Infection of Melissococcus plutonius clonal complex 12 strain in European honeybee larvae is essentially confined to the digestive tract

Melissococcus plutonius is an important pathogen that causes European foulbrood (EFB) in honeybee larvae. Recently, we discovered a group of M. plutonius strains that are phenotypically and genetically distinct from other strains. These strains belong to clonal complex (CC) 12, as determined by multilocus sequence typing analysis, and show atypical cultural and biochemical characteristics in vitro compared with strains of other CCs tested. Although EFB is considered to be a purely intestinal infection according to early studies, it is unknown whether the recently found CC12 strains cause EFB by the same pathomechanism. In this study, to obtain a better understanding of EFB, we infected European honeybee (Apis mellifera) larvae per os with a well-characterized CC12 strain, DAT561, and analyzed the larvae histopathologically. Ingested DAT561 was mainly localized in the midgut lumen surrounded by the peritrophic matrix (PM) in the larvae. In badly affected larvae, the PM and midgut epithelial cells degenerated, and some bacterial cells were detected outside of the midgut. However, they did not proliferate in the deep tissues actively. By immunohistochemical analysis, the PM was stained with anti-M. plutonius serum in most of the DAT561-infected larvae. In some larvae, luminal surfaces of the PM were more strongly stained than the inside. These results suggest that infection of CC12 strain in honeybee larvae is essentially confined to the intestine. Moreover, our results imply the presence of M. plutonius-derived substances diffusing into the larval tissues in the course of infection.     

Terrestrial prey inputs to streams bordered by deciduous broadleaved forests,conifer plantations and clear‐cut sites in southwestern Japan: effects on the abundance of red‐spotted masu salmon

Terrestrial invertebrates falling from the riparian canopy are a major energy source for fishes in headwater streams. Because quantity and quality of such allochthonous resources can vary depending on riparian conditions, conversion of riparian forests to conifer plantations may affect stream productivity. We compared falling and drifting invertebrate abundances and the diet of masu salmon (Oncorhynchus masou) among stream reaches bordered by deciduous broadleaved forests, conifer plantations (Cryptomeria japonica), and clear‐cut sites in southwestern Japan. We also examined whether among‐reach variation in salmon abundance was related to the riparian vegetation types. The results indicated that, on an annual basis, falling inputs of terrestrial invertebrates at the broadleaved reaches were 2–4 times higher than those at the plantation and clear‐cut reaches. In nonwinter seasons, terrestrial invertebrates made up 40–60% and 30–90% of drift and masu salmon diets, respectively, and drifting invertebrate abundance was higher in the broadleaved reaches than in the plantation reaches. Furthermore, a multivariate analysis of salmon abundance showed that variation in salmon biomass was explained primarily by riparian vegetation type, with broadleaved and clear‐cut reaches having higher biomass than the plantation reaches. These results indicate that terrestrial invertebrates are an important resource for masu salmon, and suggest that streams bordered by conifer plantations receive lower terrestrial prey inputs, which results in lower salmon abundance. In regions where natural forests have been extensively converted to conifer plantations, forest management that allows and facilitates recovery of natural riparian stands is important.     

New role for glyoxylate cycle enzymes in wood-rotting basidiomycetes in relation to biosynthesis of oxalic acid

The key enzymes of the glyoxylate cycle, isocitrate lyase (ICL) and malate synthase (MS), were detected in varying amounts in the mycelia of the woodrotting basidiomycetes tested, although they were grown in a glucose-rich medium. The highest specific activities of ICL (0.37 U/mg protein) and MS (0.63 U/mg protein) were measured for the brown-rot basidiomycetesLaetiporus sulphureus andFomitopsis palustris, respectively. The results indicate that the glyoxylate cycle enzymes occur in wood-rotting basidiomycetes as the seemingly constitutive enzymes at varying levels. The glyoxylate cycle enzymes, including malate dehydrogenase (MDH), and the oxalate-producing enzymes glyoxylate dehydrogenase (GDH) and oxaloacetase (OXA) were found to have good correlation with biosynthesis of oxalic acid and fungal growth, which was also confirmed by use of an ICL inhibitor. A new role for the glyoxylate cycle is discussed in relation to oxalic acid biosynthesis in wood-rotting basidiomycetes.This paper was presented in the 50th annual meeting of the Japan Wood Research Society, Kyoto, April 2000     

Survey and enzymatic formation of lignans ofAnthriscus sylvestris

Gas chromatography — mass spectrometry analysis of the -glucosidase-treated MeOH extracts ofAnthriscus sylvestris showed, based on comparison of the mass spectra and retention times with those of authentic samples, the presence of lignans, yatein, secoisolariciresinol, lariciresinol, matairesinol, hinokinin, and pluviatolide. The existence of small amounts of bursehernin was suggested by mass chromatography. In addition, nemerosin and deoxypodophyllotoxin were tentatively identified by comparing the mass spectra with those reported in the literature. Enzyme preparations fromA. sylvestris catalyzed the formation of secoisolariciresinol and lariciresinol from coniferyl alcohol. Furthermore, the enzyme preparation catalyzed the formation of lariciresinol from (±)-pinoresinols and the formation of secoisolariciresinol from (±)-lariciresinols. Thus, pinoresinol/lariciresinol reductase (PLR) activity was detected. Chiral high-performance liquid chromatography analysis showed selective formation of (+)-lariciresinol and (–)-secoisolariciresinol from (±)pinoresinols with theA. sylvestris PLR preparation, indicating that the stereochemical property ofA. sylvestris PLR-catalyzed reduction was similar to those ofForsythia PLR andArctium lappa ripening seed PLR.Part of this report was presented at the 43rd Lignin Symposium, Fuchu, October 1998     

