Suppressed mutagenicity of benzo[a]pyrend (B[a]P) in the mutagenesis test (Ames test) using liver microsomes prepared from rats treated with eugenol (4-allyl-2-methoxyphenol) was previously reported [Yokota, H., Hoshino, J., and Yuasa, A., 1986, Mutation Res. 172: 231-236]. The mechanism of this suppression was examined. Specific reduction of B[a]P hydroxylase activity in rat liver microsomes was observed by administration with eugenol. However, eugenol treatment did not change either the contents or activities of other microsomal cytochrome P-450-dependent monooxygenase system. Cytochrome P-450s were solubilized with sodium cholate from eugenol-treated rat liver microsomes, and separated into three fractions by the DEAE-TOYOPEARL column chromatography at room temperature. Cytochrome P-450 content of the fraction having B[a]P hydroxylase activity decreased in contrast to the corresponding fraction from untreated rat liver microsomes. In the Ouchterlony double diffusion analysis, the antibody against P-450c (a major 3-methylcholanthrene inducible form of P-450 highly catalytic of B[a]P hydroxylation) did not show any reaction line with untreated microsomes. B[a]P hydroxylase activity in untreated microsomes was not inhibited by the antibody against P-450c. These results suggest that specific cytochrome P-450 isozyme(s) mediating the B[a]P hydroxylase activity was reduced in eugenol-treated rat liver microsomes, and that the isozyme(s) was a different isozyme from P-450c.

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In the winter of 1985, an outbreak of the respiratory disease of dogs occurred in one kennel in Tokyo area. The common clinical signs were depression, anorexia, nasal discharge and mild to severe dry cough. At necropsy of the dogs with severe clinical signs, the pathologic changes were confined only to the respiratory system especially lungs. Canine distemper virus was not considered as the causative agent, however, two different viruses were recovered from 6 of 33 dogs, and they were identified as canine adenovirus-2 (CAV-2) and canine parainfluenza virus. They were present either alone or in combination. Bordetella bronchiseptica was also recovered in pure culture or in combination with other bacteria regarded as normal flora of the dogs. Serologically, CAV-2 was considered as the principal causative agent in the outbreak, even if most infection was silent and the association of opportunistic bacteria was necessary to show the clinical signs. It appeared that the outbreak was "Kennel cough complex", which was confirmed and analyzed for the first time in Japan.

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In the diabetic voles fed with concentrate diet and KK mice, changes in hepatic glycolytic enzyme activities and plasma insulin levels were measured. The hepatic glycolytic enzyme (glucokinase and pyruvate kinase) activities of normal voles and KK mice were about one forth and one half of that of normal C57BL mice, respectively. Both voles and KK mice had the same characteristic which showed that hepatic glycolytic ability was very low. In the slight diabetic voles, glycolytic enzyme activity increased to twice of normal level in proportion to increasing of plasma insulin level, but in the serious diabetic voles plasma insulin level and hepatic glycolytic enzyme activity decreased considerably. In young KK mice without glycosuria, plasma insulin level was considerably high, over three times higher than normal level of C57BL mice. In old KK mice with glycosuria, although plasma insulin level increased moreover, the hepatic glycolytic enzyme activity did not increase. In conclusion, the diabetes of voles is considered to be classified to insulin dependent type and that of KK mice to non-insulin dependent type.

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Cultures of Vibrio fluvialis and Vibrio furnissii isolated from the environment (40 strains) and diarrheal humans (8 strains) were examined for enterotoxicity and other biological activities. The CHO-K1 cell-elongation factor (CEF) was found in 18 and the cell-killing factor (CKF) in 39 of the 48 strains. CEF produced by two strains of V. fluvialis was appreciably neutralized by anti-cholera toxin serum (ACT) but CKF was not. Live cultures of all the human strains tested (3/3) and some environmental strains (8/16) caused fluid accumulation in rabbit ileal loops; the activity shown with concentrated (×10) filtrate was not neutralized by ACT. Cholera toxin (CT)-like substance was detected by ELISA in 28 of 38 cell-free extracts of V. fluvialis and V. furnissii. The correlation between CEF and CKF was discussed. These results and ELISA suggest that CEF has an immunological relation to CT and is distinct from CKF. None of culture filtrates of the 48 strains causcd fluid accumulation in suckling mice. Most strains had mouse-lethal and hemolytic activities.

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Serum cholesterol levels were examined during two consecutive estrous cycles in female beagles. Estrogen and progesterone were also measured simultaneously. In addition, phospholipid, triglyceride and free fatty acid were measured. The peak levels of total cholesterol were 226±25 mg/100 ml at 5.3±0.5 weeks during the first estrus and 234±20 mg/100 ml at 4.8±1.0 weeks during the second estrus. The peak levels of 17β-estradiol and progesterone were 33.0±7.8 pg/ml at just after estrus and 18.7±1.6 ng/ml 2.0±1.0 weeks after the estrus, respectively. The changes in progesterone after estrus resembled those of serum cholesterol after estrus. The increase of phospholipid levels occurred slightly later and was more moderate than that of cholesterol levels. Triglyceride levels tended to be slightly higher at estrus and after several weeks of estrus, but free fatty acid did not changc during estrous cycle in the female beagle.

