The present regulation on Pertussis Vaccine in Japan requires the following method for the “safety test ”: Each of two guinea pigs should be inoculated intraperitoneally with 5.0 cc and 5 mice 0.5 cc of a test vaccine. The animals should survive for a specified period without any pathological signs due to the vaccine, and their body weight should regain to its starting level within a specified time. This method is essentially equal to those used in many other countries.
This type of method belongs to the so-called “minimum requirement” type in which an absolute value of toxicity is defined. The definition of an absolute toxicity in terms of animal reactions (animal units) is unsatisfactory, because the animal unit is highly variable due not only to the variation of animals, but also to that in the mean response of stocks of animals in different laboratories and at different times. Even if the assay conditions are controlled as strictly as possible, it is not possible that the experimental conditions are quite the same or different only in minor factors, therefore another source of variation occurs. Thus, when the toxicity of a vaccine is sufficiently far from the specified level, no matter whether it is above or below the level there will be no trouble. It is obvious, however, that the results of assay are variable when the difference in toxicities between a test preparation and the specified level is within the range of variation of the response in the system, as we have often experienced in the routine assay for toxicity of commercial pertussis vaccines.
In contrast to the failure to make accurate measurements in terms of animal reactions, much greater success has been attained by the introduction of a method estimating relative potency, in which estimation of an unknown preparation is made by comparison with a standard (or a reference) preparation. The standards in a stable form, which was introduced by Ehrlich, made this comparative methods possible.
Besides, by the method of minimum requirement type neither quantitative determination of toxicity nor statistical analysis of the accuracy of the results are possible.
For these reasons we investigated the comparative method in order not only to establish a more reliable method in the “safety test” of pertussis vaccine, but also to reinvestigate the relationship between the toxicity of pertussis vaccine and the magnitude of reactions by the vaccine in human beings.
As there had never appeared reports concerning biological assay for toxicity of pertussis vaccine by the comparative method, the first step of this study was to select the response of animals or its metameter (Bacharach et al., 1942) biologically reasonable and suitable for the statistical analysis. Since the final step of the study should be the comparison of results between human beings and test animals, it would be desirable to select such a response as fever, which is common and measurable both in human beings and in such an experimental animal as the rabbit. However it was considered impractical to select fever as the response because of the difficulty in using adequately large numbers of rabbits for the routine work. We have long been making assays measuring the body weight changes of animals according to the current regulation. Measuring the body weight is relatively easy in practice and a large number of mice are available for the test under more or less controlled condition. Therefore, the assay method by the body weight change in mice was first investigated.
If we find no significant correlation between the magnitude of reaction in human beings and the toxicity of vaccines estimated by the method proposed, then we will have to further search for other animal responses suitable for the purpose.

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A biological assay method for toxicity of pertussis vaccine was proposed in the previous paper (Kurokawa et al., 1962) . By the method it was expected to enable a quantitative determination of toxicity and to give more reproducible estimates than the current method because it was designed to control the variation in the mean response of animals at different times or in different laboratories.
The present paper is chiefly concerned with expressing the level of toxicity passing the current regulation in terms of the relative toxicity. The considerations were made chiefly being based on the results from the experiments using mice, which have been carried out relatively in detail and were presented in the previous paper. Experimental data obtained with guinea pigs also were referred to in the considerations.

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Jensen (1933) claimed that the Dlm/ (DRM) m ratio of diphtheria toxin is constant, the value being 3, 000. No detailed reports on the relationship between Dim and DRM can be found.
Our previous finding; i. e., the Dlm/DRM ratio of fresh diphtheria toxin preparations is highly variable in a range from about 3, 000 to 100, 000 or higher (Kondo et .al. 1960), did not support Jensen's claim. Furthermore, the following two facts were found: 1) When crude or fractionated diphtheria toxin was partially detoxified by mild heating, by dilution with buffered saline or by f ormalinization, DRM increased faster than Dlm, and the Dlm/DRM ratio became smaller during the detoxificating process, approaching to 3, 000, the value given to the Dlm/DRM ratio by Jensen; 2) When fractionated with ammonium sulphate, fractions whose DRM/Lf ratios were different from that of the parent toxin but Dlm/Lf ratios remained constant. A tendency was also observed that the fractions precipitable at a lower concentration of ammonium sulphate had smaller Dlm/DRM ratios (Kondo et al., 1960) .
The present paper deals with some additional findings which together with the previous findings are expected to contribute a great deal to studies on the biological activity of diphtheria toxin.

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The literature concerning the occurrence of multiphasic cultures of Enterobacteriaceae was reviewed briefly by Fife, Edwards and Sakazski (1962) . The purpose of the present paper is to describe three additional triphasic Arizona cultures.

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The occurrence of multiple phases and complex phases in Enterobacteriaceae was reviewed by Fife et al. (1961), Edwards, Sakazaki and Kato (1962) and Edwards, McWhorter and Douglas (1962) . To the present, three Arizona serotypes which possessed three reversilbe flageller phases have been described and one culture of Salmonella inikawashima which exhibited four reversible flagellar phases has been recognized. Sakazaki, Tsujimoto, Edwards and Fife (1962) detected three additional triphasic Arizona cultures. Aside from these observations no cultures of Enterobacte riaceae have been encountered which contained more than two naturally occurring reversible H phases. The purpose of the present paper is to describe an Arizona type in which four distinct flagellar phases were detected.

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It is known that organisms of the Arizona group occur very frequently in reptiles and many different serotypes have been found among cultures isolated from snakes (LeMinor, Fife and Edwards, 1958) . The purpose of the present paper is to report the isolation of five hitherto undescribed serotypes from reptiles.

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Preliminary laboratory and field tests with a new molluscicide, Bayer 73 (5, 2-dichloro-4-nitro-salicylicanilide), were reported by Gönnert et al, in 1948. Since then, this compound has been tested on aquatic snails transmitting Schistosoma mansoni and S. haematobium, both in the laboratory and in the field. However, very little is known of the effect of the compound on amphibious snails transmitting S. japonicum.
Another chemical, Sevin (1-naphthyl N-methylcarbamate) has been used as a very promising insecticide. Data are, however, not yet available as to whether Sevin could be applied as a molluscicide against schistosome bearing snails.
The aim of this study is to examine the molluscicidal activity of both compounds, Bayer 73 and Sevin, against O. nosophora, the vector snail of Schistosoma japonicum, under laboratory and field conditions.

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