Maturation-associated changes in internal distribution and intra-ovarian microdistribution of tetrodotoxin in the pufferfish <Emphasis Type="Italic">Takifugu pardalis</Emphasis>

To clarify the maturation-associated accumulation profile of tetrodotoxin (TTX) in the female pufferfish Takifugu pardalis, specimens collected from Omura Bay, Nagasaki Prefecture, Japan, from November through December, 2014, were separated based on the histology of ovarian sections into four groups: yolk vesicle stage (Yv), yolk globule stage (Yg)-1, Yg-2, and Yg-3. TTX quantification in each tissue in each group by liquid chromatography–mass spectrometry revealed that the TTX concentration [mouse unit (MU)/g] in the ovary increased from Yv to Yg-1, and the TTX amount (MU/individual) in the ovary increased from Yg-1 through Yg-3. In Yv, TTX in the skin accounted for 86% of the total TTX in the body, whereas from Yg-1 through Yg-3, ovarian TTX accounted for 53–76% of the total. In female specimens collected from the same bay from February to May 2015, the TTX amount in the ovary markedly decreased after spawning, and the TTX amount in the liver increased. Immunohistochemical observation of the ovarian sections indicated that TTX localized mainly in the nucleus of immature oocytes, but then distributes to the yolk vesicles and yolk globules as maturation advances, and is partly transferred to the egg membrane as spawning approaches.     

Decontamination of Cs from Japanese cedar (<Emphasis Type="Italic">Cryptomeria japonica</Emphasis>) via kraft cooking

To investigate the possibility of decontaminating ¹³⁷Cs-contaminated Cryptomeria japonica wood, kraft pulping was conducted and the Cs behavior in the reaction process was examined. ¹³³Cs-treated or ¹³⁷Cs-contaminated bark, sapwood, and heartwood chips of Cryptomeria japonica were digested using an aqueous solution of NaOH and Na₂S. The pulp was washed with ultrapure water and filtered, after which the filtrate (black liquor) was collected. The black liquor was acidified to separate the supernatant and precipitation. The Cs (¹³³Cs and ¹³⁷Cs) concentrations in the chip and reaction products were measured. As for wood samples, the majority of Cs was present in black liquor, while only a minor amount of Cs was retained in the pulp (<1%). In the case of bark, although the majority of Cs was present in the black liquor, the proportion of Cs in the pulp was much higher than that in the wood pulp. In addition, the Cs in the precipitation of the bark was higher than that in the wood, possibly because the Cs in the bark was combined with some components, which is insoluble in alkaline solution. Our results suggest that ¹³⁷Cs-contaminated Cryptomeria japonica wood can be used in the pulp and paper industries.     

Spatial variations in nitrous oxide and nitric oxide emission potential on a slope of Japanese cedar (Cryptomeria japonica) forest

To quantify the spatial variation and spatial structure of nitrous oxide (N₂O) and nitric oxide (NO) emission from forest soils, we measured N₂O and NO emission rates from surface soil cores taken at 1 m intervals on a cross-line transect (65 m × 20 m) on a slope of Japanese cedar ( Cryptomeria japonica ) forest in a temperate region of central Japan and analyzed the spatial dependency of N oxide gas emissions using geostatistics. We divided N₂O emission into N₂O from denitrification and N₂O from nitrification using the acetylene inhibition method. According to the geostatistical analysis, N₂O emission rates on the slope had large spatial variation and weak spatial dependency. This weak spatial dependency was caused by the inordinately high N₂O emissions on the slope, which were derived mainly from denitrification. In contrast, NO emission rate on the slope had large spatial variation, but strong spatial dependency and a distinct spatial distribution related to slope position, that is, high in the middle of the slope and low in the shoulder and the foot of the slope. The CN ratio and water-filled pore space were the dominant factors controlling NO emission rate on a slope. Our results suggest that spatial information about topographic factors helps to improve the estimation of both N₂O emission and NO emission from forest soils.     

