Characterization of Avian Paramyxoviruses Isolated from Feral Ducks in Northern Japan: The Presence of Three Distinct Viruses in Nature

Seventy-eight strains of avian paramyxoviruses (PMV) were isolated from cloacal and/or tracheal swabs taken from 1,342 feral ducks, comprised of spot-bill ducks, mallards, pintails, teals, falcated teals, wigeons and buffie-heads, in Wakuya-cho, Miyagi Prefecture, Japan, between 1976 and 1979. Five and a half percent of the ducks were positive for virus. Serological and structural characterization indicated that three different avian paramyxoviruses arc prevalent in the Japanese feral duck population. The first group of PMV was Newcastle disease virus (NDV), and in vivo pathogenecity tests in embryonated chicken eggs and 1-day-old chicks revealed that all the NDV strains isolated were avirulent. The second and most prevalent strain was closely related to PMV-4, duck/Hong Kong/D3/75 strain. The viruses of the third group were recovered only from pintails. They cross-reacted antigenically with PMV-3 when antisera to the PMV-3 reference strains, turkey/Wisconsin/68 and parakeet/Netherlands/449/75, were employed. However, no cross-reaction was observed when antiserum to pintail/ Wakuya/20/78, the prototype of this group, was used. The viruses of the third group also differed in viral polypeptide profile from the reference strains of PMV-3.     

Self-incompatibility (S) alleles of the rosaceae encode members of a distinct class of the T2/S ribonuclease superfamily

Stylar riboncleases (RNases) are associated with gametophytic self-incompatibility in two plant families, the Solanaceae and the Rosaceae. The self-incompatibility-associated RNases (S-RNases) of both the Solanaceae and the Rosaceae were recently reported to belong to the T2 RNase gene family, based on the presence of two well-conserved sequence motifs. Here, the cloning and characterization of S-RNase genes from two species of Rosaceae, apple (Malus × domestica) and Japanese pear (Pyrus serotina) is described and these sequences are compared with those of other T2-type RNases. The S-RNases of apple specifically accumulated in styles following maturation of the flower bud. Two cDNA clones for S-RNases from apple, and PCR clones encoding a further two apple S-RNases as well as two Japanese pear S-RNases were isolated and sequenced. The deduced amino acid sequences of the rosaceous S-RNases contained two conserved regions characteristic of the T2/S-type RNases. The sequences showed a high degree of diversity, with similarities ranging from 60.4% to 69.2%. Interestingly, some interspecific sequence similarities were higher than those within a species, possibly indicating that diversification of S-RNase alleles predated speciation in the Rosaceae. A phylogenetic tree of members of the T2/S-RNase superfamily in plants was obtained. The rosaceous S-RNases formed a new lineage in the tree that was distinct from those of the solanaceous S-RNases and the S-like RNases. The findings suggested that self-incompatibility mechanisms in Rosaceae and Solanaceae are similar but arose independently in the course of evolution.     

An Activation of Synaptosomal Na+, K+-ATPase by a Novel Dibenzoxazepine Derivative (BY-1949) in the Rat Brain: Its Functional Role in the Neurotransmitter Uptake Systems

In search of factors mitigating the final outcome of ischemic and epileptic brain damage, we tested a novel dibenzoxazepine derivative (BY-1949), as the compound has been shown to be effective under these two conditions. First, using rat brain, we assessed whether or not BY-1949 affects the Na+,K(+)-ATPase activity. Although in vitro applications of either BY-1949 or its three major metabolites did not cause any apparent effects, both acute and chronic oral administrations of the compound (10 mg/kg) invariably increased the Na+,K(+)-ATPase activity in the synaptosomal plasma membranes by increasing Vmax values. Second, it was shown by this study that the drug treatment caused marked increases in the uptake of both glutamic acid and gamma-aminobutyric acid into the synaptosomes. These results suggest that the activity against ischemic/epileptic brain damage by BY-1949 is explicable, at least partly, in terms of improvement of ionic derangements across the neural membranes via Na+,K(+)-ATPase activation.     

Selective Injection System into Hippocampus CA1 via Monitored Theta Oscillation

Methods of cell biology and electrophysiology using dissociated primary cultured neurons allow in vitro study of molecular functions; however, analysis of intact neuronal circuitry is often preferable. To investigate exogenous genes, viral vectors are most commonly injected using a pipette that is inserted from the top of the cortex. Although there are few reports that describe the success rate of injection in detail, it is sometimes difficult to locate the pipette tip accurately within the CA1 pyramidal cell layer because the pyramidal layer is only 0.1 mm thick. In the present study, we have developed a system to inject viral vectors accurately into the mouse hippocampal CA1 pyramidal cell layer using a stereotaxic injection system with simultaneous electrophysiological monitoring of theta oscillation. The pipette tip was positioned reliably based on integrated values of the theta oscillation in the hippocampal CA1 pyramidal cell layer. This approach allows accurate injection of solutions and provides an efficient method of gene transfer using viral vectors into the hippocampus, which can be a useful tool for studies involving the molecular mechanisms of neuronal functions.     

