Literature concerning the mechanism of soil-transmission in soil-borne cereal mosaic viruses and in other soil-borne plant viruses has accumulated during recent years. It is believed by most workers, that perhaps soil-borne viruses preserve their activities in some soil-inhabiting microorganism which carries the viruses and introduces them into the underground parts of the plants. However, the exact mechanism of the soil-transmission is not yet understood, because no microorganism is experimentally found as a vector.
The present work was undertaken in order to ascertain the exact relation of these soil-borne viruses to the soil and to understand the nature of soil-transmission, making use of two kind of soil materials which were certainly infested with wheat yellow-mosaic virus (WYMV, Marmor tritici var. fulvum McK.) and barley yellow-mosaic virus (BYMV).
A small number of plants were infected when grown in clean soil in which ground tissues of plants affected with WYMV or BYMV had been buried about two years previously (Table 1).
Soil particles were separated mechanically by water from WYMV- or BYMV-infested soil, moreover, very small particles were separated according to Stokes' Law in soil science. Some kind of tests of the infectivity of separated soil particle fractions were made by planting seeds in them or by rubbing seedlings with them (Tables 2-4). It was found, eventually, that the infectivity of the clay fraction (‘<2μ’ particle fraction, including inorganic and organic colloidal substances) was very much stronger than the other particle fraction (‘>2μ’ particle fraction, including a small quantity of ‘<2μ’ particles) in both WYMV- and BYMV-infested soils, when the separated suspension including these particles was concentrated by centrifugation at 4000 r.p.m. (2000g) for 15-20 minutes (Tables 4 and 5). In the pile of clay particles, no special soil-inhabiting microorganism regarded as a vector was recognized under microscope. It was shown, moreover, that a small number of plants could be infected by rubbing method of inoculation with mud composed of above-mentioned clay fraction (Table 5).
These findings may support the writer's view that these viruses exist adsorbed in soil particles of clay fraction which have colloidal characteristics and ability to adsorb proteins, and that no soil-inhabiting microorganism is necessarily needed to explain the mechanism of soil-transmission in soil-borne cereal mosaic viruses.

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In the past, remarkable difference was encountered in the routine potency tests of the Japanese encephalitis vaccines of two different strains of virus, G-1 and Nakayama, challenged with Nakayama strain. Such result gave us a suggestion of strain difference which might exist in producing immunity in mice.
In order to give answer to such question, the vaccines were prepared with 2 strains, Nakayama and G-1, respectively. Both viruses were identified serologically as Japanese encephalitis viruses. Group of mice were immunized with 2 vaccines, and after the vaccination, cross challenge test was accomplished with both strains on the one hand and on the other, neutralization, complement-fixation and hemagglutination inhibition tests were carried out on sera collected from mice at 2 weeks after the initial vaccination.
As the results, protection index obtained in Nakayama-immunized mice challenged with the heterologous strain at 2 weeks was found to be lower than that obtained by the challenge with homologous one. In case of G-1-immunized mice, the protection index was found very low when challenged either with the homologous strain or with the heterologous one. However, no significant difference was recognized in term of antibody titer between the mice immunized with both strain vaccines.