Unilateral basement membrane zone alteration of the regenerated laminar region in equine chronic laminitis

Between the laminar epidermis and the laminar dermis of laminar region (LR) in equine foot, it can be observed the basement membrane zone (BMZ), which is composed of a basement membrane and its accompaniments like the hemidesmosome and anchoring fibril. Alteration in the BMZ in equine laminitis is possibly related with not only development but also recovery outcome and recurrence of this disease. However, there is little known about the structure of the BMZ during the recovery phase of this disease. To assess the condition of the BMZ of LR affected by chronic laminitis, the tissue was examined in three cases at two weeks, four weeks and three months after the onset of laminitis, using pathological, immunohistochemical and electron microscopic techniques. Histologically in all laminitis cases, there was a regenerated laminar epidermis with proliferating keratinocytes between the Stratum medium and the dermis, but it included the undeveloped secondary epidermal laminae (ud-SELs) structure in one side of the primary epidermal laminae, especially in the part of the deep area of LR. Immunohistochemical results were positive for the anti-type IV collagen, anti-type VII collagen and anti-laminin 5 antibodies in the most BMZs. However, partial BMZs adjacent to the ud-SELs were negative for the anti-type VII collagen and anti-laminin 5 antibodies. Ultrastructurally, in the BMZ of the ud-SEL, the lamina densa and the lamina lucida were present. In contrast, the anchoring fibrils and the hemidesmosomes were either absent, or present at lower than normal levels. In conclusion, the present study indicated that the part of regenerated LR in chronic laminitis was not able to fully restore to construct the BMZ for a long time, especially in the unilateral side of laminar epidermis. It might be related with recurrence of this disease.     

Detection of New Ethylene-Producing Bacteria, Pseudomonas syringae pvs. cannabina and sesami, by PCR Amplification of Genes for the Ethylene-Forming Enzyme

ABSTRACT Strains of Pseudomonas syringae (78 strains and 43 pathovars) and other strains (79) of plant and insect origin were examined for the presence of the ethylene-forming enzyme gene (efe) by polymerase chain reaction (PCR) assay. The sequence of the efe gene of P. syringae pv. phaseolicola PK2 was used to design two primer sets for amplification of the gene. In addition to P.syringae pv. phaseolicola (the "kudzu strain") and P.syringae pv. glycinea, which were efficient ethylene producers, several strains of P.syringae pvs. sesami and cannabina generated PCR products of the predicted size. A DNA probe of the efe gene, isolated from strain PK2, hybridized to these PCR products, indicating homology to the P.syringae pv. phaseolicola efe gene. PCR restriction fragment length polymorphism analyses suggested that these four pathovars harbor a similar efe gene. Furthermore, the probe hybridized to an indigenous plasmid of P.syringae pv. cannabina, suggesting that the efe gene could be located on a plasmid in this pathovar, but did not hybridize to plas-mids of P.syringae pv. sesami strains. P.syringae pvs. sesami and cannabina strains produced ethylene in King's medium B at levels similar to those of P.syringae pvs. phaseolicola and glycinea. Thus, two new ethylene-producing bacteria were detected by the PCR assay.     

Effect of Ca on Al-induced activation of antioxidant enzymes in the needles of hinoki cypress (Chamaecyparis obtusa)

The effect of CA(NO₃)₂ on the active oxygen scavenging system in hinoki cypress (Chamaecyparis obtusa) seedlings cultured in a nutrient solution containing aluminum was examined. The hinoki cypress seedlings were transferred to nutrient solutions containing 5 mM AlCl₃ together with various concentrations of Ca(NO₃)₂ in pots containing glass beads and Teflon tips. The growth in height and dry matter allocation to each organ was little influenced over a period of 12 weeks by either Al or the concentration of Ca(NO₃)₂. The activity of superoxide dismutase (SOD) in the needles was stimulated by Al, and the effect of Al was lowered significantly by simultaneous application of 25 mM Ca(NO₃)₂. At week 1, the activity of catalase (CAT) in the needles was increased by Al, but the effect was no longer observed at week 12. The Al concentration in the roots was increased by treatment with Al, whereas the Al concentration in needles was not. These results indicate that rhizospheric Al stress stimulates antioxidative enzyme activities in hinoki cypress needles and the activation of the enzymes is suppressed by addition of Ca. The transmission of Al stress to the needles, which induced a change in the enzyme activity, is not caused by the transfer of the Al ion itself from roots to needles. This work was supported in part by funding from the Japan Science and Technology Corporation, the CREST program 1996–2001, and the Center for Forest Decline Studies.     

