Bulletin of The Iwate Agricultural Research Center No.2
Summary

Studies on Fusarium Wilt of Spinach
Kazunori KATSUBE

@The outbreak of soil-borne diseases inflicts economic damage on spinach cropping. In this study, the incidence of soil-borne diseases of spinach was surveyed at major cropping areas in Iwate prefecture. It was revealed that Fusarium wilt of spinach, caused by Fusarium oxysporum f. sp. spinaciae, has been the most serious disease. Based on these results, distinctive incidence of Fusarium wilt among cropping areas was analyzed by comparing the isolation rate of pathogenic fungi and distribution of vegetative compatibility groups with nitrate non-utilizing (nit) mutants. Finally, control methods were researched without chemical fungicides. Results obtained were as follows.

1. Surveys of incidence of soil-borne diseases of spinach at major cropping areas in Iwate prefecture
@At major cropping areas such as Tono, Nishine and Yamagata, outbreaks of soil-borne diseases of spinach were observed from June to September at the harvesting periods, that is, Fusarium wilt, root rot, damping-off and foot rot. The incidence of outbreaks distinguished the areas as follows. In Tono, incidence of Fusarium wilt and root rot disease was recognized. In Nishine, incidence of root rot disease was mainly recognized, and Fusarium wilt was also observed. And in Yamagata, although incidence of soil-borne disease was not abundant, incidence of Fusarium wilt was observed. Damping-off was not abundant and foot rot was hardly observed. The relationships between these distinctive incidence and soil-type, or continuous cropping years were not founded. However, Fusarium wilt disease was most important for spinach cropping because the outbreak of this disease has been observed in different many areas.

2. Establishment of the methods for pathogenicity test of Fusarium oxysporum isolated from wilted spinach
@The methods for the pathogenicity test of F. oxysporum isolated from spinach were studied. The inoculated density to soil for disease incidence was 10
2 colony forming units / g of soil (cfu), and 104 cfu was needed for disease development. In this case, the disease development using a budcell suspension for inoculum was used as well as using conidia in infested soil. The most susceptible growth period of spinach for pathogenic fungi was at the emergence-cotyledon period, and then disease severity was higher than inoculation at the sowing period or after 2 - 4 leaves had developed. The most sensitive spinach cultivars were cv. 'Okame', 'Magic' and 'Oracle', because these varieties wilted without differences in severity against all tested pathogenic isolates. The next method of pathogenicity test was simple and was suited to test many isolates isolated from wilted spinach as follows. Spinach seedlings were grown to the emergence-cotyledon period in a nursery on a plug tray (128 cells, using mixed soil for plug seedlings), and then were drenched evenly with budcells suspension (107 - 108 budcells/ml). After 2 - 3 weeks, the pathogenicity of isolates were assessed with the simple disease rating that surveyed them on the whole of the treatment. The isolation rates of pathogenic F. oxysporum obtained from wilted spinach in major cropping areas were investigated with the above pathogenicity test. It was found that the isolation rate was high in Tono (91.7%) and Yamagata (69.0%), but the rate was low in Nishine (45.3%), a result that approximately agreed with survey incidence of fusarium wilt at major spinach dropping areas.

3. Distribution of vegetative compatibility groups of Fusarium oxysporum recovered from wilted spinach
@Geographic distribution and population structure of mono-cultured spinach of Japanese isolates of F. oxysporum f.sp. spinaciae in Japan and Iwate prefecture were studied on the basis of vegetative compatibility groups (VCGs) with nit mutants and pathogenicity tests. VCG0330 and 0331 are distributed widely in Japan. Although, VCG0332 was detected only in one prefecture from which many isolates were tested, VCG0332 must also be present in other prefectures because VCG0332 is distributed widely in Iwate prefecture. In every case@such as in Japan, Iwate, major cropping areas and a field, VCG0330 existed preferentially and VCG0331 and 0332 were minor groups. In the field, isolates of F. oxysporum f. sp. spinaciae had diversity with one major VCG and other minor groups. Seasonable variation of VCGs composition in Iwate prefecture was found. Although the composition rate of minor groups, VCG0331 and 0332, had reached a peak in summer cropping, these isolates could not be detected in autumn cropping. On the other hand, the rate of the major group VCG0330 gradually became higher as continuous cropping was increased. VCG0330 may be easy to spread with the above condition. Optimum growth temperatures of standard isolates of 3 VCGs were equal at 20 - 30Ž. Pathogenic isolates which were not compatible with the three pathogenic VCGs nor with pathogenic isolates were found widely in Iwate prefecture, Japan.

