The hemagglutination (HA) test was done on 85 strains of Branhamella catarrhalis, isolated from sputum of patients with respiratory infections; 53% were HA positive strains. Three HA positive and three HA negative strains were selected and were observed under the electron microscope. The bacterial cell wall appeared to be lobulated and its total thickness was about 38nm. The nuclear region consisted of whorls or fibrils and dense bodies. Five strains were fimbriated and one strain was nonfimbriated. The size of fimbriae was about 68nm in length and 4.5nm in width. The fimbriae of Branhamella catarrhalis were densely arranged and peritrichous in distribution. There was no change of fimbriation between broth and agar cultures.

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DNA fragments Vibrio alginolyticus chemovar iophagus, at least 7kb in length, were ligated to Escherichia coli expression vectors. Three clones of Escherichia coli HB101 (pLCO-1, pLCO-2, pLCO-3) were obtained by the colony immuno-blotting method using anti-collagenase antibody. In Escherichia coli, all these genes produced collagenase antigens which were detected with Western blotting. The amino acid sequence of chemically purified collagenase fragments was also analyzed. An approximately 2.5kb DNA fragment of the pLCO-1 clone was sequenced, and we found that portions of the deduced amino acid sequence were completely identical with the corresponding amino acid sequence of the chemically analyzed fragments. Therefore, it is highly probable that the gene studied in the present experiment is truly a collagenase structural gene.

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The present study was conducted by the use of purified glucocorticoid antagonizing factor (GAF) released in blood of endotoxemic mice to determine whether or not the factor (GAF and Ca²⁺) may play a possible role of mediator in depletion mechanism of liver glycogen in endotoxemia. The liver glycogen level in 2hr after injection with GAF plus cortisone-treated mice was markedly lower than that in cortisone alone-treated mice. However, the administration of trifluoperazine or verapamil markedly increased glycogen levels in liver of GAF plus cortisone-injected mice. On the other hand, when the mice fed a calcium-free diet were injected with GAF plus cortisone, there was merely a significant difference in liver glycogen level as compared to cortisone alone-treated mice. The level of Ca²⁺ in liver cytosol fraction in cortisone-treated mice was higher 2hr after GAF injection than that in the cortisone alone-treated one. The phosphorylase a activity in liver 2hr after injection of GAF plus cortisone did not show a significant difference as compared to that in mice treated with cortisone alone. However, the activity ratio of glycogen synthase enzyme (synthase I/synthase I+D) was decreased in GAF plus cortisone-treated mice as compared to that in cortisone alone-treated mice. These findings suggest that there are participations of Ca²⁺ and mediator GAF released from reticuloendothelial system (RES) macrophages in glucoregulation of endotoxemia. Thus, it may be speculated that intracellular Ca²⁺ may mediate glycogenesis rather than glycogenolysis in the depletion mechanism of liver glycogen during GAF-poisoning.

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Although mycobacteria grow in Dubos liquid medium showing an arithmetic linear growth, the initial few days of growth were found to correspond to an ‘induction’ period. In this period, rapid increase of the amount of growth occurred, whereas increase of the number of colony-forming units remained at a low level. This finding shows that the rapid increase of the amount of growth is accompanied by rapid death of multiplied bacteria. In a successive period, which was considered to correspond to the logarithmic growth phase, a 1:1 correspondence existed between the amount of growth and the number of colony-forming units. The induction period is not considered to be a lag phase, in which the bacteria grow slowly, but a period of unbalanced relationship between the growth and the viability. Even when we inoculated different sizes of bacteria, the amounts of growth became similar in both inoculations after several days of incubation. However, the number of colony-forming units remained always smaller in the use of small inocula than in the use of large inocula. In the use of small inocula, much more rapid increase of the amount of growth occurred. However, this rapid increase gave rise to rapid death of bacteria.

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During 1985 to 1990, serum samples were obtained from 229 healthy dogs. The dogs lived in Hokkaido, known to be infested with ticks. An enzyme-linked immunosorbent assay (ELISA) was used to detect IgG and IgM antibodies against Borrelia burgdorferi HO14 and HP3, which were isolated from Ixodes ovatus and I. persulcatus in the area. IgG antibody to B. burgdorferi HO14 was detected in 8.8% (1985), 16.4% (1987) and 18.5% (1990). IgM antibody to the bacteria was detected in 1.8% (1987) and 2.5% (1990). Antibodies to the strain HP3 of B. burgdorferi were also detected in the serum samples of dogs, but the percentage of seropositive sample to the strain HP3 was lower than that to the strain HO14. Statistical differences were not noticed between pet and street dogs. No antibody to B. burgdorferi was observed in 13 beagle dogs as experimental animal.

