Aromatic Substances Inhibiting Wheat Powdery Mildew Produced by a Fungus Detected with a New Screening Method for Phylloplane Fungi

Microorganisms isolated from wheat leaf surfaces were screened for inhibition of wheat powdery mildew. A new screening method, in which wheat leaves were inoculated with Blumeria graminis f. sp. tritici and incubated with the cultured microorganisms under non-contact conditions, was developed in the present study. Using this method, 10 phylloplane fungi that inhibited wheat powdery mildew were selected from 408 microorganisms isolated from wheat leaf surfaces. Among these 10 strains, a fungus designated as Kyu-W63 had an especially strong inhibitory effect. Kyu-W63 produced white colonies without spores when cultivated on PDA. Kyu-W63 had a strong aromatic odor when being cultured. Wheat powdery mildew was suppressed even though a membrane filter with a pore size of 0.45 μm was placed between the mycelial colony and wheat leaf segment. However, when activated charcoal was introduced, Kyu-W63 did not inhibit growth of B. graminis. It was presumed that volatile substances were involved in the inhibitory effect of Kyu-W63. GC-MS analysis was used to identify two substances produced by Kyu-W63 with molecular weights of 164 and 166. Kyu-W63 also inhibited the in vitro growth of four plant pathogenic fungi other than B. graminis. Received 19 September 2001/ Accepted in revised form 7 February 2002     

A novel culture method of canine peripheral blood lymphocytes with concanavalin a and recombinant human interleukin-2 for adoptive immunotherapy

This study was designed to develop a novel culture method for the efficient proliferation of canine peripheral blood lymphocytes (cPBL) for adoptive immunotherapy. When cPBL were cultured in the presence of concanavalin A (Con A), proliferation of cPBL was induced and expression of interleukin-2 receptor (IL-2R) which enables to respond to exogenously added IL-2 was upregulated. And then, when cPBL were cultured with recombinant human interleukin-2 (rhIL-2) in addition to Con A, proliferation was accelerated and increased to about 10-fold after 1 week. The phenotypic analysis showed that the main population of the cultured cPBL was consisted of CD8+ positive lymphocytes. Among them, CD4+CD8+ double positive (DP) lymphocytes had significantly increased, and the ratio of CD4+ single positive (SP) lymphocytes to CD8+ SP lymphocytes (CD4+SP/CD8+SP) was decreased as compared to before culturing. To evaluate the cytotoxic activity of cPBL cultured with Con A and rhIL-2, furthermore, cytotoxic assay was carried out against xenogeneic melanoma cell line (MeWo), which resulted in MHC-unrestricted cytokilling. These results suggest that the culture method of cPBL by the use of Con A and rhIL-2 may be useful for generating lymphokine activated killer cells, and also this may be beneficial for adoptive immunotherapy of tumor-bearing dogs.     

稻瘟菌水平抗性稳定性的研究——品种与小种适合度的分析

在温室条件下,对9个无垂直抗性基因粳稻品种×8个日本标准稻瘟菌株组合测定了侵染几率(IP)、病斑型比例(PB)、病斑扩展速率(LR)和产孢能力(SPC)组分,并据此确定了品种与小种的适合度。方差分析结果表明,除SPC的互作项外,其它组分及类型,方差均达到显著或极显著水平。但品种和小种主效应所占方差更大,从而证实了品种的抗性类型为水平抗性。某些品种在IP、PB及LR组分上反映了小种的专化作用,但因为不同品种在各组分上的抗性反应不同,因此,品种水平抗性的稳定性不能由组分的小种专化作用确定,而须由各组分组成的适合度的小种专化性确定。对品种X小种的适合度进行了互作的多重比较,未发现专化性互作,说明供试品种的水平抗性是稳定的。     

Experimental infection of recent field isolates of feline herpesvirus type 1

Two field isolates of feline herpesvirus type 1 (FHV-1) designated as 00-015 and 00-035, were obtained from cats diagnosed as feline viral rhinotracheitis (FVR) in Japan. To analyze the character of recent FHV-1, these two isolates and our laboratory strain C7301 were inoculated experimentally to specific-pathogen-free cats. Although all cats showed typical FVR symptoms, more severe clinical symptoms were observed on cats infected with the isolates 00-015 and 00-035 compared with those of C7301-infected cats. Severe ocular lesions including conjunctivitis were found in the cats infected with the isolates, indicating that the recent FHV-1 has a potential to induce severe FVR symptoms including ocular lesions.     

Phenotype and function of murine discrete Peyer's patch macrophage derived - dendritic cells

The phenotype and function of peritoneal cavity macrophage-derived dendritic cells (PEC-DC) was previously reported. In this study we have gone further in using our established culture system to generated discrete Peyer's patch dendritic cells (DPP-DC) from murine discrete Peyer's patch macrophages (DPP-M?), following stimulation with granulocyte macrophage colony stimulating factor (GM-CSF) plus interleukin 4 (IL-4) for 7 days. DPP-M? from murine small intestines were obtained by mechanical disruption of discrete Peyer's patches (DPP), followed by metrizamide density gradient centrifugation to remove Peyer's patch resident DC and debri, after which an overnight adherent step in tissue culture medium was carried out for macrophage enrichment. Characterization of the generated DPP-DC was carried out using well-established criteria of morphology, expression of membrane antigens and capacity for antigen presentation. Dendritic cells expressed DEC-205, F4/80 and CD34 at high levels, but exhibited very low CD11c levels. They were shown to present soluble protein antigen to CD3(+) spleen T cells. A comparison of the surface antigen expression in the progenitor DPP-M? population and the generated DPP-DC showed a significant decrease in MHC class II levels and a marked down regulation of the co-stimulatory molecule CD86 (B7-2). High expression of the haemopoietic progenitor marker CD34 indicates that the generated DC, possess a haemopoietic rather than myeloid origin. Taken together, these results may provide a better understanding of the complex network regulating mucosal immune responses.     