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The effects of a recombinant interleukin-2 (rIL-2) on active hepatitis due to low virulence mouse hepatitis virus, MHV-2-CC, in athymic nude mice, were studied. Athymic nude mice were inoculated intraperitoneally with 6×10⁵ PFU of the virus and given daily with 0.1, 1 and 10μg of rIL-2. The nude mice treated with 1μg of rIL-2 showed virus titers 1 log lower than those of untreated mice, through Days 7 to 21 of postinoculation. Liver lesions in mice treated with 1 or 10μg of rIL-2 were characterized by multiple round and circumscribed necrotizing foci without inflammatory cells, whereas inflammatory reactions were noticeable in necrotizing lesions of non-treated controls. At the periphery of the lesions of rIL-2 treated mice there were many degenerated hepatocytes being positive for viral antigen surrounded by mononuclear cells, some of which were positive with asialo-GM1.

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The hormonal influence on the carcinogenicity of methylmercury chloride (MMC) was examined in an 80-week experiment using mice. One hundred intact males, 100 intact females, 100 castrated males, and 100 spayed females were divided into 4 groups: (I)intact mice on a basal diet, (II)intact mice on MMC diet, (III)castrated or spayed mice on MMC diet, and (IV)castrated or spayed mice on MMC diet and testosterone propionate (TP) administered s.c. at 0.2 mg/head/week for the entire period. The dietary concentration of MMC was 10 ppm. The survival rate was lowest in both sexes of Group IV because of the aggravation of the toxic nephropathy and the higher incidence of amyloidosis. The incidence of renal epithelial tumors (15/50) and tubular cell hyperplasia (6/50) in Group II males was significantly increased when compared with the controls. No renal tumors or hyperplastic tubules were induced in Group II females or in either sex of Group III. In Group IV, 2 males and 3 females had renal adenocarcinomas and 3 males and 2 females showed tubular cell hyperplasias, indicating some contributing effects of TP. The present results indicate that the testis may have an important role in the induction of renal tumors in mice by MMC.

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Isolation and serological survey of Chlamydia Psittaci in sheep were performed. Complement fixation antibodies were found in 74(34.9%) out of 212 sheep tested in Hokkaido, Japan. The organisms were isolated from feces of 3 out of 8 sheep tested in March and 7 out of 16 sheep in June, 1986. The sheep with the organisms were all clinically normal. From two sheep, the organisms were isolated in both March and June. The results indicated that the organisms were wide-spread in sheep in Hokkaido and excreted in the feces for a long time.

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Thirteen slaughtered dairy cows with high serum fatty acid levels (902 to 2127μEq/l) were examined for their liver lipid components and the fatty acid composition of each component. Almost all of the cows showed parturient paresis or ketosis and had fatty degeneration or hydropic degeneration in their livers. In the diseased livers, the levels of triglyceride increased and those of cholesterol easter, phosphatidyl ethanolamine, and sphingomyelin decreased compared with those from five healthy cows. The unsaturated fatty acid components C18-1 and C18-2 in cholesterol ester and free fatty acids increased, and C18 saturated fatty acid of these lipid components decreased. Polyenoic acids such as C20-3, C20-4 and C22-4, which become precursors of chemical and physiolosical mediators such as prostagrandins, thromboxanes and leukotrienes, dramatically decreased in all phospholipid component except sphingomyelin. In this paper, the question of whether the lack of such polyenoic acid synthesis in liver may cause a clinical sign of parturient paresis in dairy cows is discussed.

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Pharyngostomum cordatum [PC] was investigated on the relation in number between the eggs detected by fecal examination and the flukes parasitic in the small intestine of four cats. Stoll's and the MGL methods were examined on the detecting ability of the eggs in the feces. The number of eggs per day per worm (EPDPW) was calculated at about 1, 000. Based on this egg count, the average detectable number of eggs was calculated at 19.1 on average (from 6.5 to 34.4) in cats parasitized by one fluke by MGL method with 0.5g of feces. From the results, the MGL method was confirmed to be highly reliable on diagnosing Pharyngostomum cordatum infections, even with a single worm.

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It was shown that laboratory-reared Afghan pika, Ochotona rufescens rufescens, was susceptible to gastro-intestinal nematodes of ruminants; Haemonchus contortus, Ostertagia circumcincta, Trichostrongylus axei and T. colubriformis. T. axei and T. colubriformis were recovered up to 70 days post-inoculation and their infection rates were high. Only two gravid females of O. circumcincta were recovered from the pikas necropsied on day 20 post-inoculation. H. contortus was recovered until 20 days post-inoculation but the recovery rate was very low and mature worm was not found. No worm was detected from 45 day onward. Strongyloides papillosus, Bunostomum trigonocephalum, Chabertia ovina, Mecistocirrus digitatus failed to become established in the pika.

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Vibrio parahaemolyticus strain D-3, positive in Kanagawa phenomenon, was found to survive at a considerable level for 21 days after oral inoculation at a level of 10⁴ viable units per gram in an estuarine gastropod mollusc, Clithon retropictus, maintained in artificial seawater with salinities of 15 and 20 grams per liter at 25°C. V. parahaemolyticus strain R-13, negative in Kanagawa phenomenon, survived in the mollusc in lower levels than strain D-3. Escherichia coli strain YS-2 survived in the mollusc in levels of less than 10³ viable units per gram. All strains were decreased to non-detectable level within 72 hr after inoculation in a marine gastropod, Nerita albicilla, maintained in water with a salinity of 35 grams per liter at 28°C. Levels of V. parahaemolyticus strains detected from C. retropictus were fluctuated largely while those of E. coli strain were rather stable. Strains D-3 and R-13 survived together in C. retropictus after mixed inoculation, in which conversion of Kanagawa phenomenon was not observed.

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