Changes in microbial biomass after continuous application of azolla and rice straw in soil

A model experiment was conducted under tropical conditions with a view to evaluating the changes in microbial biomass and nutrient dynamics in upland soil through the continuous application of azolla and rice straw (2 g C kg^-1 soil per each application). Flush decomposition of C was observed immediately after each application and the rate of mineralization did not change appreciably during this period. After flush decomposition, the rate of C mineralization from azolla was higher than that from rice straw until 9 to 13 weeks after each application and thereafter the mineralization rate was similar. The amount of inorganic N released from azolla increased following each application, whereas inorganic N in rice straw plot was immediately immobilized and the rate of immobilization increased until the 3rd application and did not increase further after the 4th application. The amounts of biomass C and N increased immediately after residue incorporation, reached the maximum level one week after each application and declined thereafter. Maximum biomass formation increased until the 2nd application and then the level remained constant. Maximum biomass N formation was higher in azolla than in rice straw after the 1st application, but after repeated applications, the difference became less pronounced: Continuous increase in biomass in a certain week after each application was observed, probably because of the cumulative effects of the previous applications. The increase suggests that continuous application of organic materials may enable to improve the amount of soil microbial biomass.     

Application of rRNA-based probe for detection of a bacterium in rhizoplane

Recently, in situ hybridization technique targeted to ribosomal RNA (Delong et al. 1989) has been widely used to detect specific microorganisms in various environments such as marine environment (Nishimura 1995), sediments (Spring et al. 1993), inside of organisms (Amann et al. 1991), and soil (Hahn et al. 1992). This technique provides information not only on microscale localization of the microorganisms but also on their metabolic activities in situ. So far, detection by in situ hybridization targeted to ribosomes in soil has been scarcely reported because two problems arise when this method is applied to soil. The first is that cells in natural soils cannot be easily stained because the cellular ribosome content is low in an oligotrophic soil environment. The second is that non-specific binding of the probes to soil particles and autofluorescence of soil components such as organic matter and mineral particles interfere with the signal from the probe specifically hybridized.     

Estimation of mycorrhizal colonization of the roots of oak seedlings inoculated with an ectomycorrhizal fungus,Laccaria amethystea

Six-month-old seedlings of Quercus serrata and Quercus glauca in a nursery were inoculated with the ectomycorrhizal fungus Laccaria amethystea encapsulated in alginate gel and grown in the nursery. The seedlings were collected at 1, 3, and 5 months after the inoculation and examined for colonization of the root system with ectomycorrhizal fungi. The roots within 5 months after the inoculation showed rudimentary ectomycorrhizal colonization. The level of colonization of the root system was estimated based on the intensity of hyphal covering on the root tips by staining with a fluorescent dye and expressed as an index of mycorrhizal colonization (IMC). IMC increased with the time after inoculation and reached values of 4 and 12% in Q. serrata and Q. glauca, respectively at 5 months after the inoculation. The determination of IMC may enable to assess the development of mycorrhizal colonization of the root system that shows rudimentary ectomycorrhizas after the inoculation.     

VALIDATION OF NEW BITE BLOCK‐TYPE HEAD‐IMMOBILIZATION DEVICES FOR RADIOTHERAPY IN DOGS

An ideal head‐immobilization method provides a high level of accuracy and reproducibility in the immobilization. Various head‐immobilization methods for radiotherapy have been published and are excellent in terms of accuracy; however, these methods are complicated to use, and labor intensive. The present study describes two new bite block‐type head‐immobilization devices designed for higher stability and lower vertical variation. The device designed in our previous study (the bite block‐type head‐immobilization device; Device A) was modified by making a groove on the top the horizontal plate (Device B) for a stable ventral‐dorsal position, or beneath the horizontal plate (Device C) for a stable dorsal‐ventral position. The three devices were objectively compared with respect to setup time, and accuracy of the computed tomography scan images by two authors independently. Five male healthy beagles were used in this study. For each device, the setup time and the variation in the coordinates were measured five times for each dog. The mean setup times for Devices A, B, and C were 3.3, 1.5, and 2.4 min, respectively, showing the groove modifications were able to reduce the setup time (in device B, by at least 50%). Moreover, three‐dimensional analysis of the computed tomography images revealed that the measurement variability of Device A (1.6 ± 1.0 mm) was significantly higher than that of Device C (0.7 ± 0.4 mm; P < 0.001). Collectively, our results show that use of a bite block‐type head‐immobilization device with a groove improves the setup time and head‐immobilization accuracy.     

