Polar cardenolide monoglycosides from stems and twigs of Nerium oleander and their biological activities

Twelve polar cardenolide monoglycosides, 1, 2, 4?C13, and oleagenin (3) were isolated from the methanol extract of stems and twigs of Nerium oleander. Among these, oleagenin (3) and cardenolide monoglycosides named cardenolide B-1 (1) and cardenolide B-2 (2) were isolated from natural sources for the first time. The in vitro antiinflammatory activity of compounds 1?C13 was examined on the basis of inhibitory activity against the induction of the intercellular adhesion molecule-1 (ICAM-1). Compounds 4?C7 were active at an IC₅₀ value of less than 0.4 ??M. The cytotoxic activity of compounds 1?C13 was evaluated against three human cell lines: normal human fibroblast cells (WI-38), malignant tumor cells derived from WI-38 (VA-13), and human liver tumor cells (HepG2). Compounds 4, 6, and 7 were active toward these three cell lines at IC₅₀ values of less than 0.7 ??M, and compounds 5 and 8 were active toward the cell lines at IC₅₀ values of less than 1.5 ??M. The multidrug resistance (MDR) cancer-reversal activity of compounds 1?C13 was evaluated on the basis of the amount of calcein accumulated in MDR human ovarian cancer 2780AD cells in the presence of each compound. Compound 1 and 12 showed significant effects on calcein accumulation.     

Seroconversion of anti-Getah virus antibody among Japanese native Noma horses around 2012

Getah virus (GETV), an arthropod-borne virus transmitted by mosquitoes, has been isolated from several animals. GETV infection in horses shows clinical signs such as fever, rash, and edema in the leg. Noma horses are one of the eight Japanese native horses. The present study aimed to clarify the occurrence of GETV infection in Noma horses. Serum samples collected from Noma horses were analyzed using a virus neutralization test and enzyme-linked immunosorbent assay and showed that the anti-GETV antibody titers in the samples collected in 2017 were significantly higher than those collected in 2012. We concluded that a seroconversion of anti-GETV antibodies was occurred in the Noma horse population around 2012, providing evidence of the GETV epidemic in Japan circa 2012.     

Seed formation in triploid loquat (Eriobotrya japonica) through cross-hybridization with pollen of diploid cultivars

As the fruits of loquat (Eriobotrya japonica, 2n = 2x = 34) carry large seeds, the breeding of seedless loquat has long been a goal. The recent creation of triploid cultivars (2n = 3x = 51) and the application of gibberellins allow commercial production of seedless loquat, but the possibility of seed formation in triploid loquats has not been carefully investigated. Through crossing experiments and cytological observations of meiosis and pollen tube growth, we found that the triploid line 3N-N28 was essentially self-sterile, but developed seeds on pollination with pollen from diploid cultivars at rates of up to 5.5%. Almost half of the seedlings survived to 5 months, and carried diploid (2n = 34), tetraploid (2n = 68), or aneuploid chromosome numbers. Our results suggest that triploid loquat cultivars might retain the risk of seed formation. Protection from pollination by diploid cultivars or the development of new triploid cultivars will be necessary to ensure the production of seedless loquat fruits.     

QTL analysis for soil‐surface roots originating from a New Plant Type rice (Oryza sativa L.)

The introgression line YTH16 harbouring chromosome segments from the New Plant Type cultivar IR65600–87–2‐2–3 with genetic background of an Indica Group rice IR 64 forms soil‐surface roots. To clarify the genetic mechanism, QTL analysis was performed using hybrid populations derived from a cross between IR 64 and YTH16. A total of eight QTLs were detected in the three introgressed segments on chromosomes 2, 5 and 7. Seven chromosome segment lines (CSLs) combining these three QTL regions were selected from the progenies. The two CSLs harbouring a single region (excluding the CSL with a region on chromosome 5) showed high scores and low means of root angle distribution in comparison with IR 64. Four CSLs harbouring two or three regions showed high scores and low means of root angle distribution in comparison with YTH16 and the CSLs harbouring a single QTL region. These results indicated that the soil‐surface rooting of YTH16 was controlled by the three QTLs’ regions and that chr. 5 particularly played a role in supporting the effect with others.     

