A new DNA marker for sex identification in purple asparagus

Garden asparagus (Asparagus officinalis L.) is an economically-important perennial crop. This plant is dioecious, as there are both male and female individuals; male individuals are preferred over females for agricultural production. To reduce the time required for garden asparagus breeding, various male-specific DNA markers are utilized. Male-specific DNA markers, such as Asp1-T7sp and MSSTS710, are currently available for sex identification in many asparagus cultivars. In the current study, we found that these markers are not suitable for sex identification in the purple asparagus cultivar ‘Pacific Purple’, as male-specific amplification of this marker was detected in some male individuals of this cultivar but not in other males. The Asp1-T7sp marker is suitable for use in sex identification in various Asparagus species related to A. officinalis, indicating that the region around this marker is conserved among these species. Thus, we isolated a DNA fragment around this marker by inverse PCR and produced a new DNA marker, MspHd, based on this sequence. However, like Asp1-T7sp and MSSTS710, MspHd was not suitable for sex identification in the cultivar ‘Pacific Purple’. Since all ‘Pacific Purple’ males have morphologically similar male flowers with functional stamens, we produced a new male-specific marker based on the sex determination gene, MSE1/AspMYB35/AspTDF1, which is responsible for stamen development. This marker, named AspMSD, is suitable for sex identification in ‘Pacific Purple’. In addition, this marker can be utilized for sex identification in various asparagus cultivars and some related Asparagus species.     

Determination of physical and chemical properties and degradation of archeological Japanese cypress wood from the Tohyamago area using near-infrared spectroscopy

Here, we evaluated the application of near-infrared (NIR) spectroscopy for estimating the degradation level of archeological wood samples from the Tohyamago area, the dendrochronological ages of which were also determined. The wood samples were radially cut from three logs obtained from the Tohyamago area. NIR reflectance spectra were measured from the tangential faces of air- and oven-dried wood samples using a Fourier transform NIR spectrophotometer. The second derivative spectra within the wavenumber range of 6400–5200 cm^?1, in which the effect of moisture content in wood is suspected to be insignificant, showed a characteristic behavior with age. By comparing the second derivative spectral change in our wood samples with that in wood degraded by aging, thermal treatment, fungal attack, and lightning reported in the literature, we found that the second derivative spectra of wood samples from one log was similar to those of wood degraded by hygro-thermal treatment, whereas those of wood samples from another log was similar to those of wood degraded by brown-rot fungi. The physical and chemical properties of archeological wood were well predicted using a combination of partial least square regression analysis and NIR spectroscopy.     

Selective assessment of duplex heat-treated wood by near-infrared spectroscopy with principal component and kinetic analyses

We selectively assessed the thermal and hygrothermal treatment times of duplex heat-treated samples from the softwood hinoki cypress (Chamaecyparis obtusa) and the hardwood Japanese zelkova (Zelkova serrata) using near-infrared (NIR) spectroscopy with principal component analysis (PCA) and spectral-kinetic analysis. Wood samples from each species were thermally or hygrothermally treated at 120, 130, 150, and 180 °C, and the second-derivative spectra of these samples in the 6300–5450 cm^?1 range, where moisture content has the smallest effect, were then subjected to PCA. The master curve that was calculated by kinetic analysis successfully explained changes in the first principal component (PC1) scores with thermal treatment time for all temperatures. The angles between the PC1 loadings that explained the spectral variation due to thermal and hygrothermal treatment were 79° for hinoki and 80° for zelkova. Thus, calculation of the inner product between the second-derivative spectra of duplex heat-treated wood and a loading vector that explained the spectral variation due to thermal or hygrothermal treatment allowed us to selectively assess the thermal and hygrothermal treatment times.     

