Using the fluorescein-labeled antibodies, slide agglutination tests were employed to detect several plant viruses under dark field illumination in a Nikon microscope connected with a mercury vapour lamp (Chiyoda AHL 250). Positive reactions were recognized with tobacco masaic virus, potato virus X and Y, and rice dwarf virus by their corresponding conjugated antisera. A drop of crude sap from infected plants was mixed with about one third to one fourth its volume of the concentrated fluorescein-labeled antibody solution on a microscope slide, stirred with the corner of a cover glass and covered with same. This method is available for detecting the antigen-antibody complexes stained specifically in the mixtures despite the weak reaction. This method is very excellent because it is easy to apply, and the quantity of both antiserum and antigen to be required is very little, and specificity of the reaction can be directly confirmed under a microscope. A high sensibility of the slide agglutination test was ascertained by experimental results with box titration using each virus and its homologous conjugated antiserum.
It has been well known that the root of Brassica rapa var. neosuguki Kitam. develops abnormally and forms a gall when the root was infected with Plasmodiophora brassicae Woronin. In order to characterize plant hormones responsible for the abnormal growth, extractions were carried out with petroleum ether, benzene, ethyl ether and ethanol successively from the infected roots. Active principle(s) were contained in benzene and ethyl ether fractions. The active substance was not considered to be gibberellin or its like substance by the gibberellin test using rice seedlings. One or possibly two active substances were found in ethyl ether extract from the benzene fraction of the infected roots by the biological tests with Avena. One of them was presumed to be indole acetic acid by Rf values of paper chromato-graphic analyses, and by the Avena tests.
1) In a survey to determine the causal viruses of tomato plants showing mosaic and/or streak, 89 samples were collected from Yamanashi, Kanagawa, Nagano, Kochi and Tottori Prefectures. These diseased plant samples were subjected to inoculation tests on a series of 8 differential hosts using carborundum. 85 samples (95 per cent) yielded tobacco mosaic virus (TMV), 21 samples (24 per cent) yielded cucumber mosaic virus (CMV) and 2 samples (2 per cent) yielded potato virus X (PVX). From 17 samples (19 per cent), both TMV and CMV were isolated, and from 2 samples both TMV and PVX were isolated. The collections were divided on basis of the original symptoms on tomato plants into mosaic group (70 samples) and streak group (19 samples). Both groups yielded almost the same kinds of viruses and isolation percentage of each virus. 2) Of the 85 samples which contained TMV, 78 samples (92 per cent) yielded TMV specialized to tomato (tomato strain of TMV), while 11 samples (13 per cent) were of the ordinary strain of TMV. From 4 samples (5 per cent), both tomato strain and ordinary strain were isolated. 3) Tomato strain produced only local necrotic lesions on tobacco (Bright Yellow), N. sylvestris and petunia, but no systemic mosaic on these plants. No local lesions were produced on bean (Otebo). Ordinary strain produced local lesions on bean, but no local lesions on the above 3 species. On these plants, ordinary strain produced systemic mosaic. 4) Tomato strain was considered to be identical with tomato streak strain of TMV described by Ainsworth (1933) and Smith (1957), judging from reactions of several plants, thermal inactivating point, cross protection test, particle size and agglutinaton reaction on slides with antiserum against ordinary strain of TMV. The name “tomato strain” seems to be more preferable than “tomato streak strain”, because this strain can be isolated from samples showing only the mosaic symptom. Hence the name “tomato strain” is preferred. 5) Multiplication of tomato strain was better than that of ordinary strain in the inoculated leaves of tomato. When these two stranis were inoculated on lower leaves of tomato respectively, tomato strain was recovered from top leaves after 3 days, while ordinary strain was recovered after 5 days. These results indicate that tomato strain is more pathogenic to tomato plant than ordinary strain.
Necrotic spot of muskmelon, a virus disease new to Japan, was found in Shizuoka and a few other Prefectures. Three types of leaf symtoms are recognized: (I) necrotic specks, appearing in abundance on later developing leaves after infection, (II) large necrotic spots, and (III) veinal necrosis. Type II and type III symptoms are usually seen on mature leaves. On the stem and petioles, necrotic streaks are produced. On mechanical inoculation, the virus incited primary local lesions and systemic necrotic spots on muskmelon, cantaloup, and makuwa (Cucumis melo var. makuwa). It produced local lesions, but no systemic infection, on watermelon, cucumber (Chicago Pickling), and oriental pickling melon (Cucumis melo var. conomon). It did not infect other cucurbits, involving squash, white gourd (Benincasa hispida), bottle gourd (Lagenaria siceraria var. hispida), bonnet gourd (Luffa cylindrica), and oriental varieties of cucumber, and 22 species of non-cucurbitaceous plants. The virus was seed-transmitted in a moderate percentage (about 20%). A test for transmission by Aphis gossypii was negative. Test with beetles has not yet been made. The virus in expressed sap was not inactivated in 10 minutes at 60°C, during 9-10 days aging in vitro, or at 1:50, 000 dilution. Purified preparation of the virus showed, by electron microscopy, spherical particles of about 31mμ in diameter (Saito, Kishi, and Iwata, 1962). Among the known viruses infective to cucurbits, wild cucumber mosaic virus described by Freitag (1952) and Lindberg et al. (1956) seems to be most closely related to the present virus, though there are slight differences in symptoms and host range.
