A bacterial strain isolated from feces of a patient with ulcerative colitis, which had been shown to produce a novel immunoglobulin A (IgA) protease (cleaving both the human IgA1 subclass and IgA2 subclass of A2m(1) allotype) extracellularly, was identified as Clostridium ramosum. By using a selective medium (proprionate-rifampicin-gentamicin-colimycin-polymyxin medium) devised for C. ramosum, analysis of the population level of this organism was performed to determine its ecology in the human intestinal tract. C. ramosum was isolated in 20 of 25 fecal samples (80%) from patients with inflammatory bowel disease (I.B.D.) and in 112 of 135 samples (83%) from patients without I.B.D. (control group). C. ramosum was also isolated from 6 of 11 biopsy samples (55%) of the inflamed rectal mucosa from patients with ulcerative colitis and from five of 15 samples (33%) from the intact mucosa of the control group. The population levels of C. ramosum in most of the biopsy samples ranged from 2.3 to 5.0 log₁₀ per gram. The IgA protease-positive C. ramosum was found in only four of 135 fecal samples (3%) and one of 15 biopsy samples (6.7%) from the control group. These results indicate that IgA protease-positive C. ramosum is not likely to play a role in the induction of I.B.D., unless the organism is first isolated from the patient with I.B.D.

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Two attenuated strains of Mycoplasma pneumoniae, P24-S1 and P24-S11, were tested for their ability as a live vaccine to confer on hamsters immune resistance against challenge infection with a virulent strain of M. pneumoniae, FH-P24. Fifty percent protection was obtained by vaccination with the P24-S1 strain administered once or twice. In contrast, only 10% of the animals were protected by the P24-S11 vaccine even when it was given three times. Vaccination with the P24-S1 strain resulted in higher humoral and cellular immune responses than the P24-S11 did. These results suggest that the P24-S1 strain has the primary qualities a vaccine which may be used for protection against human mycoplasmal infection.

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Numerical identification using the range of a species defined as (M±2s)% was shown to be suitable for the identification of slowly growing mycobacteria. In the formula, M is the mean matching coefficient for individual strains of each species to the hypothetical median organism pattern of their own species and s is the standard deviation.

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An R-form lipopolysaccharide (LPS) extracted from Klebsiella strain LEN-111 (O3-:K1-) by the phenol-chloroform-petroleum ether method was compared with that extracted by the phenol-water method in the ability to form a hexagonal assembly. The LPS which was extracted by the phenol-water method and dialyzed against tap water to remove phenol showed ribbon-like structures, and it formed a hexagonal lattice structure with a lattice constant of 14.5±0.3nm when it was precipitated by addition of two volumes of 10mM MgCl₂-ethanol. The LPS which was extracted by the phenol-chloroform-petroleum ether method and lyophilized consisted of ribbon-like structures and their fragments and it often formed small pieces of a hexagonal lattice, although the LPS before lyophilization did not form such a lattice. When the LPS extracted by the phenol-chloroform-petroleum ether method was precipitated by addition of two volumes of 10mM MgCl₂-ethanol, it formed essentially the same hexagonal lattice structure as that formed by the LPS extracted by the phenol-water method. From these results it is concluded that the ability of the LPS to form a hexagonal lattice structure does not depend upon the method of its extraction from bacterial cells.

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Various uniform salt forms of an R-form lipopolysaccharide (LPS) extracted from Klebsiella strain LEN-111 (O3-:K1-) were prepared and their ultrastructure was examined. The LPS, which was extracted by the phenol-water method, freed from contamination with RNA by treatment with RNase, and precipitated by addition of two volumes of 10mM MgCl₂-ethanol, was used as the original preparation for uniform salt forms. The original LPS preparation formed a hexagonal lattice structure with a lattice constant of 14.9±0.2nm. The LPS after electrodialysis retained the ability to form a hexagonal lattice structure, although its lattice constant was large (18.7±0.5nm) and the lattice structure of the electrodialyzed LPS was labile at pH 8.0 in contrast to that of the original LPS preparation. The magnesium salt form of the LPS formed essentially the same ordered hexagonal lattice structure (lattice constant of 15.0±0.2nm) as that of the original LPS preparation. The calcium and ammonium salt forms formed a hexagonal lattice structure, but the lattice constants of the calcium and ammonium salt forms were larger (18.6±0.6nm and 19.3±0.4nm, respectively) than that of the magnesium salt form. The sodium and potassium salt forms consisted of freely branching ribbon-like structures with an average width of 13nm and an average thickness of 9nm. The triethylamine salt form consisted principally of short rods (10nm×9-13nm).