Lignans ofChamaecyparis obtusa

Heartwood ofChamaecyparis obtusa contains significant amounts of a dibenzylbutyrolactone lignan, hinokinin (8). This investigation demonstrated that the contents of 8 and a norlignan, hinokiresinol (12), were higher in the heartwood region than in the sapwood, indicating their nature of being heartwood extractives. Eleven lignans — xanthoxylol (1), 7-oxohinokinin (2), savinin (3), dihydrosesamin (4), isoactifolin (5), sesamin (6), piperitol (7), hinokinin (8), pluviatolide (9), haplomyrfolin (10), and rnatairesinol (11) — were isolated from young shoots ofChamaecyparis obtusa cv. Breviramea. Eight lignans (1, 2, 4, 5, 7, 9,10, and11) were isolated from this plant for the first time. Chiral high-performance liquid Chromatographie analysis showed that8, 9, 10, and11, were found to be levorotatory and optically pure (>99% e.e.). Based on the chemical structures of the isolated lignans, possible biosynthetic pathways of8 are discussed.Parts of this report were presented at the 44th annual meeting of the Japan Wood Research Society, Nara, April 1994; the 46th annual meeting of the Japan Wood Research Society, Kumamoto, April 1996; and the 44th Lignin Symposium, Gifu, October 1999     

Profiling of hepatic gene expression of mice fed with edible japanese mushrooms by DNA microarray analysis: comparison among Pleurotus ostreatus, Grifola frondosa, and Hypsizigus marmoreus

To compare and estimate the effects of dietary intake of three kinds of mushrooms (Pleurotus ostreatus, Grifola frondosa, and Hypsizigus marmoreus), mice were fed a diet containing 10-14% of each mushroom for 4 weeks. Triacylglycerol in the liver and plasma decreased and plasma cholesterol increased in the P. ostreatus-fed group compared with those in the control group. Cholesterol in the liver was lower in the G. frondosa-fed group than in the control group, but no changes were found in the H. marmoreus-fed group. DNA microarray analysis of the liver revealed differences of gene expression patterns among mushrooms. Ctp1a and Fabp families were upregulated in the P. ostreatus-fed group, which were considered to promote lipid transport and β-oxidation. In the G. frondosa-fed group, not only the gene involved in signal transduction of innate immunity via TLR3 and interferon but also virus resistance genes, such as Mx1, Rsad2, and Oas1, were upregulated.     

Bone marrow stromal cells generate muscle cells and repair muscle degeneration

Bone marrow stromal cells (MSCs) have great potential as therapeutic agents. We report a method for inducing skeletal muscle lineage cells from human and rat general adherent MSCs with an efficiency of 89%. Induced cells differentiated into muscle fibers upon transplantation into degenerated muscles of rats and mdx-nude mice. The induced population contained Pax7-positive cells that contributed to subsequent regeneration of muscle upon repetitive damage without additional transplantation of cells. These MSCs represent a more ready supply of myogenic cells than do the rare myogenic stem cells normally found in muscle and bone marrow.     

Estimates of genotypic and yearly variations on fruit quality and functional traits for tetraploid table grape breeding

We estimated genetic and environmental variance components for fruit quality traits (cluster weight, berry weight, soluble solids content, total sugar content, sugar composition, free acid content, total amino acid content, amino acid composition), and a functional trait (gamma-aminobutyric acid (GABA) content) of three commercial tetraploid table grape cultivars (Kyoho, Pione, Suiho) grown in Japan over three successive years. ANOVA showed that the effect of genotype was significant (P < 0.05) for all traits except soluble solids content and total sugar content. The effect of the year was significant for all traits except amino acid composition (γ ratio) and GABA content. The variance of genotype (σ _g ²) was highest for γ ratio (63.6%), high in sugar composition (α ratio, 60.3%) and GABA content (58.8%), and negligible for soluble solids content (0.0%) and total sugar content (0.0%). The variance of among years (σ _y ²) was very high in soluble solids content (82.8%), high in total sugar content (60.2%) and negligibly small in γ ratio (8.7%) and GABA content (6.4%). Some effects of genotype × year interaction, among vines within genotypes, and vine × year interaction were statistically significant, but these components were negligibly small. Except for soluble solids content and total sugar content, significant varietal differences in the traits were recorded. Variance estimates for each trait provides useful information for optimal yearly repetition to effectively discriminate genetic differences in fruit quality and functional traits for improving future tetraploid table grape breeding programs.     

Further Studies on the Hypoglossal Nucleus in the Fowl

The delineation of the fowl's hypoglossal nucleus was determined by severing the hypoglossal nerve or its branches and observing retrograde degeneration. Birds were killed 7 or 10 days after surgery, the brains embedded in paraffin and cut at 10 μm. Transection of the hypoglossal nerve stem resulted in chromatolysis near the obex and no chromatolysis was found in the Nuclei cervicales. Severing of the laryngolingual branch led to degeneration phenomena near the obex nearly similar to the degeneration observed after severing of the hypoglossal nervestem. Cutting of the superior laryngeal branch caused a degeneration of cells caudal to the obex, and cutting of the syringeal branch resulted in chromatolysis in about the same area. The origin of the anastomosing branch between the first cervical and hypoglossal nerve was not found because no chromatolytic cells could be detected. Similarly the cutting of several upper cervical nerves did not lead to chromatolysis in the areas we investigated.