Analysis of rice <Emphasis Type="Italic">RNA-dependent RNA polymerase 1</Emphasis> (<Emphasis Type="Italic">OsRDR1</Emphasis>) in virus-mediated RNA silencing after particle bombardment

RNA-dependent RNA polymerases (RDRs) play a key role in various RNA silencing pathways in many organisms. Using the nucleotide sequence of SGS2/SDE1/RDR6 in Arabidopsis as the search query for sequences that flank the insertions of rice retrotransposon Tos17, we selected rice mutant lines (OsRDR1). RT-PCR analysis showed that OsRDR1 mRNA was undetectable in leaves and calli of the mutants, while it was detected in wild type. RNA silencing was induced by particle bombardment to investigate any effects of OsRDR1 on RNA silencing with β-glucuronidase or green fluorescent protein DNA/RNA in the mutant lines. The results showed that RNA silencing was impaired in these mutant lines by inverted repeat (IR) DNA or in vitro transcribed double-stranded RNA. Further, the mutant lines were bombarded with Brome mosaic bromovirus (BMV, a ssRNA virus) or Wheat dwarf geminivirus (WDV, a ssDNA virus), each carrying the IR sequence of a reporter gene. As a result, RNA silencing was impaired by BMV. Interestingly, however, it was not impaired by WDV. Thus we propose that OsRDR1 is required for RNA silencing mediated by Bromovirus, but not by Geminivirus in this system.     

Transport and survival of Japanese sardine (Sardinops melanostictus) eggs and larvae via particle‐tracking experiments

Particle‐tracking experiments were performed to infer the distribution of larvae of the Japanese sardine (Sardinops melanostictus) and to detect effects of transport environment on sardine recruitment, using the output of a high‐resolution ocean general circulation model and observed data of sardine spawning grounds during 1978–2004. By the 60th day following spawning, approximately 50% of the larvae had been transported to the Kuroshio Extension (KE). Whereas the spawning period and grounds changed markedly in relation to the stock level, the proportion of larvae transported to the KE remained relatively constant and no significant correlations were found between sardine recruitment and the transport proportion. Instead, the recruitment was found to be correlated with physical parameters including the mixed layer depth and the sea surface temperature along several major transport trajectories of sardine larvae. The correlations were most significant for the trajectories in the region 0.5° south to 1° north of the Kuroshio axis (defined as the location of velocity maxima at each longitude) and for larvae spawned in February and March during the high stock period (1978–94), and for larvae spawned in March and April during the low stock period (1995–2004).     

Screening and cDNA cloning of Kv1 potassium channel toxins in sea anemones

When 21 species of sea anemones were screened for Kv1 potassium channel toxins by competitive inhibition of the binding of (125)I-α-dendrotoxin to rat synaptosomal membranes, 11 species (two species of Actiniidae, one species of Hormathiidae, five species of Stichodactylidae and three species of Thalassianthidae) were found to be positive. Furthermore, full-length cDNAs encoding type 1 potassium channel toxins from three species of Stichodactylidae and three species of Thalassianthidae were cloned by a combination of RT-PCR, 3'RACE and 5'RACE. The precursors of these six toxins are commonly composed of signal peptide, propart and mature peptide portions. As for the mature peptide (35 amino acid residues), the six toxins share more than 90% sequence identities with one another and with κ(1.3)-SHTX-She1a (Shk) from Stichodactyla helianthus but only 34-63% identities with the other type 1 potassium channel toxins.     

Carbon balance in a cool–temperate deciduous forest in northern Japan: seasonal and interannual variations, and environmental controls of its annual balance

We monitored variation in seasonal and annual net ecosystem production (NEP), gross primary production (GPP), and ecosystem respiration (R _E) based on 7-year eddy covariance measurements above a cool?Ctemperate deciduous broad-leaved forest (Japanese beech forest). The 7-year means (±SD) of annual NEP, GPP, and R _E were 312?±?64, 1250?±?62, and 938?±?36?g?C?m^?2?year^?1, respectively. Variation in NEP was much larger than variation in GPP and R _E. During the growing season, the main factor controlling carbon balance was air temperature; variation in seasonal integrated NEP was regulated by accumulated air temperature (degree-day) with a significant negative correlation, whereas the seasonal ratio of R _E to GPP was correlated positively with accumulated air temperature. Because the deviation of seasonal NEP was also significantly correlated with seasonal R _E/GPP, NEP was controlled by R _E/GPP, depending on air temperature during the growing season. Seasonal R _E in the defoliation and snow seasons was also important for evaluating the annual carbon balance, because the total number of days in the two seasons was quite large owing to a long snowy winter. In the defoliation and snow seasons, we found defoliation season length was a major factor determining seasonal integrated R _E, illustrating the positive correlation between R _E and defoliation season length. The major factors controlling interannual variations in forest carbon balance are discussed.     

Canine intestinal lactic acid bacteria agglutinated with concanavalin A

Twenty-six out of 46 representative lactic acid bacteria (LAB) that we isolated from 36 dogs in a previous study were agglutinated by concanavalin A (ConA) at a concentration of 0.1563 mg/ml, while isolates did not agglutinate without the addition of ConA. Amongst the isolates, L. reuteri, L. mucosae, and E. canintestini agglutinated strongly, while L. gallinarum, L. kitasatonis, L. acidophilus, L. saerimneri, B. animalis ssp. animalis, P. acidilactici, and E. hirae did not agglutinate. ConA-agglutination of LAB was specifically inhibited by D-glucose, D-galactose, and D-mannose at a concentration of 1.563 mg/ml. Among the sugars, ConA-agglutination was strongly inhibited by D-mannose, while the inhibition level by D-glucose and D-galactose were lower than that of D-mannose. ConA- agglutination of all the LAB isolates was inhibited by D-mannose, except for L. reuteri (one species) and L. mucosae (two species). ConA-agglutination of Bifidobacterium spp. was inhibited by only D-mannose. Based on our results, ConA-agglutination of LAB seems to be strain-specific, but not species-specific.     

