Susceptibility to 13 antimicrobial agents was examined for 138 Escherichia coli isolates obtained from 192 fecal samples of wild cranes that migrated for wintering to the Izumi plain, Kagoshima prefecture in Japan. The numbers of isolates that were resistant to the antimicrobials used in this study are as follows: oxytetracycline (OTC), 22 isolates; minocycline, 7 isolates; ampicillin (ABPC), 4 isolates; nalidixic acid, 4 isolates; enrofloxacin, 2 isolates; kanamycin, one isolate. Multidrug resistant isolates exhibiting 2-4 drug resistances were obtained. All of the OTC-resistant isolates carried either the tet (A) or tet(B) gene. The bla_TEM gene was found in all of the ABPC-resistant isolates.

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Salmonella enterica includes several related serovars which have different host ranges and cause diseases of different severities. However, their pathogenic potential is unknown, and it is not clear what mechanisms are activated or inhibited during adaptation to a specific host environment. Some proteins are involved in the mechanism of pathogenicity at a molecular level and provide the functional aspects that create the diverse phenotypes. To compare proteomic analyses of the total proteins of Salmonella Enteriditis (SE), Typhimurium (ST), and Gallinarum (SG), two-dimensional gel electrophoresis (2-DGE) was performed using a pH 4-10 immobilized pH gradient (IPG) strip, and some proteins were identified by mass spectrometry (MS). After staining the gels, the proteins that were expressed at 10-fold or higher levels compared to other spots on the gel were characterized. Some of the identified proteins were related to virulence, such as β-lactamase, RfbH protein, and shikimate kinase. Additionally, there was a high level of variation between serovars despite the similarities in the expression patterns. Furthermore, this study shows that 2-DGE combined with MS is a useful tool for identifying proteins differentially expressed between serovars with different host ranges and pathogenic potential.

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It has been suggested that drinking oxygenated water may improve oxygen availability, which may increase vitality and improving immune activity. The present study evaluated the immune enhancing effects of oxygenated drinking water in broiler chicks and demonstrated the protective efficacy of oxygenated drinking water against Salmonella Gallinarum in experimentally infected broiler chicks. Continuous drinking of oxygenated water markedly increased serum lysozyme activity, peripheral blood mononuclear cell proliferation and the CD4⁺/CD8⁺ splenocyte ratio in broiler chicks. In the chicks experimentally infected with S. Gallinarum, oxygenated drinking water alleviated symptoms and increased survival. These findings suggest that oxygenated drinking water enhances immune activity in broiler chicks, and increases survivability against S. Gallinarum in experimentally infected broiler chicks.

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Interleukin (IL)-12 and IL-23 play central roles in the regulation of distinct helper T-cell subsets, i.e. Th1 and Th17, respectively. Although IL-12 and IL-23 have been well studied in human and rodent systems, little is known about their significance in other animals, including livestock mammals such as cattle and pigs. In this study, we performed molecular cloning and genetic characterization of a small component of swine IL-23, i.e., IL-23p19; in addition, we identified and performed chromosomal assignment of the genes encoding its receptor (R) subunits IL-23Rα and IL-12Rβ1. These results provide genetic information about both swine IL-23/IL-23R and IL-12/IL-12R systems, which allows for better understanding of IL-12/IL-23 systems involved in pig immunity.

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Loop-mediated isothermal amplification (LAMP) constitutes a potentially valuable diagnostic tool for rapid diagnosis of contagious diseases. In this study, we developed a novel LAMP method (seM-LAMP) to detect the seM gene of Streptococcus equi subsp. equi (S. equi), the causative agent of strangles in equids. The seM-LAMP successfully amplified the target sequence of the seM gene at 63°C within 60 min. The sensitivity of the seM-LAMP was slightly lower than the 2nd reaction of the seM semi-nested PCR. To evaluate the species specificity of the seM-LAMP, we tested 100 S. equi and 189 non-S. equi strains. Significant amplification of the DNA originating from S. equi was observed within 60 min incubation, but no amplification of non-S. equi DNA occurred. The results were identical to those of seM semi-nested PCR. To investigate the clinical usefulness of the methods, the seM-LAMP and the seM semi-nested PCR were used to screen 590 nasal swabs obtained during an outbreak of strangles. Both methods showed that 79 and 511 swabs were S. equi positive and negative, respectively, and the results were identical to those of the culture examination. These results indicate that the seM-LAMP is potentially useful for the reliable routine diagnosis of Streptococcus equi subsp. equi infections.

