From the epidemiological investigation of faeces of human being, multiple resistant Escherichia coli and Citrobacter were isolated. The carrier of Escherichia coli R4 (CM. TC. SM. SA) and R3 (CM. SM. SA) was about 1.4 percent among 1145 healthy human being and was more in patients treated with CM, afflicted with tuberculosis and dysentery among several kinds of in-patients. Detailed study of the type of multiple resistance of Escherichia coli isolated from 109 tuberculosis patients showed that many kinds of multiple resistance existed and 16 had more than two types of multiple resistance. The multiple resistance of Shigella and Escherichia coli was eliminated after 6 month's storage on cooked meat media. The contamination rate of multiply resistant Escherichia coli in faeces was described.
Staphylocoagulase has been tacitly assumed to play an important role in the virulence of Staphylococci and also it has been considered by most that M-protein might have a significant role in the Streptococcal infection. However, few direct demonstration of its role in the pathogenesis of infection with virulent, coagulase positive, and avirulent, coagulase negative staphylococci has been made. The results presented here show that when virulent, coagulase positive and avirulent, coagulase negative Staphylococci are suspended in solutions of purified coagulase prepared by Blobel's method or in solutions of purified M-protein of Group A Streptococci (Type 6, 5-43-M) prepared by Slade & Kimura's method and injected intracerebrally into mice, the slight or marked increase in the virulence of the organisms results. As the test strains of Staphylococcus, Staphy. aureus (A-II), coagulase positive virulent strain, and Staphy. epidermidis (B-2, B-7, B-12), coagulase negative avirulent strain were used. These strains tested were all isolated in the authors laboratory. The results show that virulent Staphylococcus was slightly increased by the addition of coagulase or M-protein and one out of three avirulent Staphylococci remarkably increased the virulence by the addition of coagulase and one out of two avirulent strain remarkably increased the virulence by the addition of M-protein.
The purpose of the present investigation was to clarify the nature of normal antibody responsible for the bactericidal activity of normal serum mainly against Gram-negative bacilli, as comparing with properdin, a new bactericidal entity in serum, reported by Pillemer et al. Properdin-zymosan or normal antibody-organism complexes were respectively prepared by the absorption of normal serum with zymosan, Sh. dysenteriae 1 and Sal. typhosa 0-901 W at the defined temperatures. These complexes were placed in the reservoirs of agar gel in a barbital buffer, pH 8.2, ionic strength 0.05, and then the immunoelectrophoresis was performed. The obtained results were as follows. 1. Both of the normal antibody and properdin of human serum were demonstrated as a single precipitin arcs at β1-globulin position; those of rabbit serum at α2, p1-globulin position. It was, furthermore, shown that normal antibody is substantially different from properdin. 2. By means of the mutual absorption of rabbit serum with Sal. typhosa and Sh. dysenteriae, the precipitin arcs of the normal antibodies against these bacteria were individually observed at the same position of α2, β1-globulin. Accordingly, it may be taken to indicate that normal antibodies against enteric bacteria posses a marked specifity, in contrast with properdin which is non specific. 3. The precipitin arcs of the immune antibodies against zymosan and Sh. dysenteriae, on the other hand, were clearly observed at the γ-globulin position. The above-mentioned findings suggest that normal antibody and properdin, in substance, differ from so-called immune antibody.
In a previous study, a total of 72 strains of non-pathogenic acid-fast bacilli isolated from bovine, porcine, human sputum, human smegma, human leprous lesion, leprous rat, guinea-pig and saprophytes were investigated for their morphology in an attempt to classify these strains. The present study was undertaken to investigate the biological characteristics of the 72 strains with special reference to their cultural characteristics, sensitivity to dyes, survival at 60°C, growth in various temperatures, decolorization by boiling water, neutral red test, production of urease and reduction of potassium tellurate. It was found that the biological characteristics which might furnish significant keys in the differentia tion of these acid-fast bacilli were survival at 60°C for one hour, growth at 47°C, growth in sorbit NFI4Cl media, and the production of urease. Furthermore, it seemedthat the growth feature on the methyl violet media was partially significant for identifying the strains. Seven of 72 strains showed positive reaction in neutral red test. All of such strains were isolated from bovine or porcine organs but non of saprophytes showedpositive results. Fifteen strains which did not produce urease resembled each other in colony morphology, growth feature on methyl violet media, lack of ability to grow in sorbit NFI4Cl media and weakness in heat tolerance. All of such strains were isolated from bovine, porcine, or human organs, while all strains of saprophytes produced urease.
