Hemorrhagic necrotizing splenitis in a slaughter pig infected with Arcanobacterium species

A 6-month-old barrow presented with lethargy, inappetence and dysstasia. At necropsy, multiple coalescing hemorrhagic foci were detected in the margins of the spleen. Gram-positive bacilli were isolated from the spleen, kidney, muscle and liver. Comparative 16S rDNA gene sequencing analysis of the isolates (TO16177) revealed that they would be the same species of unpublished Arcanobacterium species strain HJ57-14E (accession no. gi 18873551) (99.7% similarity based on a comparison of 675 bp). Histologic examination of the splenic tissue sections revealed extensive necrosis and inflammation, and gram-positive bacilli were discernible. Multifocal necrosis was also detected in the liver. Immunohistochemically, the isolates were cross-reacted with polyclonal antibodies against Arcanobacterium pyogenes and Actinomyces naeslundii, and the reaction was strong for the latter. Similar reactions were found in the suppurative lesions of the tonsil, and occasionally in the spleen and lymph nodes. The present results indicate that the unpublished Arcanobacterium species induced multiple organ failure accompanied by acute hemorrhagic necrotizing splenitis in this growing-finishing pig.     

Antihyaluronidase action of ellagic acid effectively prevents polyspermy as a result of suppression of the acrosome reaction induced by sperm-zona interaction during in vitro fertilization of porcine oocytes

The present study was conducted to examine the effects of three tannin relatives (tannic acid, TA; gallic acid, GA; and ellagic acid, EA) on antihyaluronidase and reactive oxygen species (ROS) scavenging activity, in vitro fertilization (IVF) parameters, and the acrosome reaction (AR) induced by sperm-zona interaction. Among the three tannin relatives, TA and EA showed the strongest potency for blocking the hyaluronidase activity of boar sperm, with concentration-dependent inhibition over the range of 2-10 microg/ml. In contrast, ROSs were effectively scavenged by TA and GA, but not EA. When cumulus-free oocytes were inseminated in IVF medium containing 5 microg/ml of the tannin relatives, polyspermy was significantly reduced by TA and EA (32 and 29%, respectively) compared with oocytes treated with or without GA (51 and 69%, respectively) under conditions that maintained a high sperm penetration rate (P<0.05). Interestingly, induction of the AR by treatment of preincubated sperm with progesterone was blocked by TA and GA as a result of their higher levels of ROS scavenging activity, while EA, which possessed weak ROS scavenging activity, did not disturb induction of the AR with progesterone. However, the incidence of AR induced by sperm-zona interaction was significantly decreased by the strong antihyaluronidase actions of TA and EA compared with that in the absence of these compounds. Treatment with the compounds caused neither a protective proteolytic modification of the zona pellucida matrix before fertilization nor a reduction in acrosomal proteolytic activity or the number of zona-bound sperm. These findings suggest that the antihyaluronidase action of EA effectively prevents polyspermy by suppression of AR functionality induced by sperm-zona interaction and that hyaluronidase intervention is therefore required during porcine IVF.     

Androgen induces production of male effect pheromone in female goats

Previously we showed that the primer pheromone responsible for the "male effect" was produced in specific skin regions of castrated male goats by androgen treatments. In the present study, we examined whether androgen can also induce production of the male effect pheromone in female goats. Capsules containing dihydrotestosterone (DHT) or testosterone (T) were subcutaneously implanted into six ovariectomized (OVX) goats for 28 days. Small skin samples were collected from the head and rump regions, and the pheromone activity of their ether extracts was examined using a bioassay that monitors the electrophysiological manifestation of the hypothalamic gonadotropin-releasing hormone pulse generator as multiple-unit activity. Behaviors of OVX goats towards ovary-intact estrous goats were also examined before and at the end of DHT or T treatment. Before androgen treatment, neither the head nor rump skin samples in OVX goats showed pheromone activity. DHT treatment induced pheromone activity in the head skin sample of six OVX goats and in the rump skin sample of two OVX goats. Similar results were obtained by T treatment. In addition, OVX goats treated with T showed masculine-type sexual behaviors such as courtship and mounting behaviors towards the estrous goats. These results demonstrate that androgen is capable of inducing primer pheromone activity in the female and suggest that the synthesis pathway of the male effect pheromone exists in both sexes in the goat.     

