Genetic homogeneity between adult and juvenile populations of <Emphasis Type="Italic">Scombrops gilberti</Emphasis> (Percoid,Scombropidae) in the Pacific Ocean off the Japanese Islands

Scombrops gilberti is a member of the percoid family Scombropidae, which includes a single genus and three to four species worldwide. Little is known about the ecology of this species. Juvenile S. gilberti have been found in the waters off northern Japan (Iwate Prefecture), whereas adults are found in the waters around the southern counterpart (Izu Islands), approximately 700 km from the northern waters. In the present study, we captured immature S. gilberti (106–248 mm standard length, SL) in the northern waters by set net at 8–80 m depth, whereas larger individuals (150–328 mm SL) were captured by trawling at 150–500 m depth. By contrast, only adult S. gilberti (422–590 mm SL) were captured in the southern waters. The genetic composition of the adult population of S. gilberti from the southern waters and of the juvenile population from the northern waters was compared using the nucleotide sequence of the mitochondrial DNA (mtDNA) cytochrome b gene. No significant differences in genetic parameters such as fixation index, neutrality test or mismatch distribution analysis were found between these geographically distinct populations of S. gilberti, showing that these populations are genetically homogeneous.     

Growth, stress tolerance and non-specific immune response of Japanese flounder Paralichthys olivaceus to probiotics in a closed recirculating system

ABSTRACT:   Effects of probiotics on growth, stress tolerance and non-specific immune response in Japanese flounder Paralichthys olivaceus were evaluated in a closed recirculating system. Survival and growth of flounder treated by supplying commercial probiotics either in the diet (the probiotic diet group), or into the rearing water (the water supply group), were higher compared to the untreated group (the control group). Water quality parameters, pH, NH₄-N, NO₂-N and PO₄-P showed lower concentration in the probiotic diet group compared with the control group and the supply group. Plasma lysozyme activity in the probiotic diet group and the water supply group was significantly higher ( P  < 0.05) than that in the control group. In heat shock stress tests, flounder in the probiotics-treated groups showed greater heat tolerance (measured by 50% lethal time, LT50) than the control group. Pathogen challenge tests with Vibrio anguillarum (2 × 10⁷ c.f.u./mL) resulted in significantly higher survival in the probiotics-treated groups than the control group. Results indicated that probiotics supplied in the rearing water and the diet of fish enhanced the stress tolerance and the non-specific immune system of Japanese flounder, providing them a higher resistance against stress conditions and pathogens.     

Dystrophin-deficient muscular dystrophy in a Toy Poodle with a single base pair insertion in exon 45 of the Duchenne muscular dystrophy gene

A 10-month-old, intact male Toy Poodle was referred for a postural abnormality. Blood biochemical tests revealed a marked increase in plasma creatine phosphokinase (CPK) concentration. The isoenzyme test showed that 99% of serum CPK consisted of CPK-MM. Histopathological evaluation of muscle biopsy samples confirmed scattered degeneration and necrosis of myofibers. Immunohistochemistry for dystrophin showed an absence of staining in muscle cells. Based on these findings, the dog was diagnosed with dystrophin-deficient muscular dystrophy. Whole genome sequencing using genomic DNA extracted from blood revealed a single base pair insertion in exon 45 of the Duchenne muscular dystrophy (DMD) gene. This is the first report on muscular dystrophy in Toy Poodles and identified a novel mutation in the DMD gene.     

Preparation of Anti-Rabies Virus N Protein IgYs by DNA Immunization of Hens Using Different Types of Adjuvants

DNA immunization has been used to study vaccination methods and for production of specific antibodies. The present study aimed to apply DNA immunization to prepare specific IgYs, which react against rabies virus N protein (RV-N) and can be used to research and diagnose rabies virus. The DNA sequence of RV-N was ligated into a pcDNA 3.1 plasmid for constructing pcDNA-N. Eight hens were divided into four groups. Group 1 comprised the control group (non-immunized). In Groups 2, 3, and 4, hens were injected intramuscularly with pcDNA-N (400 µg/hen). Eight injections were administered every other week. From the 4th week, an adjuvant was injected in addition to pcDNA-N. Freund''s complete adjuvant (FCA) and λ-carrageenan were administered to Groups 3 and 4, respectively. Eggs were collected daily, and the specific antibody activities of egg yolks were measured by ELISA. IgYs were purified from pooled egg yolks at 16–19 weeks post-administration in each group. The detection sensitivities of the RV-N were compared using purified IgY as the primary antibody for ELISA, dot blotting, and western blotting. Egg yolks from one of the two hens in Group 2 (pcDNA-N alone) and all hens in Groups 3 (pcDNA-N + FCA) and 4 (pcDNA-N + λCarra) had increased ELISA values. The combined use of λ-carrageen in DNA immunization resulted in an adjuvant effect comparable to that of FCA. Each purified specific IgY detected RV-N in the ELISA, western blotting, and dot blotting; however, the detection sensitivity differed. Higher detection sensitivity of the +λCarra IgY was observed by ELISA, whereas there was higher detection sensitivity of +FCA IgY in western blotting and dot blotting. In summary, anti-rabies virus N protein IgY was prepared through DNA immunization of hens using FCA or λ-carrageenan as adjuvants and can be used as a primary antibody to detect rabies viruses.     

