Reproductive performance and expression of imprinted genes in somatic cell cloned boars

To assess the performance of boars derived by somatic cell cloning, we analyzed various aspects of their reproductive characteristics and the expression of two imprinted genes. Cloned boars (cloned Duroc × Jinhua) were analyzed for birth weight, growth rate, age at first ejaculation, semen characteristics and fertility, in comparison with naturally bred control boars of the same strain. The expression of imprinted genes was analyzed using the microsatellite marker SWC9 for the paternally expressed gene insulin‐like growth factor ‐2 (IGF2) and with single nucleotide polymorphisms (SNPs) for the gene maternally expressed 3 (MEG3). The cloned boars had high production of semen and were nearly equal in level of fertility to conventional pigs; they showed similar characteristics as naturally bred boars of the same strains. The expression of IGF2 was partially disturbed, but this disturbed expression was not linked to a change in developmental fate or reproductive performance. These results indicate that use of cloned boars could be highly effective for proliferation of pigs with desirable characteristics, preservation of genetic resources and risk reduction against epidemic diseases, such as foot‐and‐mouth disease, through storage of somatic cells as a precautionary measure for use in regenerating pig populations after a future pandemic.     

Effects of different application timings of methane fermentation digested liquid to paddy plots on soil nitrogen and rice yield

Treatment of animal and food wastes using a methane fermentation technique is drawing considerable public attention as a suitable option for the utilization of biomass resources. The application of a fermentation byproduct (methane fermentation digested liquid) as an agricultural fertilizer has been investigated. Determining the appropriate timing required for applying digested liquid on a rice (Oryza sativa L.) paddy plot is important. The concentrations of soil nitrogen (N) components and rice yield should be considered because digested liquid contains both inorganic and organic N. This study compares the N transformation and the rice yield and growth at different application times over a period of 3 y. The effects of the timing of basal application on soil N were different and the timing that maximized the rice yield was different in each year. Days before ponding (DBP) affected soil N before mid-summer drainage, and rice growth rates at the panicle formation stage and the ear emergence stage. The effects of DBP disappeared before harvest. The results indicated that sufficient potentially mineralizable N existed regardless of DBP, and the effect of DBP lessened after the mid-summer drainage, which coincides with the period when N uptake is most active.     

High-pressure balloon valvuloplasty for severe pulmonary valve stenosis: a prospective observational pilot study in 25 dogs

Objectives

We aimed to evaluate safety and efficacy of high-pressure balloon valvuloplasty (HPBVP) for treatment of canine severe pulmonary valve stenosis (PS). A secondary aim was to provide pre-procedure predictors of success.

Characterization of Bovine herpesvirus type 4 isolated from cattle with mastitis and subclinical infection by the virus among cattle

An outbreak of contagious mastitis occurred among cattle on a farm, and bovine herpesviruses were isolated from the affected mammary tissues, scabs and abscess discharge of the cattle. A bovine herpesvirus type 4 (BoHV-4)-specific fragment was amplified from the isolates by polymerase chain reaction (PCR). Restriction endonuclease analyses demonstrated that the isolates were related to Movar-like European type BoHV-4. To determine the ratio of BoHV-4 subclinical infection in the cattle, a genomic survey was performed by PCR for cattle that were moved to the animal hygiene service station in Ibaraki prefecture. The BoHV-4 genome was occasionally detected in peripheral blood leukocytes, lymph nodes and nervous tissues. The rate of BoHV-4 subclinical infection was relatively high in the cattle.     

