Identification and characterization of Marek's disease virus serotype 1 (MDV1) ICP22 gene product: MDV1 ICP22 transactivates the MDV1 ICP27 promoter synergistically with MDV1 ICP4

A previous report [Virus Genes 6 (1992) 365-378] has shown that the US1 gene of Marek's disease virus serotype 1 (MDV1) encodes a homologue of herpes simplex virus type 1 infected cell protein No. 22 (ICP22). In the present study, we expressed and identified a product of the MDV1 US1 gene in chicken embryo fibroblasts (CEFs) with the aid of a recombinant baculovirus expressing a Flag epitope-tagged MDV1 US1 gene, under control of the SRalpha promoter (composed of the enhancer region of the simian virus 40 early promoter and the R region of the human T-cell leukaemia virus type 1 long terminal repeat). In CEF infected with the recombinant baculovirus, MDV1 ICP22 was specifically and efficiently expressed in the presence of n-butyric acid. The apparent M(r) of the expressed protein was 30,000. Reporter gene assays revealed that MDV1 ICP22 by itself transactivated an MDV1 ICP27 promoter/reporter construct weakly but specifically, and furthermore, worked synergistically with MDV1 ICP4 to efficiently up-regulate the MDV1 ICP27 promoter. MDV1 ICP22 may be a regulatory protein that stimulates viral promoters in co-operation with other viral regulatory proteins such as MDV1 ICP4.     

Cloning, expression, and characterization of a novel anti-HIV lectin from the cultured cyanobacterium, Oscillatoria agardhii

OAA, the potent anti-HIV protein from Oscillatoria agardhii NIES-204 belongs to a new lectin family, shows strict binding specificity for high-mannose N-glycans, and has an extremely high association constant in the picomolar range for recombinant gp120, an envelope protein of HIV. In this study we have cloned the gene encoding OAA from the genomic DNA of the cyanobacterium, and efficiently expressed the recombinant lectin (rOAA) in Escherichia coli. The rOAA expressed as a His-tagged fusion protein was recovered in a soluble form and purified in high yield (48 mg/1 l-culture) by metal chelate chromatography. The fusion protein was cleaved with factor Xa, and the resulting rOAA was isolated in a final yield of 14.8 mg/1 l-culture by reversed-phase HPLC. Both the N-terminal sequence and the molecular mass of rOAA were found to be identical with those of OAA. The rOAA was fully functional with the same properties as OAA, as evidenced by hemagglutination activity, hapten-inhibition test, and binding specificity for high-mannose-type N-glycans. This rOAA should be applicable as a specific probe for high-mannose N-glycans and should contribute to elucidation of the molecular basis of its strict carbohydrate-binding specificity and potent anti-HIV activity.     

Influence of sudden changes in management program on physiological and behavioral parameters in hens

The present study was designed to test the effects of sudden changes of food access and light duration on the physiological and behavioral parameters of hens. The physiological parameters investigated were heart rate (HR), body temperature (BT), and locomotor activity (LA) using a radiotelemetry system. After implantation of the telemetry transmitters, six hens were housed individually in cages under constant environmental conditions for 10 days with a photoperiod of 15 h light (04.00–19.00 hours), and food was available ad libitum at all times. After that, the same hens were subjected to a feed withdrawal trial, from 12.00 to 08.30 hours, followed by a lighting hour reduction trial by changing the time of lights‐off from 19.00 to 14.00 hours. The physiological and behavioral data were recorded for 2 days before each trial, as control data. With the feed withdrawal trial, during the light and dark periods, HR and BT were significantly lower in the hens without food access than in the control. Whereas, LA was significantly lower only during the light period in the hens without food access than in the control. Further, the time spent resting increased significantly, but the time spent feeding decreased significantly in the hens without food access than in the control. Also, the number of times the cage was pecked and pecking of the feeder occured at a significantly higher level, while the number of times beaks were wiped occured at a significantly lower level in the hens without food access than in the control. With the lighting hour reduction trial, during the light period, HR and BT were significantly higher, whereas LA was significantly lower in the ’sudden light‐off’ treated hens than in the control. In addition, during the dark period, HR, BT and LA were significantly higher in the sudden light‐off treated hens than in the control. Moreover, in the sudden light‐off treated hens, the time spent preening and feeding decreased significantly, but the time spent resting increased significantly than in the control. It is concluded that sudden changes in a management program might result in many significant differences as were found in the physiological and behavioral parameters of hens in the present study.     

Co-localization of chondromodulin-I (ChM-I) and bone morphogenetic protein-6 (BMP-6) in myoepithelial cells of canine mammary tumors

To compare the roles of chondromodulin-I (ChM-I) and bone morphogenetic protein-6 (BMP-6) in ectopic mesenchymal tissue formation in canine mammary gland tumors, 33 tumors and 2 normal mammary glands were examined. Immunohistochemical analysis revealed co-expression of ChM-I and BMP-6 in canine mammary tumors. In mixed tumors, newly formed woven bone with ossified cartilage matrix was observed in 4/9 cases. The osteoblasts lining the woven bone showed moderate immunoreactivity to ChM-I and BMP-6. Almost all chondrocytes and proliferative myoepithelial cells within the basement membrane showed intense immunoreactivity to both, and the myoepithelial cells adjacent to the mature cartilage showed the most intense immunoreactivity. The immunoreactivity to ChM-I and BMP-6 of the interstitial myoepithelial cells in the myxomatous stroma varied in each focus of mixed tumors. Similar findings were found in complex adenomas. In simple adenomas, hyperplasic myoepithelial cells within the basement membrane showed moderate immunoreactivity to both markers. Western blot analysis detected a 25 kDa band of ChM-I in fresh tissue samples from three mixed tumors. Our results support the hypothesis that proliferating myoepithelial cells with ChM-I and BMP-6 expression play important roles in mesenchymal metaplasia in canine mammary tumors.     