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A 6-year-old ShihTzu presented with tonic-clonic cluster seizure. T2-weighted magnetic resonance (MR) images showed bilateral diffuse hyperintense lesions at the cerebral cortex with enlarged sulci. Computed tomography revealed a portosystemic shunt (PSS) and azygos continuation. Based on the clinical signs, blood examinations and diagnostic images, the dog was diagnosed with hepatic encephalopathy secondary to PSS. The neurologic signs were gradually improved after medical therapy for hyperammonemia. This is the first report of hyperintensity of the cerebral cortex on T2-weighted MR images associated with acute hepatic encephalopathy in a dog.

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A serological survey of hepatitis E virus (HEV) antibody was conducted using 202 adult captive cynomolgus monkeys, who did not show any clinical signs of acute hepatitis. Out of these, 44 monkeys were sero-positive for anti-HEV IgG and all monkeys were negative for anti-HEV IgM. All positive monkeys came from either Vietnam or China, but none from the Philippines, Indonesia, or our facility. Selected 12 monkeys out of positive monkeys from Vietnum, including 9 positive and 3 negative, revealed mostly within the reference ranges for alanine aminotransferase (ALT) and asparatate aminotransferase (AST) by serum biochemistries. Their titers of anti-HEV IgG did not correlate with the concentrations of ALT and AST. Moreover, HEV-RNA could not be detected from any fecal specimens of the 12 monkeys. Thus, monkeys with anti-HEV IgG sero-positive did not seem to be source of the HEV-pollution, because 1) sero-positive monkeys did not excrete HEV-RNA from their feces, and 2) monkeys from the Philippines and Indonesia have remained to be sero-negative for anti-HEV IgG, even if the monkeys were kept in same animal room of our facility. From these results, it could be inferred that primary infection of HEV occurred in the exported countries, but not in our colony. The contamination of HEV in indoor-captive monkeys could be prevented by precise quarantine tests, including ELISA for detecting anti-HEV and RT-PCR for HEV RNA.

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Retinal ischemia is a common cause of visual impairment for humans and animals. The neuroprotective effects of lidocaine (LDC) and methylprednisolone (MP) upon retinal ischemic injury were investigated in a rat model. Sprague-Dawley rats were divided into 3 groups, the IR control, LDC and MP. A very high intraocular pressure (HIOP) and retinal ischemia were induced. In LDC group, LDC bolus (1.5 mg/kg) was IV injected 30 min before ischemia and then a constant rate infusion (CRI) with 2 mg/kg/hr was given until 60 min after reperfusion. In MP group, MP bolus (30 mg/kg) was IV administered twice at 2 min before and immediately after ischemia, respectively. The HIOP damage to retina was evaluated by electroretinogram (ERG) and morphometrical histology. The functional analysis of the retina by ERG revealed a 35.2% reduction of a-wave in the IR group, 49.7% reduction in the LDC group but no significant change in the MP group compared to normal controls. An 81.0% reduction of b-wave was observed in the IR group, 80.7% reduction in the LDC group and 17.6% reduction in the MP group. In the morphometrical histology, the retinal inner plexiform layer/outer nuclear layer (IPL/ONL) ratio was reduced to 48.8% in the IR group, 80.1% in the LDC group and 96.2% in MP group. In conclusion, the MP showed significantly good neuroprotective effects on retinal IR injury, and the LDC showed moderate neuroprotective effects demonstrated in retinal structure but not in retinal function.

