Bioenergy grass [Erianthus ravennae (L.) Beauv.] secretes two members of mugineic acid family phytosiderophores which involved in their tolerance to Fe deficiency

Ravenna grass, Erianthus ravennae (L.) Beauv. (E. ravennae) is a potential high biomass-energy crop with low input requirements. Iron (Fe) deficiency in calcareous soils is a widespread agronomic problem which reduces crop yields. Fe is sparingly soluble under aerobic conditions at high soil pH, such as in calcareous soils; therefore, plants cannot take up enough Fe. Increasing crop productivity of giant grasses, such as Ravenna grass in calcareous soil, has a positive effect by alleviating environmental problems. However, the growth character in calcareous soil and Fe homeostatic trait of Ravenna grass are largely unknown. In this study, we analyzed characteristics of Ravenna grass. The growth of E. ravennae plants were impaired in calcareous soil compared to those in the normal soil. In calcareous soil, the growth of E. ravennae plants differ among the water and fertilizer conditions; E. ravennae plants were grown better in the submerged condition adding micronutrient among conditions. These results suggested that impaired growth of E. ravennae in calcareous soil might be micronutrient shortage. We found that E. ravennae roots possess Fe reductase activities which were upregulated under Fe-deficient conditions. E. ravennae produced and secreted mugineic acid (MA) and deoxymugineic acid (DMA) to acquire Fe from the soil. The amount of MA was higher than that of DMA. Thus, E. ravennae might have both partial Strategy-I and Strategy-II Fe uptake systems. E. ravennae intercropped with transgenic rice plants producing and secreting MA through the introduction of the barley MA synthase gene showed improved growth compared to monocropped E. ravennae plants, suggesting that the increased amounts of MA enhanced their tolerance to Fe deficiency. Our results suggest that there is a considerable potential to improve the growth of E. ravennae plants in calcareous soils by enhancement of their Fe uptake systems through increase of MA production.     

Ethanol production from sugi pulp under simultaneous saccharification and fermentation using a cocktail enzyme of T. reesei and A. tubingensis produced by solid-state fermentation

The process of solid-state fermentation was used to produce a cocktail enzyme of Trichoderma reesei ATCC 66587 and Aspergillus tubingensis KRCF 700-33. Wheat bran, corncob, and sugi pulp were supplemented with ammonium sulfate as an enzyme-producing medium using T. reesei and A. tubingensis. The corncob blend ratio, duration of incubation, and ammonium sulfate concentration were optimized for enhancing cellulase production from T. reesei using a Box-Behnken design. Filter paper degrading activity more than tripled when T. reesei was grown in the optimized medium, as compared with the initial medium. The highest activity of 4.03 FPU/ml (about 29 FPU/g of dry material) was obtained with a cocktail enzyme having a 25 % content of A. tubingensis and 75 % of T. reesei. The sugi pulp was then fermented to ethanol with the cocktail enzyme and thermotolerant yeast (Saccharomyces cerevisiae BA-11) under simultaneous saccharification and fermentation (SSF) at 40 °C. An ethanol concentration of 4.48 % (w/v) was achieved using the cocktail enzyme (4 FPU/g-pulp) that was produced on-site with a substrate loading level of 12.5 wt %, which achieved an ethanol yield of 76 % after 72 h.     

Therapeutic effect of frequent injections of GnRH analogue in a beagle with knobbed acrosome abnormality of sperm

A Beagle dog (3 years old) that ejaculated high percentages (mean ± SE: 29.1 ± 1.2%) of sperm with a knobbed acrosome abnormality and a low number of sperm and that also had a low plasma testosterone (T) level was given 10 subcutaneous injections of 1 μg of gonadotropin-releasing hormone analogue (GnRH-A) at 3-day intervals. The plasma T level and number of sperm increased 12-14 weeks after the first injection. Although the percentages of sperm with knobbed acrosome abnormality did not change after the GnRH-A therapy, the number of sperm and percentage of actively motile sperm increased after the therapy, and a bitch gave birth to 5 healthy puppies after intravaginal artificial insemination with fresh semen collected 14 weeks after the first injection.     

Prevalence of Bordetella hinzii in mice in experimental facilities in Japan

To reveal the current status of the prevalence of Bordetella hinzii in mice in experimental facilities in Japan, a survey of this agent was performed by culture of tracheal swabs from a total of 12,923 mice from 1699 facilities (12,192 mice from 1572 facilities in universities and research institutes and 731 mice from 127 facilities in pharmaceutical companies) in total. In the results, 195 out of 12,192 mice (1.6%) from 44 out of 1572 facilities (2.8%) in universities and research institutes were positive for B. hinzii. No B. hinzii-positive mice were found in 127 pharmaceutical companies surveyed. Gross lesions in the lungs with isolation of B. hinzii were observed in seven mice from four universities, and the lesions were identified as bronchopneumonia histopathologically. To our knowledge, this is the first report to reveal the prevalence of B. hinzii in laboratory mice.     

