An isolate of Prototheca wickerhamii from systemic canine protothecosis

The isolate from a canine disseminated protothecosis was identified to be Prototheca wickerhamii by its morphological and biochemical characteristics. The isolate was grouped into a cluster identical to the type strain of P. wickerhamii, ATCC 16529(T) in phylogenetic trees based on the small subunit (SSU) and the 5' end of the large subunit (LSU) ribosomal DNA (rDNA); the cluster was close to that including the other Prototheca species. However, the strains of P. wickerhamii, SAG 263-11 and Pore 1283 belonged to a cluster different from the other isolates of Prototheca species and closely related to those of Auxenochorella species. Therefore, P. wickerhamii could be divided into two distinct genetic groups, one group close to the other Prototheca spp. including a standard strain of P. wickerhamii, and another group consisting of isolates previously reported to be close to the Auxenochorella species.     

Acute lipoprivation suppresses pulsatile luteinizing hormone secretion without affecting food intake in female rats

The present study examined the effect of acute lipoprivation on pulsatile luteinizing hormone (LH) secretion in both normal-fat diet, ad libitum-fed and fasted female rats. To produce an acute lipoprivic condition, mercaptoacetate (MA), an inhibitor of fatty acid oxidation, was administered intraperitoneally to ad libitum-fed or 24-h fasted ovariectomized (OVX) rats with or without an estradiol (E2) implant, that produces a negative feedback effect on LH pulses. The steroid treatment was performed to determine the effect of estrogen on lipoprivic changes in LH release, because estrogen enhances fasting- or glucoprivation-induced suppression of LH pulses. Pulsatile LH secretion was suppressed by MA administration in a dose-dependent manner in the ad libitum-fed OVX and OVX+E2 rats. LH pulses were more severely suppressed in the 24-h-fasted OVX and OVX+E2 rats compared to the ad libitum-fed rats. Estrogen slightly enhanced lipoprivic suppression but the effect was not significant. In the present study, increased plasma glucose and free-fatty acid concentrations may indicate a blockade of fatty acid metabolism by the MA treatment, but food intake was not affected by any of the MA doses. Acute vagotomy did not block lipoprivic suppression of LH pulses. Thus, the present study indicates that lipid metabolism is important for maintenance of normal reproductive function even in rats fed a normal-fat diet and lipoprivation may be more critical in fasted animals that probably rely more heavily on fatty acid oxidation to maintain appropriate metabolic fuel levels. In addition, failure of blockade of lipoprivic LH inhibition by vagotomy suggests that lipoprivic information resulting in LH suppression is not transmitted to the brain via the vagus nerve.     

Rumen fermentation,milk production and conjugated linoleic acid in the milk of cows fed high fiber diets added with dried distillers grains with solubles

The effects of corn dried distillers grains with solubles (DDGS) feeding on rumen fermentation and milk production in cows were evaluated using diets high in neutral detergent fiber (NDF, 45.9–46.6%). The control diet (Control) consisted mainly of hay, corn silage and concentrates. In the experimental diets, the concentrates were replaced with DDGS as 10% dry matter (DM) (10%DDGS) and 20% DM (20%DDGS). Eight cows were used for each 14‐day treatment period. Effect of DDGS feeding on DM intake was not significant. Ruminal volatile fatty acids and ammonia‐N at 5 h after feeding of 20%DDGS were decreased compared to Control, whereas protozoal count at 2 h after feeding of 20%DDGS was higher than that of 10%DDGS. Milk yield of cows fed DDGS diets was greater than that of Control, although percentages of milk protein and solids‐not‐fat were decreased by DDGS diets. The proportions of C10:0, C12:0, C14:0 and C16:0 in the milk fat decreased, and those of C18:0, C18:1, C18:2 and cis‐9, trans‐11 conjugated linoleic acid (CLA) increased markedly with elevated DDGS. Increase in trans‐11 C18:1 was observed in the rumen fluid at 5 h after feeding. These findings suggest that DDGS feeding enhanced milk yield, as well as CLA synthesis under a high dietary NDF condition.     

Role of noradrenergic receptors in the bed nucleus of the stria terminalis in regulating pulsatile luteinizing hormone secretion in female rats

The bed nucleus of the stria terminalis (BNST) is one of the brain areas densely innervated by noradrenergic neurons originating in the brain stem. The present study aims to determine the role of noradrenergic receptors in the BNST in regulating pulsatile luteinizing hormone (LH) secretion in female rats. Ovariectomized (OVX) or estrogen-primed OVX (OVX+E2) rats received three 1-h-interval injections of 0.05 micromol of noradrenaline (NA), phenylephrine (alpha1-adrenergic receptor agonist), clonidine (alpha2-agonist), or isoproterenol (beta-agonist) into the BNST. Injection of NA or alpha1-adrenergic agonist into the BNST strongly suppressed pulsatile LH secretion in OVX+E2 rats with a significant (P < 0.05) decrease in the mean LH level for 3 h and LH pulse frequency, but alpha2-and beta-agonists did not affect any of the LH pulse parameters. In OVX animals, alpha1- and alpha2-adrenergic agonists caused a significant change in LH pulse frequency and amplitude, respectively, though the effect was not as apparent as the NA- or alpha1-agonist-induced changes in OVX+E2 animals. These results indicate that NA inputs to the BNST suppress pulsatile LH secretion via alpha1-adrenergic receptors and that estrogen enhances this suppression.     

