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Hayashi S.,Japan National Agriculture and Food Research Organization | Shigematsu K.,Japan National Agriculture and Food Research Organization | Yamamoto S.,Japan National Agriculture and Food Research Organization | Kobayashi K.,Japan National Agriculture and Food Research Organization | And 3 more authors.
Biosystems Engineering | Year: 2010

We developed a strawberry-harvesting robot, consisting of a cylindrical manipulator, end-effector, machine vision unit, storage unit and travelling unit, for application to an elevated substrate culture. The robot was based on the development concepts of night operation, peduncle handling and task sharing with workers, to overcome the robotic harvesting problems identified by previous studies, such as low work efficiency, low success rate, fruit damage, difficulty of detection in unstable illumination and high cost. In functional tests, the machine vision assessments of fruit maturity agreed with human assessments for the Amaotome and Beni-hoppe cultivars, but the performance for Amaotome was significantly better. Moreover, the machine vision unit correctly detected a peduncle of the target fruit at a rate of 60%. In harvesting tests conducted throughout the harvest season on target fruits with a maturity of 80% or more, the successful harvesting rate of the system was 41.3% when fruits were picked using a suction device before cutting the peduncle, while the rate was 34.9% when fruits were picked without suction. There were no significant differences between the two picking methods in terms of unsuccessful picking rates. The execution time for the successful harvest of a single fruit, including the time taken to transfer the harvested fruit to a tray, was 11.5 s. © 2009 IAgrE.


Nishizaki Y.,Tokyo University of Agriculture and Technology | Yasunaga M.,Tokyo University of Agriculture and Technology | Okamoto E.,Tokyo University of Agriculture and Technology | Okamoto M.,Ehime Research Institute of Agriculture | And 5 more authors.
Plant Cell | Year: 2013

The blue color of delphinium (Delphinium grandiflorum) flowers is produced by two 7-polyacylated anthocyanins, violdelphin and cyanodelphin. Violdelphin is derived from the chromophore delphinidin that has been modified at the 7-position by Glc and p-hydroxybenzoic acid (pHBA) molecules. Modification of violdelphin by linear conjugation of Glc and pHBA molecules to a Glc moiety at the 7-position produces cyanodelphin. We recently showed that anthocyanin 7-O-glucosylation in delphinium is catalyzed by the acyl-Glc-dependent anthocyanin glucosyltransferase (AAGT). Here, we sought to answer the question of which enzyme activities are necessary for catalyzing the transfer of Glc and pHBA moieties to 7-glucosylated anthocyanin. We found that these transfers were catalyzed by enzymes that use p-hydroxybenzoyl-Glc (pHBG) as a bifunctional acyl and glucosyl donor. In addition, we determined that violdelphin is synthesized via step-by-step enzymatic reactions catalyzed by two enzymes that use pHBG as an acyl or glucosyl donor. We also isolated a cDNA encoding a protein that has the potential for p-hydroxybenzoylation activity and two AAGT cDNAs that encode a protein capable of adding Glc to delphinidin 3-Orutinoside- 7-O-(6-O-[p-hydroxybenzoyl]-glucoside) to form violdelphin. © 2013 American Society of Plant Biologists.


Nishi S.,Hiroshima University | Yamashita H.,Ehime Research Institute of Agriculture | Kawato Y.,Japan National Research Institute of Fisheries And Environment of Inland Sea | Nakai T.,Hiroshima University
Applied and Environmental Microbiology | Year: 2016