Effect of intramammary infusion of rbGM-CSF on SCC and expression of polymorphonuclear neutrophil adhesion molecules in subclinical mastitis cows

The effect of rbGM-CSF intramammary infusion on the subclinical mastitis was evaluated by the somatic cell count (SCC) and expression of adhesion molecules (CD62L and CD11b) on the surface of neutrophils (PMN) in blood and milk. Fifteen cows diagnosed to have subclinical mastitis were used in this study. Seven cows showed a decrease in the SCC (decreased group), whereas 8 cows showed an increase in the SCC (increased group) 7 days after infusion of rbGM-CSF compared to pre infusion level. The percentage of CD62+ cells tended to be lower and CD11b+cells tended to be higher at 6 h on blood PMN in the decreased group of cows. Increased group of cows showed opposite tendencies. The mean fluorescent intensity of these adhesion molecules expressed on PMN in blood and milk was similar in both groups. These results suggested some association between expression of adhesion molecules and changes in SCC by rbGM-CSF. Responsiveness of PMN adhesion molecules to rbGM-CSF might determine the changes in SCC of the subclinical mastitic cows after infusion of rbGM-CSF.     

Generation of canine dendritic cells from peripheral blood monocytes without using purified cytokines

Dendritic cells (DCs), which differentiate in vitro from peripheral blood monocytes (PBMOs) or bone marrow precursors, are a promising candidate for immunotherapy against cancer. The dog, which suffers common types of cancers along with humans, make an ideal large animal model for cancer studies. Monocyte-derived DCs in the dog have not been well characterized, however, since the appropriate condition for in vitro differentiation has not been established. To tackle this problem, we have developed a conditioned media by culturing T cells with immobilized anti-canine CD3 antibody, and sought to induce differentiation of DCs from PBMOs. When purified CD14+ PBMOs were cultured in the presence of 25% T cell conditioned medium (TCCM), the PBMOs increased size and had extended dendritic processes by day 12 of the culture. The cultured PBMOs were found to increase the expression of MHC class II and CD1a molecules, and significantly increased stimulatory activity for allogeneic T cells in the mixed leukocyte reaction. Moreover, the cells significantly increased their expression of IL-18 and IFN-gamma when stimulated with polyinosinic-polycytidylic acid (Poly (I:C)). The cells have a reduced phagocytic activity, which is a common defect in mature DCs. It follows from these results that TCCM does induce the differentiation of DCs from PBMOs.     

Immuno-magnetic separation and agar layer methods for the isolation of freeze-injured Yersinia enterocolitica O:8 from water

To develop an effective method to isolate an injured pathogenic Yersinia enterocolitica O:8 organism from environmental samples, we compared the isolation of freeze-injured and non-injured Y. enterocolitica O:8 and found that the isolation was more successful when immuno-magnetic separation (IMS) with anti-Y. enterocolitica O:8 antibody was used. Plating onto cefsulodin-irgasan-novobiocin (CIN) agar and Virulent Yersinia enterocolitica (VYE) agar by means of the agar layer method was found to be effective in isolating the injured cells. The alkali treatment which is generally used for selective detection of Yersinia organism failed to isolate freeze-injured pathogenic Y. enterocolitica O:8 cells. Recovery methods without using the alkali treatment were superior for detecting freeze-injured Y. enterocolitica O:8. Our results demonstrate that the IMS and the agar layer methods should be used to isolate injured pathogenic Yersinia organisms from environmental samples such as water.     

Annual changes in testicular size and serum and fecal testosterone concentrations in male bharals, Pseudois nayaur

The serum and fecal testosterone (T) concentrations and testicular sizes of two male bharals (Pseudois nayaur) were determined for approximately one year. The profiles of the fecal T concentrations showed a similar tendency as the profiles of serum T concentrations, and there was a significant correlation between serum and fecal T concentrations (r=0.72). T concentrations rose drastically in October and decreased gradually until January. The maximum testicular size was observed between November and January. Semen collected between December and January was excellent in quality and comparable to domestic sheep and goats. The active periods of the testes were synchronized with the early breeding season of females.     

The relationship between sperm morphology and in vitro fertilization ability in mice

In vitro fertilization (IVF) is widely used in reproduction research, but the sperm of some inbred strains of mice yield low fertilization rates in IVF. To determine the cause of this problem, we examined the effect of epididymal sperm morphology, in particular, tail bending and the presence and type of cytoplasmic droplet (CD), on fertilizability in vitro. Sperm suspensions were obtained from the following five strains: C57BL/6J, BALB/cA, C3H/HeN, DBA/2J, and 129 x 1/SvJ. The sperm were fixed in 10% formalin and three parts of the sperm, namely the head, tail, and CD, were examined. We recorded the proportion of abnormal sperm heads and hairpins at the neck; tails were categorized as straight, proximal bent, or distal bent; and the CDs were categorized as none, light-type, and heavy-type. Based on these parameters, we determined the correlations between sperm morphology and fertilizability in vitro, as judged by IVF using ICR oocytes. The proportion of sperm with a hairpin neck was higher in strain C57BL/6J, while abnormal head morphology occurred significantly more often in strain BALB/cA. The percentage of sperm with a proximal bent tail was highest in strain DBA/2J and lowest in strain 129 x 1/SvJ. A heavy-type CD was observed more frequently in the 129 x 1/SvJ and C57BL/6J strains than in the other three strains in which a light-type CD predominated. The rank order of the fertilization rates was 129 x 1/SvJ < C57BL/6J < C3H/HeN < BALB/cA < DBA/2J. In addition, fertilization rate was positively correlated with a proximal bent tail, but negatively correlated with a heavy-type CD and distal bent tail. This new classification system establishes that the morphological characteristics of epididymal sperm differ among inbred strains of mice and that tail and CD morphology are closely related to fertilization ability in IVF. Thus, our results provide a novel method for assessing the quality of mouse sperm used for IVF.