Guinea pigs were conferred immunity by the inoculation of Bacterium tularense vaccines. The degree of protection thus conferred on the animals was higher in the case of the vaccines made from living organisms than those made from killed culture. The immunized animals survived the challenge with the organisms and the pathological changes caused by the challenge were found slight in every aspect as compared with those of control animals, When the immunized animals were challenged with highly virulent strain, the agglutinin titers having been produced in the animals declined once, but they tended to increase again several days later, showing the evidence that the animals which recovered the rise in the agglutinin titers could survive and those which failed to do so were brought to death. It seems that the agglutinin has some relation to the degree of animal protection but had no protective properties against Bacterium tularense.
Malignant tumor cells (HeLa strain, L strain, chicken sarcoma and human uterine cancer cells) were dispersed with trypsin and inoculated onto the chorioallantoic membrane (CAM) of embryonated chicken eggs. HeLa and L cells formed tumors on CAM. Some of factors affecting the tumor formation were age of eggs used and number of cells inoculated. In the present experiments, in which 9 to 13-day old eggs were used and each egg received 5-6×105 or more cells, the rate of tumor formation was practically 100%. The tumors became visible 3 or 4 days after the inoculation of cells and increased in size during the additional 3 to 5 days. The maximum size was about 15×15×5mm. The in vitro restoration or egg passage of the original cell strains was possible. While the tumors produced by either kind of cells were of the similar macroscopic appearance, their histological architectures were different. HeLa cells in the tumors were arranged in aciniform grouping, and the acini gathered together to form larger conglomerates. The L cell tumors were characterized by diffuse infiltration of round or a littleelongated cells without showing specific arrangement; this resembled the pattern of tumors produced by the chicken sarcoma cells which were more infiltrative and typically'sarcomatous.' Trypsinized cells from human uterine cancer tissues formed tumors when inoculated onto CAM. It was shown histologically that the tumors included acinous clusters of cells with undifferentiated characters; the type of cellulargrouping had a resemblance to the specific arrangement of HeLa cells, on one hand, and to that of the cells found in the original cancer tissues, on the other. Some concepts regarding growth of cancer cells are discussed, and a potentiality ofthe technique for studying cancer cells is suggested.
A yellow pigment producing organism of Gram-negative, flagellated bacillus was isolated from the blood of a feverous patient. The serum of the patient taken on the fifth disease day showed comparatively high titer (1: 640) in the agglutination test with this organism. On isolation, this organism had possessed Vi-antigen, since the living and heated bacilli or this strain had been agglutinated by anti-Vi sera to full titer, but not by anti-S. typhi 0901 serum. However, on successive subculture on agar, the Vi-antigen was lost in 2 months after isolation and the organism showed reaction with anti-S. typhi 0901 serum. Contray to the first apperance that the organism seemed to be a strain of S. typhi, it was identified as Bact. typhiflavum by its culturai and biochemical properties (Cf. Cruickshank: J. Hyg., 1935, 35, 354). It formed so-called “Symplasmata” discribed by Kathe (1933) on agar slant, especially in the part of condensed water. As to the serological relationship between Bact. typhi-flavum and Salmonella groups, no satisfactory description could be found in the literature. Therefore, an attempt was made to investigate it using the isolated organism and more than 30 different Salmonella serotypes. After the many cross-absorption tests, it was found that this strain of Bact. typhi-flavum possessed such 0-antigens as Salmonella 0122, 13, and 34. Besides, it contained some partial 0-antigens common to such Salmonella serotypes as S. newington, S. albany, S. hoi-sham, S. minnesota and S. tel-aviv, respectively. By slide agglutination tests, the organism showed no reaction with any anti-H standard serum of Salmonella. On the cultivation in bile containg broth, the organism grew easily a variant strain producing no yellow pigment. Serologically this strain was identical to the original strain, but different in the fact that it did not ferment maltose and formed no symplasmata. Although the pathogenicity of the organism for mice was so weak, that on M. L. D. was about 7mg of culture by intraperitoneal inoculation, the present study, together with the literature survey, suggests that Bact. typhi-flavum would cause mild typhoid fever in man depending on conditions.
Semisynthetic medium, containing vitamin-free casein acid hydolysate as nitrogen source, is generally superior, in growth-promoting ability, to pure synthetic medium, containing amino acids and inorganic salts approximately in the same composition as the former. According to the reports of Demain, it was assumed that a minute amount of fatty acid, which is present in hydrolysate, would exert growth-promoting action. In order to ascertain this assumption, casein acid hydrolysate was treated with ether, and the ether-soluble portion was examined for growth-promoting action. As the result, in ether-soluble portion of vitamin-free Casamino acids (Difco), it was found a factor which exerted accelerative effect on the growth of Streptococcus hemolyticus S8, though not Escherichia coli, Salmonella pullorum, Vibrio El Tor, Brucella melitensis, Staphylococcus aureus and Diplococcus pneumoniae. Paper chromatography demonstrated that about 0.01-0.001% stearic acid was contained in Casamino acids. In view of this, pure fatty acids were investigated for growthpromoting effect on Streptococcus hemolyticus S8, and palmitic, stearic and arachidic acid were found effective. Especially, stearic acid was most effective, promoting the bacterial growth even in concentrations below 1.0γ/ml. Oleic, linoleic and 1 inolenic acid did not show any accelerative effect, and in higher concentrations, all of them were conversely inhibitory. It was confirmed inthis way that vitaminfree Casamino acids contained a minute amount of saturated fatty acid which promoted the growth of Streptococcus hemolyticus S8.