Genome sequence of the clover-nodulating Rhizobium leguminosarum bv. trifolii strain TA1

Rhizobium leguminosarum bv. trifolii strain TA1 is an aerobic, motile, Gram-negative, non-spore-forming rod that is an effective nitrogen fixing microsymbiont on the perennial clovers originating from Europe and the Mediterranean basin. TA1 however is ineffective with many annual and perennial clovers originating from Africa and America. Here we describe the features of R. leguminosarum bv. trifolii strain TA1, together with genome sequence information and annotation. The 8,618,824 bp high-quality-draft genome is arranged in a 6 scaffold of 32 contigs, contains 8,493 protein-coding genes and 83 RNA-only encoding genes, and is one of 20 rhizobial genomes sequenced as part of the DOE Joint Genome Institute 2010 Community Sequencing Program.     

The Role of NF-κB Signaling in the Maintenance of Pluripotency of Human Induced Pluripotent Stem Cells

NF-κB signaling plays an essential role in maintaining the undifferentiated state of embryonic stem (ES) cells. However, opposing roles of NF-κB have been reported in mouse and human ES cells, and the role of NF-κB in human induced pluripotent stem (iPS) cells has not yet been clarified. Here, we report the role of NF-κB signaling in maintaining the undifferentiated state of human iPS cells. Compared with differentiated cells, undifferentiated human iPS cells showed an augmentation of NF-κB activity. During differentiation induced by the removal of feeder cells and FGF2, we observed a reduction in NF-κB activity, the expression of the undifferentiation markers Oct3/4 and Nanog, and the up-regulation of the differentiated markers WT-1 and Pax-2. The specific knockdown of NF-κB signaling using p65 siRNA also reduced the expression of Oct3/4 and Nanog and up-regulated WT-1 and Pax-2 but did not change the ES-like colony formation. Our results show that the augmentation of NF-κB signaling maintains the undifferentiated state of human iPS and suggest the importance of this signaling pathway in maintenance of human iPS cells.     

Human immune responses elicited by an intranasal inactivated H5 influenza vaccine

Intranasally administered influenza vaccines could be more effective than injected vaccines, because intranasal vaccination can induce virus-specific immunoglobulin A (IgA) antibodies in the upper respiratory tract, which is the initial site of infection. In this study, immune responses elicited by an intranasal inactivated vaccine of influenza A(H5N1) virus were evaluated in healthy individuals naive for influenza A(H5N1) virus. Three doses of intranasal inactivated whole-virion H5 influenza vaccine induced strong neutralizing nasal IgA and serum IgG antibodies. In addition, a mucoadhesive excipient, carboxy vinyl polymer, had a notable impact on the induction of nasal IgA antibody responses but not on serum IgG antibody responses. The nasal hemagglutinin (HA)-specific IgA antibody responses clearly correlated with mucosal neutralizing antibody responses, indicating that measurement of nasal HA-specific IgA titers could be used as a surrogate for the mucosal antibody response. Furthermore, increased numbers of plasma cells and vaccine antigen-specific Th cells in the peripheral blood were observed after vaccination, suggesting that peripheral blood biomarkers may also be used to evaluate the intranasal vaccine-induced immune response. However, peripheral blood immune cell responses correlated with neutralizing antibody titers in serum samples but not in nasal wash samples. Thus, analysis of the peripheral blood immune response could be a surrogate for the systemic immune response to intranasal vaccination but not for the mucosal immune response. The current study suggests the clinical potential of intranasal inactivated vaccines against influenza A(H5N1) viruses and highlights the need to develop novel means to evaluate intranasal vaccine-induced mucosal immune responses.     

Efficacy of Dietary Lipid Control in Healing High-Fat and High-Cholesterol Diet-Induced Fibrotic Steatohepatitis in Rats

Nonalcoholic steatohepatitis is related to lifestyle, particularly to dietary habits. We developed diet-induced fibrotic steatohepatitis model stroke-prone spontaneously hypertensive 5/Dmcr (SHRSP5/Dmcr) rats showing steatosis, hepatic inflammation, and severe fibrosis induced by high-fat and -cholesterol (HFC) diet feeding. We aimed to clarify the efficacy of dietary intervention on the disease before and after the appearance of fibrosis. Male SHRSP5/Dmcr rats were divided into 9 groups; of these, 6 groups were fed control or HFC diet for several weeks and the remaining 3 groups represented the dietary intervention groups, which were fed the control diet after HFC diet feeding for 2 (before the appearance of fibrosis) or 8 (after the appearance of fibrosis) weeks. Dietary intervention before the appearance of fibrosis significantly improved the steatosis and reset the increased serum aspartate aminotransferase (AST), alanine aminotransferase (ALT), and serum total cholesterol (TC) levels. However, dietary intervention after the appearance of fibrosis was unable to reset the levels of hepatic TC, serum ALT, and fibrogenesis-related markers and had only a minor influence on hepatic fibrosis, although it reset the increased expression of transforming growth factor (TGF)-β1 and α-smooth muscle actin (SMA). It was noted that dietary intervention improved the increased AST levels; however, aggregated CD68-positive cells were still observed around the fibrosis area, which may be related to the findings of inflammatory cytokine mRNAs. Taken together, dietary intervention for fibrotic steatohepatitis improved steatosis, although it could not completely improve fibrosis.