Profile of bordered pit aspiration in Cryptomeria japonica using confocal laser scanning microscopy: pit aspiration and heartwood color

Nine trees of Cryptomeria japonica from six elite tree clones with a broad range of heartwood colors were selected. The profiles of pit aspiration percentage (ASP) of earlywood and latewood from pith to bark for green and air-dry conditions were determined to study the relationship between heartwood color and pit aspiration. Confocal laser scanning microscopy (CLSM) observations showed that the ASP of earlywood was low in sapwood and high in heartwood in the green condition. Pit aspiration increased in intermediate wood when compared with sapwood. On the other hand, latewood pits did not aspirate during heartwood formation. Comparing the air-dry condition with the green condition, sapwood pits aspirated during drying in both earlywood and latewood; however, there was no significant difference in pit aspiration of heartwood. There was no significant difference between samples with red and black heartwoods for ASP. The difference in ASP between individual trees was larger than that by heartwood color. The general advantage of CLSM over light microscopy is that serial optical sections along the Z axis can be obtained for any moisture condition, without the need for thin sectioning or embedding.     

Modulation of the tissue reninangiotensin-aldosterone system in dogs with chronic mild regurgitation through the mitral valve

OBJECTIVE: To investigate whether the tissue and plasma renin-angiotensin-aldosterone system (RAAS) is activated in dogs with mild regurgitation through the mitral valve and determine the contribution of chymase and angiotensin-converting enzyme (ACE) to the activation of the RAAS and potential production of angiotensin II during the chronic stage of mild mitral valve regurgitation. ANIMALS: 5 Beagles with experimentally induced mild mitral valve regurgitation and 6 clinically normal (control) Beagles. PROCEDURES: Tissue ACE and chymase-like activities and plasma RAAS were measured and the RAAS evaluated approximately 1,000 days after experimental induction of mitral valve regurgitation in the 5 dogs. RESULTS: Dogs with experimentally induced mitral valve regurgitation did not have clinical signs of the condition, although echocardiography revealed substantial eccentric hyper- trophy. On the basis of these findings, dogs with mitral valve regurgitation were classified as International Small Animal Cardiac Health Council class Ib. Plasma activity of renin and plasma concentrations of angiotensin I, angiotensin II, and aldosterone were not significantly different between dogs with mitral valve regurgitation and clinically normal dogs. Tissue ACE activity was significantly increased and chymase-like activity significantly decreased in dogs with mitral valve regurgitation, compared with values in clinically normal dogs. CONCLUSIONS AND CLINICAL RELEVANCE: The tissue RAAS was modulated without changes in the plasma RAAS in dogs with mild mitral valve regurgitation during the chronic stage of the condition. An ACE-dependent pathway may be a major route for production of angiotensin II during this stage of the condition.     

The role of neighboring infected cattle in bovine leukemia virus transmission risk

A cohort study was conducted to evaluate the risk of bovine leukemia virus (BLV) transmission to uninfected cattle by adjacent infected cattle in 6 dairy farms. Animals were initially tested in 2010–2011 using a commercial ELISA kit. Uninfected cattle were repeatedly tested every 4 to 6 months until fall of 2012. The Cox proportional hazard model with frailty showed that uninfected cattle neighboring to infected cattle (n=53) had a significant higher risk of seroconversion than those without any infected neighbors (n=81) (hazard ratio: 12.4, P=0.001), implying that neighboring infected cattle were a significant risk factor for BLV transmission. This finding provides scientific support for animal health authorities and farmers to segregate infected cattle on farms to prevent spread of BLV.     

One-step immunochromatography assay for detection of high-level canine serum trypsin-like immunoreactivity

We developed a one-step immunochromatography assay kit to measure high levels of canine trypsin-like immunoreactivity (cTLI) for bedside estimation of canine pancreatitis. The serum cTLI level can be determined within 10 min by visual comparison of color strengths in the test and reference zones. The serum cTLI levels determined by this method correlate well with canine TLI-ELISA and can be classified into 3 categories: cTLI levels higher than 60 ng/ml were considered positive; 20-60 ng/ml, weakly positive; and less than 20 ng/ml, negative. Twelve dogs suspected of pancreatitis were examined using this method; 4 dogs were positive, 2 were weakly positive, and 6 were negative. This test can detect a high level of serum cTLI and a positive result in the TLIH test will provide critical information for evaluation of pancreatitis in dogs.     