From the first to fifth reports successively, the establishment of oral infection of paratyphoid B was confirmed experimentally using various methods. The following observations were obtained. The sign of sickness appeared in almost all mice. The discovering of bacilli harboring in mice organs continued until the 15th week. The blood picture changed. The serum agglutinin titer against paratyphoid B O and H antigen raised significantly. All strains of mice examined were infected though grade varied with strain. Histological examination showed several proofs of infection also. In this report, mice were treated with some methods before infection to change the mode of infection, and the following results were obtained. 1) When carbon tetrachloride was injected intraperitoneally before infection, more infecting bacilli were discovered in mice organs. 2) When tripanblue was combined with carbon tetrachloride pretreatment, many mice died of paratyphoid B bactremia. 3) After Bordetella pertussis intracerebral inoculation, paratyphoid B oral infection was established in almost all mice. 4) When Freund's adjuvant prepared by mice intestines cut out from duodenum to rectum and that contents had been washed out was injected intramuscularly for sensitization to infection site namely intestines, the rate of bacilli retention in organs rather reduced. 5) The quantity of dead tubercle bacilli contained in Freund's adjuvant, the frequency of adjuvant injection to mice as pretreatment, and the interval between abjuvant injection and paratyphoid B infection were changed respectively. The change of quantity of dead tubercle bacilli showed no influence upon experiment. The prolongation of interval between adjuvant injection and bacilli administration resulted in increase of bacilli harboring in organ culture.
Three different fractions, cell wall, insoluble granules and soluble part, were separated from the cultured cells of a human type tubercle bacillus H37Rv and the tuberculin allergy and hemagglutination were investigated in guinea pigs immunized with the fractions obtained, the results are as follows. 1. An induced.tuberculin allergy was obtained in the animal immunized with the cell-wall fraction and the antigen manifested was found to be considerably heat-stable. 2. No tuberculin allergy was induced by the other fractions. 3. Hemagglutination antibodies was produced in the animals immunized with each of the fractions.
We have elucidated in a previous study that, in mice actively immunized with killed vaccine or passively immunized with anti-O serum, intraperitoneally challenged virulent organisms of S. enteritidis were cleared from their abdominal cavities during an early stage of the infection, although these immunizing. procedures did not save mice from death after the infection. In the present study it was clarified that the clearance of infecting bacteria in the peritoneal cavity was also achieved by transfer of cells from killed vaccine-immunized mice. The tentative conclusion we have reached, is that the phenomenon is mediated by the participation of transferred antibody producing, cells, and the synthesis of O-antibodies in the recipients is an essential feature of this phenomenon. This conclusion was supported by the following facts. 1. Peritoneal exudate cells, especially rich in macrophages, obtained from killed vaccine-immunized. mice conferred the most potent “clearing capacity” on recipient mice. Cells from the spleen or lymph nodes of immunized mice also could transfer the clearing capacity, but those from the liver could not. 2. Intravenous, intraperitoneal and subcutaneous routes were all effective in transferring the capacity. 3. The capacity was transferred only by live cells, being inactivated by treatments of cells with. freezing-thawing, mild heating, and sonic oscillation. 4. The clearance was effective only when the antigen used for immunizing donors and challenging bacteria had common specific O-antigen. 5. Circulating antibody of more than 1: 20 agglutinin titer appeared in the serum of recipients 1 to. 4 days after transfer, when the cells collected from donors were secondarily stimulated in vitro by incubation with the heat killed Salmonella before transfer. 6. The clearing capacity transferred by cells decreased in its intensity within a week or so after transfer, when the transfer was made into homologous adult mice.
Even by the use of enough high titar of bacteriophage, the phage typing of staphylococcus for the examination of staphylococcus infection or food poisoning encounters several times the difficulty of typing due to inhibition or host range spreading etc. This difficulty induced us to try the use of adaptive phage in the examination of staphylococcus typing. Several examinations showed that the use of adaptive phage is very satisfactory for the typing of this bacteria, and some examples of the examinations were presented.