4. Evaluation of spinach cultivars for resistance to Fusarium wilt disease applying pathogenicity test
@The established method for the pathogenicity test of F. oxysporum isolated from wilted spinach was applied to the evaluation of spinach cultivars for resistance to Fusarium wilt. The differentiation of variety resistance was observed with the inoculation method with the following conditions. Inoculum density was prepared at 10
5 - 106 budcells/ml, and then the seedlings were inoculated with isolates using a repeating syringe. Resistance was classified into 5 levels by a frequency distribution with an average disease index of 25 cultivars, inoculated with 5 isolates of pathogenic fungi, respectively. Comparative cultivars were decided at respective resistant levels for relative evaluations and were as follows. 'Highly resistant': 'Ujo' ; 'Lightly resistant': 'Atras' or 'Solomone' ; 'Intermediate': 'Baltic' and 'Active' ; 'Slightly susceptible': 'Munsterland' and 'Okame' ; 'Highly susceptible': 'King of Denmark' and 'Magic'. The host-pathogen interaction was statistically significant at the 1% level on the analysis of variance in disease severity. However, the interactive variance ratio was small in comparison with variations in fungal virulence and host resistance.

5. Control methods of Fusarium wilt of spinach
[1] Biological control of Fusarium wilt of spinach with non-pathogenic F. oxysporum

@Many isolates of F. oxysporum were recovered from spinach grown in the field infested with Fusarium wilt pathogen. Non-pathogenic strains were included in these isolates. Non-pathogenic F. oxysporum isolates S3HO3, S1HI1-w, 970211, 970481, 970576 and 970600 showed cross-protection against F. oxysporum f. sp. spinaciae when isolates were inoculated by soil-application before infestation with the pathogen. F. oxysporum isolate S3HO3 was mainly used for development of control methods. Non-pathogenic F. oxysporum isolate must be inoculated into soil before infestation with pathogenic fungi. An inoculum density of more than 104 bud-cells / g of soil and 10 - 100 times the pathogen inoculum density was required for the soil-application of bud-cell suspension. This cross-protection, however, did not continue through the harvesting. In order to extend the duration of disease suppression, pre-treatment of spinach seedlings with non-pathogenic F. oxysporum S3HO3 was combined with subsequent transplanting to reduce disease development. The most effective pre-treatment method for seedlings was mixing a bud-cell suspension (10
6 cfu/dry@soil g)@with nursery soil before sowing and then growing seedlings for 15 days before transplanting. Disease suppression then lasted through harvesting with a high protective value when pre-treated seedlings were transplanted in infested soil. Non-pathogenic F. oxysporum S3HO3 was not pathogenic for spinach and many other vegetable crops. A practical control test of Fusarium wilt of spinach by transplanting seedlings pre-treated with non-pathogenic F. oxysporum was assessed in the natural infested field. The protective effect of this method was even greater than that obtained by direct sowing, and more effective than transplanting without non-pathogenic F. oxysporum. And that was as effective as solar heating sterilization in a closed green plastic house in summer. The protective effect lasted through harvesting. These results suggest that control of Fusarium wilt of spinach by transplanting culture with non-pathogenic F. oxysporum should be practical. In addition, non-pathogenic F. oxysporum was able to suppress not only Fusarium wilt but also Pythium damping-off on spinach and other pathogenic fusaria of cucumber, melon, watermelon, radish and garland chrysanthemum when non-pathogenic F. oxysporum was added to the soil before inoculation of respective plant pathogens. Furthermore, the mechanisms of disease suppression by non-pathogenic F. oxysporum were studied. The mechanical plugging of wounds with bud-cell pellets of non-pathogenic F. oxysporum induced the cross-protection in spinach plants. The culture broth filtered with a 0.45ƒΚm mesh membrane filter had the ability to inhibit pathogen growth, such as the inhibition of conidia germination or the elongation of germtubes. Finally, the result of the prosperity and decay of both densities of non-pathogenic F. oxysporum and pathogen in root tissue suggests that competition with each fungus has appeared in spinach root tissue. This is because colony formation of non-pathogenic F. oxysporum recovered from healthy spinach root tissue was more than that of pathogen, but colony formation of non-pathogenic F. oxysporum recovered from diseased root tissue was less than that of pathogen. In addition, the detected period of pathogen from root tissue inoculated with non-pathogenic F. oxysporum was later than the one without non-pathogenic F. oxysporum. These result suggest that competition has appeared in spinach root between non-pathogenic F. oxysporum and pathogenic F. oxysporum.

