Establishment of embryo transfer in the musk shrew (Suncus murinus)

The present study established techniques to induce pseudopregnancy, in vitro oocyte cultures from pronuclear to 2- to 4-cell stages, and embryo transfer in musk shrews, a reflex ovulator. Offspring were subsequently obtained by transferring in vivo-developed or in vitro-cultured embryos. Female musk shrews received human chronic gonadotropin (hCG), with or without mating stimuli, from vasectomized males to produce pseudopregnant recipients. Embryos at the 2- to 4-cell stage were collected 44–48 h after mating. Another set of embryos was collected 26–27 h after mating and then cultured for 20 h from the pronuclear to 2- to 4-cell stages. Subsequently, embryos were transferred into the oviducts of pseudopregnant recipients 24 or 48 h after the induction of pseudopregnancy. Offsprings were successfully obtained from recipients that received hCG 24 h before embryo transfer, regardless of mating stimuli. These techniques may be valuable for producing transgenic musk shrews.     

Factors affecting premature chromosome condensation of cumulus cell nuclei injected into rat oocytes

To date, production of cloned rats by somatic cell nuclear transfer (NT) has not yet been successful. Inducing premature chromosome condensation (PCC) of injected cell nuclei in recipient cytoplasm is considered essential for successful mouse cloning by the Honolulu method. In the present study, some factors affecting PCC of rat cumulus cell nuclei injected into rat oocytes were examined. Wistar female rats (young: 4 to 5-week-old, mature: > or =10-week-old) were superovulated by injections of eCG and hCG, and oocytes recovered 14 or 17 h after hCG injection were received with cumulus cell nuclei using piezo-driven micromanipulator. When the oocytes were recovered 14 h post-hCG injection from young rats and the nuclear injection into oocytes was completed within 45 min, PCC was observed in 44-49% of NT oocytes. In the case of oocytes from mature rats, PCC occurred in 11-19% of the NT oocytes. Oocytes recovered 17 h post-hCG injection did not support PCC of the injected nuclei (0-7%) regardless of the donor age. Treatment of oocytes with a neutral cysteine protease inhibitor, N-acetylleucylleucylnorleucinal, slightly increased the incidence of PCC (48 vs 37%). Comparison of rat strains for oocyte donors indicated that proportions of NT oocytes undergoing PCC in Wistar and LEW oocytes (41-46%) were higher than those in Donryu and F344 oocytes (17-25%). Thus, ability of rat oocytes to promote PCC of the injected nuclei is dependent on the characteristics of oocytes, such as age or strain of donor rats, and timing of oocyte recovery.     

Infection of Melissococcus plutonius clonal complex 12 strain in European honeybee larvae is essentially confined to the digestive tract

Melissococcus plutonius is an important pathogen that causesEuropean foulbrood (EFB) in honeybee larvae. Recently, we discovered a group of M.plutonius strains that are phenotypically and genetically distinct from otherstrains. These strains belong to clonal complex (CC) 12, as determined by multilocussequence typing analysis, and show atypical cultural and biochemical characteristicsin vitro compared with strains of other CCs tested. Although EFB isconsidered to be a purely intestinal infection according to early studies, it is unknownwhether the recently found CC12 strains cause EFB by the same pathomechanism. In thisstudy, to obtain a better understanding of EFB, we infected European honeybee(Apis mellifera) larvae per os with awell-characterized CC12 strain, DAT561, and analyzed the larvae histopathologically.Ingested DAT561 was mainly localized in the midgut lumen surrounded by the peritrophicmatrix (PM) in the larvae. In badly affected larvae, the PM and midgut epithelial cellsdegenerated, and some bacterial cells were detected outside of the midgut. However, theydid not proliferate in the deep tissues actively. By immunohistochemical analysis, the PMwas stained with anti-M. plutonius serum in most of the DAT561-infectedlarvae. In some larvae, luminal surfaces of the PM were more strongly stained than theinside. These results suggest that infection of CC12 strain in honeybee larvae isessentially confined to the intestine. Moreover, our results imply the presence ofM. plutonius-derived substances diffusing into the larval tissues inthe course of infection.     

Donor and recipient rat strains affect full-term development of one-cell zygotes cultured to morulae/blastocysts

The present study was conducted to examine the developmental potential to offspring of rat embryos cultured from 1-cell to morula/blastocyst stage. Pronuclear zygotes from Wistar x Wistar or (SD x DA) x Wistar strains were cultured in modified rat 1-cell embryo culture medium (mR1ECM) for 96 h in 5% CO(2) in air at 37 C. The proportion of the 3-way cross hybrid zygotes developing into morula/blastocyst stage (74%) was higher than that of the Wistar zygotes (66%). Day-5 morulae/blastocysts developed in vitro were transferred into Day-3 or -4 pseudopregnant recipients of Wistar or SD x DA strain. The transfer of cultured embryos resulted in the birth of offspring at 13-59%, while that of non-cultured control blastocysts showed birth rates of 35-65%. The best offspring rate of cultured embryos (59%) was obtained when the hybrid 1-cell zygotes were cultured in mR1ECM medium and transferred into the 2-days earlier uteri of SD x DA recipients. These results suggest that genetic background of recipients as well as donors is a possible factor affecting full-term development of rat morulae/blastocysts derived from 1-cell stage zygotes cultured in vitro.     