Evaluation of the immunogenicity of Mycobacterium avium subsp. paratuberculosis (MAP) stress-associated recombinant proteins

The aim of this study was to assess the humoral immune responses in MAP-infected and uninfected sheep against 27 MAP stress-associated recombinant proteins that were regulated in in vitro models of physiological stress. These include evaluation of 5 proteins, which were previously reported by Gumber et al. (2009b), using serum samples from sheep with a wide range of disease stages. For purification of recombinant his-tagged proteins expressed as an insoluble protein, on-column refolding purification was applied as well as one-step denaturing purification. All purifications together resulted in a total of 48 recombinant antigen preparations. In antibody ELISA tests, 23 of these, representing 18 MAP proteins, showed significant differences in responses between infected and uninfected sheep. Recombinant antigen preparations MAP2281c, MAP3555 (refolded form), and MAP0711c (refolded form) when incorporated in an ELISA, had similar sensitivity to a commercial antibody ELISA test at the cutpoint of 90% specificity, and showed relatively high values in receiver operating characteristic (ROC) curve analysis. However, as some of the sera from uninfected sheep also reacted to recombinant antigens, further development of the assays is necessary prior to practical application. Compared to the commercial antibody ELISA, MAP0593c, MAP2281c, MAP2411, MAP3555, and MAP3200 detected more infected sheep with a lower grade of lesion, suggesting that these proteins identified in the in vitro models of stress were also expressed in vivo in MAP-infected sheep at an early stage of infection. This is consistent with the hypothesis of latency or dormancy in subclinical mycobacterial infection.     

Valdensia leaf blight of highbush blueberry caused by Valdensinia heterodoxa, a new fungal disease in Japan

In July 2003, a new disease occurred on leaves of highbush blueberry (Vaccinium corymbossum L.) in Iwate, Japan. Leaves initially had brownish spots, which subsequently developed into large lesions with concentric rings, resulting in premature defoliation. Teardrop-shaped conidia infecting leaves were visible by the naked eye as small protuberances in the center of lesions. Star-shaped conidia were sporadically produced on large lesions. The causal fungus was identified as Valdensinia heterodoxa, based on cultural, morphological and genetic studies. Inoculation tests showed that the fungus reproduced lesions on detached young leaves of highbush blueberry. A field survey in 2009 indicated that symptoms initially appeared on the lower leaves of basal shoots in late May, and the disease rapidly progressed on leaves of basal shoots, eventually spreading to lateral shoots from late June to late July. Removal of all basal shoots in late June significantly reduced disease incidence on lateral shoots in late July.     

Biological activity of recombinant bovine interferon tau using an Autographa californica nuclear polyhedrosis virus expression system

Bovine interferon (bIFN) tau, which plays a key role in maternal-fetal recognition of pregnancy, was expressed by an Autographa californica nuclear polyhedrosis virus expression system. cDNA coding bIFNtau was derived from cultured trophoblast cells. The recombinant (r) bIFNtau had high antiviral activity (1 x 10 (8) IU/mg) and the molecular weight of rbIFNtau was estimated to be 23 kDa by Western blotting analysis. We investigated the biological effect of rbIFNtau on prostaglandin (PG) F(2alpha) synthesis in cultured bovine endometrial epithelial cells in the presence or absence of oxytocin (OT, 100 nM). rbIFNtau suppressed basal and OT-induced PGF(2alpha) production in a dose-dependent manner (1-1,000 ng/ml). These results showed that biologically active rbIFNtau was produced in the baculovirus expression system, and that rbIFNtau had the ability to suppress the synthesis of PGF(2alpha) from bovine endometrial epithelial cells.     

High level expression and purification of bioactive bovine interleukin-18 using a baculovirus system

Bioactive recombinant bovine interleukin-18 (rboIL-18) was expressed using a baculovirus system. Normally, IL-18 is translated as a precursor form of a 24kDa polypeptide and processed by IL-1beta converting enzyme (ICE) to a mature bioactive form of 18kDa protein. Hence, to express active form IL-18, we constructed two recombinant baculoviruses containing boIL-18 and human ICE (hICE) genes, respectively, and superinfected these viruses into insect cells. Superinfection of both recombinant viruses into the cells resulted in the expression of a 24kDa precursor form and an 18kDa mature form detectable in the supernatant by immunoblotting using anti-porcine IL-18 antibody. Culture supernatant from the superinfected cells showed a synergistic effect with recombinant boIL-12 for production of interferon-gamma (IFN-gamma) in bovine peripheral mononuclear cells. By addition of histidine hexamer at the C-terminal of boIL-18, the mature IL-18 was purified. Bioactivity remained after purification.