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It was assumed that the Japanese encephalitis vaccine of mouse brain type might give rise preventive effect on the disease in nature in approximately 75 per cent. However, such a brain type vaccine was proved to contain possibly some factor to produce demyelinization following inoculation with the vaccine combined with Freund's adjuvant into guinea pigs. Accordingly the minimum requirement of the vaccine was revised to be prepared from the supernatant of 2 per cent emulsion of infected mouse brain following centrifugation at 3, 000 rpm for 30 min. Encephaligenic factor of such vaccine was remarkably minimized so that no more experimental demyelinization in guinea pig was encountered with this.
Herewith an attempt was made to elucidate immune response following injection of formalin killed mouse brain type vaccine of G-1 strain which was prepared following the method of the minimum requirement. G-1 strain of Japanese encephalitis virus was isolated from an encephalitis patient by us in 1949 and had been passed through mouse brain roughly by 200 generations. To criticize immune response by the vaccine, the vaccine was diluted to 1:1, 1:16 and 1:128 or 1:1, 1:10, 1:100 and 1:1, 000 and a group of mice was immunized with each dilution. Those immunized mice were divided into subgroups, and challenged with a series of 10 fold dilution of G-1 strain intracerebrally at 2 and 3 weeks after the initial immunization. On the other hand, neutralization, complement-fixation (CF) and hemagglutination inhibition (HAI) tests were carried out with pooled mouse sera from each group at 2 and 3 weeks after the initial vaccination. By comparing protection index (PI) with neutralization index (NI), CF and HAI antibody titers in mice at 2 and 3 weeks after the immunization with each dilution of the vaccine, those may be said:
1) The less PI and antibody titers were found in mice immunized with the higher dilution of vaccine at 2 weeks.
2) There was no significant difference in immune responses of mice vaccinated with both original and 10 times diluted vaccines. The minimum effective dose of the vaccine in mice was computed to be between 10 and 100 times dilution of the vaccine.
3) Comparing with immune responses each other in mice at 2 and 3 weeks, both PI and CF antibody titers reached their peaks at 2 weeks and fell down by 3 weeks, while both neutralizing and HAI antibody titers were found almost the same.
From the foregoing results it may be suggested that the effect of vaccine should be criticized on the basis of results obtained in mice first immunized with dilutions of vaccine of some range and then challenged with a series of virus dilution.

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The present paper deals with the results of the studies carried out under 15° to 20°C, on the mechanism of aphid (Myzus persicae Sulz.) transmission of mosaic disease of Japanese radish using radioactive phosphorus 32.
1) Aphids were fed for 5 (Table 3) and 10 minutes (Table 1) each on the virus diseased Japanese radish (Raphanus sativus L.) seedling which had been cultured with the solution containing radioactive phosphorus 32 (200μc/ml). They were then transferred to each of 10 healthy radish seedlings successively and fed for 5 and 10 minutes respectively. Results showed that aphids could transmit the virus disease up to the 6th seedling, while all of 10 seedlings on which aphids were fed were found to be radioactive when determined by autoradiography.
2) Aphids fed for 10 minutes (Table 2), 1 minute (Table 4), and 30 seconds (Table 5) respectively on the diseased radish seedling containing radioactive phosphorus 32, were transferred to each of 8 to 15 healthy radish seedlings successively and fed for 1 minute or 30 seconds. Results showed that in the case of short infection feeding time such as 30 seconds or 1 minute no appreciable differences were observed between infectivity of the virus and transportation of phosphorus 32 by one aphid when 8 to 15 plants were infected successively.

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In order to study on causal relation between polio virus and Guillain-Barré Syndrome encountered in the Setouchi area in 1953-1955, the comparative investigation of the isolation of polio virus from feces of patients with paralytic poliomyelitis and with G-B syndrome were attempted by means of roller tube cultures of human embrionic skin muscle.
7 strains of polio virus were isolated from feces of 9 polio patients and, of these, 5 were idemtified as type 1 and 2 as type 3; No viruses of the type 2 group were isolated from the material studied.
Only one strain of virus was isolated from feces of 9 G-B syndrome patients. However, this virus was not polio virus but ECHO virus. No polio virus of any type was isolated from feces of all G-B syndrome patients.
By referring to the study on the presence of the polio neutralizing antibody in the group of patients with paralytic poliomyelitis and of patients with the polyradiculoneuritis so-called G-B syndrome by Oyama, the Study on the interference phenomena between polio virus and Coxsackie virus by Ohara and above mentioned results, the pathogenesis of G-B syndrome was discussed.

