Analysis of protective mechanisms in cultured ayu, Plecoglossus altivelis Temminck and Schlegel, administered Vibrio vaccine by the immersion method

Abstract. The protective effect of Vibrio angnillarum immunization administered by the direct immersion technique in ayu, Plecoglossus altivelis Temminck and Schlegel, was investigated. The activities of phagoeytic cells were suppressed at the time of vaccination by previous bath exposures to a cyclophosphamide solution containing a concentration of 500 mg/kg/ml on days 2 and 3 before vaccination. An artificial challenge was carried out 3 weeks after vaccination by bath exposure. The survival rate of the vaccine group pretreated with cyclophosphamide was 96%. In the group given traditional vaccine only, 73% survived; and in the non-vaccine control group, 10% survived. These results suggest that protective immunity of ayu vaccinated by the direct immersion technique may not implicate the early activities of phagoeytic cells.     

High GUS expression in protoplasts isolated from immature peach fruits

A protocol for transient expression analysis was developed using protoplast isolated from immature peach fruits. The uid A gene for β-glucuronidase (GUS) was introduced into the protoplast as a reporter gene to evaluate the protoplast activity. Protoplasts isolated from immature fruits at 28–32 days after full bloom (DAFB) showed high GUS activity under the cauliflower mosaic virus 35S promoter. The highest GUS activity was obtained from the protoplasts isolated at 32 DAFB, but it was difficult to isolate protoplasts showing high GUS activity from 34 DAFB. A comparison of the effect of promoter cassette on gene expression showed that the promoter containing the tobacco mosaic virus Ω sequence enhanced GUS activity by at least 10-fold in the peach protoplasts relative to the 35S promoter. The level of GUS activity under the 35S promoter in the peach protoplasts was 0.47-fold as that obtained from maize protoplasts isolated from young greening leaves, indicating that the GUS activity of immature peach protoplast is sufficient for gene expression analysis. Since stable transformation and evaluation of fruit traits in peach transformants are difficult, this transient expression system could be useful for the characterization of genes expressed in peach and other Rosaceae fruit species.     

Rhizopus Rot of Mulberry-grafted Saplings Caused by Rhizopus oryzae

A new disease on stocks of mulberry-grafted saplings was found in Tsukuba in 1999. A representative isolate of the Rhizopus species that was consistently isolated from rotted tissues was pathogenic to healthy mulberry stocks. The causal fungus was identified morphologically as Rhizopus oryzae Went & Prinsen Geerligs. Rhizopus rot (“Naefuhaibyou” in Japanese) was proposed for the name of the disease. This is the first report on a mulberry disease caused by R. oryzae. Received 30 March 2001/ Accepted in revised form 4 June2001     

Properties of Cross‐Linked Starch from Waxy Mutant Wheat Tanikei A6599‐4

Native starch from waxy mutant wheat Tanikei A6599‐4 is known to exhibit more stable hot paste viscosity than a typical waxy wheat (Tanikei H1881) and waxy corn. The objective of this study was to investigate the starch paste properties of Tanikei A6599‐4 after cross‐linking and compare with Tanikei H1881 and waxy corn. As an example of cross‐linking, the reaction (at 30, 60, 120, and 360 min) with sodium trimetaphosphate was used. In Rapid Visco Analyser (RVA) measurement, the unique characteristic was maintained in Tanikei A6599‐4 starch cross‐linked at low reaction time (<120 min) levels. Cross‐linking at a high reaction time (360 min) level suppressed the swelling of both Tanikei A6599‐4 and Tanikei H1881 starches but not waxy corn starch. Although unmodified Tanikei A6599‐4 starch showed the lowest paste clarity among unmodified waxy starches, this defect became unremarkable when starch was cross‐linked for ≥120 min. In gel‐dispersed dynamic viscoelasticity measurement, the order of G′ and G″ values was always Tanikei A6599‐4 > Tanikei H1881 > waxy corn. This indicates that cross‐linked Tanikei A6599‐4 and Tanikei H1881 starches have different starch properties and that swollen Tanikei A6599‐4 starch granules are more rigid than swollen Tanikei H1881 starch granules.     

Interactions Among Wx Genes and Effects of a Mutated Wx‐D1e Allele on Starch Properties in Wheat

The waxy mutant wheat Tanikei A6599‐4 contains little amylose and exhibits a stable hot paste viscosity. It has null alleles at the Wx‐A1 and Wx‐B1 loci and a mutated allele at the Wx‐D1 locus (Wx‐D1e). From the cross‐combination of Kanto 123 (normal amylose line) and Tanikei A6599‐4, 51 DH (doubled haploid) lines were produced and their genotypes were determined by SDS‐PAGE and a derived cleaved amplified polymorphic sequence (dCAPS) analysis. Eight genotypes were obtained at the expected ratio. Using a Rapid Visco Analyser (RVA), all the lines with the same genotype as Tanikei A6599‐4 showed a stable hot paste viscosity. The other lines did not show a stable hot paste viscosity regardless of the presence of the Wx‐D1e allele. When two genotypes with the same Wx‐A1 and Wx‐B1 alleles were compared, the one with Wx‐D1e contained less amylose and exhibited a lower final viscosity and a lower setback with RVA. Although the Wx‐D1e allele produces an almost inactive Wx protein, these findings suggest that this allele contributes at some level to starch synthesis.     