An inoculation experiment of Japanese <Emphasis Type="Italic">Bursaphelenchus</Emphasis> nematodes on Japanese black and red pine, <Emphasis Type="Italic">Pinus thunbergii</Emphasis> and <Emphasis Type="Italic">P. densiflora</Emphasis>

The pinewood nematode (PWN) Bursaphelenchus xylophilus is an invasive pathogen that was introduced from North America to Asian countries and Portugal and is devastating native pine forests. Some native European and Asian Bursaphelenchus nematodes also have weak to moderate pathogenicity to native pine species. To evaluate the potential risk of native Bursaphelenchus species, we inoculated ten Japanese Bursaphelenchus species into native pine species (the dominant forest species) in Japan, and evaluated their pathogenicity using mortality and tracheal tissue damage as indices. Inoculation was conducted on August 3, 2007, and the symptoms were observed every 2 weeks until February 1, 2008. None of the inoculated trees, excluding the pathogenic PWN inoculated control, showed external disease symptoms; however, four species [a less pathogenic PWN isolate, B. luxuriosae, Bursaphelenchus sp. NK215 (undescribed), and NK224 (undescribed)] caused tracheal tissue damage in inoculated seedlings and showed weak pathogenicity. Therefore, we conclude that there are some potentially pathogenic native species of nematodes distributed in Japan. Interestingly, two of these weakly pathogenic species, B. luxuriosae and NK215, are not associated with Pinaceae trees, suggesting that nematode pathogenicity may be a pre-adaptive character. More experimental studies under different conditions are necessary to accurately evaluate the potential risk of these pathogens.     

Two-dimensional 1H-13C nuclear magnetic resonance (NMR)-based comprehensive analysis of roasted coffee bean extract

Coffee was characterized by proton and carbon nuclear magnetic resonance (NMR) spectroscopy. To identify the coffee components, a detailed and approximately 90% signal assignment was carried out using various two-dimensional NMR spectra and a spiking method, in which authentic compounds were added to the roasted coffee bean extract (RCBE) sample. A total of 24 coffee components, including 5 polysaccharide units, 3 stereoisomers of chlorogenic acids, and 2 stereoisomers of quinic acids, were identified with the NMR spectra of RCBE. On the basis of the signal assignment, state analyses were further launched for the metal ion-citrate complexes and caffeine-chlorogenate complexes. On the basis of the signal integration, the coffee components were successfully quantified. This NMR methodology yielded detailed information on RCBE using only a single observation and provides a systemic approach for the analysis of other complex mixtures.     

Polyphenol composition of a functional fermented tea obtained by tea-rolling processing of green tea and loquat leaves

Phenolic constituents of a new functional fermented tea produced by tea-rolling processing of a mixture (9:1) of tea leaves and loquat leaves were examined in detail. The similarity of the phenolic composition to that of black tea was indicated by high-performance liquid chromatography comparison with other tea products. Twenty-five compounds, including three new catechin oxidation products, were isolated, and the structures of the new compounds were determined to be (2R)-2-hydroxy-3-(2,4,6-trihydroxyphenyl)-1-(3,4,5-trihydroxyphenyl)-1-propanone 2-O-gallate, dehydrotheasinensin H, and acetonyl theacitrin A by spectroscopic methods. In addition, theacitrinin A and theasinensin H were obtained for the first time from commercial tea products. Isolation of these new and known compounds confirms that reactions previously demonstrated by in vitro model experiments actually occur when fresh tea leaves are mechanically distorted and bruised during the production process.     

Isolation and Caenorhabditis elegans lifespan assay of flavonoids from onion

The main flavonoids were isolated from three selected onion cultivars. Three phenolic compounds were obtained by reverse-phase HPLC, and their structures were elucidated by multiple NMR measurements. There were two known compounds, quercetin and quercetin 3'-O-β-D-glucopyranoside (Q3'G), and one novel compound, quercetin 3-O-β-D-glucopyranoside-(4→1)-β-d-glucopyranoside (Q3M), which was identified in onion for the first time. These flavonoids were found to be more abundant in the onion peel than in the flesh or core. Their antioxidative activities were tested using the DPPH method, and their antiaging activities were evaluated using a Caenorhabditis elegans lifespan assay. No direct correlation was found between antioxidative activity and antiaging activity. Quercetin showed the highest antioxidative activity, whereas Q3M showed the strongest antiaging activity among these flavonoids, which might be related to its high hydrophilicity.     

One-week antihypertensive effect of Ile-Gln-Pro in spontaneously hypertensive rats

The antihypertensive effect of an angiotensin I-converting enzyme (ACE) inhibitory peptide Ile-Gln-Pro (IQP), whose sequence was derived from Spirulina platensis , was investigated in spontaneously hypertensive rats (SHRs) for 1 week. The weighted systolic blood pressure (SBP) and diastolic blood pressure (DBP) of the peptide IQP-treated group were significantly lower than those of the negative control group from the third and fourth days, respectively. Accompanying the blood pressure reduction, a significant regulation of the expression of major components of the renin-angiotensin system (RAS) was found in the treatment group, including downregulation of the mRNA levels of renin, ACE, and the angiotensin II type 1 (AT1) receptor in the kidney, as well as serum angiotensinogen (Ang), ACE, and angiotensin II (Ang II) concentrations. The treatment group also showed upregulation of mRNA expression of the angiotensin II type 2 (AT2) receptor in the kidney. Our findings suggested that IQP might be of potential use in the treatment of hypertension.