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Difficulties were often encountered in carrying out hemagglutination inhibition test with Asian influenza virus, for there were non-specific inhibitors against Asian influenza virus in sera of many animals. In addition, the properties of the inhibitor appeared somewhat different from those of inhibitors so far described.
Following experiments were made in order to elucidate the nature of the inhibitor and to estimate its content in sera from various animals.
1) Hemagglutination inhibition titers of egg white and normal sera from nine species of animals against various strains of influenza virus were determined. Changes in their inhibition titers, after various treatments were also examined. A/Adachi/2/57, now used as a standard strain of Asian influenza virus in Japan, was readily inhibited by many of normal animal sera, especially in high titer by horse, swine and guinea pig sera.
2) The inhibitory activity was heat stable, destroyed neither by RDE (crude filtrate of the culture of V. cholerae) treatment nor by trypsin treatment, but was readily destroyed by KIO₄ treatment. Marked difference in its inhibition titer against live virus and indicator virus of A/Adachi/2/57 was not observed. A/Adachi/2/57 was neutralized by normal horse serum in embryonated eggs. The neutralizing activity was also inactivated by KIO₄ treatment. Based upon these results, the inhibitor against A/Adachi/2/57 is considered to be distinct from α and β inhibitors, and the term γ inhibitor was proposed. Comparison of properties of α, β and γ inhibitors and their contents in sera of various animals were diagramatically shown.
3) A correlation was found between hemagglutination inhibition by human antisera and that by inhibitor, at least with six strains of Asian influenza virus so far examined. A/Kumamoto/Y5/57, which was not inhibited by γ inhibitor in hemagglutination, was not also neutralized by γ inhibitor in embryonated eggs.
4) Addition of two volumes of M/100 KIO₄ to one volume of sample was recommended as a useful procedure in practice in order to remove γ inhibitor in hemagglutination inhibition test. This treatment did not seriously affect specific antibody titer.

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Variations in the sensitivities of lysogenic bacteria to X-ray during the growth cycle were investigated using two strains, lysogenic Shigella flexneri strain KA (βL) and nonlysogenic strain KA. The main results obtained were as follows.
(1) The radiosensitivity of the colony forming ability of rapidly dividing cells was markedly higher than that of resting cells in stationary phase.
(2) On the contrary, the sensitivity of the phage inducibility of the lysogenic strain KA (βL) to X-ray was found to be considerably lower when it was in logarithmic growth phase than in stationary phase.
(3) The survival curves measured by counting the visible colonies of both strains were similarly approximately single-hit type, indicated by their exponential inactivation kinetics.
(4) However the radiosensitivity of lysogenic strain KA (βL) for colony formation seemed to be superior than that of nonlysogenic KA, suggesting that the β phage induction may occur independently of the lethal mutation in the irradiated bacteria.

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The nature of ectromelia virus inactivated photodynamically was examined using the system of the virus-Ehrlich ascites cells.
1) The ectromelia virus is very sensitive to the photodynamic action of methylen blue.
2) The virus which has already entered the cells becomes resistant to the photodynamic action.
3) The inactivated virus has no interfering property.
4) The inactivated virus retains its immunogenical property.
5) The formation of inclusion bodies and the development of the inclusion bodies were suppressed in the immunized mouse even when the infection had already progressed for several hours in the normal mouse.