Feasibility of improved slotted bolted connection for timber moment frames

This note examines the feasibility of an improved slotted bolted connection for timber moment frames. In the improved connection, steel tubes are inserted into drill holes in glulam and fixed to the glulam with resin injection. Aluminum splice plates with curved slots, or curved elongated holes, are fastened mechanically by using high-strength bolts that go through the steel tubes. Since the compression due to bolt tension is fully supported by the steel tubes, the reduction of bolt tension due to shrinkage of the glulam can be avoided. The use of slotted aluminum splice plates allows stable energy dissipation due to smooth sliding between the aluminum splice plates and the end surfaces of the steel tubes within the specified range of rotation angle. Through quasistatic cyclic loading tests of two connection specimens, it was demonstrated that stable and nearly rigid-plastic hysteresis loops were obtained whose equivalent viscous damping ratio was more than 30% in the range of rotation angle close to or greater than 1/50 radian. Although further improvement is necessary, the experimental results demonstrate the feasibility and potential of the present connection.     

Cardiac hamartoma in a young squirrel monkey who died suddenly

A case of cardiac hamartoma in a 2-month-old squirrel monkey is reported. The monkey showed a loss of appetite and died suddenly. Microscopically, an encapsulated nodular lesion was found at the right atrial wall. The lesion consisted of irregularly shaped, slender myocytes intermingled with a few fibroblasts and collagen fibers. Neither nuclear atypia nor inflammatory cell infiltrate was seen. The constituting cells had stratified striations in the cytoplasm and reacted immunohistochemically for desmin, indicating the nature of myocytes. Based on the above findings, a diagnosis of cardiac hamartoma was made. This is the first case of cardiac hamartoma in this species.     

Genetic characterization and susceptibility on poultry and mammal of H7N6 subtype avian influenza virus isolated in Japan in 2009

From February to March 2009, six strains of H7N6 subtype avian influenza virus were isolated from quails in three farms in Aichi prefecture in Japan. The isolates were shown to be low pathogenic for chicken by the examination performed using the "Manual of Standards for Diagnostic Tests and Vaccines" by World organisation for Animal Health (OIE). The deduced amino acid sequence at the cleavage site was PE (I/Q/L) PKRR (nucleotide sequences were cct gaa (a/c) (t/a) a cc (a/g) aaa aga aga), suggesting persistence in domestic poultry for some time. The direct putative ancestor strain could not be elucidated by phylogenetic analysis of all genome segments of the quail isolates. Diverged date from a putative common ancestor in a non-rooted phylogenetic tree among quail viruses was estimated between March 2002 and July 2004. Three putative N-linked glycosylation sites resided in the vicinity of the receptor binding pocket of HA1 region. They are considered to decrease the reactivity of neutralizing antibody against the virus. Experiments for the infectivity and pathogenicity of a quail strain to poultry indicated that the quail isolate had higher infectivity to quails than chickens and ducks. Direct and dust-borne and/or droplet-borne transmissions among quail were proven in quails with and without direct contact with experimentally infected quails. The virus is seldom transmitted among chickens either directly or indirectly, and indirect transmission from infected quails to chickens was not observed. The pathogenicity of the quail strain for mammalian, pig and mouse was low, although it could replicate in those animals.     

Successful management with CHOP for pulmonary lymphomatoid granulomatosis in a dog

A 3-year-old, spayed female miniature dachshund was presented for vomiting and anorexia. Thoracic radiographs and CT scan revealed abnormal pulmonary opacities at bilateral caudal lobe. Cytological analysis of the pulmonary mass revealed the presence of large lymphohistiocytic cells and small lymphocytes with occasional neutrophils and plasma cells. An open lung biopsy was performed and a diagnosis of pulmonary lymphomatoid granulomatosis (LYG) was made. The dog was administered CHOP based therapy (modified UW-25), and it survived for 1,022 days after admission. Immunohistochemistry revealed pulmonary lesions consisted of many CD79a positive B cells aggregation and proliferation with prominent angiocentric pattern. This was the first case of canine pulmonary LYG managed by CHOP chemotherapy.     

Non-CpG methylation occurs in the regulatory region of the Sry gene

The Sry (sex determining region on Y chromosome) gene is a master gene for sex determination. We previously reported that the Sry gene has tissue-dependent and differentially methylated regions (T-DMRs) by analyzing the DNA methylation states at CpG sites in the promoter regions. In this study, we found unique non-CpG methylation at the internal cytosine in the 5'-CCTGG-3' pentanucleotide sequence in the Sry T-DMR. This non-CpG methylation was detected in four mouse strains (ICR, BALB/c, DBA2 and C3H), but not in two strains (C57BL/6 and 129S1), suggesting that the CCTGG methylation is tentative and unstable. Interestingly, this CCTGG methylation was associated with demethylation of the CpG sites in the Sry T-DMR in the developmental process. A methylation-mediated promoter assay showed that the CCTGG methylation promotes gene expression. Our finding shows that non-CpG methylation has unique characteristic and is still conserved in mammals.