In vitro detection of equine arteritis virus from seminal plasma for identification of carrier stallions

Equine arteritis virus (EAV) was readily isolated in RK-13 cell monolayers by plaque assay from seminal plasma of experimental carrier stallions when they contained high titers of virus regardless of the presence of non-viral cytotoxicity in the seminal plasma. The cytotoxicity interfered with virus isolation from seminal plasma which contained virus at titers less than 10 PFU/ml. However, it was possible to detect the virus in seminal plasma pretreated with PEG (#6000). EAV was consistently identified by RT-PCR from crude seminal plasma which contained virus at titers of more than 10(2.7) PFU/ml. In vitro detection of EAV by virus isolation supplemented with RT-PCR using seminal plasma was proved to be an effective alternative to the standard test mating as a diagnostic method for carrier stallions.     

Screening of extracts of Japanese woods for melanin biosynthesis inhibition

The effects of 120 methanol extracts prepared from bark and heartwood of 69 types of Japanese wood on the melanin production of B16 melanoma cells were examined. The melanin content of B16 melanoma cells was determined spectrophotometrically at 405nm. The extracts were also examined for their effects on cell viability. We found that the methanol extracts of Fagus crenata (buna, wood, 100μg/ml), Sapium sebiferum (Nankinhaze, wood, bark, 10μg/ml), and Zelkova serrata (keyaki, wood, 10μg/ml) greatly inhibited the melanin production of B16 melanoma cells without significant cytotoxicity. However, these extracts did not inhibit tyrosinase activity at the concentration of 100μg/ml. These findings indicate that the depigmenting mechanism of these extracts involves the suppression of some pigmenting signals in stimulating melanogenesis rather than the inhibition of tyrosinase activity. Part of this study was presented at the 53rd Annual Meeting of the Japan Wood Research Society, Fukuoka, Japan, March 2003     

Separation of sardine oil without heating from surimi waste and its effect on lipid metabolism in rats

Sardine oil was obtained by centrifugation of surimi wastewater without heating or chemical refining. This oil (CE) showed light yellow color and the peroxide value was less than 1.0 meq/kg. The main lipid class of CE was triacylglycerol (TG) (>99%). These features indicate that CE can be directly used as food materials without further purification. Commercial sardine oil (CO) is usually prepared via some kind of refining process with high temperature (250 degrees C) and chemical treatment. The comparative study on the physiological effects of these sardine oils (CE and CO) revealed that the dietary sardine oils were more effective in reducing abdominal fat pads, plasma total cholesterol, and TG levels of rats than was a soybean oil diet (control). Furthermore, these effects were greater in CE than CO, although there was little difference in the fatty acid composition of both oils. Although the main lipid class of CE was TG (>99%), CE was prepared by centrifugation from surimi waste and directly used as dietary fat without further purification. Therefore, CE may contain some kinds of minor components, which could be attributed to the higher physiological activity of CE. To reveal the involvement of the minor compounds in CE, we prepared TG from CE by column chromatography and measured its effect on lipid metabolism of rats. TG from CE also showed the reducing effects on abdominal fad pads and plasma lipid levels. The effect of TG from CE was almost the same as that of original CE, suggesting that the higher nutritional activity of CE than CO may not be due to the minor compounds in CE.     

Growth and maturation of Pacific saury Cololabis saira under laboratory conditions

This study details the growth and maturation processes of Pacific saury Cololabis saira from eggs to first spawning under laboratory conditions. They were reared at 20°C, and fed almost to satiation every day. There was no significant difference in the knob length (KnL) between males and females, and so data for both sexes were combined in the following Gompertz growth formula: KnL _t  = 277.1 exp[?exp(?0.015 (t ? 83.8))]. The first spawning was observed 243 days after hatching (DAH). At the beginning of spawning, when the mean KnL was about 250 mm, only several hundred eggs were spawned at most. The spawning continued, and the number of spawned eggs increased notably after 260 DAH. Correlation and stepwise multiple regression analysis of the gonad somatic index (GSI) of Pacific saury versus KnL, CF, and DAH revealed that only DAH (R = 0.88 and 0.72 for males and females, respectively) was significantly correlated with GSI of Pacific saury (P < 0.001). This result suggests that DAH is one of the most influential factors for maturation in this species.     