Changes in expression and localization of GPRC5B and RARalpha in the placenta and yolk sac during middle to late gestation in mice

The mRNA expression of GPRC5B, an orphan G protein-coupled receptor, is induced by retinoic acid (RA). Because RA plays critical roles in embryonic development, reproductive functions, metabolism and homeostasis, GPRC5B is also considered crucial in these physiological events. We investigated the changes in expression of GPRC5B and RA receptor (RAR) alpha mRNAs and immunohistochemical localization of their proteins in the murine placenta and yolk sac at 13.5, 15.5 and 17.5 days post coitus. Stable levels of GPRC5B and RARalpha mRNAs were detected in the placenta and yolk sac. In the placenta, GPRC5B was present in maternal and fetal vascular endothelial cells, stromal cells, fibroblast-like cells and glycogen cells. A strong reaction to RARalpha was detected in maternal and fetal vascular endothelial cells and stromal cells. The levels of GPRC5B and RARalpha proteins in maternal and fetal vascular endothelial cells decreased with gestation. In the yolk sac, GPRC5B and RARalpha proteins were detected in vascular endothelial cells, but their levels did not change during the gestation period. These findings indicate that GPRC5B is involved in RA-dependent morphogenesis/angiogenesis and regulation of extracellular matrix synthesis in the murine placenta and yolk sac.     

Evaluation of an indirect enzyme-linked immunosorbent assay (ELISA) using recombinant toxin for detection of antibodies against Pasteurella multocida toxin

To facilitate the control of progressive atrophic rhinitis (PAR) of swine caused by toxigenic Pasteurella multocida, an enzyme-linked immunosorbent assay (ELISA) and a serum neutralization test (NT) have recently been developed to detect antibodies against the P. multocida dermonecrotic toxin (PmDNT). However, the NT is a cumbersome and time-consuming technique. To overcome these drawbacks, we developed an indirect ELISA, using recombinant PmDNT expressed in Escherichia coli, for the detection of antibodies to PmDNT in serum samples from pigs. The practical usefulness of this ELISA was compared with the NT using serum samples obtained from experimentally infected and naturally infected pigs. In the pigs experimentally inoculated with vaccine including PmDNT toxoid, the ELISA and neutralization antibodies were detected at almost the same time, and a good correlation was demonstrated between both tests (P<0.01, R(2)=0.807). Therefore, the ELISA can be used to evaluate the immune reaction of pigs after vaccination with P. multocida toxoid. In a survey conducted on a field herd with a history of clinical AR, the seropositivity by ELISA in pigs of age 4.5-6 months was increased even though the NT was negative, and the correlation was low between the results obtained with the two tests (P<0.01, R(2)=0.38). Therefore, the results indicated that this ELISA might be a useful alternative to the NT currently used to detect the antibody to PmDNT after vaccination or infection with P. multocida.     

Ovarian activity and pregnancy in the Siberian tiger, Panthera tigris altaica, assessed by fecal gonadal steroid hormones analyses

Feces were collected from two female and one male Siberian tigers, Panthera tigris altaica. Steroid hormones were extracted from lyophilized feces and quantified by enzyme immunoassay. The fecal contents of estradiol-17beta (E(2)) and testosterone in the females and male, respectively, changed markedly throughout the year. The fecal E(2) contents of females Nos. 179 and 238 increased at 26.4 +/- 8.0 and 28.0 +/- 14.2 day intervals, respectively. However, the fecal contents of progesterone (P(4)) in the female kept alone did not change. In contrast, the other female, which was kept with a male, had increased fecal P(4) contents after copulation. The fecal progesterone levels of the pregnant female remained high during her 106-day pregnancy.     