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The antigenic relationships among 11 strains of Japanese encephalitis (JE) virus were analyzed by using monoclonal antibodies (NARMA) against the Nakayama-RFVL strain in hemagglutination-inhibition (HI) and neutralization (Nt) tests. Of the 14 JE virus-specific HI antibodies, all except NARMA 5 showed Nt reactivity with the homologous strain. The HI and Nt titers of these antibodies were not parallel. The 14 antibodies included the following characteristic antibodies: NARMA 3 is a species-specific antibody with HI and Nt reactivities against JE virus, NARMA 13 is a species-specific HI antibody, NARMA 6 is a Nakayama strain-specific antibody with HI and Nt reactivities, and NARMA 5 is a Nakayama strain-specific HI antibody. The 11 strains of JE virus were divided into four major antigenic groups. However, slight antigenic differences were found among some strains of the same group. Furthermore, competitive binding assays were performed to determine the distribution of antigenic determinants by enzyme-linked immunosorbent assay. The results suggest the existence of at least five HI sites on the JE virus virion, and indicate that the JE species-specific HI site and the flavivirus genus-specific HI site are topologically distinct.

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The level of antibody to AIDS-associated virus (LAV) in sera from patients with AIDS or AIDS related diseases (AIDS related complex; ARC) and Japanese hemophiliacs was studied using indirect immunofluorescence. Titer of anti-LAV antibody in sera from 89 patients with AIDS or ARC ranged between 10 and 40, 960 (median: 1, 280) and that of 83 Japanese hemophiliacs ranged from 20 to 20, 480 (median: 640). The distribution of the level of anti-LAV antibody in hemophiliacs was similar to that of patients with AIDS or ARC, and no correlation between the titer of antibody and the presence of symptoms of AIDS was observed. Our results suggested that hemophiliacs in Japan might have been infected with live LAV rather than been immunized with inactivated LAV by injection of factor VIII or IX concentrate, and more hemophiliacs in Japan might show symptoms of AIDS in the future as in the United States.

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Intraperitoneal infection of mice with two strains of Yersinia enterocolitica resulted in an inflammatory response and immunomodulation which appeared to be related to the invasive properties of the bacteria. The primary antibody response to sheep erythrocytes was enhanced by noninvasive cultures of Y. enterocolitica (serotype O:4-33 grown at 22C and at 37C, and serotype O:3 grown at 37C), when given at the same time or two days after the antigen (invasiveness was tested on HeLa cells). In contrast, invasive cultures of serotype O:3 grown at 22C, injected three days before the antigen suppressed the antibody response; enhancement was caused by these cultures only when given on the day of immunization. Delayed-type hypersensitivity to sheep erythrocytes was also suppressed by invasive cultures of Y. enterccolitica. These data indicate that the temperature of growth as well as some serotype-linked factors play a role in immunomodulation by Y. enterocolitica.

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A high molecular weight proteinaceous factor in the cell extract of sarcoma 180 (S-180) was found to inhibit phorbol myristate acetate (PMA)-triggering of macrophage H₂O₂ release. This factor (S-180 factor) was stable at 56C for 1hr and resistant to ultraviolet-irradiation. The S-180 factor inhibited the specific binding of PMA to macrophages and this was accompanied by a parallel reduction of PMA-triggered H₂O₂ release. S-180 factor preferentially depressed macrophage H₂O₂ release in response to phorbol diesters including PMA, 4β-phorbol 12β, 13α-diacetate, 4β-phorbol 12β, 13α-didecanoate, 4β-phorbol 12β, 13α-dibenzoate, and 4-o-methyl-PMA rather than the H₂O₂ release triggered by wheat germ aggulutinin or by phagocytosis of latex particles. The S-180 factor failed to affect the PMA-elicited macrophage cell spreading and macrophage phagocytic activity against latex beads with or without PMA-mediated stimulation. A similar inhibitory factor was found in the extracts of some other murine tumor cells (Ehrlich carcinoma and thymic leukemia) and normal cells (liver, spleen, and peritoneal exudate cells).

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Direct stimulations of murine B lymphocytes with synthetic lipid A analogs and synthetic muramyl dipeptide (MDP) derivatives were studied using a limiting dilution assay system. Synthetic lipid A analogs, GLA-27 and GLA-40, when conjugated with bovine serum albumin (BSA) had the ability to induce B cell clonal expansion of a single B cell from the spleen or bone-marrow. Their activities were almost the same as those of naturally obtained lipid A, but were lower than that of bacterial lipopolysaccharide (LPS). Addition of dextran sulfate (DXS) enhanced the effect of lipid A analogs. In contrast, synthetic MDP and its derivatives, although they had many biological and immunological activities in experimental animals, could not stimulate a single B cell to induce clonal expansion regardless of the presence or absence of DXS. These results suggested that lipid A analogs can directly cause the proliferation of B cells, but MDPs can not.

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