γ-HCH-Decomposing ability of several strains of Pseudomonas paucimobilis

Abstract

Extract

We succeeded in isolating a γ-HCH (γ-1,2,3,4,5,6-hexachlorocyclohexane)-decomposing aerobic bacterium from an upland field where γ-HCH had been repeatedly applied for more than 10 years. The bacterium was identified as Pseudomonas paucimobilis (Oyaizu and Komagata, 1983; Senoo and Wada 1989; Wada et al. 1989). We assumed that P. paucimobilis was able to thrive in the upland field by acquiring the γ-HCH-decomposing ability (Tu 1975, 1976) based on the following mechanism:     

Gross nitrification rates in four Japanese forest soils: heterotrophic versus autotrophic and the regulation factors for the nitrification

Measurements of gross NH ₄ ⁺ and NO ₃ ^? production in forest soils were conducted using the ¹⁵N pool dilution method. Mineral topsoils (0?C10?cm depth) were collected from four forests from northern to southern Japan with a natural climate gradient to elucidate the mechanisms regulating gross nitrification rates in forest soils. Additionally, we attempted to evaluate the relative importance of heterotrophic nitrification in gross total nitrification using acetylene as a specific inhibitor of autotrophic nitrification. Distinct differences were found among sites in the gross rates of NH ₄ ⁺ production (3.1?C11.4?mg?N?kg^?1?day^?1) and gross total nitrification (0.0?C6.1?mg?N?kg^?1?day^?1). The rates of gross heterotrophic nitrification were low in this study, indicating that heterotrophic nitrification is of minor importance in most forest mineral topsoils in Japan. Significant relations were found between gross autotrophic nitrification and gross NH ₄ ⁺ production, soil N, and soil C concentrations, but none was found between gross autotrophic nitrification and soil pH. We determined the critical value of the gross NH ₄ ⁺ production rates for gross autotrophic nitrification under which no gross autotrophic nitrification occurred, as well as the critical soil C/N ratio above which gross autotrophic nitrification ceased. Results show that tight coupling of production and consumption of NH ₄ ⁺ prevents autotrophic nitrifiers from utilizing NH ₄ ⁺ as long as NH ₄ ⁺ availability is low.     

Genomic variations among Bartonella henselae isolates derived from naturally infected cats

The purpose of this study was to understand the mechanisms of persistent infection with Bartonella henselae in cats. Blood samples were collected from three naturally infected cats for 24 months. These cats were confirmed to be persistently infected with B. henselae by serological and bacteriological examination. Relapsing bacteremia was found in all three cats with intervals of 3-19 months. Following the peaks of bacteremia, increases of specific antibody titer were observed in these cats. To examine the genetic differences among the isolates derived from the first and following bacteremia, the genome DNA patterns of the restriction enzyme fragment length polymorphism (RFLP) of the isolates were examined by pulsed field gel electrophoresis. The isolates derived from the first bacteremia showed an identical RFLP pattern in each of the three cats. The isolates derived from the following peaks, however, showed 1-3 of different RFLP patterns in these cats. Furthermore, the isolates showing different RFLP patterns from those of the first bacteremia were also detected at the following bacteremic peaks in all three cats examined. The 16S ribosomal RNA (rRNA) gene type of all isolates was found to be 16S rRNA type I. The emergence of genetically distinct organisms at various peaks of bacteremia may contribute to the establishment of persistent infection in the naturally infected cats.     

