The chemical nature of the green fraction of P type humic acid

P type humic acid has absorption bands near 615, 570 and 450mµ in the visible region, and is a common component of many soils in north temperate regions (1, 2, 3). Its unusual spectral characteristics are due to a green fraction (Pg), which can be separated from the humic acid dissolved in aqueous alkali by chromatography on Sephadex or cellulose powder, and is precipitated as a reddish-brown substance on acidification (2). In aqueous alkali, it has λmax at 613, 568, 448, 430 (infl.) and 281mµ. The distribution of Pg in different soils is probably related to the presence of fungal sclerotia. The highest content of Pg in p type humic acid is about 10%. We have now obtained evidence that the characteristic colour of Pg is due to the presence of a derivative of 4,9 - dihydroxyperylene - 3,10 - quinone.     

Amino acid metabolism in plant leaf

¹⁴C-labelled sodium bicarbonate and ¹⁵N-labelled ammonium sulfate were simultaneously vacuum-infiltrated into detached sunflower leaves, and the incorporation of ¹⁴C and ¹⁵N into free amino acids was chased during 60-min period in the light and in the dark.

In the light, the ue specific activity of aspartic acid, alanine, serine and glycine rapidly increased for 5 min and thereafter decreased. On the other hand, that of gultamic acid continued to increase slowly during the entire 60-min period. In the dark, aspartic acid most actively incorporated ¹⁴C. The difference of changes in ¹⁴C specific activity between glutamic acid and other amino acids was also observed in the dark as in the light. These results suggest that the carbon skeleton of glutamic acid is synthesized from aspartic acid, alanine, serine and glycine.

¹⁵N content of glutamine was the highest of all amino acids investigated in the light, and it was followed by glutamic acid. alanine, aspartic acid, serine and glycine, in this order. In the dark, ¹⁵N content of glutamic acid fell remarkably and was lower than that of alanine up to 5 min. From these ¹⁵N tracer experiments, it is suggested that the incorporation of ammonium into glutamic acid is strictly dependent on light and that alanine incorporates ammonium by the direct amination besides the transamination from glutamic acid.     

The chemical nature of components of PG (Green fraction of P type humic acid)

P type humic acids showing characteristic absorption maxima at 615, 575, and 450 nm have been found in various types of soils around the world (3, 5, 7,8). P type humic acid was fractionated into a brown fraction (Pb) and a green fraction (Pg) by column chromatography using cellulose powder (4) or Sephadex gel (3). The Pg fraction which has a strong absorption maxima at 615, 575, and 450 nm causes the characteristic absorption pattern of P type humic acid.     

Root-shoot interaction as a limiting factor of biomass productivity in new tropical rice lines

The objective of the present study was to identify the limiting factors in biomass productivity of new tropical rice lines (panicle weight type) by analyzing the relationship between root and shoot activity. Five field experiments using three new lines, IR65598-112-2, IR65600-42-5-2, and IR68544-29-2-1-3-1-2, and one of the highest-yielding indica rice varieties (panicle number type), IR72, were conducted at the International Rice Research Institute from 1997 to 2002. Specific absorption rate of N (SARN , mg N g^-1 root dry weight d^-1), specific stem sap exudation rate (sap exudation rate, g exudate g^-1 root dry weight 12 h^-1) and specific root respiratory rate (root respiratory rate, μmol O₂ g^-1 root dry weight h^-1) were determined as indices of root activity. Relative growth rate (RGR, mg g^-1 d^-1) was used as an index of shoot activity. Compared with IR72, the new lines showed lower RGR and SAR_N values during the early growth stages. In contrast, during the late growth stages, these activities were higher in the new lines than in IR72. The SAR_B and sap exudation rate showed a linear correlation with RGR at successive growth stages, regardless of the genotypes and growth conditions. These findings indicated that active root-shoot interaction was maintained throughout the growth period in high-yielding tropical rice, including the new lines and IR72. Therefore, it was considered that SAR_N and the sap exudation rate are useful indices of root activity regulating RGR in the new lines. However, the root respiratory rate could not be used for selection, because the relationship between RGR and the root respiratory rate changed with the growth conditions. Our findings support the hypothesis that root and shoot activities during maturation are important factors affecting the productivity of the new lines, which have not yet been able to attain the targeted yield.     

