Horizontal transmission and phylogenetic analysis of bovine leukemia virus in two districts of Miyazaki,Japan

Horizontal transmission is recognized as a major infection route for bovine leukemia virus (BLV), and cattle with high viral loads are considered to be a major infectious source in a herd. However, a correlation between viral loads and the risk of infection has been insufficient to use as a foundation for BLV control strategies. In this report, we examined the epidemiology of BLV infection and the infectious source in a local area. In 2013–2014, BLV infection was investigated in 1,823 cattle from 117 farms in two adjacent districts, Miyazaki, Japan. Seropositive samples for BLV were detected with 88 cattle and in 14 farms. Phylogenetic analysis revealed that 94% of the isolates clustered into genotype I and the remaining isolate into genotype III. Among genotype I, genetically distinct strains were spread at each farm, and cattle infected with less than 3 copies/100 cells did not transmit BLV to other cattle for more than thirty months. This is the first report of concrete data of viral load in relation to viral horizontal transmission under the field condition. The data facilitate farmers and veterinarians understanding the status of BLV infected cattle. This research contributes to BLV infection control and the development of effective BLV eradication programs.     

Expression of cell adhesion molecules in canine choroid plexus tumors

Choroid plexus tumor (CPT) is a primary intracranial neoplasm of the choroid plexus epithelium in the central nervous system. In the current World Health Organization classification, CPT is classified into two categories; choroid plexus papilloma (CPP) and carcinoma (CPC). In the present study, we investigated immunohistochemical expressions of N-cadherin, E-cadherin and β-catenin in 5 canine CPT cases (1 disseminated CPC, 2 CPCs and 2 CPPs). One CPP case was positive for N-cadherin and β-catenin, but negative for E-cadherin. The disseminated CPC case was positive for E-cadherin and β-catenin, but negative for N-cadherin. The other cases were positive for the three molecules examined. These results suggest that loss of the N-cadherin expression might associate with the spreading of CPC cells.     

Intraperitoneal administration of synthetic microRNA-214 elicits tumor suppression in an intraperitoneal dissemination mouse model of canine hemangiosarcoma

Canine hemangiosarcoma (HSA) has an extremely poor prognosis, making it necessary to develop new systemic treatment methods. MicroRNA-214 (miR-214) is one of many microRNAs (miRNA) that can induce apoptosis in HSA cell lines. Synthetic miR-214 (miR-214/5AE), which showed higher cytotoxicity and greater nuclease resistance than mature miR-214, has been developed for clinical application. In this study, we evaluated the effects of miR-214/5AE on stage 2 HSA in a mouse model. Mice intraperitoneally administered with miR-214/5AE (5AE group) had significantly fewer intraperitoneal dissemination tumor foci (median number: 72.5 vs. 237.5; p?<?0.05) and a lower median foci weight (0.26 g vs. 0.61 g; p?<?0.05). Mice in the 5AE group had increased expression of p53 and cleaved caspase-3, and a significantly lower proportion of Ki-67-positive cells, than those in the non-specific miR group. Notably, no significant side effects were observed. These results indicate that intraperitoneal administration of miR-214/5AE exhibits antitumor effects in an intraperitoneal dissemination mouse model of HSA by inducing apoptosis and suppressing cell proliferation. These results provide a basis for future studies on the antitumor effect of miR-214/5AE for HSA.

     

Identification of processed Japanese green tea based on polymorphisms generated by STS-RFLP analysis

In Japan, tea is generally sold blended, though 90% of the total production is clonal. Due to the increasingly strict consumer need and taste, however, more and more Japanese green teas are being sold under their particular cultivar name. Moreover, tea made from Yabukita, a much appreciated cultivar originally developed in Japan, has recently been produced and imported from a neighboring country. This paper describes a simple and inexpensive methodology capable of identifying fresh and processed Japanese green teas to discourage its fraudulent commercialization. The study was based on 46 main tea cultivars, and polymorphism detected through STS-RFLP analysis of the coding and noncoding DNA regions of three genes, namely phenylalanine ammonia-lyase, chalcone synthase, and dihydroflavonol 4-reductase, for which nucleotide information was available. All 46 tea cultivars analyzed could be easily distinguished using a combination of codominant DNA markers. Yabukita displayed a unique profile when PAL intron was digested with DdeI, thus allowing its rapid authentication at low cost.     

Glutelin is partially degraded in globulin-less mutants of rice (Oryza sativa L.)

Multigenic glutelins and monogenic globulin are major storage proteins accumulating in vacuole-derived protein body (PB-II) of rice (Oryza sativa L.) seeds. Because their interplay in PB-II formation was scarcely known, the effect of globulin-less mutation on glutelin accumulation was investigated. In globulin-less mutants, no phenotypic defect was found in seed and plant growth, while PB-II was deformed and apparent glutelin composition was changed, producing new glutelin α polypeptides X1–X5. 2D-PAGE of different combinations of globulin-less and glutelin subunit mutations suggested that the X1/X2, X3, and X4/X5 were derived from glutelin GluB1/GluB2/GluB4, GluA3, and GluA1/GluA2 subunits, respectively. Western blot with glutelin GluB4 subunit-specific and its variable region discriminable antibodies indicated at least in part the new spots X1/X2 are partially degraded products of GluB4 α polypeptides by the removal of 2–39 residues from C-terminus. Time course experiments with maturing seeds indicated the partial degradation of GluB4 occurred earlier (from 7 days after flowering) and higher than that of GluA1/GluA2. Considering the above results together with the fact that globulin accumulates at the periphery of PB-II and its absence produces deformed PB-II, globulin protects glutelins from proteinase digestion and thereby facilitates stable glutelin accumulation.     

