The Complete Nucleotide Sequence of a Japanese Isolate of White clover mosaic virus Strain RC

The complete nucleotide sequence was determined for genomic RNA of White clover mosaic virus (WClMV-RC) isolated from red clover (Trifolium pratense) in Japan, It is 5843 nucleotides in length, excluding the poly(A) tail at the 3' terminus. Similar to other potexviruses, it contains five open reading frames (ORFs 1 through 5), which putatively encode an RNA-dependent RNA polymerase (RdRp) (147 kDa), a triple gene block (TGB) (26 kDa/13 kDa/7 kDa), and a coat protein (CP) (22 kDa), respectively. The deduced amino acid sequence of the WClMV-RC CP was identical to that of WClMV-O, one of two New Zealand isolates, but only 85% identical to that of WClMV-M, the other New Zealand isolate, because of heterogeneity in the C-termini of CP amino acid sequences. The implication of this CP heterogeneity is discussed. Received 30 August 2001/ Accepted in revised form 11 January 2002     

Effect of postactivation treatment with latrunculin a on in vitro and in vivo development of cloned embryos derived from kidney fibroblasts of an aged clawn miniature boar

The objective of this study was to examine the effect of postactivation treatment with latrunculin A (LatA), an actin polymerization inhibitor, on in vitro and in vivo development of somatic cell nuclear transfer (SCNT) embryos derived from kidney fibroblasts of an aged Clawn miniature boar (12 years old). After electric activation, SCNT embryos were treated with 0, 0.5 or 1 μM LatA and cultured in vitro. The rate of blastocyst formation was significantly higher (P<0.05) in SCNT embryos treated with 0.5 μM LatA (38%) than those in control (14%). When cloned embryos treated with 0.5 μM LatA were transferred into the oviducts of two recipient miniature gilts to assess their development in vivo, both recipients became pregnant; one maintained pregnancy to term, and a live piglet (weighing 220 g) was delivered by Caesarean section. The results of this study indicated that the postactivation treatment with LatA was effective in improving in vitro developmental capacity of SCNT miniature pig embryos derived from kidney fibroblasts of an aged animal and that miniature pig cloned embryos treated with LatA had the ability to develop to term.     

A retrospective study of inflammatory colorectal polyps in miniature dachshunds

Medical records of dogs with colorectal polyps were retrospectively reviewed, and clinical presentation of inflammatory colorectal polyps in miniature dachshunds was evaluated. Of 33 dogs found to have colorectal polyps, miniature dachshunds were markedly over-represented with 16 dogs (48%), of which 12 (75%) were found to have inflammatory polyps. Multiple polyps localized between the rectum and the descending colon was the most common finding in miniature dachshunds with inflammatory polyps. Twenty dogs (80%) out of 25 miniature dachshunds with inflammatory colorectal polyps responded to immunosuppressive therapy using prednisolone and cyclosporine. The results of this study indicate that miniature dachshunds are predisposed to develop inflammatory colorectal multiple polyps, for which immunosuppressive therapy may be a treatment option.     

Patch isolation only matters for specialist butterflies but patch area affects both specialist and generalist species

Over the past four decades, many researchers have applied the theory of island biogeography (IBT) to predict and understand species loss and distribution in fragmented landscapes. Recent studies found that specialist species were more affected by fragment size and isolation than generalists. However, the mechanisms underlying different effects of area and isolation among specialists and generalists are unknown. We tested the predictions of IBT on butterfly assemblages in Tokyo, Japan, and hypothesized that the effects of fragment size and isolation would be stronger for specialists than for generalists. We classified butterfly species into specialists and generalists for each of two dimensions (food range and voltinism) and according to tolerance to the matrix. We recorded 26 feeding specialists and 27 generalists, 24 seasonal specialists and 29 generalists, 32 low matrix-tolerant species and 21 high matrix-tolerant species in 20 forest fragments. We used generalized linear models to relate the number of species in a fragment to fragment size and isolation (distance to the mainland). The averaged models based on AICc showed that fragment size had positive and significant effects on both specialist and generalist and high matrix-tolerant butterfly species richness. However, the negative effects of isolation on species richness were only found in specialist and low matrix-tolerant species. Our results demonstrate that patch isolation only affects specialist species. This suggests that when applying IBT to terrestrial fragmented landscapes, researchers should be careful not to overlook patch area and isolation effects on specialists.     

Mastitis caused by Mycobacterium kansasii infection in a dog

A 2‐year, 7‐month‐old female Chihuahua was admitted for a mammary mass measuring one cm in diameter. The dog had a history of demodicosis for 4 months and showed signs of pseudopregnancy at the time of the visit. Cytologic examination of an aspirate of the mass revealed a large number of macrophages containing nonstaining bacterial rods, which were acid‐fast in a Ziehl–Neelsen stain, suggesting mycobacterial infection. Histologic examination of the mass revealed a pyogranulomatous mastitis characterized by an infiltration with macrophages containing acid‐fast bacteria. Mycobacterium kansasii was subsequently cultured and identified by PCR. Surgical excision of the mass resulted in the growth of other dermal masses, but antimycobacterial treatment with rifampin and clarithromycin resolved these masses within 1 month. Three months after discontinuation of the treatment, similar organisms were found in aspirates of the enlarged bilateral inguinal lymph nodes by cytologic examination. Despite antimycobacterial treatment for another 4 months, there was no improvement and demodicosis also recurred. The dog eventually died of lymphoma 5 months after the relapse of mycobacterial infection. Although M kansasii is considered an important pathogen for pulmonary and cutaneous disease in people, there is only one report in a dog with an infection in a pleural effusion. As both adult‐onset demodicosis in dogs as well as mycobacterial infection in people have been associated with T‐lymphocyte deficiency, the M kansasii infection in this dog may have been associated with a condition of immune compromise.     

