Proliferation of diversified clostridial species during biological soil disinfestation incorporated with plant biomass under various conditions

Biological soil disinfestation (BSD) involves the anaerobic decomposition of plant biomass by microbial communities leading to control of plant pathogens. We analyzed bacterial communities in soil of a model experiment of BSD, as affected by biomass incorporation under various conditions, to find out the major anaerobic bacterial groups which emerged after BSD treatments. The soil was treated with Brassica juncea plants, wheat bran, or Avena strigosa plants, irrigated at 20 or 30 % moisture content and incubated at 25–30 °C for 17 days. The population of Fusarium oxysporum f. sp. spinaciae incorporated at the start of the experiment declined markedly for some BSD conditions and rather high concentrations of acetate and butyrate were detected from these BSD-treated soils. The polymerase chain reaction-denaturing gradient gel electrophoresis analysis based on the V3 region of 16S rRNA gene sequences from the soil DNA revealed that bacterial profiles greatly changed according to the treatment conditions. Based on the clone library analysis, phylogenetically diverse clostridial species appeared exceedingly dominant in the bacterial community of BSD soil incorporated with Brassica plants or wheat bran, in which the pathogen was suppressed completely. Species in the class Clostridia such as Clostridium saccharobutylicum, Clostridium acetobutylicum, Clostridium xylanovorans, Oxobacter pfennigii, Clostridium pasteurianum, Clostridium sufflavum, Clostridium cylindrosporum, etc. were commonly recognized as closely related species of the dominant clone groups from these soil samples.     

A New Method Using a Modified Mayer's Hemalum at pH 6 for Demonstrating Mucous Neck Cells

The mucous neck cells of gastric glands were stained with a modified Mayer's hemalum adjusted to pH 6 with saturated aqueous lithium carbonate. One gram of hematoxylin was dissolved in 1000 ml distilled water and 200 mg sodium iodate, 3 g potassium alum, 50 g chloral hydrate and 1 g citric acid were added to the solution. Prior to staining, the solution was adjusted to pH 6 with saturated aqueous lithium carbonate. Bromine oxidation and urea abolished the alum hematoxylin reactivity of the mucous neck cells.     

Heat Shock Proteins in the Human Periodontal Disease Process

The production of HSP by periodontopathic Gram-negative bacteria was examined by SDS-PAGE, two dimensional gel electrophoresis, and Western blotting using monoclonal antibodies against HSPs. Strains of Actinobacillus actinomycetemcomitans, Eikenella corrodens, Fusobacterium nucleatum, Prevotella intermedia, Prevotella nigrescens, Prevotella melaninogenica, and Treponema socranskii species produced HSP which reacted with anti-Yersinia enterocolitica HSP 60 and/or mycobacterial 65-kDa HSP monoclonal antibodies. It was found that gingival homogenate samples from patients with adult periodontitis reacted with anti-human HSP 60 and bovine brain HSP 70 monoclonal antibodies. Antibodies which reacted with bacterial HSP were also found in a serum sample from a periodontitis patient. The present study suggests that HSPs are implicated in the human periodontal disease process.     

T Cell-Dependent Activation of Macrophages and Enhancement of Their Phagocytic Activity in the Lungs of Mice Inoculated with Heat-Killed Cryptococcus neoformans: Involvement of IFN-γ and Its Protective Effect against Cryptococcal Infection

Previous investigations have demonstrated that macrophages play a critical role in the first-line cellular defense mechanism against infection with Cryptococcus neoformans. In the present study, to elucidate the way in which anticryptococcal activity of macrophages is regulated at the site of infection, pulmonary intraparenchymal macrophages were directly analyzed for expression of their surface molecules and their phagocytic activities against the organism, and the effects of depletion of T cells and endogenous IFN-γ in vivo on these parameters were examined. In the lungs of mice intratracheally inoculated with heat-killed C. neoformans, macrophages were activated, as indicated by augmented expression of MHC class II, intercellular adhesion molecule-1 (ICAM-1) and Fc receptor (FcR), and about two-thirds of macrophages were found to have ingested an average of 3.77 ± 0.12 yeast cells per macrophage. In mice depleted of both CD4⁺ and CD8⁺ T cells by injecting the specific monoclonal antibodies (mAbs) or anti-IFN-γ mAb, not only augmentation of the expression of macrophage activation markers but also phagocytosis of C. neoformans was significantly reduced. These results suggest that anticryptococcal activity of macrophages is regulated by IFN-γ endogenously produced by T cells. Additionally, treatment with IFN-γ were shown to significantly prolong the survival time of mice infected with viable C. neoformans. Additionally, preimmunization with heat-killed C. neoformans significantly prolonged the survival time of mice which received the following infection.     

The plastid aldolase gene fromChlamydomonas reinhardtii: Intron/exon organization, evolution, and promoter structure

Genomic clones encoding the plastidic fructose- 1,6-bisphosphate aldolase ofChlamydomonas reinhardtii were isolated and sequenced. The gene contains three introns which are located within the coding sequence for the mature protein. No introns are located within or near the sequence encoding the transit-peptide, in contrast to the genes for plastidic aldolases of higher plants. Neither the number nor the positions of the three introns of theC. reinhardtii aldolase gene are conserved in the plastidic or cytosolic aldolase genes of higher plants and animals. The 5 border sequences of introns in the aldolase gene ofC. reinhardtii exhibit the conserved plant consensus sequence. The 3 acceptor splice sites for introns 1 and 3 show much less similarity to the eukaryotic consensus sequences than do those of intron 2. The plastidic aldolase gene has two tandemly repeated CAAT box motifs in the promoter region. Genomic Southern blots indicate that the gene is encoded by a single locus in theC. reinhardtii genome.     