The Negative Relationship between Fouling Organisms and the Content of Eicosapentaenoic Acid and Docosahexaenoic Acid in Cultivated Pacific Oysters,Crassostrea gigas

Bivalves serve as an important aquaculture product, as they are the source of essential fatty acids, such as eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), in our diet. However, their cultivation in the wild can be affected by fouling organisms that, in turn, affect their EPA and DHA content. The effects of fouling organisms on the EPA and DHA contents of cultivated bivalves have not been well documented. We examined the effects of fouling organisms on the EPA and DHA contents and condition index of cultured oysters, Crassostrea gigas, in an aquaculture system. We sampled two-year-old oysters from five sites in Shizugawa Bay, Japan, in August 2014. Most of the fouling organisms were sponges, macroalgae, and Mytilus galloprovincialis. A significant negative relationship existed between the DHA content in C. gigas and the presence of sponges and macroalgae. A lower C. gigas EPA content corresponded to a higher M. galloprovincialis fouling mass and a lower C. gigas condition index. This can be explained by dietary competition between C. gigas and M. galloprovincialis for diatoms, which were the main producer of EPA in our study sites. Our findings indicate that fouling organisms likely reduce the EPA and DHA content in cultivated oysters. Therefore, our results suggest that the current efforts to remove fouling organisms from oyster clusters is an effective strategy to enhance the content of EPA and DHA in oysters.     

Seasonal variations in the incidence of pine wilt and infestation by its vector, Monochamus alternatus, near the northern limit of the disease in Japan

During the last few decades, pine wilt disease has spread to cool-climate regions in Japan and, more recently, the potential risk of it spreading into the European midwest has also become a concern. In a coastal pine stand (84.7?ha) in Akita, near the northern limit of pine wilt disease in Japan, we investigated seasonal variations in the incidence of damage caused by the disease to trees and oviposition by the disease’s insect vector, Monochamus alternatus, during a two-year period. Foliage discoloration occurred throughout each year, and its seasonal variation showed a bimodal pattern in Pinus thunbergii (a higher peak in May–June and a smaller peak in October) and a clear peak in June in P. densiflora, which differed from the patterns in seasonal variation seen for warm-climate regions. Oviposition scars by M. alternatus were found in 40–45% of the trees damaged each year. The percentage of trees that had oviposition scars was higher in P. thunbergii than in P. densiflora. This appeared to reflect the difference in seasonal discoloration pattern between the two species. Analysis of the oviposition risk showed that trees that exhibited discoloration starting between July and October had a significantly higher risk or significantly higher oviposition scar densities, particularly for those that became discolored between August and September (2.5–14.6-fold higher risk than during other months). Oviposition scar densities per damaged tree were similar within the period of higher oviposition risk. Considering both oviposition risks and scar densities, we concluded that trees with discoloration that become apparent between July and October are important targets for preventing the spread of pine wilt disease in Akita.     

Trapped neutrophil syndrome in a Border Collie dog: clinical, clinico-pathologic, and molecular findings

Trapped neutrophil syndrome (TNS) is an autosomal recessive inherited neutropenia known in Border Collies since the 1990's. Recently, the causative mutation has been identified in the canine VPS13B gene and a DNA-based diagnosis has now become available. The present paper describes clinical and clinico-pathologic findings in a Border Collie with TNS that was molecularly diagnosed for the first time in Japan. In a 10-week-old male Border Collie with microgenesis and symptoms related to recurrent infections, a hematological examination revealed severe leukopenia due to neutropenia, suggesting the dog to be affected by inherited neutropenic immunodeficiency. Direct DNA sequencing demonstrated that the dog was homozygous for the causative mutation of TNS and both its parents were heterozygous carriers. In addition, a simple and rapid polymerase chain reaction-based length polymorphism analysis coupled with microchip electrophoresis was developed for the genotyping of TNS. This assay could discriminate clearly all genotypes, suggesting that it was suitable for both individual diagnosis and large-scale surveys for prevention.     