Effects of momilactone on the protein expression in Arabidopsis germination

Although momilactone is known to play an important role in the allelopathy of rice (Oryza sativa L.), there is no information available about the mode of action of momilactone on the allelopathy of Arabidopsis. The present research describes the allelopathic activity of momilactone A and B on the germination of Arabidopsis thaliana and the effects of momilactone A and B on protein expression during germination. Momilactone A and B inhibited the germination of Arabidopsis at concentrations >30 and 10 μmol L^?1, respectively. The protein bands of 18, 19 and 21 kDa on sodium dodecyl sulphate‐polyacrylamide gel electrophoresis (SDS‐PAGE) were abundant in the momilactone A‐ and B‐treated Arabidopsis compared with the control. The protein bands of 18, 19 and 21 kDa were identified by the N‐terminal amino acid sequencing as cruciferin 2, cruciferina and cruciferin 3, respectively. Cruciferins and cruciferina are important storage proteins and play an important role as the initial source of nitrogen for seed germination. These results suggest that momilactone A and B may inhibit the germination of Arabidopsis seeds by inhibiting the degradation process of cruciferin and cruciferina.     

Stem-specific cadmium accumulation in cadmium-tolerant Polygonum thunbergii

Polygonum thunbergii was selected among wild plants as a cadmium (Cd)-tolerant plant because of the high accumulation of Cd in the stems. However, the concentration of Cd in the leaves was low, which suggested the existence of an inhibitory mechanism of Cd translocation from stems to leaves. Observation of Cd distribution in stem-petiole-lamina in P. thunbergii plants that had absorbed Cd, using SEM-EPMA, revealed that Cd accumulation occurred mainly in the nodes, junctions of petioles, and vessels in stems, with minimal translocation in petioles. A detection method for tissue distribution of Cd was developed. Resin-fixed tissue tips were stained with a fluorescent probe BTC-5N and then sites of Cd location were detected with a fluorescence microscope. It was found that Cd in the stem tissues of P. thunbergii was distributed at specific sites of the cell walls of vessels and surrounding cells. The results were consistent with the findings of the investigation of Cd distribution in stem cells obtained by the cell fractionation method, which showed that most of Cd was present in the cell wall and cytoplasm. These results suggest that P. thunbergii accumulates Cd in specific cell walls of the stems and inhibits Cd-translocation to the leaves.     

Preferential translocation of boron to young leaves in Arabidopsis thaliana Regulated by the BOR1 Gene

A mutant of Arabidopsis thaliana, bor1-1 (Noguchi et al. 1997: Plant Physiol., 115, 901–916) requires high levels of boron (B) for normal growth. We analyzed the B-deficiency symptoms of bor1-1 mutant plants in detail. A low B supply retarded the growth of the mutant plants more evidently in leaves than in roots. In particular, cell expansion and formation of air spaces were severely impaired by B deficiency in young rosette leaves. Such defects in growth were correlated with the reduced contents of B in leaves. These defects were not observed when a sufficient amount of B was supplied. Uptake experiments with ¹⁰B-enriched tracer B demonstrated that B taken up through roots was preferentially transported to young leaves compared to old leaves in the wild-type plants under a low B supply. Such a preferential transport to young leaves was not evident in the mutant plants. In conclusion, our data demonstrated that in A. thaliana plants B is preferentially transported to young organs under a low B supply and that this transport process is controlled at least in part by the BOR1 gene.     