[2] Control of Fusarium wilt of spinach by solar heating sterilization in the closed plastic house
@In Iwate prefecture, solar heating sterilization has not succeeded before. The reason for this failure was considered to be the difficult of raising the soil temperature. In order to address this problem, mulching plus plastic film used to raise the effective soil temperature@(40Ž at depth of 10cm), and no cultivation after sterilization to avoid confusion of sterilized soil and non sterilized soil, were studied in the naturally infested field. Solar heated sterilization was applied from June 26 to July 25 and there were 131 hours of the effective soil temperature. High protective effect was observed for this treatment before summer harvesting. The daily hours of the effective soil temperature correlated with the sunshine duration time of the meteorological observation point in Tono (AMeDAS observation point). The above result suggests that solar heating sterilization is also a practical control method during June and July in Iwate prefecture.

[3] Effect and durability of disease suppression of Fusarium wilt of spinach by soil-application with composted chitin materials
@Composted crab shells were used for suppression of Fusarium wilt of spinach in this study. In pot level and the field level studies, disease suppression with soil application of this material after soil sterilization has been higher than without soil sterilization and with no application. In another field, the applying condition was studied. The most effective disease suppression was observed through harvesting when the composted crab shell material has been applied to soil every year. The soil sterilization was treated in the first year, but has not been needed after the next year. An increase in disease was observed in the no application year when an alternative year application system was used. These results suggest that composted crab shells must be applied to the soil every year to suppress Fusarium wilt of spinach in a field.

The Breeding Process and Characteristics of Five New Clonal Lines "Ai Star" of Limonium sinuatum Mill
Toshiichi Takahashi, Zenbi Naitoh, Hiroshi Satoh and Tatsuo Yoshida

@With the purpose of breeding new high quality Limonium sinuatum,@"Sopia" was selectively cultivated as breeding material. Individual selections were carried out from 1995 to 1997, the characteristics of sixteen chosen clones were researched from 1996 to 1998, and the superior lines were selected. After five of the "Ai Star" clonal lines were checked and confirmed for stability, they were given identification numbers in 1998 and were entered to be officially registered the following year, 1999. The clonal lines were found to be suitable for cropping in the [1]spring and summer, [2]autumn and winter, [3]deferment of autumn and winter. The characteristics of each of these clonal lines are as follows. "Ai Star Rosy Pink" has a sepal color of bright red purple and has a high output of quality flowers. It has a long sepal diameter as well as a long flower spike length, and therefore seems to be of generally larger volume than the other types. "Ai Star Lilac" has a light purple sepal color and a small stem wing. The sepal diameter is of medium size and the number of produced flowers is large. The flower spike form seems to be good for various cropping types. The sepal color for "Ai Star Lilac Blue" is light purple and has a good output of quality flowers. It has a long sepal diameter as well as a long flower spike and good volume. The sepal color of "Ai Star Lavender" is light purple and the tip of the sepal is a bright bluish purple. The stem wing is small. The flower spike is wide and the form is suitable for various cropping types. The sepal color of "Ai Star Mauve" is a vivid red purple. The branch length and the flower spike length are medium, and the flower spikes are good for various cropping types. As for a low temperature requirement for bolting in "Ai Star Rosy Pink", it was found a 50 to 60 day period at 15Ž, and to induce high bolting rate, and to yield the largest crop. The rooting rate in response to cutting propagation was found to differ between the lines, but even in the more difficult lines, a high rooting rate was produced with the use of a chemical rooting agent. For "Ai Star Lilac" and "Ai Star Lilac Blue", autumn and winter cropping types which used rooted herbaceous cutting, crops were produced by raising the seedlings naturally.