Prediction of dry veneer stiffness using near infrared spectra from transverse section of green log

This study examined the feasibility of near infrared spectroscopy as a novel technique for log assessment on the basis of wood property. Near infrared (NIR) spectra were obtained from the transverse section of green log and multivariate regression analysis was carried out to predict the stiffness of veneer processed from the log. The stiffness of the veneer was dynamic modulus of elasticity measured using ultrasonic method. The calibrations of veneer stiffness had moderate relationships between measured and NIR-predicted values, with regression coefficients ranging from 0.84 to 0.88. The calibration equations were applied to the test set and it was found that predictions were also well fitted, with regression coefficients ranging from 0.67 to 0.89. The results indicate that the variation of wood stiffness within the logs could be assessed using the NIR spectra from the cross-section of logs. The spectra were obtained from green condition of the log and the stiffness of veneer was measured after kiln drying. Thus, the results imply that the wood stiffness in dry condition could be predicted using the spectra collected from green logs. If the models obtained in this study put into the imaging system, the two-dimensional map of the stiffness would be visualized on the cross-section of logs. The NIR spectroscopy coupled with imaging system could compensate the weak point of the traditional methods for log assessment.     

Blockade of bovine PD-1 increases T cell function and inhibits bovine leukemia virus expression in B cells in vitro

Programmed death-1 (PD-1) is a known immunoinhibitory receptor that contributes to immune evasion of various tumor cells and pathogens causing chronic infection, such as bovine leukemia virus (BLV) infection. First, in this study, to establish a method for the expression and functional analysis of bovine PD-1, hybridomas producing monoclonal antibodies (mAb) specific for bovine PD-1 were established. Treatment with these anti-PD-1 mAb enhanced interferon-gamma (IFN-γ) production of bovine peripheral blood mononuclear cells (PBMC). Next, to examine whether PD-1 blockade by anti-PD-1 mAb could upregulate the immune reaction during chronic infection, the expression and functional analysis of PD-1 in PBMC isolated from BLV-infected cattle with or without lymphoma were performed using anti-PD-1 mAb. The frequencies of both PD-1⁺ CD4⁺ T cells in blood and lymph node and PD-1⁺ CD8⁺ T cells in lymph node were higher in BLV-infected cattle with lymphoma than those without lymphoma or control uninfected cattle. PD-1 blockade enhanced IFN-γ production and proliferation and reduced BLV-gp51 expression and B-cell activation in PBMC from BLV-infected cattle in response to BLV-gp51 peptide mixture. These data show that anti-bovine PD-1 mAb could provide a new therapy to control BLV infection via upregulation of immune response.     

Linkage analysis of rice bacterial blight resistance gene xa20 in XM6, a mutant line from IR24

Bacterial blight caused by Xanthomonas oryzae pv. oryzae (Xoo) is an important disease constraining rice (Oryza sativa L.) production worldwide. The XM6 line was induced by N-methyl-N-nitrosourea from IR24, an Indica cultivar that is susceptible to Philippine and Japanese Xoo races. XM6 was confirmed to carry a recessive gene named xa20, resistant to six Philippine and five Japanese Xoo races. The chromosomal gene location was found using 10 plants with the shortest lesion length in an F₂ population consisting of 298 plants from a susceptible Japonica variety Koshihikari × XM6. Analysis using PCR-based DNA markers covering the whole rice genome indicated the gene as located on the distal region of the long arm of chromosome 3. The IKC3 line carries IR24 genetic background with Koshihikari fragment on chromosome 3 where a resistance gene was thought to be located. The F₂ population from IKC3 × XM6 clearly showed a bimodal distribution separating resistant and susceptible plants. Further linkage analysis conducted using this F₂ population revealed that xa20 is located within the 0.8 cM region flanked by DNA markers KIC3-33.88 (33.0 Mb) and KIC3-34.06 (33.2 Mb). This study yields important findings for resistance breeding and for the genetic mechanism of Xoo resistance.     

Genetic analysis of ion-beam induced extremely late heading mutants in rice

Two extremely late heading mutants were induced by ion beam irradiation in rice cultivar ‘Taichung 65’: KGM26 and KGM27. The F₂ populations from the cross between the two mutants and Taichung 65 showed clear 3 early: 1 late segregation, suggesting control of late heading by a recessive gene. The genes identified in KGM26 and KGM27 were respectively designated as FLT1 and FLT2. The two genes were mapped using the crosses between the two mutants and an Indica cultivar ‘Kasalath’. FLT1 was located on the distal end of the short arm of chromosome 8. FLT2 was located around the centromere of chromosome 9. FLT1 might share the same locus as EHD3 because their chromosomal location is overlapping. FLT2 is inferred to be a new gene because no gene with a comparable effect to that of this gene was mapped near the centromere of chromosome 9. In crosses with Kasalath, homozygotes of late heading mutant genes showed a large variation of days to heading, suggesting that other genes affected late heading mutant genes.