Influence of sickness condition on diurnal rhythms of heart rate and heart rate variability in cows

Parameters of heart rate variability would explain changes in heart rate during the disease status in cows and to evaluate whether such changes might provide a more sensitive and quantitative indicator of these conditions than crude indices. For this purpose, we recorded electrocardiograms for 24 hr using a Holter-type electrocardiograph and applied power spectral analysis of heart rate variability in both five clinically healthy and four hospitalized cows. The significant findings of the current investigation were that the diurnal variations of autonomic nervous function are abolished in cows that are sick. This abnormal rhythm was induced by predominant parasympathetic inhibition in these cows. Therefore, the heart rate variability may be a useful indicator of sickness condition in cows.     

Relationship between crack propagation and the stress intensity factor in cutting parallel to the grain of hinoki (<Emphasis Type="Italic">Chamaecyparis obtusa</Emphasis>)

The method of digital image correlation (DIC) was applied to the digital image of orthogonal cutting parallel to the grain of hinoki, and the strain distribution near the cutting edge was evaluated. The wood fracture associated with chip generation was considered as mode I fracture, and the stress intensity factor K_I for fracture mode I was calculated from the strain distribution according to the theory of linear elastic fracture mechanics for the anisotropic material. The calculated K_I increased prior to crack propagation and decreased just after the crack propagation. The change in K_I before and after crack propagation, ΔK_I, decreased in accordance with the crack propagation length, although the variance in ΔK_I should depend on the relationships between the resolution of DIC method and the dimensions of cellular structure. The calculated K_I in this study was almost on the same order as reported in the literatures. It was also revealed, for the case of chip generation Type 0 or I, the stress intensity factor for fracture mode II could be negligible due to the higher longitudinal elastic properties of wood in the tool feed direction than the one radial ones, and the mode I fracture was dominant.     

Expression of genes encoding xyloglucan endotransglycosylase/hydrolase in ‘Saijo’ persimmon fruit during softening after deastringency treatment

Persimmon (Diospyros kaki Thunb.) fruit undergoes intensive cell wall modification during postharvest fruit softening. Xyloglucan metabolism is important in cell wall disassembly. We cloned cDNAs for two xyloglucan endotransglycosylase/hydrolase genes (DkXTH1 and DkXTH2) from ‘Saijo’ persimmon fruit treated with dry ice to remove astringency. In order to determine the ethylene dependence of XTH gene expression, fruit were exposed to 1-methylcyclopropene (1-MCP), an inhibitor of ethylene action, prior to removal of astringency. Ethylene production increased in mature control and 1-MCP-pretreated fruit after dry-ice treatment, and flesh firmness decreased to the same extent during dry-ice treatment in the control and 1-MCP-pretreated fruit. After dry-ice treatment, control fruit softened completely, but fruit firmness was maintained in 1-MCP-pretreated fruit. Accumulation of DkXTH1 mRNA was induced simultaneously with commencement of ethylene production in mature control fruit. Pretreatment with 1-MCP delayed accumulation of DkXTH1 mRNA. DkXTH2 expression also coincided with fruit softening but was intensified by 1-MCP treatment during the deastringency treatment. These results indicate that fruit softening was related to both DkXTH1 and DkXTH2 expression in ‘Saijo’ persimmons.     

Morphology and heredity of tepal spots in Asiatic and Oriental hybrid lilies (Lilium spp.)

Petal spots constitute an important trait for ornamental flowers, because they generate variation in flower colour and pigmentation patterning. However, only a few reports have described the morphologies and inheritances of petal spots. Two types of spots—raised spots and splatters—appear on the flower tepals of lily (Lilium spp.). Here, we microscopically analysed the morphologies of raised spots and splatters, and compared them with those of petal spots in other species. The raised spots of lily showed an increase in the numbers of parenchymal and epidermal cells, and accumulation of anthocyanin pigments in these cells. We did not observe this type of morphology in the petal spots of rhododendron or Tricyrtis macropoda, indicating that the morphology observed in the raised spots is unique to lily. In the splatters of lily, anthocyanin pigments accumulated only in epidermal cells, and the shape of pigmented cells did not differ from that of unpigmented cells. These features were also observed in spots of T. macropoda. We subsequently investigated the inheritance of raised spots and splatters in an F₁ segregating population. We revealed that F₁ plants with raised spots, splatters, raised spots and splatters, or no spots showed a 1:1:1:1 segregation ratio, indicating that the two types of spots are genetically independent. Such knowledge will facilitate efficient selection during breeding programs focused on the development of cultivars with (or without) spots.     

Continuous plasma outflows from the edge of a solar active region as a possible source of solar wind

The Sun continuously expels a huge amount of ionized material into interplanetary space as the solar wind. Despite its influence on the heliospheric environment, the origin of the solar wind has yet to be well identified. In this paper, we report Hinode X-ray Telescope observations of a solar active region. At the edge of the active region, located adjacent to a coronal hole, a pattern of continuous outflow of soft-x-ray-emitting plasmas was identified emanating along apparently open magnetic field lines and into the upper corona. Estimates of temperature and density for the outflowing plasmas suggest a mass loss rate that amounts to approximately 1/4 of the total mass loss rate of the solar wind. These outflows may be indicative of one of the solar wind sources at the Sun.