Expression of CD4 T cell cytokine genes in the colorectal mucosa of inflammatory colorectal polyps in miniature dachshunds

Inflammatory colorectal polyps (ICRPs) in miniature dachshunds are recently recognized as a major cause of large bowel diarrhea in this dog breed in Japan. ICRPs are characterized by the formation of multiple small polyps and a space-occupying large polyp in the colorectal area, and are thought to be a novel form of inflammatory bowel disease (IBD). In humans, specific cytokine patterns attributed to T helper (Th)1, Th17 and regulatory T cells have important roles in the pathogenesis of IBD. Thus, the aim of the present study was to assess the gene expression of cytokines of T cell subsets in the colorectal mucosa from dogs with ICRPs. Colorectal mucosal specimens from 10 dogs with ICRPs and 14 control dogs were used in this study. Interferon (IFN)-γ, interleukin (IL)-4, IL-17A and IL-10 mRNA expression was assessed using quantitative real-time PCR. IL-17A mRNA expression was significantly increased in large polyps compared to small polyps and controls. IFN-γ and IL-10 mRNA expression in large polyps were significantly higher than in controls. There was no significant difference in IL-4 mRNA expression among the three groups. IL-17A is thought to play important roles in the pathogenesis of ICRPs. IL-10 up-regulation could oppose the proinflammatory function of IL-17A.     

The Effects of Frequent Electroejaculation on the Semen Characteristics of a Captive Siberian Tiger (Panthera tigris altaica)

Artificial insemination (AI) can help to avoid inbreeding and genetic degeneration for sustaining genetically healthy populations of endangered species in captivity. Collection of a sufficient quantity of viable sperm is an essential first step in the AI process. In the present study, we examined the effects of frequent electroejaculation on semen characteristics in a Siberian tiger. We collected semen in all 17 trials during 6 breeding seasons (6 years). The mean number of sperm and the percentage of motile sperm were 294.3 ± 250.2×10⁶/ejaculate and 82.4 ± 11.4%, respectively. The number of motile sperm tended to increase during frequent electroejaculation in the same breeding season. Semen collection by electroejaculation can be performed effectively up to the fourth sequential ejaculate, which contained the most sperm in the study. In conclusion, frequent collection of sperm by electroejaculation from tigers may be effective for collection of a large number of motile sperm.     

Cytotoxicity of benzyl isothiocyanate in normal renal proximal tubular cells and its modulation by glutathione

In the present study, we examined the toxicity of benzyl ITC (BITC) and its urinary mercapturic acid metabolite (BITC-NAC), using a normal renal proximal tubular cell line, pig LLC-PK1. BITC increased cell death with an IC(50) value of about 7 μM, whereas the cytotoxic effect of BITC-NAC was five times weaker than that of BITC. We observed a significant necrosis of the compounds on LLC-PK1 cells with oxidative stress. In the presence of 5 mM glutathione (GSH), comparable to physiological levels, the cytotoxicity of BITC-NAC as well as BITC was significantly reduced. Furthermore, the increase in intracellular GSH levels by pretreatment with NAC before the BITC treatment resulted in inhibition of the BITC-induced necrotic events as well as intracellular oxidative stress. These results suggest that GSH is a determinant of cellular resistance against the BITC-mediated and oxidative stress-dependent cytotoxicity in renal proximal tubular cells.     

Localization of putative pituitary stem/progenitor cells in female dairy cattle

Research on sex-determining region Y-box 2 (SOX2)-positive pituitary stem/progenitor cells, as a source of hormone-producing cells, is progressing rapidly in rodents. However, the stem/progenitor cells supplying hormone-producing cells that are essential for growth, reproduction, and lactation in bovines have not yet been identified. In this study, we characterized SOX2-positive cells in the pituitary gland of dairy cattle (Holstein heifers) after sexual maturity. Immunofluorescence analysis revealed that the localization pattern of SOX2-positive cells in the dairy cattle pituitary gland was similar to that observed in the rodent pituitary gland; the marginal cell layer (MCL), dense cell clusters, and single cells scattered in the parenchyma of the anterior lobe. Furthermore, most of the SOX2-positive cells were positive for the pituitary stem/progenitor cell niche markers E-cadherin and cytokeratin 8+18, which have been reported in rodents. In addition, in the MCL of the anterior lobe, there was a subpopulation of SOX2-positive cells positive for paired-related homeobox 1 and 2, whereas negative for S100β. Moreover, in the parenchyma of the anterior lobe, co-localization of SOX2 and pituitary hormones was infrequent. In summary, this study reveals the localization of putative pituitary stem/progenitor cells positive for SOX2 in dairy cattle. These results provide valuable information to support further investigation of cell supply in the dairy cattle pituitary gland.