Invasion and colonization of mature apple fruit by Erwinia amylovora tagged with bioluminescence genes

Invasion and colonization of mature apple fruit by a transformant of Erwinia amylovora tagged with bioluminescence genes from Vibrio fischeri was examined. The transformant was deposited on cut surfaces of fruit stems, wounds on the shoulders and calyces, injured fruit-bearing twigs of harvested apple fruit, and cut fruit flesh. After incubation in closed stainless steel or plastic boxes at 25°C, fruit were periodically observed with a two-dimensional luminometer. The presence of the transformant in luminous areas was confirmed by isolating it on selective media. E. amylovora, when deposited in fruit stems: (1) can invade mature as well as immature apple fruit; (2) vertically and horizontally spreads and colonizes along vascular bundles, increasing its population; (3) reaches the calyx end and the flesh just under the exocarp within 3–4 days after inoculation; (4) when deposited on cut fruit flesh, irrespective of its maturity, can easily increase its population and survive 2–4 weeks or more at 25°C; and (5) even at the time of fruit maturation, can migrate within twigs rapidly and reaches the abscission layers between fruit-bearing twigs and fruit stems.     

Proliferation of canine intervertebral disk chondrocytes in three-dimensional alginate microsphere culture

Proliferation of chondrocytes from nucleus pulposus (NP) and anulus fibrosus (AF) was confirmed in three-dimensional culture using alginate microspheres. Cells isolated from NP and AF were incorporated in microspheres and cultured for 14 days. Round mononuclear cells of 20-25 microm in diameter proliferated and formed aggregates. At day 14, alcian blue positive matrix surrounded the proliferating cells. The cells had cytoplasmic vacuoles stained positively by toluidine blue. On electron microscopy, the cells contained proteoglycan vacuoles and lipid droplets in the cytoplasm and synthesized collagen fibrils and electron dense granules surrounding the cell. These features of the cells were characteristic for chondrocytes. This culture system should be useful to further investigate metabolic activities of intervertebral disk chondrocytes.     

Development of novel clubroot resistant rapeseed lines (Brassica napus L.) effective against Japanese field isolates by marker assisted selection

Clubroot is an important disease infectible to cruciferous plants and a major threat to rapeseed production in Japan. However, no clubroot resistant rapeseed cultivars have been released. We surveyed pathotype variation of six isolates collected from rapeseed fields and found they were classified as pathotype groups 2 and 4 using Japanese F₁ Chinese cabbage cultivars. We produced the resynthesized clubroot resistant Brassica napus harboring two resistant loci, Crr1 and Crr2, by interspecific crossing and developed resistant rapeseed lines for southern and northern regions by marker-assisted selection and backcrossing. We improved the DNA marker for erucic acid content to remove linkage drag between Crr1 and high erucic acid content and successfully selected lines with clubroot resistance and zero erucic acid for northern regions. A novel line, ‘Tohoku No. 106’, suitable for southern regions showed stable resistance against all six isolates and high performance in infested fields. We conclude that Crr1 and Crr2 are important genes for CR rapeseed breeding and marker-assisted selection is effective in improving clubroot resistance.     

Analysis of mitochondrial DNA of an endangered beech species,Fagus hayatae Palibin ex Hayata

Mitochondrial DNA restriction fragment length polymorphisms were used to examine cytoplasmic diversity within a relic-like population of Fagus hayatae, located in northern Taiwan. Fifteen trees were surveyed for three restriction endonucleases (BamHI, EcoRI and HindIII) and five mitochondrial gene probes (atpA, atp6, atp9, coxI and coxII). The analysis failed to reveal any polymorphisms, an observation that suggests cytoplasmic uniformity in the F. hayatae population examined. It is also interesting to note that the chondriome type of our F. hayatae samples is very close to that characteristic of F. crenata populations in the southernmost area of Japan.     

Determining fertilizer-induced NO emission ratio from soils by a statistical distribution model

To reduce uncertainties in the highly variable estimates of NO emission from N fertilizer, we compiled and analyzed available reports of field measurements on fertilizer-induced NO emission. Three statistical distribution models, lognormal, gamma and Weibull, were used to fit the observation data. Results show that while all three models fit the observation data statistically, the lognormal model is not applicable to this data set. A mean fertilizer-induced NO emission ratio of 0.71% was derived from a Weibull distribution, which is the besting fitting model.     

