Japan Horticultural Production and Research Institute

Matsudo, Japan

Japan Horticultural Production and Research Institute

Matsudo, Japan
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Momma N.,Japan Horticultural Production and Research Institute | Momma M.,Japan Horticultural Production and Research Institute | Kobara Y.,Japan National Institute for Agro - Environmental Sciences
Journal of General Plant Pathology | Year: 2010

Better soil disinfestation methods, such as biological soil disinfestation (BSD), that are environmentally safe are increasingly been developed and used because of rising concerns related to environmental risks. We evaluated the efficacy of soil disinfestation using ethanol to control the fungus Fusarium oxysporum f. sp. lycopersici, which causes fusarium wilt of tomato. Survival of bud cells and chlamydospores declined markedly in soil saturated with diluted ethanol solution in the laboratory. In field trials, artificially added nonpathogenic Fusarium oxysporum and indigenous F. oxysporum were both strongly suppressed in soil saturated with 1% ethanol solution; a wheat bran treatment was not as effective. The artificially added fungus was not detected in three of four sites treated with ethanol but was detected in three of four sites amended with wheat bran. Using ethanol in pre-autoclaved soil was not suppressive; thus native microorganisms are essential for the suppression. This ethanol-mediated biological soil disinfestation (Et-BSD) temporarily increased the number of anaerobic bacteria, but the number of fungi and aerobic bacteria was stable. Polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) analysis revealed slight but apparent differences in bacterial community structures in the soil treated with Et-BSD compared with the structure in soils after other treatments such as water irrigation and in the control soil, which received neither organic amendment nor irrigation after 15 days. Et-BSD is a potentially effective and easy soil disinfestation method, and its impact on native, beneficial microorganisms is moderate. © 2010 The Phytopathological Society of Japan and Springer.


Momma N.,Japan Horticultural Production and Research Institute | Kobara Y.,Japan National Institute for Agro - Environmental Sciences | Momma M.,Japan Horticultural Production and Research Institute | Momma M.,Japan National Food Research Institute
Journal of General Plant Pathology | Year: 2011

Here Fusarium oxysporum was killed in exudates obtained from soil biologically disinfested with ethanol, indicating that physical interaction with soil microorganisms was not essential. Because acetic acid was confirmed to accumulated during the treatment, we evaluated the effect of acetic acid amendment against the pathogen in plastic containers. A drop in the soil redox potential seemed to be correlated with the fungicidal efficacy of acetic acid. Under reductive soil conditions, metal ions such as Mn2+ and Fe2+ formed, and the pathogen was effectively suppressed in Mn2+ and Fe2+ solution. Therefore, Fe2+ and Mn2+ may be the agents that induce suppression of the pathogen during biological soil disinfestation. © 2011 The Phytopathological Society of Japan and Springer.


Ogawa D.,Japan Horticultural Production and Research Institute | Ishikawa K.,Japan Horticultural Production and Research Institute | Mii M.,Chiba University
Scientia Horticulturae | Year: 2012

Polysomaty status of fruit pericarp was compared between a diploid cultivar of pepper (Capsicum annuum 'Shishitou No. 562') and its artificially induced triploid and tetraploid plants. In pepper fruits of different ploidy levels, fruit size as assessed by fruit length reached a plateau from 3 to 4 weeks after anthesis, and the maximum fruit length of triploid and tetraploid fruits was about 43% and 88% of the diploid counterpart, respectively. However, the maximum fruit diameter of both triploid and tetraploid plants was almost the same as that of diploid plant. Polysomaty status of fruit pericarp increased during fruit development and the maximum DNA content of the pericarp cells in the diploid, triploid and tetraploid fruits showed 64C, 96C and 128C, respectively. These results indicate that the same number of endoreduplication is programmed to occur in pepper fruits irrespective of the difference in the ploidy level of plants. © 2011 Elsevier B.V.


Ogawa D.,Japan Horticultural Production and Research Institute | Ishikawa K.,Japan Horticultural Production and Research Institute | Nunomura O.,Japan Horticultural Production and Research Institute | Mii M.,Chiba University
Journal of the Japanese Society for Horticultural Science | Year: 2010

The correlation between several fruit characters and the degree of polysomaty, i.e., the number of peaks in flow cytometric analysis, was examined using mature fruits of 12 genotypes from three species of Capsicum. Capsicum chacoense PI260429, which had the smallest fruit with the thinnest pericarp, showed the least number of peaks in ploidy levels, i.e., four ploidies ranging from 2C to 16C, whereas Capsicum annuum 'Édes alma' had the thickest pericarp and the highest peak numbers of eight ploidy levels, ranging from 2C to 256C. Among the morphological traits of fruit examined, pericarp thickness showed the highest correlation with the degree of polysomaty (r = 0.88). © 2010 JSHS.