Piscine nodavirus (betanodavirus) is the causative agent of viral nervous necrosis (VNN) in a variety of cultured fish species, particularly marine fish. In the present study, we developed a sensitive method for cell culture isolation of the virus from seawater and applied the method to a spontaneous fish-rearing environment. The virus in seawater was concentrated by an iron-based flocculation method and subjected to isolation with E-11 cells. A real-time reverse transcriptase PCR (RT-PCR) assay was used to quantify the virus in water. After spiking into seawater was performed, a betanodavirus strain (redspotted grouper nervous necrosis virus [RGNNV] genotype) was effectively recovered in the E-11 cells at a detection limit of approximately 105 copies (equivalent to 102 50% tissue culture infective doses [TCID50])/liter seawater. In an experimental infection of juvenile sevenband grouper (Epinephelus septemfasciatus) with the virus, the virus was isolated from the drainage of a fish-rearing tank when the virus level in water was at least approximately 105 copies/liter. The application of this method to sevenband grouper-rearing floating net pens, where VNN prevailed, resulted in the successful isolation of the virus from seawater. No differences were found in the partial sequences of the coat protein gene (RNA2) between the clinical virus isolates of dead fish and the cell-cultured virus isolates from seawater, and the viruses were identified as RGNNV. The infection experiment showed that the virus isolates from seawater were virulent to sevenband grouper. These results showed direct evidence of the horizontal transmission of betanodavirus via rearing water in marine aquaculture. © 2016, American Society for Microbiology. All Rights Reserved.


PubMed | Hiroshima University, Japan National Research Institute of Fisheries And Environment of Inland Sea and Ehime Research Institute of Agriculture
Type: Journal Article | Journal: Applied and environmental microbiology | Year: 2016

Piscine nodavirus (betanodavirus) is the causative agent of viral nervous necrosis (VNN) in a variety of cultured fish species, particularly marine fish. In the present study, we developed a sensitive method for cell culture isolation of the virus from seawater and applied the method to a spontaneous fish-rearing environment. The virus in seawater was concentrated by an iron-based flocculation method and subjected to isolation with E-11 cells. A real-time reverse transcriptase PCR (RT-PCR) assay was used to quantify the virus in water. After spiking into seawater was performed, a betanodavirus strain (red spotted grouper nervous necrosis virus [RGNNV] genotype) was effectively recovered in the E-11 cells at a detection limit of approximately 10(5)copies (equivalent to 10(2)50% tissue culture infective doses [TCID50])/liter seawater. In an experimental infection of juvenile sevenband grouper (Epinephelus septemfasciatus) with the virus, the virus was isolated from the drainage of a fish-rearing tank when the virus level in water was at least approximately 10(5)copies/liter. The application of this method to seven band grouper-rearing floating net pens, where VNN prevailed, resulted in the successful isolation of the virus from seawater. No differences were found in the partial sequences of the coat protein gene (RNA2) between the clinical virus isolates of dead fish and the cell-cultured virus isolates from seawater, and the viruses were identified as RGNNV. The infection experiment showed that the virus isolates from seawater were virulent to seven band grouper. These results showed direct evidence of the horizontal transmission of betanodavirus via rearing water in marine aquaculture.


Nishizaki Y.,Tokyo University of Agriculture and Technology | Sasaki N.,Tokyo University of Agriculture and Technology | Yasunaga M.,Tokyo University of Agriculture and Technology | Miyahara T.,Tokyo University of Agriculture and Technology | And 4 more authors.
Journal of Experimental Botany | Year: 2014