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The penetration of gentamicin and polymyxin B into the inside of Bacillus subtilis spores was examined by an immunoelectron microscopy method with colloidal gold-immunoglobulin G (IgG) complex. The colloidal gold particles were located predominantly in the coat, region of both gentamicin-treated and polymyxin B-treated spores and were hardly observed in the other regions, i.e., the cortex and core regions. When these antibiotic-treated spores were subsequently treated with CaCl₂, the number of gold particles bound to the coat region was greatly decreased. These results suggest that these two antibiotics are able to penetrate into the spore coat but not into the cortex or core, that is, the primary permeability barrier to them exists between the coat and the cortex regions.

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Ia restriction specificity involved in T cell proliferative responses to keyhole limpet hemocyanin (KLH) has been analyzed using a variety of allogeneic bone marrow chimeras. The chimeric mice were prepared by reconstituting irradiated AKR, SJL, B10.BR and B10A(4R) mice with bone marrow cells from B10 mice. When such chimeric mice had first been primed with KLH in complete Freund's adjuvant (CFA), T cells from H-2 incompatible fully allogeneic chimeras showed significantly higher responses to KLH in the presence of antigen-presenting cells (APC) of donor strain (B10) than APC of recipient strain. However, in H-2 subregion compatible chimeras, [B10→B10.A(4R)], which were matched at the H-2D locus and at minor histocompatible loci, the T cells could mount vigorous responses to KLH with antigen-presenting cells (APC) of either donor or recipient type. The same results were obtained as well with chimeras that had been thymectomized after full reconstitution of lymphoid tissues by donor-derived cells. A considerable proportion of KLH-specific T cell hybridomas established from [B10→B10.A(4R)] chimeras exhibited both I-A^b and I-A^k restriction specificities. The present findings indicate that the bias to donor Ia type of antigen specific T cells is determined by donor-derived APC present in the extrathymic environment but that cross-reactivity to the recipient Ia is influenced to some degree by histocompatibility between donor and recipient mice, even though the histocompatible H-2D locus and minor histocompatibility loci seem not to be directly involved in the I-A restricted responses studied herein.

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Chromosomal DNAs were extracted from toxigenic three Clostridium botulinum type E strains isolated from food-borne botulism. After digestion by EcoRI, the fragments were cloned into Escherichia coli by using bacteriophage lambda gt11 and screened with monoclonal antibody recognizing the light chain component of botulinum type E toxin. The fragments (about 1kbp size) cloned from each strain were recloned into a plasmid vector pUC118. The E. coli cells transformed with the recombinant plasmids produced 33kDa protein with or without IPTG (isopropyl-β-D-thiogalactopyranoside) which reacted with the monoclonal antibody. The nucleotide sequences of the cloned EcoRI fragments from the three type E strains were identical and contain the 5'-terminal region of the type E toxin gene. It was also found that there exist several highly homologous nucleotide sequences among the botulinum types A, C and E, and tetanus toxin genes in both translated and untranslated regions.

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Chemical analysis was carried out on lipopolysaccharides from Vibrio cholerae bio-serogroup Hakata 487-85. The O-specific chain of the phenol-soluble lipopolysaccharides was demonstrated by ¹³C-NMR spectroscopy and methylation analysis to contain a linear homopolymer of α(1→2) linked N-acetylperosamine (4-acetamido-4, 6-dideoxy-D-mannopyranose), which was closely similar to but not identical to a linear α(1→2) linked N-3-deoxy-L-glycerotetronyl (S-2, 4-dihydroxybutyryl) perosamine-homopolymer constituting that of O1 Vibrio cholerae lipopolysaccharides.

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To know the biological significance of the HIV variation, we investigated the interaction between two different HIV clones. Two distinct clones were mixed and propagated by infecting MT-4 cells. After passaging the mixture viruses 8 times, and cloning by the plaque method, we obtained not only viruses of homogeneous type like each parental clone but also heterogeneous-type viruses which showed a mixture of two parental viruses with regard to phenotype and genotype. To further segregate a single virus from the mixture, we recloned the heterogeneous viruses using the plaque method. We found that about 40% of recloned viruses from heterogeneous-type viruses were still heterogeneous.

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