Growing porcine oocyte-granulosa cell complexes acquired meiotic competence during in vitro culture

The aim of this study was to establish a culture system to improve the meiotic competence of porcine oocyte-granulosa cell complexes (OGCs) obtained from preantral or early antral follicles. Porcine OGCs were recovered from follicles with diameters of 230-300 (preantral follicles), 300-500, and 500-700 mum (early antral follicles) using scalpels. The OGCs were cultured for 2 weeks in culture medium. We examined the effects of the sizes of the follicles from which OGCs were recovered, the concentrations of polyvinylpyrrolidone (PVP, 0-8%) in the culture medium, and 2 types of culture dish (Falcon 3002 vs 1007) on formation of the antrum of OGCs. After culture, the oocytes were matured for 44 h to assess their meiotic competence. OGCs recovered from small follicles (230-500 microm) required longer (P<0.05) than larger follicles to form the antrum structure. The percentage of OGCs forming the antrum structure that were cultured in 2% PVP (31%) was higher (P<0.05) than for those cultured in other PVP concentrations (0-11%). The percentages of antrum-structure formation for OGCs cultured on Falcon 3002 (83% for 2% PVP and 60% for 4% PVP) were higher (P<0.05) than those cultured on Falcon 1007 (47% for 2% PVP and 9% for 4% PVP). Furthermore, all of the intact oocytes that were obtained from culture of OGCs and that formed an antrum were in the GV stage (n=28). When these immature oocytes were cultured for 44 h, the percentage of oocytes that reached the metaphase II stage (25%, n=68) was higher (P<0.0001) than that of oocytes matured without culture (0.7%, n=137). The results of the present study show that porcine OGCs obtained from preantral or early antral follicles acquire meiotic competence in vitro.     

Aging of recipient oocytes reduces the development of cloned embryos receiving cumulus cells

Parthenogenetic activation is an important factor in successful production of cloned mammals. Because it has been reported that aged oocytes are more sensitive to parthenogenetic activation than young oocytes, the present study examined the effects of oocyte aging on the in vitro and in vivo developmental potential of nuclear-transferred (NT) mouse oocytes receiving cumulus cells. The potentials of young NT oocytes (14 h after human chorionic gonadotrophin [hCG] injection) to develop into blastocysts was, however, significantly higher than that of aged oocytes (20 h after hCG injection; 16% vs 6%). When the nuclei of NT oocytes at the 2-cell stage were fused with enucleated fertilized 2-cell embryos, the potentials of the serial NT embryos to develop into blastocysts were no different for both young and aged oocytes (74% vs 74%). Live young, however, were obtained only after transfer of serial NT blastocysts developed from young NT oocytes (2%). In contrast to a report using embryonic nuclei as the nuclear donors, the results of the present study indicate that young oocytes are superior to aged oocytes as a source of recipient cytoplasm for mouse somatic cell cloning.     

Prebiotic effect of glycerol galactoside isolated from color-faded nori in rats

Glycerol galactoside (GG; floridoside: 2-O-glycerol-α-d-galactopyranoside, and isofloridoside: 1-O-glycerol-α-d-galactopyranoside) is known to be a component of edible red seaweed nori. Recently, we have found that low-quality nori (color-faded nori), which has a low protein content, contains a large quantity of GG. From further studies, we have also found that GG has prebiotic characteristics in vitro. In this study, we evaluated the in vivo prebiotic activity of GG in rats. Dietary GG selectively increased the cecal Bifidobacterium count in rats. Other indices of prebiotics, such as pH of cecal content, organic acid concentrations, and fecal weight, also supported the existence of prebiotic activity of GG. The present data will also contribute to the development of a new method of utilizing color-faded nori as a health-promoting foodstuff.     