In delphiniums (Delphinium grandiflorum), blue flowers are produced by the presence of 7-polyacylated anthocyanins. The polyacyl moiety is composed of glucose and p-hydroxybenzoic acid (pHBA). The 7-polyacylation of anthocyanin has been shown to be catalysed by two different enzymes, a glucosyltransferase and an acyltransferase; both enzymes utilize p-hydroxybenzoyl-glucose (pHBG) as a bi-functional (Zwitter) donor. To date, however, the enzyme that synthesizes pHBG and the gene that encodes it have not been elucidated. Here, five delphinium cultivars were investigated and found to show reduced or undetectable 7-polyacylation activity; these cultivars synthesized delphinidin 3-O-rutinoside (Dp3R) to produce mauve sepals. One cultivar showed a deficiency for the acyl-glucose-dependent anthocyanin 7-O-glucosyltransferase (AA7GT) necessary for mediating the first step of 7-polyacylation. The other four cultivars showed both AA7GT activity and DgAA7GT expression; nevertheless, pHBG accumulation was significantly reduced compared with wild-type cultivars, whereas p-glucosyl-oxybenzoic acid (pGBA) was accumulated. Three candidate cDNAs encoding a UDP-glucose-dependent pHBA glucosyltransferase (pHBAGT) were identified. A phylogenetic analysis of DgpHBAGT amino acid sequences showed a close relationship with UGTs that act in acyl-glucose synthesis in other plant species. Recombinant DgpHBAGT protein synthesized pHBG and had a high preference for pHBA in vitro. Mutant cultivars accumulating pGBA had very low expression of DgpHBAGT, whereas expression during the development of sepals and tissues in a wild cultivar showed a close correlation to the level of accumulation of pHBG. These results support the conclusion that DgpHBAGT is responsible for in vivo synthesis of pHBG in delphiniums. © 2014 The Author.


PubMed | Japan National Research Institute of Fisheries Science, Japan National Research Institute of Fisheries And Environment of Inland Sea, Ehime Research Institute of Agriculture and Tokyo University of Marine Science and Technology
Type: Journal Article | Journal: Marine biotechnology (New York, N.Y.) | Year: 2016

To initiate breeding programs for kelp grouper (Epinephelus bruneus), the establishment of genetic linkage maps becomes essential accompanied by the search for quantitative trait loci that may be utilized in selection programs. We constructed a high-resolution genetic linkage map using 1055 simple sequence repeat (SSR) markers in an F1 family. Genome-wide and chromosome-wide significances of growth-related quantitative trait loci (QTLs) (body weight (BW) and total length (TL)) were detected using non-parametric mapping, Kruskal-Wallis (K-W) analysis, simple interval mapping (IM) and a permutation test (PT). Two stages and two families of fish were used to confirm the QTL regions. Ultimately, 714 SSR markers were matched that evenly covered the 24 linkage groups. In total, 509 and 512 markers were localized to the female and male maps, respectively. The genome lengths were approximately 1475.95 and 1370.39 cM and covered 84.68 and 83.21% of the genome, with an average interval of 4.1 and 4.0 cM, in females and males, respectively. One major QTL affecting BW and TL was found on linkage group EBR 17F that identified for 1% of the genome-wide significance and accounted for 14.6-18.9 and 14.7-18.5% of the phenotypic variance, and several putative QTL with 5% chromosome-wide significance were detected on eight linkage groups. Furthermore, the confirmed results of the regions harboring the major and putative QTLs showed consistent significant experiment-wide values of 1 and 5% as well as a chromosome-wide value of 5%. We identified growth-related QTLs that could be applied to find candidate genes for growth traits in further studies, and potentially useful in MAS breeding.


Irie K.,Ehime University | Kawaguchi M.,Ehime University | Mizuno K.,Ehime Research Institute of Agriculture | Song J.-Y.,Ehime University | And 3 more authors.
Marine Pollution Bulletin | Year: 2011

Heavy oil (HO) on the sea surface penetrates into fish eggs and prevents the normal morphogenesis. To identify the toxicological effects of HO in the context of the egg types, we performed exposure experiments using floating eggs and sinking eggs. In the course of development, HO-exposed embryos of floating eggs showed abnormal morphology, whereas early larva of the sinking eggs had almost normal morphology. However, the developing peripheral nervous system of sinking eggs showed abnormal projections. These findings suggest that HO exposed fishes have problems in the developing neurons, although they have no morphological malformations. Through these observations, we conclude that HO is strongly toxic to floating eggs in the morphogenesis, and also affect the neuron development in both floating and sinking eggs. © 2011 Elsevier Ltd.