Differentiation of stem progenitor CD9/SOX2-positive cells is promoted with increased prolactin-producing and endothelial cells in the pituitary

Sex-determining region Y-box 2 (SOX2)-positive cells are stem/progenitor cells in the adenohypophysis, comprising the anterior and intermediate lobes (AL and IL, respectively). The cells are located in the marginal cell layer (MCL) facing Rathke’s cleft (primary niche) and the parenchyma of the AL (secondary niche). We previously demonstrated in vitro that the tetraspanin superfamily CD9 and SOX2 double-positive (CD9/SOX2-positive) cells in the IL-side MCL migrate to the AL side and differentiate into hormone-producing and endothelial cells in the AL parenchyma. Here, we performed in vivo studies to evaluate the role of IL-side CD9/SOX2-positive cells in pregnancy, lactation, and treatment with diethylstilbestrol (DES; an estrogen analog) when an increased population of prolactin (PRL) cells was observed in the AL of the rat pituitary. The proportions of CD9/SOX2-, CD9/Ki67-, and PRL/TUNEL-positive cells decreased in the primary and secondary niches during pregnancy and DES treatment. In contrast, the number of CD9/PRL-positive cells increased in the AL-side MCL and AL parenchyma during pregnancy and during DES treatment. The proportion of PRL/Ki67-positive cells increased in the AL-side MCL and AL parenchyma in response to DES treatment. Next, we isolated CD9-positive cells from the IL-side MCL using an anti-CD9 antibody. During cell culture, the cells formed free-floating three-dimensional clusters (pituispheres). Furthermore, CD9-positive cells in the pituisphere differentiated into PRL cells, and their differentiation potential was promoted by DES. These findings suggest that CD9/SOX2-positive cells in the IL-side MCL may act as adult stem cells in the AL parenchyma that supply PRL cells under the influence of estrogen.     

Role of polyamines in peach fruit development and storage

To investigate the role of polyamines in pre- and post-harvest fruit development of 'Akatsuki' peach (Prunus persica (L.) Batsch.) we measured polyamine concentrations, activities of polyamine biosynthetic enzymes and expression of genes encoding these enzymes. Concentrations of the free polyamines, putrescine (Put), spermidine (Spd) and spermine (Spm) in pre-harvest fruit peaked 16 days after full bloom (DAF) and then progressively decreased until harvest with the exception of Put, which showed a second peak at 94 DAF, just before the onset of ethylene production. In post-harvest fruit, minor changes in concentrations of Spd and Spm were observed, whereas Put concentration peaked on the harvest day, followed by an abrupt decrease and a subsequent 2-fold increase, which was opposite to the fluctuating pattern of ethylene production. Activities of arginine decarboxylase (ADC) and ornithine decarboxylase (ODC) peaked during the first stage of fruit development and then decreased until 80 DAF, after which the activities were below detection limits, suggesting that Put is synthesized during the early stage of fruit development. Activity of S-adenosylmethionine decarboxylase (SAMDC) decreased progressively until the end of S2. Expression levels of five putative polyamine biosynthetic genes, ADC, ODC, SAMDC, spermidine synthase (SPDS) and spermine synthase (SPMS), in pre-harvest and post-harvest fruit did not coincide precisely with the observed changes in enzymatic activities and polyamine concentrations. The possible role of polyamines during peach fruit development and the relationship between polyamines and ethylene biosynthesis are discussed.     

Effect of phosphogypsum application on the growth and Ca uptake of Komatsuna Brassica rapa L. cv. Natsurakuten

(pp. 1–7)
Application of phosphogypsum PG and limestone to Andisol were compared in terms of their effects on the growth and uptake of Ca in Komatsuna Brassica rapa L. cv. Natsurakuten. The studies were carried out in soil pot cultures, and both PG and limestone were applied to the Andisol at the rates of 0, 0.30, 0.75, 1.50 g kg⁻¹ in the form of CaO.

• 1) 

  Limestone application resulted in an increase in the soil pH. On the other hand, no change was observed in the case of PG application at pH 6.1 ± 0.1 regardless of the application rate. Both soil EC and water-soluble Ca increased in proportion to the PG application rate, and this increase was approximately 5 times greater than that observed when limestone was applied at 1.50 g kg⁻¹. The w-Ca/ex-Ca ratio (water-soluble Ca: 1 M acetic acid-extractable Ca) was clearly increased from 0.12 up to 0.26 in PG, while the value was slightly decreased from 0.12 to 0.05 in limestone.