies appeared, but later predominantly 7S γ-globulin (γ2-globulin) antibodies. One hundred thirty seven sera which were sent to our laboratory for FTA test, were tested also by RPCF, W.R. and VDRL tests, comparing with the results of this test. It was found that discrepancies of 20%, were encountered between FTA test and several other tests for syphilis (RPCF, W.R. and VDRL tests). The present results show that FTA test is found to be most specific compared with the other three tests for syphilis, and also show the most promising results also in the cases of biological false positive reaction.
The most effective immunity against mouse typhoid is known to be attained by immunization with “live” vaccine but not with “killed” vaccine. To clarify the relation between the effectiveness of the immunization and the rate of multiplication or persistence of immunizing microorganisms in the host, the present study was conducted using a strain of streptomycin (SM)-dependent Salmonella enteritidis, lld, as a “live” vaccine. About 8×104 organisms of lld were injected intsaperitoneally into a numper of mice, and the mice were divided into 3 groups according to the schedule of administration of streptomycin. To the mice of the first group, 8mg of SM was daily injected intraperitoneally for 50 consecutive days starting from the day of infection. To the mice of the second group, the same daily dosis of SM was administered for 12 days. The mice of the third group were not administered with SM at all, but injection with lid was repeated 2 or 3 times during the period for immunization. A number of mice of each group were sacrificed at intervals and the number of viable lld organisms in the mice was calculated. Thus, the homogenates of the lymph nodes, spleen, liver and remaining carcuss, except the digestive tract, were cultivated quantitatively on agar plates containing SM. To evaluate the effectiveness of the immunization, 51 days after the infection with lld a number of mice of each group were challenged intraperitoneally with about 8×103 organisms of virulent S. enteritidis and their mortality was observed for 21 days. The results were as follows. 1. In the mice of the first group, lld, the immunizing organisms, increased for 2 weeks and then dicreased gradually. The mice acquired a significant resistance to the challenge with virulent S. enteritidis. 2. In the mice of the second group, lld organisms as well increased for 2 weeks as in teose of the first group, but decreased rapidly thereafter, and they could not be recovered from any parts of body on the 40th day of the immunization or thereafter. The aquired resistance against virulent infection of the mice of this group was inferior to that of the first group, and the mice challenged on the 18th day of the immunization were well protected from the virulent infection but those challenged on the 51 the day were protected at lower grade than those of the first group. 3. In the mice of the third group, no increase but rapid decrease of lld organisms was observed after every injections with lld. The mice were not protected from the virulent challenge at all.