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Studies on the site of action of various antivirals including caprochlorone, chartreusin, 2, 5-dimethyl-benzimidazole, 5, 6-dichlor-1-b-ribofuranosyl-benzimidazole, myxoviromycin, canavanine sulfate and dinitrophenol were conducted against PR8 virus growth in chorioallantoic membrane and Sendai virus growth in L cells.
Firstly, these antivirals were tested against the growth of PR8 virus in a tissue culture system, consisting of 2.0ml of Hanks'solution and 4 pieces of chorioallantoic membrane together with the attached shell.
The results obtained in the system are as follows:
1) These antivirals in fairly high concentrations did not show any inhibitory tendency of viral hemagglutination. They were not virocidal and their inhibitory effect on the adsorption stage was also excluded as far as tested with 2 or more times of minimum inhibitory concentrations. Thus the inhibition of virus synthesis within the cell was suggested as the site of antiviral activities.
2) Determining the sensitive stage of the virus growth to these antivirals, chartreusin, 2, 5-dimethyl-benzimidazole and 5, 6-dichlor-1-b-ribofuranosyl-benzimidazole were grouped as an early active agent, and caprochlorone, canavanine sulfate, dinitrophenol and myxoviromycin were grouped as the late active antivirals. In other words, latter antivirals were active even when added at late stage of the incubation.
3) On the basis of calculating chemotherapeutic index, myxoviromycin was thought to be the best among these antivirals.
Secondly, these antivirals were examined their effect on the growth of Sendai virus in Earle's L cells.
The results obtained in the system may be summarized as follows:
1) All these antivirals showed the toxicity to L cells and the concentration which inhibited the growth of virus was around the vicinities of their toxic concentrations.
2) Both myxoviromycin and caprochlorone inhibited the cytopathogenic effect of the Sendai virus in addition to the inhibition of hemagglutinin production at critical concentrations. With other antivirals tested here, cytopathogenic effect of the virus was not affected even when the hemagglutinin production was impaired.
3) Myxoviromycin, in a wide range of concentration, showed the toxicity of low grade and within this range, the antibiotic inhibited the growth of the Sendai virus.

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Multiplication of poxvirus in vitro including ectromelia virus, vaccinia virus and variola virus were studied using chick embryo fibroblast, strain L cells and HeLa cells as host cells.
Relationships could be established among the growth cycle of virus determined by infectivity titrations, the cellular alterations under the conventional light microscopy and the presence of virus or materials of components associated with virus within the cells seen in the electron microscope.
In growth cycle experiments no increase in the amount of cell-associated virus was observed during the first 6 hours in any of the infectious systems used. A peak virus titer was attained in 15-to 18-hour-infection. Ectromelia and variola virus adsorped on the host cells were photographed using ultrathin sectioning. Intracytoplasmic inclusions (matrix) appeared 3 hours following infection, The first recognizable viral form, so-called developmental form, were exclusively observed within the matrix area in 6-hour-infection. Most of them were limited by a double membrane. Structural appearances of the nucleoid or viroplasm in stricted area were almost the A same as those of matrix material. Coiled thread measuring about 20 to 30Å in width were seen in them. Developmental form is matured infective virus or a vegetative phase of virus from the point of view of dynamic infectivity. At later stage of infection in ectromelia-fibroblast or-L cell system another quite different intracytoplasmic inclusions appeared which did not play a necessary role in viral multiplication.
Mature forms were observed in the matrix area in a later stage of infection. The structures of them show quite characteristic appearances and it is very easy to distinct from the developing form by the presence of two kinds of double membranes, by their shape, by smaller size and by rather stronger density in thicker sections. Experiments showed that so-called mature form appearing in a later stage was resting form of virus.

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1) During the course of the studies on the acute respiratory infection among children which were found in Tokyo area during the period between January, 1956 and February, 1958, isolation of causative virus from part of the sick children was attempted. Attempts were made on the throat washings and throat mucus collected from 5 cases who were diagnosed HVJ infection serologically. Two strains of HVJ were thus isolated.
Hazeyama strain was isolated by direct intraamniotic inoculation of the throat washing collected on the 1st day of disease from an 11 year-old boy who was suffering from acute laryngitis and exacerbation of acute nephritis, The identification of this strain to HVJ was made at the Fukumi Laboratory of the National Institute of Health.
Sakai strain was isolated by direct intraamniotic inoculation of throat mucus collected on the 5th day of disease from a 4 year 11 months old girl who was suffering from acute bronchitis. This strain was identified to HVJ by cross hemagglutination inhibition tests with antisera against influenza A and B viruses, mumps virus and HVJ. The reconvalescent serum of this case was negative in the HVJ complement fixation test even at the dilution of 1:4, but it was HVJ hemagglutination inhibition test positive up to the dilution as high as 1:1024.
2) Neutralization tests by fertilized hens'eggs were conducted by serum dilution method on 5 paired specimens randomiy selected from the specimens from cases of HVJ infection. In all cases, the convalescent serum showed distinct rise in the neutralizing antibody titer.
Within the 3rd week of the disease, neutralizing antibody titer, complement fixation titer and hemagglutination inhibition titer were found rising in parallel with each other, though complement fixation titer showed a tendency of falling thereafter. In the Case No. 3, the serum collected on the 144th day of disease showed negative results in the complement fixation test and hemagglutination inhibition test, but the neutralizing antibody titer was found still remarkably high. The above suggests that the neutralizing antibody titer and hemagglutination inhibition titer fluctuate not necessarily in parallel each other.