Immunohistochemical and histoplanimetrical study on the spatial relationship between the settlement of indigenous bacteria and the secretion of bactericidal peptides in rat alimentary tract

To clarify the regulatory mechanism by bactericidal peptides secretion, the secretion of bactericidal peptides was immunohistochemically and histoplanimetrically compared with the degree of Gram-positive/negative bacterial colonization throughout the rat alimentary tract. In the associated exocrine glands from the oral cavity to the stomach, no comparable differences were observed under the changes of development of indigenous bacterial colonies. In the small intestine, immunopositive granules for lysozyme and secretory phospholipase A2 (sPLA2) were markedly decreased, whereas immunopositive vacuoles in the Paneth cells were more increased at sites with hyper-development of indigenous bacterial colonies in the intervillous spaces than at sites with no or less development. No changes in exocrine glands were observed in the large intestine because of the constant existence of large quantities of bacteria. Gram-positive bacterial colonies on the mucosal surfaces were dominant from the oral cavity to the stomach. Gram-negative bacteria were dominant in the large intestine, and the distributions of both Gram-positive and negative bacteria were intermediate in the small intestine. These findings suggest that lysozyme and sPLA2 secreted from the Paneth cells contribute to the regulation of the proliferation of indigenous bacteria in the intervillous spaces of the small intestine, and that the inversion of distributions of Gram-positive and -negative bacteria in the alimentary tract might be caused by the secretion of lysozyme and sPLA2 in the small intestine.     

Relationships between the addition rates of cellulase or glucose and silage fermentation at different temperatures

The influence of the application rates of cellulase preparation and glucose on silage fermentation at different temperatures was studied with the straw of naked barley (Hordeum vulgare L. emand Lam) and guineagrass (Panicum maximum Jacq.). Addition rate of cellulase and glucose, temperature and their interaction had significant effects on pH value, lactic acid content, butyric acid content and propionic acid content of naked barley straw silage and significant effects on all the parameters of guineagrass silage (P < 0.01). Temperature and interaction had significant effect on acetic acid content (P < 0.05) and no significant effect on NH₃‐N content of naked barley straw silage (P > 0.05). Under all the temperatures, the pH values of barley straw and guineagrass silages were reduced by cellulase and glucose addition even at the lowest rate (P < 0.05), compared with their corresponding control. Lactic acid contents of silages were the highest within the same temperature and same additive when glucose and cellulase were added at the highest rates, whereas the effect of cellulase and glucose addition on butyric acid production varied with their application rates and silage storage temperature. The addition rate of restricting butyric acid fermentation was lower at 20°C than that at 30°C, and it was the lowest at 40°C where cellulase and glucose addition restricted butyric acid fermentation even at 0.1 g/kg and 10 g/kg, respectively, when compared to the control. While the addition rate was lower than the above level, cellulase and glucose addition also promoted butyric acid fermentation.     

Partial characterization of structure and function of a xylanase gene from the rumen hemicellulolytic bacterium Eubacterium ruminantium

A gene encoding for xylanase activity in the rumen hemicellulolytic bacterium Eubacterium ruminantium was cloned into pBR322 in Escherichia coli (E. coli ). The primary clone had a 5.7 kb insert produced by Eco RI partial digestion. Subcloning followed by sequencing allowed for the discovery that this enzyme has a glycosyl‐hydrolase family 10 catalytic domain with a family 9 carbohydrate binding module at C‐terminus and a region partially homologous to a family 22 carbohydrate binding module at N‐terminus. Cloned xylanase is specifically active against xylan and oligoxyloside to produce xylobiose and xylotriose, showing optimal pH and temperature at 7.0 and 50°C, respectively. Molecular size of the xylanase (91 kDa) was confirmed by zymogram analysis of the E. coli clone, which agreed with the predicted size from the DNA sequence. Functions of the two modules at C‐ and N‐termini were evaluated by using xylanase variants with and without the respective module and the C‐terminal module was found to be functional in binding to acid‐swollen cellulose and insoluble oat‐spelt xylan, whereas the N‐terminal module was inactive for binding them.