We have already shown that the hemagglutination test (TPHA) using the antigen from a pathogenic treponema strain was useful for the serodiagnoss of syphilis. However, the test was positive with a few non-syphilitic sera, especially with the specimens from pregnant women. Experiments were, therefore, directed to eliminate such non-specific positive reaction. Attempts were also made to improve the FTA test. The results obtained are as follows: 1) Most of the non-specific reaction could be excluded when the serum specimens were pretreated with Reiter TP antigen and rabbit testes powder. 2) The new method was compared with that of FTA-200 (or the FTA-ABS), the RPCF test or the STS on 230 syphilitic and 319 non-syphilitic sera. The results obtained by the TPHA agreed very well with those by the FTA-ABS. Most of the “biological false positive reaction” by cardiolipin antigen was easily excluded by the TPHA test. The method is very simple and the antigen can be supplied as a stable lyophilized product. 3) Non-specific positive reactions of the FTA could be eliminated by various methods, but the FTAABS described by Deacon et al. was most effective
The bacteriolytic agent for Staphylococcus epidermidis and some others was obtained from the culture filtrate of a newly isolated soil bacillus.The bacillus was similar to Bacillus subtilis morphologically. There have been many studies on bacteriostatic or bacteriolytic agent produced by a B. subtilis since subtilin. subtilin C, mycosutilin, subtenolin, subtilydin and endosubtilydin were reported by E. F. Jansen in 1944. The authors have also studied the properties of these organisms, the condition of lytic agent production and methods of purification. The results were as follows. 1) The isolated organism, which was gram positive bacillus, grew well and formed a typical R. type colony on ordinary nutrient agar plate. The bacillus formed spore rapidly, fermented glucose, produced acid and gas and liquefied gelatin. 2) Lytic agent affected Staphylococcus, B. cereus, B. anthracis and B. subtilis. 3) The synthetic medium containing 1.0% glucose, 0.6% sodium-glutamate, 0.14% Na2HPO4, 0.1% KH2PO4, 0.2% NaCl, 0.01% MgSO4, adjusted to pH 6.0-7.2 was found to be the best for production of lytic agent. Incubation was usually continued for 6 days at 37°C. 4) Physico-chemical properties of the lytic agent were relatively heat-stable, nondialysable through cellophane membrane and easily adsorbed on a strong acidic or basic ion exchanger column (IR 112, IR 120 or IR 400) but not adsorbed on active carbon. A purified fraction was obtained through Sephadex G-50 column. 5) This agent acted on cell wall of several microorganisms at pH 5.0-6.0 and formed protoplast.
Biological characteristics, growth inhibition and double gel diffusion tests were studied on 115 strains of mycoplasmas isolated from 73 rats suffering from the chronic respiratory disease from five different colonies. Of the 115 strains, 31 were isolated from lung, 39 from bronchus, 41 from nose and 4 from mouth. One hundred and fourteen of the 115 strains were identified as Mycoplasma pulmonis, and the rest strain resembled to M. ahthritidis biologically, but did not show any characteristic correspondence to the species in growth inhibition test. They made large colonies (1 mm to 2 mm in diameter) on modified Edward's medium (AM-1) as reported previously. Central zones of standard strains, PG-22 and T, from mice were less remarkable, but that of rat strains were, generally, slightly remarkable. Surface of the colonies of rat strains were more granular and darker than the standard strains. They required serum for growth, did not grow at 22°C, produced “film and spots” on horse serum agar medium (AM-1), and were beta hemolytic with sheep and guinea pig erythrocytes and alpha hemolytic with horse erythrocytes. They grew well on agar medium aerobically and anaerobically. In semi-solid agar medium, they grew preferably near the surface. They reduced tetrazolium blue and methlene blue, and did not produce hydrogen sulfide and indole. The fermented glucose, mannose, maltose, glycogen and dextrin, but not galactose, lactose, sucrose, salicin and dulucitol. Variations of hemolysis and fructose were discussed. Production of “film and spots” was irregular according to cultural conditions. In growth inhibition test, some antisera did not inhibit or inhibited weakly the growth of heterologous strains, and one antiserum for strain R-1 inhibited the growth of homologous strains weakly and some heterologous strains strongly. Fifty-two out of 110 strains were inhibited by all of 6 anti-M. pulmonissera, but the remaining 58 strains were not inhibited by some of the antisera. Strain T was highly antigenic, that is, this strain was inhibited by all the 6 antisera, and antiserum of this strain inhibited all 110 strains. This may be one of the reasons why the strain PG-22 had not been identified as M pulmonisuntil quite recent days. This also indicates that adequate selection of standard strains in serological studies of Mycoplama must be done practically. By double gel diffusion test, using, the antisera that were absorbed with the liquid medium to remove the antibodies against the ingredients of this medium, especially horse serum, M. pulmonis was shown to have one to three common antigens. It was shown that there were some antigenic differences within M. pulmonis by growth inhibition and double gel diffusion tests.