Efficient generation of canine bone marrow-derived dendritic cells

Because of their unsurpassed potency in presenting antigens to naive T cells, dendritic cells are considered to be an important candidate in the development of immunotherapeutic strategies. Despite the high potential of dendritic cell-based immunotherapy, as a so-called dendritic cell vaccination, few clinical approaches using dendritic cell vaccination have been performed in the dog because of very limited information regarding the generation of canine dendritic cells and their functional properties. We therefore established a protocol for the efficient generation of dendritic cells from canine bone marrow cells using recombinant feline granulocyte-macrophage colony-stimulating factor and canine interleukin-4. Dendritic cells were generated efficiently: a yield of 1-9 x 10(6) cells per approximately 0.5 ml of canine bone marrow aspiration was achieved. These dendritic cells showed features shared with mouse and human dendritic cells: dendrite morphology, expression of surface markers MHC class II and CD11c, and up-regulation of molecules related to antigen presentation (MHC class II, B7-1, and B7-2) by activation with lipopolysaccharide. Moreover, the dendritic cells demonstrated phagocytic activity, processing activity of pinocytosed proteins, and activation of allogeneic T cells far more potent than that by macrophages. Our findings suggest that the bone marrow-derived dendritic cells are functional for the capturing and processing of antigens and the initiation of T cell responses.     

A case of adenocarcinoma of the endometrium extending into the leiomyoma of the uterus in a rabbit

In a pet rabbit, 2 tumor masses one on each horn were macroscopically seen in the wall of the uterus. On light microscopic examination, the right horn mass consisted of an admixture of neoplastic epithelial and mesenchymal element. The epithelial element was composed of neoplastic epithelial cells with numerous mitotic figures and formed varied sizes of acini, glandular, and solid structures. The tumor was diagnosed as an adenocarcinoma of the endometrium. The mesenchymal element was composed of well-differentiated smooth muscle cells and was diagnosed as a leiomyoma. While adenocarcinoma cells formed a protrusive mass in the uterine lumen, they also showed an extension into the leiomyoma of the myometrium. By immunohistochemistry, adenocarcinoma stained positive for cytokeratin (MNF116) and leiomyoma stained positive for smooth muscle actin, showing a substantial difference in the cytological nature of these tumor cells. The results may give a further evidence supporting the narrative of the tumor development that an adenocarcinoma of the endometrium extended into leiomyoma of the uterus. To the author's knowledge, this is the first report describing this type of combination of two independent tumors in a pet rabbit.     

Changes of multi-drug resistance pattern in Salmonella enterica subspecies enterica serovar typhimurium isolates from food-producing animals in Japan

A total of 153 isolates of Salmonella Typhimurium derived from food-producing animals in Japan between 2002 and 2005 were investigated for antimicrobial resistance and phage types related to definitive phage type 104 (DT104). The predominant resistance type was resistance to ampicillin, dihydrostreptomycin, kanamycin, and oxytetracycline in bovine (45.2%, 48/104) and resistance to dihydrostreptomycin and oxytetracycline in porcine isolates (58.7%, 27/48). DT104-related phage type was found in 32 of 104 bovine isolates, two of 48 porcine isolates, and one of eight isolates from poultry, showing that the proportion of the phage type in S. Typhimurium isolates from cattle and pigs significantly (P<0.01) decreased from 71.9% and 31.4% in 1999-2001 to 30.8% and 4.1% in 2002-2005, respectively.     

Expression of O6-methylguanine-DNA methyltransferase causes lomustine resistance in canine lymphoma cells

The DNA repair protein O⁶-methylguanine-DNA methyltransferase (MGMT) causes resistance to nitrosoureas in various human cancers. In this study, we analyzed the correlation between canine lymphomas and MGMT in vitro. Two of five canine lymphoma cell lines required higher concentrations of lomustine to inhibit cell growth by 50%, but their sensitivity to the drug increased when they were cultured with an MGMT inhibitor. Fluorometric oligonucleotide assay and real-time polymerase chain reaction of these cell lines revealed MGMT activity and high MGMT mRNA expression, respectively. We analyzed the methylation status of the CpG islands of the canine MGMT gene by the bisulfite-sequencing method. Unlike human cells, the canine lymphoma cell lines did not show significant correlation between methylation status and MGMT suppression levels. Our results suggest that in canine lymphoma MGMT activity may influence sensitivity to nitrosoureas; thus, inhibition of MGMT activity would benefit nitrosourea-resistant patients. Additional studies are necessary to elucidate the mechanism of regulation of MGMT expression.     

Antifungal activities of the combination of tacrolimus and itraconazole against Trichophyton mentagrophytes

Tacrolimus was reported to have no antifungal activity against dermatophytes, but it might show a synergistic fungicidal activity with azoles against dermatophytes as in the case of pathogenic yeasts. Therefore, we examined the antifungal activity of tacrolimus combined with itraconazole (ITZ) against five strains of Trichophyton mentagrophytes by measuring cell growth and performing MIC (minimum inhibitory concentration) testing. The mean colony diameter of T. mentagrophytes on 1/10 diluted Sabouraud dextrose agar (dSDA) with tacrolimus combined with ITZ was significantly shorter than that on dSDA with tacrolimus or ITZ. MICs of tacrolimus, ITZ, and tacrolimus combined with ITZ against T. mentagrophytes were determined, respectively. The results revealed a synergistic activity of tacrolimus in combination with ITZ against T. mentagrophytes.