Seroprevalence of bovine immunodeficiency virus and bovine leukemia virus in dairy and beef cattle in hokkaido

Serological survey of bovine immunodeficiency virus (BIV) and bovine leukemia virus (BLV) infection was conducted in dairy cattle from 10 different regions of Hokkaido, Japan. Among 390 cattle, 11.0% of cattle were BIV-seropositive and 3.3% were BLV-seropositive. Moreover, in two dairy farms, where bovine leukosis has been reported, prevalence of BIV infections were 6.4 and 9.1%, respectively. In contrast, among 150 beef cattle, 16.6% were BIV-seropositive while none was BLV-seropositive. Dual infections with BLV and BIV in dairy cattle were tested by using 107 BLV-seropositive sera, and 20 sera were found BIV-positive (18.7%). These results indicate that BIV infection was widespread in Hokkaido.     

Plasmodium coatneyi-infected erythrocytes bind to C32 amelanotic melanoma cells under static and flow conditions

The ability of Plasmodium coatneyi-infected red blood cells (IRBCs) to bind to C32 amelanotic melanoma cells was examined under static and physiologic flow conditions in vitro. Six blood samples obtained from P. coatneyi-infected Japanese macaques (Macaca fuscata) with severe manifestations of disease were used in the static adhesion assay. All blood samples constantly exhibited binding of IRBCs to C32 cells under static conditions. Immunofluorescence staining with anti-CD36 mAb revealed a positive reaction at the surface of C32 cells with the infected erythrocytes, while the reaction with C32 cells without IRBCs was negative. To further examine the specificity of the interaction between P. coatneyi-infected erythrocytes and C32 cells, we carried out the binding assay under physiological flow conditions. In flow adhesion assay, three blood samples were used. Adhesion and rolling of IRBCs on C32 cells were detected at several rates of shear stress under flow conditions. At a shear stress of 1.0 dyne/cm(2), the number of IRBCs adherent to C32 cell averaged 5 to 6, and the number of IRBCs rolling on C32 cells averaged 6 to 11. The anti-CD36 mAb OKM5 inhibited 75-100% of IRBC adhesion and rolling, while the inhibitory effect of anti-ICAM-1 mAb 84H10 varied between 20-40%. The combination of anti-CD36 and anti-ICAM-1 mAb resulted in 83-100% inhibition of rolling and 100% inhibition of adhesion. These findings suggest that CD36 is one of the principal adhesion receptors of P. coatneyi-infected erythrocytes.     

New melanin biosynthesis inhibitors and their structural similarities

Relationships between their activities as blast control agents, and their abilities to inhibit mycelial melanisation on a nutrient agar, are described for 103 substituted benzothiazol-2(3H)-ones, benzoxazol-2(3H)-ones, indolin-2-ones, quinolin-2(1H)-ones, 1,2,3,4-tetrahydroquinolin-2-ones, benzo-1,4-thiazin-3(2H)-ones and benz-1,4-oxazin-3(2H)-ones, and some corresponding thiones. Several compounds in the respective series had a high protective activity and an antimelanisation activity against the blast fungus Pyricularia oryzae; furthermore, there was a good correlation in both of these activities, indicating that these compounds belong to the group of melanin biosynthesis inhibitors. Structural similarities of these compounds can be identified as follows: (a) having a benzo-bicyclic ring system; (b) containing a nitrogen atom in one ring at a position alpha to the benzene ring system; and (c) substitution, at the ring nitrogen atom, at the peri position in the aromatic ring relative to the nitrogen atom, and at the position alpha to the nitrogen atom in the ring system, with a double bond such as in carbonyl and thiocarbonyl groups. Among the compounds that have been proposed as melanin biosynthesis inhibitors, the chemical structures of tricyclazole, pyroquilon, 4,5-dihydro-4-methyltetrazolo[1,5-a]quinazolin-5-one (PP-389), [1,2,4]-triazolo[4,3-a]quinoline and 1-methylquinolin-2(1H)-one exhibit the structural similarities described above; however, 4,5,6,7-tetrachlorophthalide, 2,3,4,5,6-pentachlorobenzyl alcohol and N-substituted-2,3,4,5-tetrachloro-6-(hydroxymethyl)benzamides do not have such similarities in their chemical structures.     