Genome‐wide association study and genomic evaluation of feed efficiency traits in Japanese Black cattle using single‐step genomic best linear unbiased prediction method

The objectives of this study were to better understand the genetic architecture and the possibility of genomic evaluation for feed efficiency traits by (i) performing genome‐wide association studies (GWAS), and (ii) assessing the accuracy of genomic evaluation for feed efficiency traits, using single‐step genomic best linear unbiased prediction (ssGBLUP)‐based methods. The analyses were performed in residual feed intake (RFI), residual body weight gain (RG), and residual intake and body weight gain (RIG) during three different fattening periods. The phenotypes from 4,578 Japanese Black steers, which were progenies of 362 progeny‐tested bulls and the genotypes from the bulls were used in this study. The results of GWAS showed that a total of 16, 8, and 12 gene ontology terms were related to RFI, RG, and RIG, respectively, and the candidate genes identified in RFI and RG were involved in olfactory transduction and the phosphatidylinositol signaling system, respectively. The realized reliabilities of genomic estimated breeding values were low to moderate in the feed efficiency traits. In conclusion, ssGBLUP‐based method can lead to understand some biological functions related to feed efficiency traits, even with small population with genotypes, however, an alternative strategy will be needed to enhance the reliability of genomic evaluation.     

Plant growth regulation effects of triterpenoid saponins

 To investigate structure–activity relations between the sugar chain structures of triterpenoid saponins and their plant growth regulation effects, several monodesmosidic saponins with betulin as an aglycon were synthesized by chemical and enzymic reactions. Three triterpenoids (betulin, betulinic acid, oleanolic acid) and synthesized betulin glycosides were submitted to germination and growth regulation tests on alfalfa seeds. We concluded the following. Betulin had a slight growth inhibitory effect on alfalfa radicles. Betulin glycosides exhibited stronger effects than betulin, and betulin glycosides with two to four glucose residues as a sugar moiety had the greatest inhibitory activity. These characteristics of growth inhibitory effects were considerably different from those of phenolic compounds so far reported. Some betulin glycosides also showed a significant growth regulation effect on alfalfa hypocotyls. However, hypocotyl growth was less affected than radicle growth for all betulin glycosides. Among the triterpenoids, betulinic acid had stronger growth inhibitory effects on alfalfa radicles than betulin, suggesting the importance of the carboxyl group at the C-28 position for the inhibitory effects of lupane-type triterpenoids. On the other hand, no germination regulation effects on alfalfa seeds were observed for any of the betulin glycosides or triterpenoids examined. Received: October 9, 2001 / Accepted: February 15, 2002 Acknowledgments This work was supported in part by a research grant (Development of Highly Functional Materials by Structural Modification of Carbohydrates) from the Ministry of Agriculture, Forestry, and Fisheries of Japan. We thank Saori Kudo for her assistance in isolating compounds by chromatography. Correspondence to:S. Ohara     

EVALUATION OF IOHEXOL AS A GASTROINTESTINAL CONTRAST MEDIUM IN NORMAL CATS

The non-ionic, iodinated contrast medium, iohexol (240 mg I/ml) was evaluated as a gastrointestinal (GI) contrast medium in cats. Iohexol, both undiluted and diluted with tap water, was administered via a percutaneous endoscopically-placed gastrotomy (PEG) tube to 4 mature clinically normal cats. The dilution of contrast medium administered was 1:1, 1:2, and 1:3, and doses were 10 ml/kg and 5 ml/kg body weight. All combinations of dilution and dose of iohexol provided adequate visualization of the contrast medium column within the GI tract, and results were not significantly different than those observed using 30% w/v barium sulfate. Dehydration and diarrhea were not observed after contrast medium administration, but vomiting occurred within 15–30 minutes after administration of undiluted iohexol in all experimental cats. Renal opacification did not occur on exposures made through a 2 hour period, and dilution in transit was not apparent.     

ULTRASONOGRAPHIC APPEARANCE OF CHRONIC HYPERTROPHIC PYLORIC GASTROPATHY IN THE DOG

Chronic hypertrophic pyloric gastropathy was diagnosed using ultrasound in six dogs that were presented for chronic vomiting. An evenly thick hypoechoic layer surrounding the pyloric lumen was visualized ultrasonographically in dogs with grades 1 or 2 chronic hypertrophic pyloric gastropathy. This was histologically found to correspond with a thickened muscular layer. Gastric wall thickness was greater than or equal to 9 mm in all six dogs. In dogs with grade 1 or 2 chronic hypertrophic pyloric gastropathy, the hypoechoic muscular layer was greater than or equal to 4 mm.     

Frequencies of PrP genotypes in meat breeds of Japanese sheep and trail of selective breeding in experimental sheep flock

The selection of sheep with scrapie-resistant PrP genotypes is one of the control measures for transmissible spongiform encephalopathies in ruminants. In this study, we investigated the frequencies of PrP genotypes in meat breeds in Japan. The nationwide surveillance revealed that nearly half of the Suffolk sheep, a major meat breed in Japan, carried scrapie-susceptible AQ/AQ and AQ/VQ genotypes. In addition, the VQ haplotype, which confers high susceptibility to scrapie within sheep, was also found in Poll Dorset sheep. A trial of selective breeding using sires with scrapie-resistant PrP genotypes AQ/AR and AR/AR could raise the ratio of scrapie-resistant sheep from less than 50% to 80% within 3 years. However, the use of sires with the AR/AR genotype and the selection of ewes would be required to achieve a higher ratio of scrapie-resistant sheep.