Identification of Trichoderma SKT-1, a biological control agent against seedborne pathogens of rice

Trichoderma SKT-1 was previously reported as a powerful biological control agent against seedborne pathogens of rice, but the taxonomic disposition of the fungal isolate was not clear. Trichoderma SKT-1 produced irregular pyramidal warts on conidia and had an optimum growth temperature of 30°C. Morphological characteristics and colony growth were identical to those of known species of Trichoderma, including the newly recognized species T. asperellum. The 5.8S rDNA with the internal transcribed spacer (ITS) region (ca. 514 bp) of the fungus was compared with those of known species to determine the phylogenetic placement of the fungus. The length and sequence of the regions from Trichoderma SKT-1 were completely identical to those of an isolate of T. asperellum NRRL 5242 (AJ230669). On the basis of these results, we concluded that Trichoderma SKT-1 was T. asperellum.     

Evaluation of cytokine messenger RNA expression in peripheral blood mononuclear cells from dogs with canine demodicosis

Using RT-PCR and semi-quantitative PCR, mRNA expression for canine interferon (IFN)-gamma, interleukin (IL)-2, IL-4, IL-5, IL-10, tumor necrosis factor (TNF)-alpha and transforming growth factor (TGF)-beta in peripheral blood mononuclear cells (PBMCs) was examined in dogs with or without demodicosis. mRNA expression for IFN-gamma as well as TNF-alpha in dogs with demodicosis (localized (LD) and generalized (GD)) was slightly lower than those in dogs without demodicosis (healthy controls). Expression of IL-5 mRNA in dogs with demodicosis was higher than that in control dogs, but there were no significant differences in IL-4 and IL-10 mRNA expression levels among the three groups. On the other hand, expression levels of TGF-beta mRNA in dogs with GD were higher than those in control dogs and dogs with LD. The expression levels of IL-5 and TGF-beta mRNA decreased in all three dogs with GD which showed resolution of the clinical signs. Taken together, these results suggest that the Th2-like response in PBMCs from dogs with demodicosis is up-regulated, and that subsequent increased expression of IL-5 and TGF-beta mRNA in dogs with GD is reversible after treatment. Therefore, these cytokines, particularly IL-5, might be a useful clinical index of the clinical course in demodicosis. Also, increased TGF-beta mRNA expression might be a key factor for revealing the difference in the mechanism of onset between LD and GD.     

Comparison of macrophage scavenger receptor-A knockout mice with wild type ones in the immune response against repeated infestation with Haemaphysalis longicornis

Using macrophage scavenger receptor-A knockout (SRKO) mice, we examined the role of macrophage class A scavenger receptors (MRS-A) on the immune response and acquisition of host resistance against repeated infestation with Haemaphysalis longicornis. Except for one batch of nymphs that infested one of the SRKO (SR-/-) mice and showed no appreciable reduction in body weight, all the other groups of nymphs manifested significant decrease in body weight. Both SR-/- and wild type (SR+/+) mice showed a sustained increase in anti-tick antibody titers, but SR+/+ mice showed significantly higher titers. The IFN-gamma assayed in SR-/- mouse immune sera was substantially less compared with that in SR+/+ mice. Immune sera from SR-/- and SR+/+ mice recognized the 51 and 44 kDa, and 44 kDa proteins, respectively, of the salivary gland antigen. The difference in the level of anti-tick resistance manifested by both groups of mice may be influenced by less efficient trapping and processing of tick antigens by macrophages in mice lacking for the macrophage scavenger receptors, and consequently affected the cascade of Th1 and Th2 responses. We have thus obtained valuable data that strongly infer the role of MSR-A in enhancing host defense against repeated infestation with H. longicornis.