The competitive saprophytic ability of Rhizoctonia solani Kühn was compared among three different strains: i.e., isolate F-16 which belongs to the spring strain, F-20 which belongs to the summer strain, both being obtained in the previous experiments (Ui et al, 1963), and B-5, a sugar beet strain which causes damping off of sugar beet seedlings and root rot of mature plants. For the assessment of the competitive saprophytic ability, the Cambridge method (Butler, 1955) and it's agar plate modification (Rao, 1959) were adopted. The substrate for saprophytic colonization of the inoculant were mature flax stem pieces and water agar plate respectively. From the results, as illustrated in the figures presented, F-20 may be regarded as a vigorous saprophyte, followed by B-5 and F-16. When the same estimations were applied to the soil-inoculum mixtures which were previously incubated for two weeks at room temperature before the addition of substrates, the saprophytic ability of F-20 showed little changes, while that of B-5 and F-16 decreased, particularly the latter behaved as a poor saprophyte. From the results of the experiments, the authors concluded as follows: R. solani has been regarded as one of the typical soil-inhabiting fungi, whereas some of the strains such as F-16, behave as a poor saprophyte when it was previously incubated for several days under competition of soil micro-organisms without a suitable substrate for colonization.
The purpose of this work is to evaluate the controlling effect of 36 3-methylpyrazole derivatives upon rice blast, sheath blight and bacterial leaf blight of rice and cucumber anthracnose in greenhouse. The relationship between the structural modifications of 3-methylpyrazole derivatives and their protective effect upon rice blast was intimate, but not on sheath blight of rice and cucumber anthracnose, and also their therapeutic effect on bacterial leaf blight of rice was not intimate. Some 3-methylpyrazole derivatives substituted at 1-position with p-nitrophenyl or p-halogenophenyl group, at 4-position with SCN or without SCN, and at 5-position with CH3 or C6H5 showed good protective effect upon rice blast, and substitution with SCN at 4-position was more effective than that without SCN, and both 3-methyl-4-thiocyanatopyrazole derivatives substituted at 5-position with CH3 and those at 5-position with C6H5 were similarly effective. Three pyrazole derivatives, i.e. 3, 5-dimethy-1-(2-nitrophenyl)-4-thiocyanatopyrazole, 3-methyl-1, 5-diphenyl-4-thiocyanatopyrazole, and 3, 5-dimethyl-1-(4-nitrophenyl)-4-thiocyanatopyrazole, showed good protective effect and a little less sporulation inhibiting effect on rice blast, while they were not effective in the therapeutic test. In the seedling bed test in field these three componds were less effective than blasticidin S as a blast control agent.
The present investigation was undertaken to evaluate the controlling effect of 39 pyrazole derivatives substituted at 1-, 3-, 4- and 5-postiion with various substituents upon rice blast, sheath blight and bacterial leaf blight of rice in greenhouse. The structural modifications of pyrazole derivatives had intimate relation with their protective action against rice blast, but no relation with their protective action against sheath blight of rice and with their therapeutic action against bacterial leaf blight of rice. Some 5-hydroxy-3-methylpyrazole derivatives substituted at 1-position with aryl group showed better protective effect upon rice blast than those at 4-position with SCN, but 3-amino-5-hydroxy-1-phenyl-4-thiocyanatopyrazole, which substituted methyl radical at 3-position with amino radical, gave good protective effect. Some 1-(4-thiocyanatophenyl)-pyrazole derivatives at 4-position with CN or COOC2H5 and at 5-position with NH2 or C6H5 also showed good protective effect. Protective effect of 1-(4-chlorophenyl)-5-hydroxy-3-methylpyrazole on sheath blight of rice and therapeutic effect of 5-amino-1-phenyl-4-thiocyanatopyrazole on bacterial leaf blight of rice were evaluated highly but controlling effects of both compounds were estimated less than those of the positive control.