First report of ‘Candidatus Phytoplasma asteris’ infecting hydrangea showing phyllody in Japan

Phytoplasma-induced floral malformations such as virescence, phyllody, and proliferation were observed on hydrangeas in Gunma Prefecture, Japan. Phylogenetic analyses based on 16S rRNA, secY, groEL, and amp gene sequences indicated that the affected hydrangea plants were associated with phytoplasmas belonging to ‘Candidatus Phytoplasma asteris’, but not to ‘Ca. P. japonicum’, which occurs in hydrangeas showing phyllody in Japan. This is the first molecular evidence of an association of ‘Ca. P. asteris’ with hydrangea plants in Japan.     

Magnetic resonance imaging assessment of pituitary posterior lobe displacement in dogs with pituitary-dependent hyperadrenocorticism

The displacement and signal intensity (SI) of the pituitary posterior lobe were evaluated on T1 weighted magnetic resonance (MR) images in 28 dogs with pituitary dependent hyperadrenocorticism (PDH). Compared with normal dogs, the posterior lobe was displaced dorsally in the pituitary of the PDH dogs. Correlation between the pituitary height/brain area (P/B) ratio and the displacement of the posterior lobe in the PDH dogs suggests that dorsal displacement of the posterior lobe increases in accordance with enlargement of the pituitary. As to the SI of the posterior lobe, the PDH dogs showed significantly lower SI in comparison to the normal dogs. Taken together, these results suggest that assessment of the displacement and SI of the posterior lobe of the pituitary on T1-weighted MR images is useful for the diagnosis of pituitary adenoma. In pituitary microadenoma that presents no apparent tumorigenesis on MRI, evaluation of these values may be useful for diagnosis and selection of an appropriate therapy.     

Localization of CCK-1R in the omasum and role of CCK in the regulation of omasal contractions in sheep

The present study examined localization of cholecystokinin receptor (CCK-R) mRNA in the muscle layer of the ovine omasum and role of CCK-R type 1 (CCK-1R) in the regulation of muscle contraction of the omasum. We demonstrated that not only CCK-R type 2 (CCK-2R) mRNA but also CCK-1R mRNA is highly expressed in the muscle layer of the ovine omasum. Application of CCK-8 to muscle strips of the greater curvature of the ovine omasum at 1-100 nM induced tonic contraction in a concentration-dependent manner, and the contractile effect of CCK-8 was inhibited by both CCK-1R antagonist lorglumide (IC(50) 2.7 and 7.9 microM in the longitudinal and circular muscle, respectively) and CCK-2R antagonist PD135,158 (IC(50) 51.4 microM in the longitudinal muscle), indicating that not only CCK-2R but also CCK-1R is functionally expressed in the plasma membrane of smooth muscles in the omasum and mediates action of exogenous CCK. Contractile effect of intravenous infusion of CCK-8 (1-30 pmol/kg/min) on omasal contraction was also confirmed in the in vivo experiments using conscious sheep in the absence and presence of atropine infusion (14.4 nmol/kg/min), and showed that circulating CCK increases omasal electromyographic (EMG) activity at lower plasma concentration than that it inhibits ruminal contractions. Taking account of our previous results in the in vivo study using other CCK-1R antagonist, it is suggested that circulating CCK, even at normal range of plasma concentration, plays a physiological role as a regulator of omasal contractions in sheep and CCK-1R mediates the action of CCK.     

Influence of earthworm activity and rice straw application on the soil microbial community structure analyzed by PLFA pattern

The earthworm Pheretima hilgendorfi, one of the most common anecic species in Japan, abounds in soils with applied rice-straw residues. The influences of the worm’s activity and/or those of rice-straw application on the soil microbial community structure were studied using a microcosm approach. Low Humic Andosol was incubated with or without earthworms in a jar for a month after the following treatments: 1) no treatment, 2) chopped rice straw top-dressed on the soil and 3) rice straw incorporated into the soil. The soil NO₃^–-N level increased with the earthworms’ presence even in the soil without rice straw. The soil NH₄⁺-N level was by far the highest in the soil treated with worms and no rice straw. Amounts of total and bacterial PLFAs increased due to the earthworms’ presence when rice straw was incorporated. The proportion of unsaturated fatty acids increased in the earthworm treatment while the saturated fatty acids decreased, suggesting an increase in the Gram-negative bacterial proportion. The results of principal component analysis indicated that the rice straw and the earthworms affected the soil microbial community structure independently. Any direct influence of the PLFAs contained in the worms’ bodies and in the rice straw on the soil’s PLFA profile was thought to be small.     

乌桕毛状根培养与次生代谢产物累积研究

利用发根农杆菌ATCC 15834诱导获得乌桕毛状根,研究了不同培养条件对乌桕毛状根生长及次生代谢产物累积的影响.结果表明:乌桕毛状根生产量随培养时间的增加而增加,在第6周时达到最大产量(0.6668 g/瓶,DW);1～4周内暗培养更利于毛状根的生长,其后则光照培养更好;在相同培养时间内,毛状根在B5和1/2MS基本培养基上产量大于1/4MS、1/8MS及WP培养基;HPLC测定结果表明,1/4MS较其它培养基更利于多酚类代谢产物的累积.