Induced systemic resistance of Chinese cabbage to bacterial leaf spot and <Emphasis Type="Italic">Alternaria </Emphasis>leaf spot by the root endophytic fungus, <Emphasis Type="Italic">Heteroconium chaetospira</Emphasis>

 The root endophytic fungus Heteroconium chaetospira isolate OGR-3 was tested for its ability to induce systemic resistance in Chinese cabbage against bacterial leaf spot caused by Pseudomonas syringae pv. maculicola and Alternaria leaf spot caused by Alternaria brassicae of the foliar diseases. Chinese cabbage seedlings planted in soil infested with an isolate of H. chaetospira were incubated in a growth chamber for 32 days. The first to fourth true leaves of the seedlings were challenge-inoculated with P. syringae pv. maculicola or A. brassicae. Chinese cabbage planted in soil infested with H. chaetospira showed significant decreases in the number of lesions of bacterial leaf spot or Alternaria leaf spot when compared to the control plants not treated with H. chaetospira. The results indicated that colonization of roots by H. chaetospira could induce systemic resistance in Chinese cabbage and reduce the incidence of bacterial leaf spot and Alternaria leaf spot. Received: April 24, 2002 / Accepted: August 9, 2002     

Comparison and characterization of biosorption by Weissella viridescens MYU 205 of periodic group 12 metal ions

Because heavy metals cause various health hazards, we studied biosorption by Weissella viridescens MYU 205. MYU 205 showed high biosorption for Cd (II) and Hg (II) and was low for Zn (II). The Hg (II) biosorption rate was high at about 80%. Different biosorptions were shown for each metal after successive incubation. About 20% of the Zn (II) biosorption was observed after 3 h. Cd (II) biosorption increased in a time‐dependent manner until 3 h, then gradually decreased. Hg (II) was immediately sorbed at 79.6 ± 4.7% and decreased at 3 h to 52.9 ± 2.6%, and then gradually increased to 77.8 ± 3.6%. Using heat‐killed cells, the rate of biosorption of Zn (II) and Cd (II) decreased whereas Hg (II) tended to increase. The metal resistance was high, that is Zn (II) > Cd (II) > Hg (II); while the affinity was opposite where MYU 205 showed high affinity to Hg (II) and low affinity to Zn (II). Our data shows lactic acid bacteria may be powerful heavy metal sorbents for detoxification.     

Establishment of a laboratory colony of taiga tick Ixodes persulcatus for tick-borne pathogen transmission studies

Ixodes persulcatus Schulze (I. persulcatus) is distributed in Russia and Far East Asia including Japan, and has been implicated as the vector of several human pathogens. In particular, I. persulcatus acts as the only tick vector for human lyme borreliosis in Japan. In order to elucidate the mechanism of transmission of I. persulcatus-borne pathogens, we developed a laboratory colony of I. persulcatus. Ticks were fed on Syrian hamster and engorged ticks that had dropped off the animals were collected and maintained to allow them to molt. Tick rearing was performed in incubator at 20 degrees C with 95% relative humidity and 12-hour light/dark photo-period regimen. We found out that adult females fed for 8 +/- 2 days and had a pre-oviposition period lasting for 7 +/- 2 days. The minimum egg incubation period was 1 month with the hatched larvae feeding for 3 +/- 1 days and molting to nymphs 3-4 months thereafter. Meanwhile, the nymphs fed for 4 +/- 1 days and molted to adult 2-3 months thereafter. For future analysis of gene expression profiles in I. persulcatus, we cloned and sequenced the actin gene (a housekeeping gene), and found that it is 92.7% to 98.6% homologous to the published sequences of related ixodid ticks. This laboratory colony of I. persulcatus will facilitate investigations on the role of tick-derived molecules on the transmission of I. persulcatus-borne pathogens and will be important for identification of potential anti-tick vaccine and acaricide target molecules.     