Breeding of a New Green Soybean Variety,"Takikei-C8 (named "Chage-maru")"
Takuya TAKAHASHI, Kazuo SAKUYAMA, Yuichi IMURA, Yutaka KIUCHI, Kazuya OIKAWA, Tsutomu SASAKI, Yoshinori KAMIYAMA, Masashi NITTA, Hisashi KOWATA, Yoshiaki OHGI and Hiroshi ISHIKAWA

  A new green soybean variety "Takikei-C8(named "Chage-maru")" was developed from a cross between "Fukura", which is good-taste, medium-maturing and the central variety in Iwate, and "Chusei-chamame", which is good-taste and late-maturing, at the Iwate Prefectural Agricultural Experiment Station in 1986, for the purpose of breeding of a good-taste, well-yield and medium-maturing green soybean.
@"Takikei-C8" belongs to the medium maturity group that is later than "Fukura" about 5 days, and if it is sowed between the middle and the late in May, it is able to harvest between the middle and the late in August at the middle and the south region in Iwate. The sum of shells is more than "Fukura", and it is very well-yield among medium-maturing green soybeans. And, again, the rate of complete shells and the rate of shells that possess more than 2 grains are high. And furthermore, it's sweetness and smell are very good, which are better than "Fukura"'s.

Breeding of a New Green Soybean Variety,"Takikei-C11 (named "Pukkurako")"
Takuya TAKAHASHI, Kazuo SAKUYAMA, Yuichi IMURA, Yutaka KIUCHI, Kazuya OIKAWA, Tsutomu SASAKI, Yoshinori KAMIYAMA, Masashi NITTA, Hisashi KOWATA, Yoshiaki OHGI and Hiroshi ISHIKAWA

  A new green soybean variety "Takikei-C11(named "Pukkurako")" was developed from a cross between "Fukura", which is good-taste, medium-maturing and the central variety in Iwate, and "Kinshu", which have large-shells and which is well-yield and late-maturing, at the Iwate Prefectural Agricultural Experiment Station in 1986, for the purpose of breeding of a having large and good-appearance shells and medium-maturing, whose harvesting season is the 4th~5th five days of August at the middle and the south region in Iwate, green soybean.
@"Takikei-C11" belongs to the medium maturity group that is later than "Fukura" about 8 days, and if it is sowed between the middle and the late in May, it is able to harvest between the middle and the late in August at the middle and the south region in Iwate. The rate of complete shells and the rate of shells that possess more than 2 grains are high. And, again, its appearance quality is good, because it's shells is large and deep green. And furthermore, it is good-taste.

Breeding of a New Rice Variety "Gin-otome" Suitable for Sake Brewing
Hitoshi HATAKEYAMA, Hiromi SUGAWARA, Tsutomu SASAKI, Hiroya ODANAKA, Shinsuke NAKAJYO, Masahiro TAKAHASHI, Nobuo TAKAHASHI, Syoji URUSHIBARA, Hisashi KOWATA, Yoshiaki OHGI, Hiroko NAKANO, Akikazu NAKANISHI and Tsuyoshi UWANO
@A new rice variety "Gin-otome", suitable for sake brewing, was developed from a cross between "Akita-Sake 44" and "Tohoku 141(named "Kokoromachi")" at the ex-Iwate Agricultural Experiment Station (the current Iwate Agricultural Research Center) in 1990.
@This variety was developed with intent of breeding an early-maturing variety for sake brewing.