A role for balance of interferon-gamma and interleukin-4 production in protective immunity against Neospora caninum infection

A suitable balance in the production of Th1/Th2-type cytokines has a crucial role in the control of microbial infections. We investigated cytokine production patterns and effects during Neospora caninum infection, based on two mouse models and an in vitro system. In the acute infection of N. caninum, BALB/c-background IFN-gamma-deficient mice that were sensitive to the N. caninum infection showed high levels of IL-10 production, whereas significant levels of interferon-gamma (IFN-gamma) and interleukin-4 (IL-4) production were observed in resistant wild type mice. BALB/c mice vaccinated with recombinant vaccinia virus expressing N. caninum surface protein NcSRS2 resisted parasite spread throughout the body, low levels of IFN-gamma production and high levels of IL-4 production were observed compared to unvaccinated animals. The treatment of N. caninum-infected cells with IFN-gamma or IL-10 decreased the host-cell viability in an in vitro system using mouse macrophage J774A.1 cells. On the other hand, IL-4, but not IL-10 administration, increased the viability of N. caninum-infected and IFN-gamma-treated cells. In the light of the balance of Th1/Th2-type cytokine production, an IFN-gamma/IL-4 balance may have a crucial role for the control of cellular responses against the parasite invasion.     

Suppressive effect of liquid potassium silicate on powdery mildew of strawberry in soil

A soil drench of soluble silicon (liquid potassium silicate) was tested for the control of powdery mildew of strawberry in soil for 2 years in a vinyl house under growers' conditions on two cultivars: very susceptible Toyonoka and slightly susceptible Sachinoka. Soluble silicon suppressed the disease more effectively as a preventive control than as a control to reduce initial incidence. The suppressive effect against this disease was more effective for Toyonoka than for Sachinoka. The control value of soluble potassium silicate was 85.6% for Toyonoka in the first year and 60.2% in the second year, while for Sachinoka it was 58% in the first year and 40.6% in the second year. Strawberry leaf hardness was measured for the control and silicate-treated leaves. Although silicate-treated leaves seemed to be harder than control leaves, leaf hardness did not vary significantly between controls and silicate-treated leaves in either of the 2 years, and the mode of preventive action of liquid potassium silicate remains unknown. A factor other than a physical action may be involved.     

Seedborne fungi detected on stored solanaceous berry seeds and their biological activities

We isolated 629 fungi from 1296 berry seeds of solanaceous plants, including tomato (Lycopersicon esculentum), eggplant (Solanum melongena), bell pepper (Capsicum annuum), and red pepper (Capsicum annuum var. annuum) preserved for long and short terms. The isolates were classified into 22 genera excluding unidentified fungi, and the fungal floras were divided into two types: the tomato–eggplant and pepper groups. The results of cluster analysis with unweighted pair-group method with arithmetic average also supported these groups. Most tomato seeds infested with Geotrichum candidum germinated and grew the same as uninfested seeds. Cladosporium sphaerospermum and Arthrinium sp. isolated from eggplant seeds strongly suppressed germination, and Penicillium variabile suppressed seminal root elongation on eggplant. Alternaria alternata, Botrytis cinerea, and Myrothecium verrucaria detected from red pepper or bell pepper seeds were pathogenic to the fruits and the seedlings after artificial inoculation.     

Evaluation of three phytoplankton species as food for the pearl oyster <Emphasis Type="Italic">Pinctada fucata</Emphasis>

In the pearl cultivation farms of the Ehime Prefecture, Japan, mass mortalities of the pearl oyster Pinctada fucata have occurred since 1994. The occurrences of mass mortality roughly coincided with a shift of the dominant phytoplankton from Skeletonema and Chaetoceros to Chaetoceros and Nitzschia all of which belong to Bacillariophyceae. Hence, we evaluated Nitzschia, together with Chaetoceros and Isocrysis, as food for the oyster. Wet weights, lengths, widths, glycogen contents, and growth rates in terms of wet weight of the oysters in all the feeding treatments were significantly higher than those in the non-feeding treatment. The highest glycogen content (2.34%) and growth rate (2.21 g month⁻¹) were found in the Chaetoceros treatment. Growth rate in the Isocrysis treatment (1.63 g month⁻¹) was also high, although glycogen content in this treatment (0.41%) was low. In the Nitzschia treatment, growth rate of the oyster (0.94 g month⁻¹) was the lowest and glycogen content (0.83%) was also low relative to that in the Chaetoceros treatment. Chlorophyll a concentration in fecal pellets was lowest in the Nitzschia treatment (<2.7 μg mg⁻¹), suggesting more complete digestion of Nitzschia by the oyster. Thus, Nitzschia was edible and digestible but not assimilated by P. fucata. We propose the following scenario for the relationship between Nitzschia dominance and mass mortality. When Nitzschia dominates in a culture area, the physiological condition of P. fucata deteriorates due to low assimilation of Nitzschia by the oyster, followed by susceptibility of the oyster to infection by agents lethal to the oyster.