Long-lasting enhancement of CYP activity after discontinuation of repeated administration of phenobarbital in dogs

We investigated how long in vivo hepatic cytochrome P450 (CYP) activity is enhanced even after discontinuation of repeated oral administration of phenobarbital (PB) in dogs using antipyrine clearance, which reflects hepatic CYP activity. A single antipyrine (5 mg/kg) was administered intravenously before and 34 days after the repeated oral administration of PB (5 mg/kg, bid) and 2, 4, 6, and 8 weeks after the discontinuation of PB in 5 dogs. Antipyrine clearance was increased by the repeated administration of PB, and remained increased 2 and 4, but not 6 and 8 weeks after the discontinuation of PB. The result suggests that hepatic CYP activity was enhanced by the repeated administration of PB, and this enhancement may last for at least 4 weeks even after its discontinuation.     

Origin of waxy common millet, Panicum miliaceum L. in Japan

We developed PCR-based markers for genotyping Waxy genes in common millet (Panicum miliaceum L.) and proposed a new hypothesis on the origin and dispersal of waxy type of Japanese landraces. I₂-K staining indicated that almost all the Japanese landraces had waxy endosperm. Based on the PCR and dCAPs analyses of two waxy loci on different genomes we concluded that there were at least two possible lineages in Asian continent that had independent dispersal pathways into Japan. Most Japanese common millet had the similar genotypes with those in Korean Peninsula but a few particularly from northern part of Japan had common genotypes in northeastern China and Primorskaya Province of Russia.     

Geographic distribution of Waxy gene SNPs and indels in foxtail millet, Setaria italica (L.) P. Beauv.

To elucidate diversity and evolution of the Waxy gene in foxtail millet, Setaria italica, we analyzed sequence polymorphism of Waxy gene in 83 foxtail millet landraces collected from various regions covering the entire geographical distribution of this millet in Europe and Asia. We found a unique geographic distribution pattern at the sequence level of gene haplotypes and also found a large diversity in East Asian landraces. We also found a higher degree of genetic polymorphism in a non-waxy phenotype than in other low amylose types, supporting the hypothesis that low amylose types recently originated from non-waxy type.     

Getah virus as an equine pathogen.

Getah virus is a member of the genus Alphavirus in the family Togaviridae and has been frequently isolated from mosquitoes. Seroepizootiologic studies indicate that the virus is mosquito-borne and widespread, ranging from Eurasia to southeast and far eastern Asia, the Pacific islands, and Australasia. The natural host animal of the virus was not known until the first recognized occurrence of Getah virus infection among racehorses in two training centers in Japan in 1978. Outbreaks of clinical disease due to Getah virus infection occur infrequently, and only one outbreak has been reported outside Japan; this was in India in 1990. Clinical signs of the disease are mild and nonlife-threatening and are characterized by pyrexia, edema of the hind limbs, swelling of the submandibular lymph nodes, and urticarial rash, as reported in the 1978 epizootic. The morbidity was 37.9% (722 of 1903 horses) in one training center, with 96% of 722 affected horses making a full clinical recovery within a week without any significant sequelae. Antibodies against Getah virus were detected in 61.2% (172 of 281) and 55.8% (254 of 455) of horses at two training centers, respectively. Virus isolation can be attempted in VERO, RK-13, BHK-21, and many other cell lines as well as in suckling mouse brain. Blood plasma collected from suspect cases of infection at the onset of pyrexia is the specimen of choice. A diagnosis of Getah virus infection can also be confirmed serologically based on testing acute and convalescent phase sera by using SN, CF, HI, and ELISA tests. An inactivated vaccine is available for the prevention and control of Getah virus infection in horses in Japan.