Phylogenetic relationships among natural populations of perennial buckwheat, Fagopyrum cymosum Meisn., revealed by allozyme variation

Twenty natural populations of F. cymosum, an insect-pollinated allogamous wild species of Fagopyrum, were investigated for their chromosome number and allozyme variation at 10 Ioci encoding 8 enzymes. Diploid populations were obtained in Sichuan, Yunnan and Tibet in China, whereas tetraploid populations were obtained in Tibet, the Himalayan hills, Thailand and southern China. Both diploid and tetraploid populations maintained a large amount of allozyme variation. The average heterozygosity, H_e, ranged from 0.045 to 0.389 (0.213 on average). Tetraploid populations showed higher H_e values than diploid populations. Natural populations of F. cymosum were locally well differentiated (G_ST = 0.322), probably due to reproductive isolation between the two ploidy levels and to distribution over wide areas from southern China to the Himalayan hills. The phylogenetic tree constructed by the neighbor-joining method based on allozyme variation clarified two distinct groups of diploid populations, the Sichuan and Yunnan groups. As for the tetraploid populations, polyploidization occurred twice independently, once in Yunnan and once in eastern Tibet. Tetraploid F. cymosum diffused to warmer areas such as southern Yunnan and Thailand as well as to cooler areas of Tibet and the high Himalayan hills and became the most widely distributed of the Fagopyrum species.     

Immunolocalization of <Emphasis Type="Italic">Pepper mild mottle virus</Emphasis> in developing seeds and seedlings of <Emphasis Type="Italic">Capsicum annuum</Emphasis>

The location of Pepper mild mottle virus (PMMoV) within seeds as they developed on inoculated seedlings of pepper (Capsicum annuum) was followed over time by detecting the viral coat protein using immunofluorescence microscopy. Seedlings were inoculated with PMMoV when the flower buds on the first and second branching nodes were in bloom. Fluorescence indicating the presence of PMMoV was first observed around immature seeds and placentas in the ovaries on the fourth branching node at 20 days post-anthesis (20 DPA), which corresponded to 39 days post-inoculation (39 DPI). The area with fluorescence gradually expanded from the placenta into the integument and the parenchyma, and finally reached the tip of the immature seeds by 34 DPA (53 DPI). The embryo or endosperm beyond the endothelium never fluoresced during the experiment [i.e., ending at 81 DPA (102 DPI)]. For visualizing viral routes of invasion from seeds into new seedlings, PMMoV-infected C. annuum seeds that were heterozygous for the L ³ tobamovirus-resistance gene were sown in soil at 30°C. After ~2 weeks, the cotyledon developed virally induced necrosis. These findings shed light on the infection cycle of PMMoV through vertical transmission in C. annuum.     

Comparison of Iron Availability in Leaves of Barley and Rice

Iron (Fe) is an essential trace element in all eukaryotes. In higher plants, Fe deficiency causes interveinal chlorosis in young leaves. However, in barley and rice, both of which are "Strategy II" plants, the degree and the pattern of Fe-deficiency symptoms differ. In the present study, barley and rice plants were grown in the same container, i.e., by "coculturing," to compensate for the amount of mugineic acids in rice in the nutrient solution. We examined the differential availability of Fe for distribution and retranslocation in shoots between barley and rice without considering the difference in the iron acquisition ability, which is affected by the differential mugineic acid secretion between barley and rice. Although the Fe concentration of young barley leaves had decreased under the coculture conditions, the SPAD value was similar to that in monocultured barley. In contrast, although there was an increase in the Fe concentration of the young leaves of cocultured rice, the SPAD value decreased, as in the case of monocultured rice. Rice accumulated Fe in old leaves, whereas in barley Fe was efficiently distributed to young leaves. Therefore, the SPAD value of the second leaf in rice remained constantly high. The Fe concentration of the second leaf in barley decreased under Fe-deficient coculture conditions, the SPAD value decreased and the senescence of the second leaf become accelerated. ⁵⁹Fe pulse-labeling experiments suggested that in barley Fe was more efficiently retranslocated from old leaves to young leaves than that in rice. As a result, the level of Fe present in the fraction with a molecular weight lower than the 10,000/water-soluble Fe ratio was higher in the old leaves of barley than in the old leaves of rice under Fe-deficient conditions. Based on the results obtained, we suggest that the distribution and retranslocation characteristics of internal Fe in barley may be well adapted to Fe deficiency.     