Establishment of a BRAF V595E-mutant canine prostate cancer cell line and the antitumor effects of MEK inhibitors against canine prostate cancer

Canine prostate cancer (cPCa) is a malignant neoplasm with no effective therapy. The BRAF V595E mutation, corresponding to the human BRAF V600E mutation, is found frequently in cPCa. Activating BRAF mutations are recognized as oncogenic drivers, and blockade of MAPK/ERK phosphorylation may be an effective therapeutic target against BRAF-mutated tumours. The aim of this study was to establish a novel cPCa cell line and to clarify the antitumor effects of MEK inhibitors on cPCa in vitro and in vivo. We established the novel CHP-2 cPCa cell line that was derived from the prostatic tissue of a cPCa patient. Sequencing of the canine BRAF gene in two cPCa cell lines revealed the presence of the BRAF V595E mutation. MEK inhibitors (trametinib, cobimetinib and mirdametinib) strongly suppressed cell proliferation in vitro, and trametinib showed the highest efficacy against cPCa cells with minimal cytotoxicity to non-cancer COPK cells. Furthermore, we orally administered 0.3 or 1.0 mg/kg trametinib to CHP-2 xenografted mice and examined its antitumor effects in vivo. Trametinib reduced tumour volume, decreased phosphorylated ERK levels, and lowered Ki-67 expression in xenografts in a dose-dependent manner. Although no clear adverse events were observed with administration, trametinib-treated xenografts showed osteogenesis that was independent of dosage. Our results indicate that trametinib induces cell cycle arrest by inhibiting ERK activation, resulting in cPCa tumour regression in a dose-dependent manner. MEK inhibitors, in addition to BRAF inhibitors, may be a targeted agent option for cPCa with the BRAF V595E mutation.     

Molecular characterization of a novel bovine group A rotavirus

In this study, by partial sequence analysis of the genome segments encoding VP5* and VP7, we characterized a novel bovine group A rotavirus, namely, Tak2, that was detected from adult cattle diarrhea in Tochigi Prefecture, Japan. The nucleotide (nt) and deduced amino acid (aa) sequences of the genome segments encoding VP5* and half of the amino terminal portion of VP7 of Tak2 revealed a low identity with those of group A rotaviruses carrying previously published P and G type specificities (VP5*: nt identity, 61.6%-67.6% and aa identity, 58.0%-71.4%; half of the amino terminal portion of VP7: nt identity, 57.8%-73.5% and aa identity, 61.2%-70.9%). Additionally, phylogenetic analysis of the nt sequences of the genome segments encoding VP5* and half of the amino terminal portion of VP7 revealed that Tak2 formed a branch separate from the established P and G types. These results suggested that Tak2 could possess novel P and G types yet not reported among group A rotaviruses.     

Interferon-gamma expression associated with suppression of bovine leukemia virus at the early phase of infection in sheep

Immunological control of bovine leukemia virus (BLV)-infection has been reported as dependent on the expression balance of types 1 and 2 cytokines. In this report, mRNA expression of interferon (IFN)-gamma and interleukin (IL)-2 (type 1 cytokines), and of IL-4 and IL-10 (type 2 cytokines) were evaluated in concanavalin A-stimulated peripheral blood mononuclear cells (PBMC) from BLV-infected sheep. Despite the same dose of BLV-infection, the extent of viral propagation was markedly different between eight individual sheep by 12 weeks post infection. The virus did not propagate well in three sheep, which showed augmented mRNA expression of IFN-gamma, a strong indicator of cell-mediated immunity, immediately after BLV-infection. Among the other five sheep having more than 2% of BLV-infected cells among PBMC at 12 weeks post infection, four sheep developed B-cell leukemia or lymphoma within 2 years after infection. These observations indicate IFN-gamma expression may play an important role in the protective mechanism against BLV propagation at the early phase of the infection.     