Ishida M.,Ehime University | Nishi K.,Ehime University | Watanabe H.,Ehime Research Institute of Agriculture | Sugahara T.,Ehime University
Food Chemistry | Year: 2013

The inhibitory effect of an aqueous extract from spinach on degranulation of RBL-2H3 cells is herein reported. The extract significantly suppressed antigen-induced degranulation in a dose-dependent manner without affecting cell viability. Active substances in the extract were heat-stable and trypsin-resistant with molecular weights ranging from 500 Da to 14 kDa. The extract inhibited elevation of the intracellular Ca2+ concentration caused by stimulation by antigen, while not suppressing degranulation induced by a calcium ionophore A23187. Immunoblot analysis revealed that the inhibitory effect results from downregulation of phosphorylation of both Syk kinase and phosphatidylinositol 3-kinase in the signalling pathways involved in degranulation caused by the antigen-antibody interaction. Taken together, these findings suggest that aqueous spinach extract has an anti-allergic activity that controls degranulation. © 2012 Elsevier Ltd. All rights reserved.


Kadowaki K.,Ehime University | Kurisaka N.,Ehime Research Institute of Agriculture
IEEJ Transactions on Fundamentals and Materials | Year: 2013

This paper presents an experimental study on control of germination for Arabidopsis seed with a barrier discharge produced by polarity-reversed voltage pulses. 100 ns-width polarity-reversed pulses are applied to the seeds between plane electrodes covered by glass barriers. The electrode configuration allows the barrier discharge propagation along the seed surface at a relatively low voltage. After the discharge treatment, the seeds are incubated for 2 or 3 days to measure the germination rate. Relationships between the cumulative input energy density into plasma and the germination rate are investigated. Germination rate is significantly reduced by 20 J/cm3. However, it increases when the cumulative energy density is up to 50 J/cm3 and then it decreases again by the further energy input. These results propose a superposition of two effects, stimulation and inhibition, of the discharge treatment on the germination of Arabidopsis seed. Results of evans-blue dyeing for the discharge-exposed seeds indicate that necrosis of the seed coat is caused by the discharge treatment. © 2013 The Institute of Electrical Engineers of Japan.


Thanasaksiri K.,Tokyo University of Marine Science and Technology | Sakai N.,Tokyo University of Marine Science and Technology | Yamashita H.,Ehime Research Institute of Agriculture | Hirono I.,Tokyo University of Marine Science and Technology | Kondo H.,Tokyo University of Marine Science and Technology
Fish and Shellfish Immunology | Year: 2014

Influence of temperature on the susceptibility of fish against virus infection has been studied for a decade. Recent reports have been shown the effects of rearing temperatures on the fish immune system against virus infection. However, the roles of temperature in regulation of type I interferon (IFN) system has not yet been investigated. Thus, the effects of temperature on type I IFN response were investigated in this study using poly (I:C) injection in sevenband grouper and Mx gene was used as a marker for type I IFN expression. Quantitative real-time PCR (qPCR) result showed that Mx expression profiles were moderately different between temperatures. The highly up-regulated Mx transcripts at 3hpost injection (hpi) were observed in high temperatures (25°C and 30°C) but not in low temperatures (15°C and 20°C). Meanwhile, low temperatures (15°C and 20°C) could detect the highly up-regulated Mx transcripts at 24hpi. Expression of Mx transcripts was also observed at 72hpi at 15°C. Poly (I:C)-injected fish were challenged with RGNNV after 72 and 168hpi. At 72hpi, 100% of fish survived at all temperatures, whereas 95% survival rate was observed at 168hpi at 25°C during 14 days of observation. To further verify the duration period of an antiviral state at different temperatures, qPCR and endpoint dilution assay were used to quantify the number of virus in fish challenged with RGNNV. The reduction of viral copy numbers and viral titers could be observed at 72 and 168hpi. However, high viral copy numbers and viral titers could be detected at 168hpi at 30°C. These results demonstrate that temperatures influenced on the Mx expression profiles and the duration period of an antiviral state efficiently interfered with virus replication at different temperatures. © 2014 Elsevier Ltd.

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