Nitrogenous nutrient uptake by phytoplankton and ammonium regeneration by microbial assemblage in Lake Biwa

In situ rates of nitrate, ammoniwn and urea uptake by the phytoplanktonassemblage, and the regeneration rate of ammonium by the microbialassemblage, in Lake Biwa were measured using the nitrogen 15tracer method from 1985 to 1987. The rate of total nitrogen(sum of ammonium, nitrate and urea) uptake was in the rangeof 62–594 ng N^–1 r^–1 h^–1. The percentagecontribution of ammonium uptake was 41–92%, that of urea4–58% and that of nitrate <1–28% of total uptake.The annual mean new production which was supported by nitrateuptake was 18% of the total production in 1986. The phytoplanktonassemblage in Lake Biwa preferentially utilized regeneratednitrogen, such as ammonium and urea, whose concentration wasmuch lower than that of nitrate throughout the observation penodwithout in summer. The in situ nitrogen uptake rate was almostsufficient to meet the nitrogen requirement of the phytoplanktonassemblage, except in midsummer when the nitrate concentrationwas below the detection limit of 0.3 µg N r^–1. Inthe trophogemc layer, the rate of ammonium regeneration was66–272 ng N 1^–1 h^–1 Although the ambient ammoniumconcentration in the trophogenic layer was maintained at aroundthe half-saturation constant for ammonium uptake kinetics, theammomum uptake rates were always highly correlated with ammoniumregeneration rates. From the size fractionation experimentsand estimates from the literature, it was suggested that themicrobial assemblage <1 µm may have been the most importantagent responsible for the ammonium regeneration processes inthe trophogenic layer.     

Long-term ploidy stability of shoot primordium cultures and produced plants of melon

The chromosome number of cells in the shoot primordium aggregates and produced plants of melon [Cucumis melo L. 'Prince' (2n=2x=24)] was examined. Shoot primordium aggregates were induced from shoot-tips cultured in liquid medium and shaken at low speed (2 rpm). They were maintained by subculturing small pieces (5mm<) every 4 weeks. Shoot primordium aggregates just after induction contained about 97 diploid and 3 tetraploid cells, which was similar to those maintained in shoot primordium cultures for 6 years. This indicates that the ploidy level was maintained stably. On the other hand, plants produced from the shoot primordium aggregates just after induction were either diploid, tetraploid or mixoploid with both diploid and tetraploid cells. These ploidies were again observed among plants produced from shoot primordium cultures that were 2, 3 or 4 years old. A majority of produced plants were diploid while the total frequency of tetraploids and mixoploids was less than 8 of plants produced from all ages. Therefore, the frequency of somaclonal variation with respect to ploidy among plants produced from shoot primordium aggregates is likely to be stable at a low level over the long term.     

Isolation and characterization of hemolytic genes from Actinobacillus actinomycetemcomitans

Abstract Periodontopathic Actinobacillus actinomycetemcomitans produces hemolysin and other leukotoxins. In the present study, two distinct clones which lysed horse erythrocytes were isolated by screening genomic DNA libraries of A. actinomycetemcomitans ATCC 43718 on blood agar plates. DNA hybridization analysis indicates that there were two distinct hemolytic genes present. Sonicated extracts from both Escherichia coli clones possessed hemolytic activities on horse, sheep and human erythrocytes, but not those of rabbit. Rabbit antiserum to A. actinomycetemcomitans ATCC 43718 whole cells inhibited the hemolytic activities of these clones.     

Activation of Unusual Mac-1+CD4− CD8− T Cells Bearing αβ Antigen Receptor in Murine Lungs during Infection with Mycobacterium bovis BCG: Regulation by Interferon-γ

We have recently (Kawakami et al, Immunol. Lett. 1995;46: 143) demonstrated that unusual Mac-1⁺CD4^?CD8^? T cells bearing αβ antigen receptor (Mac-1⁺ αβ T cells) reside in a considerable proportion in murine lungs. The present study was performed to examine the dynamics of accumulation of these cells in the lungs following intravenous administration of Mycobacterium bovis BCG (BCG). Mac-1⁺ αβ T cells accumulated rapidly 24 hr after infection, followed by a gradual increase over the observation period of 15 days. Furthermore, the expression of Ia, ICAM-1 and FcγR II/III on their surface intensified dramatically after BCG infection. The kinetics of enhancement of Ia expression was slower than that of ICAM-1, with the maximum level attained in one day in the latter molecule but in two weeks in the former. Neutralization of endogenous IFN-γ by specific mAb completely blocked the augmented expression of Ia on Mac-1⁺ αβ T cells after BCG infection, but did not have any significant effect on that of ICAM-1. In contrast, in vivo administration of IFN-γ enhanced the expression of ICAM-1 as well as that of Ia. Our results indicate that accumulation of Mac-1 αβ T cells within the lung is associated with a differential change in the expression of surface antigens, and suggest that these cells may play a role in the host defense against mycobacterial infection.