Large-scale survey of frequency of forest walking and related factors in a Japanese population inhabiting a large city, and comparison of an urban area and a rural area

The frequency of forest walking among the general population is a major topic of study in forest science. The objectives of this study were threefold: to assess the frequency of forest walking among Japanese residents of Nagoya, a large city; to evaluate differences among frequency of forest walking by participants from a large city, an urban area (Shizuoka; 4,666 participants), and a rural area (Yakumo Town, Hokkaido; 397 participants) in previous studies; and to examine factors related to frequency of forest walking. The survey, by self-administered questionnaire, in the major city of Nagoya was conducted between June 2008 and May 2010. In all, 5,158 participants (M/F, 1,466/3,692; mean age ± SD [range], 52.5 ± 10.3 [35–69] years) were included in the analysis. The proportions of frequency of forest walking ≥ once/month and ≥ once/year were 10.9 % (M/F, 15.1/9.3 %) and 46.1 % (51.0/44.1 %), respectively. In multiple logistic regression analysis, significant differences were noted among study sites for the adjusted odds ratio for frequency of forest walking. Overall, the order of highest to lowest frequency of forest walking was Shizuoka > Nagoya > Yakumo. Factors related to frequency of forest walking were common among the three study sites. Higher frequency of forest walking was associated with male sex and older age; the most relevant factor related to frequency of forest walking was its enjoyment level. Further studies will be required to clarify why these factors are related to frequency of forest walking.     

Mediator-type biosensor for real-time wireless monitoring of blood glucose concentrations in fish

We developed a wireless mediator-type biosensor system to rapidly monitor blood glucose concentrations in free-swimming fish. Glucose oxidase was used to determine the glucose concentration, but fluctuations in the oxygen concentrations in the samples affected the measurements. In this work, we used a mediator, a synthetic electron acceptor, as a substitute in the oxidation–reduction reaction, thus ensuring that changes in the concentration of oxygen do not affect the measurements of glucose concentration performed by the sensor. We especially focused on the use of ferrocene as a mediator. The resulting enzyme sensor utilized a Pt–Ir wire (diameter 0.178 mm) as the working electrode and Ag/AgCl paste as the reference electrode. Chitosan was used to fix ferrocene and glucose oxidase to the working electrode. The characteristics of the sensor were tested, and it was found to be capable of measuring glucose independent of the oxygen concentration of the sample. Its reproducibility was also tested, and the results showed that the sensor was suitable for performing measurements in vivo. The sensor was then inserted into to eyeball interstitial sclera fluid in order to wirelessly monitor the glucose concentration in free-swimming fish. Measurements were taken for a total of 48 h. During these measurements, artificial stress was applied to the fish. The glucose concentrations of fish rise with their stress levels. The output current from the sensor gradually increased during the application of stress, which hinted that the stress was monitored by this system.     

Nickel-catalyzed carbonization of wood for coproduction of functional carbon and fluid fuels II: improved fuel quality of oil fraction and increased heating value of gas fraction

The yields and properties of oil and gas fractions coproduced during carbonization of larch wood loaded with Ni 2%, Ni 2%+Ca 1%, and Ni 4% and without catalyst (None) at 700°–900°C were examined to clarify the catalytic effect in terms of conversion into fluid fuels. The net calorific value of oil occurred mainly below 500°C and increased in the order None < Ni 2% < Ni 4% < Ni 2%+Ca 1%, while the yield decreased in this order. The same order held for the production of gases enriched with hydrogen at 500°–700°C. Even above 800°C, markedly promoted evolution of hydrogen took place for all catalyst systems. These observations confirmed the effectiveness of nickel-catalyzed carbonization at 900°C, particularly Ni 2%+Ca 1%, for both upgrading of oil and gaseous fractions, although the quality of oil was not satisfactory. The catalysis of nickel with and without calcium is discussed on the basis of the modified Broid-Shafizadeh scheme, and the scheme was altered to adapt to the high temperature region where oil was no longer produced. Part of this study was presented at the 14th Annual Meeting of the Japan Institute of Energy, Suita, August 2005, and at the 18th Symposium, Session D, of the Materials Research Society of Japan, Tokyo, December 2007     

PCR-based assays to detect and quantify <Emphasis Type="Italic">Phomopsis sclerotioides</Emphasis> in plants and soil