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Employing HVJ (Obayashi strain) vaccine inactivated with 1:10, 000 dilution of MERZONIN (sodium ethylmercurithiosalicylate), the pattern of the antibody production was observed on 50 healthy children at the age ranging from 8 to 15 years. The following results were thus obtained. In this instance, 50 healthy children matched by age were taken as controls.
1) Initial Immunization: Two intradermal injections of 0.1cc at the interval of 1 week demonstrated a significant rise in the hemagglutination inhibition titer (HVJ-HIT) in 5 out of 48 vaccinated cases (10.4%). In all of the cases, the complement fixation titer (HVJ-CFT) was negative even at the dilution of 1.2.
2) 1st Booster Immunization: As the results of the intradermal injection of 0.5cc 3 months after the initial immunization, a significant rise in the HVJ-HIT was recognized in 20 out of 47 vaccinated cases (42.6%). In all cases, no significant fluctuation in the HVJ-CFT was recognized.
3) 2nd Booster Immunization: As the results of 2 intradermal injections of 0.5cc each at the interval of 1 week 7 months after the 1st booster immunization, a significant rise in the HVJ-HIT was noted 17 days later in 21 out of 39 vaccinated cases (53.8%). Besides, a significant rise in the HVJ-CFT was noted in 13 out of 39 vaccinated cases (33.3%). One month later, both the HVJ-HIT and HVJ-CFT showed a tendency of falling, and all cases showed almost the same titer as that prior to the immunization 4 months later.
4) No statistically significant correlation was recognized between the infection of influenza A/Asia/57, which occured immediately after the 2nd booster immunization, and the antibody production effect, and between the mumps infection in the past and the antibody production effect of HVJ vaccine.
5) As side reaction of the immunization, excepting 1 case each of fever and vomiting after the 1st booster immunization, no serious general reaction was recognized at all. Further, all of the local reactions were only transient.
6) As stated in the above, in view of the high rate of antibody production effect and only slight side reactions, the inoculation of Merzonin inactivated HVJ (Obayashi strain) vaccine gave a favorable results comparable to those reported in the past about influenza vaccine.

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In order to clarify the correlationship in the antibody production against mumps virus and HVJ which was observed in the adult case of mumps, cases of HVJ infection, cases of mumps, cases immunized with both viruses, cases of cross infection, cases cross immunized with vaccine and clinical cases of various combinations of infection and immunization with vaccine were examined for the determination of the fluctuation of antibody level. As the results, the following findings were obtained.
1) In the cases of HVJ infection, regardless of the past infection of mumps, no significant fluctuation in the antibody level against mumps was recognized at all, though the subjects were all children. However, in the cases inoculated with HVJ vaccine, rise in the antibody level against mumps virus was recognized in 3 out of 5 children who experienced mumps in the past.
2) In the cases of mumps infection, regardless of the past infection of HVJ, no fluctuation in the antibody level against HVJ was recognized in the children, while in the adult cases of mumps, remarkable rise in the antibody level against HVJ was recognized in 1 out of 2 cases. In 2 cases of mumps who had been immunized with HVJ vaccine in the past, rise in the antibody level against HVJ was clearly recognized. Further. when children who had been immunized with HVJ vaccine in the past were immunized with mumps vaccine, rise in the antibody level against HVJ was recognized in 4 out of 7 cases. In 4 child cases who had clinically contracted mumps after the immunization with HVJ vaccine prior to the immunization with mumps vaccine, rise in the antibody level against HVJ was recognized when they were immunized with mumps vaccine.
3) In view of the above, the rise in the antibody level against HVJ which was seen in the adult mumps cases was considered to be due to the fact that the antibody level acquired at the time of HVJ infection rose as a group reaction when the antibody level against mumps virus rose.
4) Although numerous studies may be required in future to clarify the correlationship in the antibody production of HVJ and mumps virus, the serological diagnosis of HVJ infection in children is considered to be done far more readily almost without error when compared with that in adult cases.