Nitrogen flow on an organically managed beef farm in Hokkaido,Japan

Nutrient recycling should be effective at balancing nutrient flows in Japanese animal production. This means replacing imported feed with self-produced feed. The Yakumo Experimental Farm of Kitasato University has produced commercial beef under ‘organic’ management, without the use of agricultural chemicals or imported feed, since 2005. Using a data set obtained from 220 ha of grassland and 250 head of cattle over the 5 years from 2008 to 2012, we estimated nitrogen (N) flow. During 2011 and 2012, we measured grass production, cattle production (selling out), soil parameters and atmospheric deposition (from precipitation and atmospheric ammonia concentrations). To determine N fixation by clover (white clover, Trifolium repens L.), we compared grass + clover plots with grass-only plots. Averaged over the period, N components on the 220 ha of grassland comprised 1952 Mg soil N stock, 3.2 Mg N yr^?1 in living livestock, 14.3 Mg N yr^?1 uptake by grass growth (including 8.6 Mg yr^?1 of N fixed by clover), 15.7 Mg N yr^?1 applied in composted manure, 1.7 Mg N yr^?1 in imported bedding material, 2.8 Mg N yr^?1 in deposition and 1.41 Mg N yr^?1 in meat production. N in composted manure equaled about 0.8% of the huge soil N stock; N in grass production equaled about 0.7%, of which clover fixation supplied 60%; N deposition was not negligible; and N export by meat production was minor. These results show that on this organically managed farm, soil N stock increased gradually (by 8.6 Mg N yr^?1 [220 ha]^?1 = 39 kg N ha^?1 yr^?1 = 0.02% of the soil N stock) and N export was small. Our findings show that it is possible to balance N inputs with N outputs in a beef cattle enterprise without the need for feed or fertilizer imports.     

Density-dependent selfing and its effects on seed performance in a tropical canopy tree species, Shorea acuminata (Dipterocarpaceae)

In the conservation and management practices of natural forests, sound reproduction and regeneration form the basis of the maintenance and viability of the tree populations. To obtain and serve biological information for sustainable forest management, we investigated reproductive biology and inbreeding depression in seeds of an important dipterocarp tree species, Shorea acuminata (Dipterocarpaceae), by both field and laboratory experiments. Results of parental analysis of immature and mature seeds showed that selfing rates varied greatly, from 7.6 to 88.4% among eight mother trees, and the mean overall selfing rate was 38.3%. Observed outcrossing events within a 40-ha study plot were predominantly (76.5%) short-distance events with a mating distance (md) ≤ 100 m. Since the selfing rate sharply decreased with increase in the number of flowering conspecifics (i.e., individuals of the same species) within a 100-m radius from the mother trees, the local density of flowering conspecifics appears to be the key factor determining the outcrossing rate in S. acuminata. However, the extremely high selfing rate (88.4%) observed for one tree could not be simply explained by the low local density of flowering conspecifics. Instead, differences in its flowering phenology (its flowering peaked ca. a week earlier than most of the other examined individuals) may have severely limited its receipt of pollen from other conspecifics, and thus promoted selfing. Since there were no significant differences in the proportion of selfed progeny between immature and mature seed stages, there was no evidence of selective abortion of selfed seeds during seed development. However, the seed mass of outcrossed progeny was heavier than that of selfed progeny, and heavier seeds showed higher success rates at germination and seedling establishment. These results suggest that inbreeding depression resulted in reductions in seed mass and may reduce the fitness of selfed seeds in S. acuminata. In addition, the outcrossing rate of S. acuminata was more sensitive to low local conspecific flowering-tree densities than that of a sympatric bee-pollinated dipterocarp species with greater pollination distances. These results suggest that the management of local adult-tree densities is important for avoiding selfing and inbreeding depression in future generations, especially in a species like S. acuminata with predominantly short-distance pollination.     

Aluminum toxicity in plant cells: Mechanisms of cell death and inhibition of cell elongation

Aluminum (Al) ions are a major constraint for crop productivity in acidic soils. The root apex is the most sensitive plant part to Al, which inhibits root elongation and causes cell death. To elucidate the mechanisms of these toxic events, Al responses have been investigated in cultured cell lines of tobacco (Nicotiana tabacum), SL and BY-2. These cells at the logarithmic growth phase serve as a model system of meristematic cells at the root apex. Our research group has revealed three types of cell death mechanisms triggered by Al: (i) Enhancement of iron (Fe)-mediated lipid peroxidation leading to a loss of plasma membrane integrity (plasma membrane pathway); (ii) dysfunction of mitochondria accompanied by ROS production (mitochondria pathway); and (iii) upregulation of NtVPE1 encoding a vacuolar processing enzyme (VPE), which leads to vacuolar collapse and the loss of plasma membrane integrity (vacuole pathway). Mechanisms (ii) and (iii) have been confirmed in root systems of pea and tobacco seedlings, respectively. The inhibition of elongation (expansion) in SL cells was detected as a decrease in water content, together with decreased osmolality and soluble sugar content, which was partly due to the inhibition of sucrose uptake by Al. The inhibition of root elongation by Al due to the inhibition of the sucrose transporter (SUT) NtSUT1 localized at the plasma membrane was confirmed in tobacco seedlings, in which overexpression of NtSUT1 mitigated both the inhibition of elongation and cell death at the root apex under Al stress.