Changes in size of soil seed bank in Robinia pseudoacacia L. (Leguminosae), an exotic tall tree species in Japan: Impacts of stand growth and apicultural utilization

This study examined the factors that influence the size of a soil seed bank in Robinia pseudoacacia stands. We proposed two hypotheses: the amount of buried seeds of R. pseudoacacia is proportional to stand biomass rather than stand age, and apicultural utilization influences the size of the soil seed bank. R. pseudoacacia generally produces seeds with various degrees of physical dormancy. In addition, this tree is short-lived, and fecundity declines after 30–40 years in relation to tree vigor. However, R. pseudoacacia is intensively used in apiculture as an important honey source, and supply of honeybees by beekeepers could influence seed crops. We investigated 25 plantations in Hokkaido (Central, Hidaka, and Oshima) and in Nagano, Japan, where many naturally regenerated R. pseudoacacia stands occur. We found great variation in the size of the soil seed bank among stands, ranging from 13,757 to 6.4 seeds m⁻² per plot. A generalized linear mixed model revealed that both basal area (BA_Rp) and apicultural utilization best explained the size of the soil seed bank. Both the positive effect of BA_Rp and small contribution of stand age to the model implied that the soil seed bank is transitory, and will not persist for very long time. The large contribution of apicultural utilization to the size of the soil seed bank implied pollinator limitation under natural conditions. R. pseudoacacia often has large flower crops, and native pollinators can not keep up with the demand for pollination. Thus, the supply of honeybees by beekeepers should improve the pollination success of R. pseudoacacia. From these results, we conclude that we must consider stand history with respect to apicultural utilization if the land is harvested or a stand declines.     

Competition as a predisposing factor of crown dieback in a secondary forest of <Emphasis Type="Italic">Betula maximowicziana</Emphasis> in Hokkaido,northern Japan

We analyzed the probability that Betula maximowicziana Regel (monarch birch) would suffer crown dieback (crown-dieback probability) and the basal area growth rate (GB), which was found to be a predisposing stress factor making birch trees susceptible to crown dieback. First, we analyzed the relationship between the probability that birch trees would suffer from crown dieback in 1999 and GB from a period prior to the occurrence of crown dieback (1985–1987), using a data set of repeated measurements on 217 trees. Logistic regression analysis revealed that monarch birch had a larger crown-dieback probability when GB was low in the preceding period. Hence, there were predisposing stress factors that reduced GB and continued to affect trees for at least a decade. Next, we analyzed GB in the same period in relation to symmetrical and asymmetrical competition between trees and found that GB was reduced by symmetrical competition, suggesting that this was one of the predisposing factors for crown dieback. Based on these results, we used selected models for crown-dieback probability and GB to calculate crown-dieback probabilities for individuals with different initial basal areas and experiencing different intensities of symmetrical competition. The predicted crown-dieback probability decreased with decreasing symmetrical competition between trees. We discuss a possible process of crown dieback to death for monarch birch and the use of thinning as a method to reduce the risk of crown dieback.     

Toxicity of chlorothalonil to marine organisms

We evaluated the acute toxicity of the antifouling biocide 2,4,5,6-tetrachloro-1,3-benzenedicarbonitrile (chlorothalonil, TPN) to an alga (Skeletonema costatum), crustacea (Tigriopus japonicus, kuruma prawn Marsupenaeus japonicus), and a teleost fish (red sea bream Pagrus major) commonly found in the coastal waters of Japan. We also performed acute toxicity and early-life-stage tests for TPN using mummichog Fundulus heteroclitus and used the results to predict the chronic toxicity of TPN to red sea bream. The acute toxicity values of TPN (based on measured concentrations) for S.?costatum, T.?japonicus, M.?japonicus, red sea bream, and mummichog were 0.95, 16, 290, 35, and 61???g/l, respectively. The early-life-stage test with mummichog embryos revealed that hatchability and growth were the most sensitive indicators, and the lowest- and no-observed-effect concentrations, based on the measured concentrations, were 32 and 11???g/l, respectively. We estimated the chronic toxicity value for red sea bream to be 11???g/l based on these toxicity values. Considering the current measured concentrations of TPN in coastal waters of Japan, its environmental impact would appear to be limited.