Evolutionary autoploidy in the sweet potato (Ipomoea batatas (L.) Lam.) and its progenitors

Summary The role of polyploidy in the evolution of the sweet potato. I. batatas (2n=6x=90), became more clear in 1971 when wild species with 30, 60, and 90 chromosomes were discovered. These species, I. leucantha (2x), I. littoralis (4x) and I. trifida (6x), are the progenitors of the sweet potato (6x), in an autopolyploid series with doubling of the I. leucantha B genome.In the present study the hypothesis of the origin of the sweet potato was confirmed by comparative studies on some plant characters, sexual compatibility, and the behavior of artificial hexaploids produced from I. leucantha (2x) and I. littoralis (4x).Since induced hexaploid I. leucantha exhibits weak growth, a few multivalents are formed at meiosis in the sweet potato, and there are differences in morphological and physiological characteristics between the artificial hexaploids and I. batatas, raw autoploidy seems unlikely, but a balanced or diploidised autoploidy could have been achieved by genic and chromosomal changes in the course of evolution. However, there is still sufficient homology so that meiotic pairing occurs usually between regular partners but also between homoeologous chromosomes in a certain situation.     

Genomic Affinity Relationships in the Genus Avena

Many interspecific hybrids between diploid, tetraploid and hexaploid Avena species were obtained without artificial embryo cultures. Most of the hybrids showed variable chromosome pairing with univalents, bivalents and multiple associations, even in PMC in the same hybrid, at first metaphase; making ii very difficult to understand the degree of genome affinity between parent species, and especially between different species. The basic or theoretical chromosome pairing consisting of bivalents and univalents is presented, in which the chromosome ratios (%) statistically agree with observations. In addition, the per cent frequency of chromosomes forming multivalents is calculated. On the bases of these numerical data, the genomic affinity relationships are discussed for present and previously reported interspecific hybrids or Avena.     

Changes in Chemical Species of Copper Added to Brown Forest Soil in Japan

Heavy metals are present as various chemical species in soil environments. Chemical species of heavy metals determinate their behavior in soil, so they must be more important concern than total contents in treating problems about heavy metal polluted soil. Microbial activity is thought to be one of the factors that affect chemical speciation of heavy metals. Although some possible microbial mechanisms have been suggested, their effects on heavy metals have not previously been thoroughly studied. In this study, we investigated whether microorganisms contribute to the fractionation of Cu in a forest soil. Copper-enriched, sterilized forest soil samples were buried at their places of origin, and subsamples were collected for analysis 3, 10, 17, 30, 60, and 90 days after burial. Tessier’s sequential extraction was employed to analyse Cu fractionation. Until two months after burial, the added Cu remained nearly unchanged. But a dramatic change was found in soil subsamples after three months. In both the A and B horizons, the carbonate fraction of Cu increased and the residual fraction decreased. We also found that clay minerals and persistent organic matter are important binding substances of Cu in the residual fraction. The change from residual to carbonate fraction could be caused by microbial activity.     

Evaluation of transgenic event CBH 351 (StarLink) corn in broiler chicks

The influence of transgenic event CBH 351 (StarLink; SL)-derived hybrid corn on the growth performance, health condition and physiological function in broiler chicks, as well as the possible transfer of the cry9C gene and Cry9C protein to blood, liver and muscle were examined in comparison with chicks fed on a diet with non-transgenic corn (SL-F). Bodyweight gain and feed conversion ratio in the chicks fed on a diet with SL were significantly greater than in chicks fed on a diet with SL-F during the starter phase (0–3 weeks of age), but this significant difference disappeared during the finisher phase (4–7 weeks of age). No abnormalities in health condition in either SL or SL-F groups were observed, and livability did not differ significantly between SL and SL-F groups. Moreover, no significant differences in serum biochemical and hematological values, histopathological observation and necropsy findings were observed between SL and SL-F groups at the end of the experiment. The cry9C gene and Cry9C protein were not detected in blood, liver and muscle of chicks at 3, 5 or 7 weeks of age. The results indicate that feeding SL does not influence growth performance, health condition or physiological function in broiler chicks, and the cry9C gene and Cry9C protein are not transferred to the blood, liver and muscles of broilers.