In June of 1966, a new bacterial disease was found causing brownish streaks on the foliage of orchardgrass (Dactylis glomerutaL.) in Japan. The causal organism was identified as Xanthomonas translucens f. sp. hordei Hagborg, considering its bacteriological characters, symptom on the leaf-blade showing the characteristic translucency and pathogenic specialization. It is the first time that X. translucens has been found to attack orchardgrass naturally and some of wheatgrasses (Agropyronspp.) artificially. The bacteriological characters of the pathogen are as follows: Cylindrical rods rounded at ends, Solitary or in pairs; 0.6×1.2-2.4μ in size, motile with a single flagellum; capsulate, Gram negative, no spores, aerobic. Superficial colonies, yellow, round, convex, smooth, shining, opaque, with entire margin. Liquefies gelatin slowly; digests milk slowly without coagulation; nitrates not reduced; hydrogen sulphide and ammonia produced, but not indol or acetoin; starch hydrolysed variable. Acid but no gass produced from arabinose, xylose, glucose, fructose, galactose, mannose, lactose, sucrose and glycerol: neither acid nor gas from rhamnose, maltose, raffinose, starch, inulin, dextrin, mannitol, sorbitol or salicin. Aesculin hydrolysed; oxidase-positive; lipolytic. According to Hugh-Leifson's method, acid produced aerobically from glucose. Optimum temperature 30°C.
The results of our experiments presented here support the conclusion that an early and specific effect of sodium fatty acid is a change of permeability resulting in loss of Saccharomyces cerevisiae cell content. Several sodium fatty acids were examined during the course of an investigation on induced weight loss in Saccharomyces cerevisiae. The weight loss was caused by sodium caprylate, caprate, undecylenate and laurate at concentration of 6×10-2 M and the pattern of weight change with time induced by them were very similar. Fig. 1 indicates that the effect of the sodium fatty acid (undecylenate in this experiment) was relatively independent on the age of the mycelium usedas “inoculum” since the fatty acid caused essentially the the same weight loss after five hours in all cases. The nature of the weight loss caused by sodium undecylenate was examined more detail. The decrease in weight clearly began at concentration of 0.2×10-2 M of sodium undecylenate. Potassium, sodium and strontium ion did not infuluence entierly on sodium undecylenate action. Calcium, Magnesium and Barium ion repressed and on the contrary ammonium ion accelerated the weight loss with sodium undecylenate. The electronmicroscopic observation of whole cell or thin section showed that these agents intensively caused plasmolysis of the cell. Sodium caprylate, caprate, undecylenante and laurate showed strong inhibition on the growth ofSaccharomyces cerevisiaeat 3.7×10-3 M, 9.4×10-4 M, 1.8×10-4 M and 3.7×10-4 M but sodium acetate, valerianate and caproate did not show inhibition even at 6.0×10-2 M.
An extracellular polysaccharide isolated from the supernatant of broth culture of a Crepidotus B-145 was soluble in water, giving a highly viscous solution. Estimation of molecular weight with light scattering gave the value of 1.52×166, while the calculation from determination of reducing endogroup gave the value of 1.6-3.6×104, suggesting the formation of molecular aggregates in solution. Analyses of its component sugar with thin layer chromatography and gas-liquid chromatograhy revealed that the polysaccharide was composed largely from glucose. By gas chromatographic analysis of its permethylation-methanolysis products, it was found that the polysaccharide has 1-3 glucosidic linkage in its structure. Infra red spectroscopy indicated that the linkage is of β-configuration. The presence of β 1-3 linkage was further confirmed by the cleavage with β 1-3 glucanase. Interestingly, the treatment with the polysaccharide was found to suppress the growth of Sarcoma 37 in mice as expected from the antitumor activity of yeast glucan, the active structure of which was said to be β 1-3 linkage.