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The kinetics of phage induction in some lysogenic strains of E. coli and Shigella was investigated by means of ultraviolet light (UV) irradiation. The main evidence to be noticed is the following.
(1) The kinetics of β phage induction in A34 (β, γ) was observed to be a kind of two-hit phenomenon, whereas that in an artificial lysogenic Shigella, strain KA (βL), single-hit phenomenon. The radiosensitivity of both lysogenic strains with respect to β phage induction was nearly equal each other.
(2) Under suitable UV dose, more than 80 per cent of the bacteria irradiated were induced with both two lysogenic strains.
(3) The number of β prophage in KA (βL) and A34 (β, γ) was estimated to be about 2 and 3-4 per bacterium, respectively.
(4) The descending portion of the induction curves is composed of two different parts, each of which is distinguished by its inclination, suggesting that the inactivation of phage production in overirradiated cells occurs by at least two different mechanisms.
(5) The UV sensitivity of bacteria tested as to phage induction and colony forming ability was markedly depend upon the physiological state of bacteria, for instance, inanition of the bacterial cells or growth phase of the culture, at the time of irradiation.

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A simple method for the staining of viral inclusion bodies was described.
Procedure:
(1) Fix in CARNOY's fluid, 10% formalin or absolute alcohol.
(2) Prepare paraffin sections in routine.
(3) Stain sections in 0.5% fuchsin S for 5 minutes.
(4) Wash off exess reagant in water for a moment.
(5) Treat with 1% phosphowolframic acid in destilled water for 20 minutes.
(6) Wash in water for a moment.
(7) Place in MAYER's hemalaum (hematoxylin) for 5 minutes.
(8) Wash in running water for 10 minutes.
(9) Dehydrate, clear and mount as usual.
Smear preparations can also be stained, but mount in Canada balsam or examine under microscope with oil immersion lens.
The ground substance of type A intracytoplasmic inclusion body (MARCHAL body) appeared deep scarlet in EHRLICH, YOSHIDA and sarcoma 180 ascites tumor cells, the chorioallantoic membrane of chick embryo, EHRLICH ascites tumor cells transplanted onto C. A. M., and the liver, other organs and tissues of mouse infected with ectromelia virus. The elementary bodies and the outline of type A intracytoplasmic inclusion body (BOLLINGER body) appeared deep scarlet in the C. A. M. of chick embryo and the skin of chick infected with fowlpox virus. The ground substance of this inclusion body was stained with neither fuchsin S nor hematoxylin. The type B intracytoplasmic inclusion bodies of these two kinds of viruses were not distinguished from the cytoplasm with this staining method. The classical type A intranuclear inclusion bodies appeared scarlet in the bronchial epithelium of monkey infected with measles virus and in the C. A. M. and the liver of chick embryo infected with herpes simplex virus. The inhalt of swollen nucleus infected with herpes simplex virus was stained slightly purple. Red blood cells and granules of eosinophilic leucocytes were stained deep scarlet. Nucleoli, some parts of degenerated cytoplasm and some of degenerated nuclei were also stained scarlet. Nuclei were stained deep blue. All others appeared faint purple.
By means of this staining the acidophilic inclusion bodies could be easily perceived.

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We described that the phages which control Salmonella O[I] antigen have transduced the sugar-fermentative ability to strains lacking that factor (chiefly, dulcitol-fermentative factor at this experiment). We also showed some experiments on the different types of transduction and discussed on its mechanism.

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