@Breeding was conducted using the "bulk method" in its early generations. After individual selection in F
4, line selections, performance test and tests of specific character were conducted.
@After line selections of F
5, evaluation of the suitability for sake brewing was conducted by Iwate Industrial Research Institute. As the results, "Iwate-Sake52" was selected from the F7 lines. In F8, the test brewing was conducted by Iwate Sake Brewing Association. "Iwate-Sake 52" was released in Iwate prefecture as a recommended variety suitable for sake brewing and named "Gin-otome" in 2000. The major characteristics of "Gin-otome" are as follows.

(1) It belongs to the early maturing group, and its date of heading and maturity are same as or slightly earlier than that of "Takaneminori".
(2) It is partial panicle plant type. Its culm is shorter and panicle number is fewer than "Takaneminori".
(3) The lodging resistance is slightly inferior to "Takaneminori", superior to "Miyamanishiki".
(4) It seems to have true resistance genes of Pi a, i. The field resistance for leaf and panicle blast is both higher than "Miyamanishiki".
(5) It has moderate tolerance to sterility caused by low temperature before heading.
(6) The yielding ability is less than "Takaneminori".
(7) The quality of grain is slightly lower than "Miyamanisiki".
(8) The wight of 1000 grains is heavier than "Miyamanishiki".
(9) The suitability for sake brewing of "Gin-otome" is equal to "Miyamanishiki" judging from the results of examination that was conducted by Iwate Industrial Research Institute and Iwate Sake Brewing Association.
Early and Easy Diagnostic Methods and Control of Apple Violet Root Rot Caused by Helicobasidium mompa Tanaka in Dwarfed Apple Trees
Fusaharu NAKATANI, Giichi ANDOH and Satoru TAKAHASHI
@Pannose mycelial mat of violet root fungus forms in and around the base of dwarf apple trees, which at first appear to be healthy, but which decline after a few years. This can then spread along a row of apple trees. Purple spots appear on the leaves of the "Fuji" apple tree, when affected by violet root rot, in summer and autumn and the spots are in proportion to the severity of the disease. In the case of the "Tsugaru", many of the buds become deformed and die, which is a new symptom, similar to that of valsa cancer, observed after acute defoliation in summer. Effective fungicides have been screened through injecting solution into the roots of the apple trees that had developed mycelial mats the previous autumn. The success of these control measures was examined by comparing mycelial mats forming on the trunks of dwarf roots in the autumn of the year when the fungicides were injected. As for the baiting method of H. Monpa from the soil, inserting into the branch was found to be more effective than the busying method, and this was used for the evaluation of fungicides' evaluation too. After screening, tolclofos-metyl solution (40 liters/tree) into the soil (at a depth of 0-30cms) within a radius of 1m2 in the spring could prevent the reoccurrence of new mycelial mats in the autumn and the development of the disease. The trees made a remarkable recovery from the damage and produced the same amount of fruit as the healthy trees. Applying composed chitin materials (Neoappu: 7.5kg/tree) after removing the root soil from the trees in spring, could prevent the formation of mycelial mat a little, but the combined treatment of amobam solution could prevent the formation of mycelial mat in autumn and the development of the disease. As a result of this the trees recovered.
Effect of Growth on Weight of Seeds in Cabbage
Fumihiro KANNO, Katsuo KODAMA and Hidenori SUGAWARA
@Weight of cabbages seeds has nearly normal distribution. The light seed and heavy seed are brought up to a big set and a small set, respectively. The lighter seed growth is not uniform. A deference of the growth affects it's a taking root. The deference remains after planting at a field. Therefore, we need to make uniform weight of the seed for a uniformity culture of cabbage. Additionally, we should remove small seeds, because it's include a lot of un-mature seeds that tend to growth un-uniformly because of weak germination of the seed.
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