Normal calves produced after transfer of embryos cultured in a chemically defined medium supplemented with epidermal growth factor and insulin-like growth factor I following ovum pick up and in vitro fertilization in Japanese black cows

The objective of this study was to examine whether high concentrations of epidermal growth factor (EGF) and/or insulin-like growth factor I (IGF-I) would have a beneficial effect on bovine embryo development in vitro and to obtain normal calves by using an ovum pick up method and embryo culture in a chemically defined medium. When compared with controls, EGF (100 or 200 ng/ml) or IGF-I (50 or 100 ng/ml) significantly increased the rate of embryos that developed into blastocysts during an 8-day culture after the in vitro fertilization of oocytes obtained from ovaries from a slaughterhouse. IGF-I induced a dose-dependent increase in cell number in both the inner cell mass and the trophectoderm, whereas EGF stimulated proliferation only in the inner cell mass. A combination of EGF (100 ng/ml) and IGF-I (50 ng/ml) produced an additive effect, and embryos developed into blastocysts at a comparatively high rate (27.9%) compared with controls (12.0%). A similar rate of development was achieved using a combination of EGF and IGF-I in the culture of embryos following ovum pick up by ultrasound-guided transvaginal follicular aspiration and in vitro fertilization, and 5 blastocysts that developed after the culture were transferred into uteri; two embryos implanted, and normal calves were born. These results suggest that the combined use of EGF and IGF-I makes bovine embryo culture in a chemically defined medium a practical and useful procedure for producing blastocysts, and its application to embryo culture following ovum pick up and in vitro fertilization could be useful for producing normal calves.     

Establishment of a potency test by ELISA for a rabies vaccine for animal use in Japan

The ELISA we developed was able to determine the antigen content and was suitable for a potency test, and we described a relative potency assay method which determines the potency of test vaccines by comparing the ELISA value of a test vaccine to that of a reference vaccine. In the present study, we standardized the reference vaccine used for determining the potencies of test vaccines, and established a potency test by ELISA. We evaluated the proposed reference vaccine by the neutralizing antibody responses in dogs after vaccination, by the challenge protection test in guinea pigs (GP potency test), which is the earlier official potency test used in Japan, and by the NIH potency test, which is widely used throughout the world. The results showed that a 4-fold dilution of the proposed reference vaccine induced sufficient immunity in dogs. A 3-fold dilution of the proposed reference vaccine passed the GP potency test. The international units (IU) calibrated by the NIH potency test were 3.7 IU/dose. From the results and the WHO recommendation that veterinary rabies vaccines should have a potency of at least 1.0 IU/dose, we determined to dilute the proposed reference vaccine by 3 fold and regarded it as the reference vaccine. Finally, we confirmed that there is a good agreement between the results of the potency test by ELISA and the results of the GP potency test. The establishment of the potency test by ELISA has made it possible to monitor the potency in the production process and has contributed to the stable production of the vaccine.     

Growth Complementation of hrpXo Mutants of Xanthomonas oryzae pv. oryzae by Virulent Strains in Rice Cultivars Resistant and Susceptible to the Parental Strain

Xanthomonas oryzae pv. oryzae (X. o. pv. oryzae) T7174 is virulent on rice cultivar IR24 and avirulent on IR-BB2. From recent reports, some virulence and avirulence factors of plant pathogenic bacteria are transferred to plant cells through the hrp-dependent type III secretion system. In this study, we investigated the involvement of hrp genes in the compatible and the incompatible interactions between rice and X. o. pv. oryzae after co-inoculation with hrpXo mutants derived from T7174 and virulent strains. Growth of the mutants, named 74ΔHrpXo and 76ΔHrpXo, was repressed in IR24 when the mutants were applied alone. However, growth of the mutants was complemented by co-inoculation with virulent strains. Growth of bioluminescent hrpXo mutant 76ΔHrpXo in IR24 and its growth in IR-BB2 after co-inoculation with T7133, which is virulent on both cultivars, was equally complemented, as detected by bioluminescence from the mutant. On the other hand, only partial complementation of growth of T7174L76, which is a bioluminescent and pathogenic derivative of T7174, by T7133 was observed in IR-BB2. Thus, growth of the hrpXo mutant of X. o. pv. oryzae was complemented by virulent strains in both susceptible and resistant rice leaves with the parental strain. Received 21 July 2000/ Accepted in revised form 26 October 2000