Shishido M.,Chiba University | Sato K.,Japan Horticultural Production and Research Institute | Yoshida N.,Chiba University | Tsukui R.,Chiba University | Usami T.,Chiba University
Journal of General Plant Pathology | Year: 2010

We developed polymerase chain reaction (PCR) assays to detect and quantify Phomopsis sclerotioides, the causal agent of black root rot of cucurbits. We used internal transcribed spacers 1 and 2 of the ribosomal DNA (rDNA) from representative isolates to search for target sequences. Primer pairs were selected after testing against 40 fungal isolates including 13 Ph. sclerotioides isolates, 9 Phomopsis isolates other than Ph. sclerotioides, and 18 soilborne fungi that were either pathogenic or nonpathogenic to cucurbits. Conventional PCR assays with the primer pair of CPs-1 (forward) and CPs-2 (reverse) produced target DNA amplicons from all Ph. sclerotioides isolates but none of the other isolates tested. From soil and root samples collected from fields naturally infested with black root rot of cucumber and melon, the CPs-1/CPs-2 primer pair successfully amplified target DNA fragments in conventional PCR assays. Moreover, we applied the CPs-1/CPs-2 primer pair in a real-time PCR assay with SYBR Green I, and PCR-amplified products were successfully quantified by reference to a standard curve generated by adding known amounts of target DNA. Target Ph. sclerotioides DNA fragments were similarly detected in artificially inoculated roots of cucumber, melon, pumpkin, and watermelon, but quantities of Ph. sclerotioides DNA in their hypocotyls of the hosts varied as follows: melon ≥ cucumber ≥ watermelon > pumpkin. These results suggest that Ph. sclerotioides infection is not species-specific but the rate of infection may differ among host species. © 2009 The Phytopathological Society of Japan and Springer.


Genda Y.,Japan Horticultural Production and Research Institute | Sato K.,Japan Horticultural Production and Research Institute | Nunomura O.,Japan Horticultural Production and Research Institute | Hirabayashi T.,Japan Horticultural Production and Research Institute | Tsuda S.,Japan National Agricultural Research Center
Journal of General Plant Pathology | Year: 2011

The location of Pepper mild mottle virus (PMMoV) within seeds as they developed on inoculated seedlings of pepper (Capsicum annuum) was followed over time by detecting the viral coat protein using immunofluorescence microscopy. Seedlings were inoculated with PMMoV when the flower buds on the first and second branching nodes were in bloom. Fluorescence indicating the presence of PMMoV was first observed around immature seeds and placentas in the ovaries on the fourth branching node at 20 days post-anthesis (20 DPA), which corresponded to 39 days post-inoculation (39 DPI). The area with fluorescence gradually expanded from the placenta into the integument and the parenchyma, and finally reached the tip of the immature seeds by 34 DPA (53 DPI). The embryo or endosperm beyond the endothelium never fluoresced during the experiment [i. e., ending at 81 DPA (102 DPI)]. For visualizing viral routes of invasion from seeds into new seedlings, PMMoV-infected C. annuum seeds that were heterozygous for the L 3 tobamovirus-resistance gene were sown in soil at 30°C. After ~2 weeks, the cotyledon developed virally induced necrosis. These findings shed light on the infection cycle of PMMoV through vertical transmission in C. annuum. © 2011 The Phytopathological Society of Japan and Springer.


Jitsopakul N.,Mahidol University | Thammasiri K.,Mahidol University | Ishikawa K.,Japan Horticultural Production and Research Institute
Acta Horticulturae | Year: 2011

Vanda coerulea is the most popular Vanda species of Thailand that has become endangered because of the wild orchid trade, deforestation and environmental changes. To preserve this orchid, droplet-vitrification was studied for the cryopreservation of protocorm-like bodies. Protocorm-like bodies were developed from one protocrom in ND liquid medium supplemented with 1 mg/L BA in combination with 0.5 mg/L NAA and 30 g/L maltose. Protocorm-like bodies (3.0 mm in diameter) were precultured in ND liquid medium supplemented with 0.5 M sucrose for 1 day on a shaker, then dehydrated with loading solution for 15 min and exposed to PVS2 solution for 30 min at 25°C. The plant materials were then immersed in liquid nitrogen, rewarmed rapidly and cultured on solidified ND medium. After cryopreservation, the survival frequency of cryopreserved protocorm-like bodies was 5%. The assessment of the genetic stability of cryopreserved protocorm-like bodies was developed using RAPD markers, size and sequence of the trnL (UAA) non-coding region of cpDNA. Results showed the same RAPD patterns of non-cryopreserved and cryopreserved protocorm-like bodies. The size and sequences of the trnL (UAA) non-coding region of cpDNA for noncryopreserved and cryopreserved protocorm-like bodies were not different. There was no difference in morphology and similar patterns of ploidy in plantlets developed from non-cryopreserved and cryopreserved protocorm-like bodies.