We developed polymerase chain reaction (PCR) assays to detect and quantify Phomopsis sclerotioides, the causal agent of black root rot of cucurbits. We used internal transcribed spacers 1 and 2 of the ribosomal DNA (rDNA) from representative isolates to search for target sequences. Primer pairs were selected after testing against 40 fungal isolates including 13 Ph. sclerotioides isolates, 9 Phomopsis isolates other than Ph. sclerotioides, and 18 soilborne fungi that were either pathogenic or nonpathogenic to cucurbits. Conventional PCR assays with the primer pair of CPs-1 (forward) and CPs-2 (reverse) produced target DNA amplicons from all Ph. sclerotioides isolates but none of the other isolates tested. From soil and root samples collected from fields naturally infested with black root rot of cucumber and melon, the CPs-1/CPs-2 primer pair successfully amplified target DNA fragments in conventional PCR assays. Moreover, we applied the CPs-1/CPs-2 primer pair in a real-time PCR assay with SYBR Green I, and PCR-amplified products were successfully quantified by reference to a standard curve generated by adding known amounts of target DNA. Target Ph. sclerotioides DNA fragments were similarly detected in artificially inoculated roots of cucumber, melon, pumpkin, and watermelon, but quantities of Ph. sclerotioides DNA in their hypocotyls of the hosts varied as follows: melon ≥ cucumber ≥ watermelon > pumpkin. These results suggest that Ph. sclerotioides infection is not species-specific but the rate of infection may differ among host species.     

Suppressing effect of neocarrabiose 4-O-sulfate on thermal and freeze denaturation of myosin subfragment-1 and myofibrils

Suppressive effects of neocarrabiose 4-O-sulfate on the thermal and freeze denaturation of myosin subfragment-1 (S-1) and on myofibrils were investigated. The compound strongly suppressed the thermal denaturation of S-1. Its suppressive effect was greater than that of sorbitol and similar to that of maltose. However, it tended to accelerate the denaturation of myofibrils, suggesting a loss of protection by F-actin upon its addition. The compound suppressed the freeze denaturation of myofibrils and S-1. The effect was similar to that of sorbitol or maltose, and completely different from that of Na-sulfate. The compound solubilized myofibrils at concentrations similar to KCl. Therefore, it was concluded that neocarrabiose 4-O-sulfate behaved as an ionic salt in the thermal treatment process, whereas it behaved as a sugar in the frozen storage process.     

Kinetics of bacterial death by heated dolomite powder slurry

The kinetics of the bactericidal action of dolomite powders heated at 600-1000 °C against Escherichia coli and Staphylococcus aureus were investigated. Dolomite powder heated to at least 700 °C exhibited bactericidal action, and the process of bacterial death in the heated dolomite powder slurries followed first-order reaction kinetics. The value of the death rate constant (k) increased with dolomite powder concentration, and the dilution coefficient (n), which indicates the dependence of k on the reagent concentration, was measured. The n values of the powder heated at 700 °C and at temperatures >900 °C were almost identical to those of MgO and CaO, respectively. This suggests that the first emergence of bactericidal action at 700 °C corresponds to generation of MgO while that at temperatures >900 °C is due to generation of CaO. The slurry temperature significantly affects the bactericidal action. The slope of the Arrhenius plot of k for E. coli and S. aureus grown at 37 °C exhibited a discontinuous point at approximately 22 °C, where a change in the value of activation energy for bacterial death occurred. This temperature corresponds to that of the phase transition of cell membrane lipids.     

Prominent expression of spinocerebellar ataxia type-1 (SCA1) gene encoding ataxin-1 in LH-producing cells, LbetaT2

The LH-producing cell line, LbetaT2, and non LH-producing cell line, alphaT3-1 cells, established from a pituitary tumor, were employed for cDNA subtraction cloning to identify genes with expression unique to LH producing cells. Several cDNAs that code for known as well as for many unidentified clones were discovered. Most clones were the spinocerebellar ataxia type-1 (SCA1) gene encoding ataxin-1, the abnormality of which causes neurodegeneration and loss of cerebellar Purkinje cells. We examined whether the expression of SCA1 gene in LbetaT2 cells is related to hormone production. We also compared the expression of SCA1 with that in various other pituitary tumor derived cell lines, and confirmed the prominent expression of SCA1 in LbetaT2 cells. The effect of gonadal factor(s) for SCA1 gene expression was examined. The expression level in female rats was low and did not change during the estrus cycle, but increased significantly after ovariectomy and did not return to the normal level under low and high doses of estrogen. In the male pituitary SCA1 gene expression increased markedly after castration and was not decreased by estrogen or testosterone. The Ontogeny of SCA1 gene expression was investigated in porcine fetal and postnatal pituitaries and revealed biphasic and sexually dimorphic expression. Transient expression of SCA1 gene was observed at fetal day 50 and 65 in males and day 40 in females, followed by a decline and increased expression before birth in both genders. Thus the expression of SCA1 gene is prominent in LH-producing cells and is not under direct control of gonadal factor(s) in both genders. In addition to the variable expression of SCA1 gene during the fetus period, the present results provide a novel aspect to the understanding of Boucher-Neuhauser syndrome (Ataxia Hypogonadism Choroidal Dystrophy).