Within- and between-site variations in leaf longevity in hinoki cypress (Chamaecyparis obtusa) plantations in southwestern Japan

We investigated mean leaf retention time in order to elucidate the factors affecting regional and local variations in stand-level leaf longevity in hinoki cypress (Chamaecyparis obtusa) plantations. Our study sites consisted of six stands at a low elevation (320–370 m a.s.l.) and 12 stands at a high elevation (850–970 m a.s.l.) in southwestern Japan. We also used published data on leaf longevity in stands at various elevations to clarify the regional-scale variations in leaf longevity and their relationships to environmental factors. At the regional scale, leaf longevity increased with increasing elevation and with decreasing air temperature, growing season length, and Kira’s warmth index across sites. Similar relationships were obtained for the variation in leaf mass. At the local scale, leaf longevity did not show a clear relationship with topographic position, soil water content, or soil C/N ratio. Contrary to our expectation, leaf longevity was negatively correlated with the leaf C/N ratio at both study sites, although the significance level was marginal. This indicates greater leaf longevity with better leaf nutrient status. Our results suggest that responses of leaf longevity in hinoki cypress stands to environmental factors would be more prominent at the regional scale than at the local scale, although large variation was detected at the local scale. Air temperature and growing season length appear to be the main drivers of the variation in leaf longevity at the regional scale, whereas the causal factors are unclear at the local scale.     

Oolong tea theasinensins attenuate cyclooxygenase-2 expression in lipopolysaccharide (LPS)-activated mouse macrophages: structure-activity relationship and molecular mechanisms

Oolong tea theasinensins are a group of tea polyphenols different from green tea catechins and black tea theaflavins. The present study reports the inhibitory effects of oolong tea theasinensins on the expression of cyclooxygenase-2 (COX-2) and underlying molecular mechanisms in lipopolysaccharide (LPS)-activated murine macrophage RAW264 cells. The structure-activity data revealed that the galloyl moiety of theasinensins played an important role in the inhibitory actions. Theasinensin A, a more potent inhibitor, caused a dose-dependent inhibition of mRNA, protein, and promoter activity of COX-2. An electrophoretic mobility shift assay (EMSA) revealed that theasinensin A reduced the complex of NF-κB- and AP-1-DNA in the promoter of COX-2. Signaling analysis demonstrated that theasinensin A attenuated IκB-α degradation, nuclear p65 accumulation, and c-Jun phosphorylation. Furthermore, theasinensin A suppressed the phosphorylation of MAPKs, IκB kinase α/β (IKKα/β), and TGF-β activated kinase (TAK1). These data demonstrated that the down-regulation of TAK1-mediated MAPKs and NF-κB signaling pathways might be involved in the inhibition of COX-2 expression by theasinensin A. These findings provide the first molecular basis for the anti-inflammatory properties of oolong tea theasinensins.     

Insulin-like growth factor I of Japanese eel, Anguilla japonica: cDNA cloning, tissue distribution, and expression after treatment with growth hormone and seawater acclimation

In an attempt to understand growth regulation in the Japanese eel, Anguilla japonica, we cloned insulin-like growth factor-I (IGF-I) cDNAs and examined their mRNA expression in several tissues. Two eel IGF-I (eIGF-I) cDNAs encoding preprohormones, eIGF-I-Ea1and eIGF-I-Ea2, were cloned from the liver by polymerase chain reaction (PCR). The preproIGF-Is were identical in signal peptide and mature IGF-I, but different in the E domain—eIGF-I-Ea2 mRNA was 36 bp longer than eIGF-I-Ea1 mRNA. Eel IGF-I was 83–94% identical with that of teleosts, 71% identical with that of dogfish, 87% identical with that of bullfrog and chicken, and 83% identical with that of humans. In both males and females the highest eIGF-I-Ea1 mRNA levels were observed in the liver, with detectable levels also found in the gills, heart, stomach, spleen, kidney, intestine, swim-bladder, muscle, and gonads. eIGF-I-Ea1 mRNA levels in the liver were higher in females than in males whereas in the intestine they were lower than in males. eIGF-I-Ea2 mRNA was detected in all the tissues examined and at similar levels in males and females. In this experiment higher eIGF-I-Ea1 mRNA levels were observed in the liver of larger glass eels than in those of smaller fish. eIGF-I-Ea2 mRNA levels were also higher in larger eels, although they were lower than IGF-I-Ea1 mRNA levels. Both eIGF-I mRNA levels in liver were positively correlated with the body size of the␣glass eels. Intraperitoneal injection of recombinant eel GH (reGH), 0.25 μg g⁻¹ body weight, into glass eels resulted in a significant increase in both eIGF-I mRNAs in the liver 1 day after injection compared with control fish, but no elevation was observed 2 days after injection. Incubation of liver slices with reGH at concentrations of 10, 100, and 1,000 ng mL⁻¹ for 24 h resulted in a significant concentration-dependent increase in the levels of both eIGF-I mRNAs. Higher levels of eIGF-I-Ea1 and Ea2 mRNA were observed in the gills ofseawater-reared eels than in those of freshwater-reared fish, but no differenceswere observed in the whole kidney. These results suggest that IGF-I is involved in the regulation of somatic growth and also in adaptation of the Japanese eel to seawater.