The expression of sialyl Lewis X in canine and feline mammary gland tumors

The expression of sialyl Lewis X (sLe(x)) in 93 canine and 15 feline mammary gland tumors (MGT) obtained by surgical resection at Veterinary Medical Center, the University of Tokyo was examined by immunohistochemistry. Their clinicopathological features and prognosis were also reviewed. Approximately 60% of MGT tissues showed sLe(x) positive expressions, while all normal mammary gland tissues were negative. However, its expression was not correlated with clinicopathological features and prognosis significantly. This study suggests that sLe(x) may be a tumor-associated antigen in canine and feline MGTs.     

Examination of the rat eye at the early stage of development with osmium tetroxide staining

This communication describes the benefit of osmium tetroxide (OsO4) staining on the examination of the eye during the early stage of organogenesis of rat embryos. The embryos were obtained by laparotomy on embryonic day 12 (ED 12) and were stained with OsO4 for examination of the ocular tissues with a binocular stereo-microscope, light microscope and scanning electron microscope. At the binocular stereo-microscopic level, the invaginated lens placode, lens pit and optic cup were clearly distinguished. The osmium-stained lens placode and the optic cup were light brown and dark brown in color, respectively. Light microscopic examination revealed that OsO4 postfixation could provide superior paraffin-embedded embryonic sections. Scanning electron microscopic examination revealed the lens pit as a round opening between the lateral nasal prominence and maxillary prominence. Thus, a rapid technique by which the ocular tissues of rat embryos can be examined under a binocular stereo-microscope was developed. This OsO4 staining method will provide a useful tool for research on organogenesis and ocular development.     

Characterization of ISPsy2 and ISPsy3, Newly Identified Insertion Sequences in Pseudomonas syringae pv. eriobotryae

Two new active insertion sequences, ISPsy2 and ISPsy3, were isolated from Pseudomonas syringae pv. eriobotryae, the causal agent of stem cankers of loquat trees. ISPsy2 is 1194-bp long, has 16-bp imperfect terminal inverted repeats, and generates a 4-bp target site duplication upon insertion into the selective cartridge of the entrap vector pSHI1063. The nucleotide sequence of ISPsy2 is completely identical with that of the previously identified IS-like element located adjacent to the virulence gene psvA of Pseudomonas syringae pv. eriobotryae NAE6. The single open reading frame of ISPsy2 encodes a 323-amino-acid protein that has similarity to the transposase of the IS5 subgroup of the IS5 family. The ISPsy3 belonging to the IS91 family is 1507 bp in length, does not duplicate its target sequence, GAAC, and presents an 81% sequence homology with IS801 in P. s. pv. phaseolicola. The transposase of ISPsy3 possesses the conserved amino acid motifs found in the rolling-circle replication protein. Southern blot analysis indicated that multiple copies of ISPsy2 and ISPsy3 are present in the genomes of P. s. pv. eriobotryae and some of the other P. s. pathovars tested. Received 16 August 2001/ Accepted in revised form 19 October 2001     

Purification and characterization of a lipolytic enzyme active at low temperature from Norwegian Typhula ishikariensis group III strain

An extracellular lipolytic enzyme active at low temperature was purified from the culture filtrate of the snow mold fungus, Typhula ishikariensis group III, strain 6-1-1. The molecular mass of enzyme was approximately 83 kDa (SDS-PAGE). The lipolytic enzyme was most active for p-nitrophenyl palmitate at 30 –C (pH 9.0). The lipolytic activity was 23.4% of the maximum at 4 –C, the temperature of culture. Thus, the isolated T. ishikariensis lipolytic enzyme is thought to represent a new member of a group of enzymes active at low temperature.