Sopalun K.,Mahidol University | Thammasiri K.,Mahidol University | Ishikawa K.,Japan Horticultural Production and Research Institute
Cryo-Letters | Year: 2010

Three vitrification-based methods for the cryopreservation of Grammatophyllum speciosum protocorms were invesigated: droplet-vitrification, encapsulation-dehydration and encapsulation-vitrification. Protocorms, 0.1 cm in diameter, developed from 2-month-old germinating seeds were used. For droplet-vitrification, protocorms were precultured on filter paper soaked in half strength Murashige and Skoog medium (1/2MS) containing 0.4 M sucrose at 25 ± 2°C for 2 d, followed by soaking in loading solution (2 M glycerol and 0.4 M sucrose in 1/2MS liquid medium) for 20 min and then dehydrated with PVS2 solution [30% (w/v) glycerol, 15% (w/v) ethylene glycol and 15% (w/v) dimethyl sulfoxide in 1/2MS liquid medium containing 0.4 M sucrose at pH 5.7] for 30 min. For encapsulation-dehydration, encapsulated protocorms were precultured in 1/2MS liquid medium containing 0.4 M sucrose on a shaker (110 rpm) at 25 ± 2°C for 2 d, followed by soaking in the same loading solution for 20 min and then exposed to a sterile air-flow at 2.5 inches/water column from the laminar air-flow cabinet for 8 h. For encapsulation- vitrification, encapsulated protocorms were precultured in 1/2MS liquid medium containing 0.4 M sucrose for 1 or 2 d, followed by soaking in the same loading solution for 20 min and then dehydrated with PVS2 solution for 60 min. For all three methods, preculturing with 0.4 M sucrose for 2 d resulted in a significant induction of dehydration and freezing tolerance. The cryopreservation results showed highest protocorm regrowth after droplet-vitrification (38%), followed by encapsulation-dehydration (24%) and encapsulation-vitrification (14%). Plantlets developed from these three methods did not show any abnormal characteristics or ploidy level change when investigated by flow cytometry. © CryoLetters.


Sopalun K.,Mahidol University | Thammasiri K.,Mahidol University | Ishikawa K.,Japan Horticultural Production and Research Institute
World Academy of Science, Engineering and Technology | Year: 2010

The effects of chitosan, a biodegradable polymer, were studied in Grammatophyllum speciosum protocorm-like bodies (PLBs) in vitro culture. The chitosan concentration of 0, 5, 10, 15, 20, 25, 50 or 100 mg/l were supplemented in half-strength Murashige and Skoog (1/2 MS) liquid or on agar media containing 2% (w/v) sucrose. The results showed that liquid medium supplemented with 15 mg/l chitosan showed the highest relative growth rate (7-fold increase) of PLBs. On 1/2 MS agar medium supplemented with 25 mg/l chitosan gave the highest relative growth rate (4-fold increase). The relative growth rate of G. speciosum PLBs on agar medium was significantly lower than that in liquid medium. Moreover, chitosan, supplemented to agar medium promoted shoot formation but not rooting. However, supplementation at too high a level, such as 100 mg/l can inhibit growth and kill PLBs.


Sopalun K.,Mahidol University | Thammasiri K.,Mahidol University | Ishikawa K.,Japan Horticultural Production and Research Institute
Plant Cell, Tissue and Organ Culture | Year: 2010

Protocorm-like bodies (PLBs) were induced from shoot tips of Grammatophyllum speciosum, a Thai orchid. The highest frequency of PLBs (93%) were observed on explants incubated on 1/2-Murashige and Skoog (MS) liquid medium containing 2% (w/v) sucrose without any plant growth regulators (PGRs). Tests with different carbon sources compared to sucrose revealed that maltose promoted the highest relative growth of G. speciosum PLBs (7-fold increase), while trehalose and sucrose yielded 5-fold and 4-fold increases, respectively. In 1/2 MS liquid medium, addition of 15 mg/l of chitosan promoted a 7-fold increase in PLB growth while 25 mg/l promoted a 4-fold increase. However, the relative growth rate in solid culture was significantly lower than that in liquid culture. In addition, chitosan supplementation in solid medium promoted shoot formation but not rooting. Plantlet regeneration was induced using a combination of NAA and BA supplementation in 1/2 MS solid medium with optimum induction shoot and root formation at 2.0 mg/l NAA and 1.0 mg/l BA. Using this protocol, approximately 8 months was required to obtain a hundred plantlets from one shoot tip. The plantlets showed no changes in ploidy when tested by flow cytometry. © 2010 Springer Science+Business Media B.V.

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