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The seroprevalence of B Virus (BV, Macacine herpesvirus 1) in bred cynomolgus monkeys (Macaca fascicularis) in Cambodia was investigated. Blood samples were collected by venous puncture between 2007 and 2011, and a commercially available enzyme-linked immunosorbent assay (ELISA) kit was used to detect IgG antibodies against BV. A total of 1710 blood samples were examined, and the overall prevalence of BV antibodies was 33.92% (580/1710). The young adult (2-6 years) monkeys had a seroprevalence of 30.57% (497/1626), and the older (>7 years) monkeys had a seroprevalence of 98.81% (83/84). The present investigation revealed that the prevalence of BV antibodies in bred cynomolgus monkeys was highly correlated with their age, with higher prevalence in older monkeys in Cambodia.

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Nicotinamide exerts neuroprotective effects against focal cerebral ischemic injury. Phosphoprotein enriched in astrocytes 15 (PEA-15) is prominently expressed in astrocytes that exert broad anti-apoptotic functions. This study investigated whether nicotinamide modulates PEA-15 and levels of two phosphorylated PEA-15 (Serine 104 and 116) in an animal model of middle cerebral artery occlusion (MCAO)-induced injury. Adult male rats were treated with vehicle or nicotinamide (500 mg/kg) 2 hr after the onset of MCAO and cerebral cortices were collected at 24 hr after MCAO. In a proteomic approach, MCAO induced decreases of PEA-15 levels, while nicotinamide treatment attenuated the injury-induced decrease in PEA-15. The results of Western blot analysis suggest that nicotinamide prevented injury-induced reduction in phospho-PEA-15 (Serine 104) and phospho-PEA-15 (Serine 116) levels. The phosphorylation of PEA-15 exerts anti-apoptotic functions, and reduction of PEA-15 phosphorylation leads to apoptotic cell death. These results suggest that nicotinamide exerts a neuroprotective effect by attenuating the injury-induced decreases of PEA-15 and phospho-PEA-15 (Ser 104 and Ser 116) proteins.

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The Japanese Fancy Mouse 1 (JF1) has a characteristic coat color similar to a very old mutant, piebald. The mutation in JF1 and the classic piebald was previously thought to be the same recessive allele in the endothelin B receptor gene (Ednrb) according to the haplotype pattern, which is insufficient for this conclusion. In this study, we identified the same insertion of a retroposon-like element in intron 1 of the Ednrb gene in JF1 as in the classic piebald mutation by PCR. Further, we investigated whether the intestine shows neuronal intestinal malformations such as hypoganglionosis and immaturity of ganglion cells by histochemical staining. Though it has been assumed that the defect of neural crest-derived lineages is restricted to melanocytes in JF1, we found that the enteric innervation and neuronal density were impaired throughout the whole colon in JF1 mice.

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A 3-year-old female common marmoset was euthanatized because of wasting. Grossly, a perforating lesion was present in the jejunum and hemorrhagic lesions in the cecum and colon. Histopathologically, these gross lesions were a perforated jejunal ulcer and necrotizing colitis, respectively. Necrotizing colitis was characterized by extensive mucosal necrosis along with numerous ribbon-shaped aseptate hyphae. These aseptate hyaline hyphae were positively stained with PAS and GMS, and reacted immunohistochemically with the antibody against the family Mucoraceae. This case was diagnosed as intestinal mucormycosis. This is the first report on mucormycosis in a common marmoset.

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The kidneys of a 3-month-old female cat were examined. The cat which had been raised under controlled conditions with no history of any poisoning showed progressive weight loss with increases in blood BUN and creatinine concentrations. At necropsy, both kidneys were firm in consistency with formation of focal scars. Histopathologically, widespread deposition of crystals was observed in the renal tubules (in both dilated lumina and degenerative epithelia) accompanying mild interstitial fibrosis with lymphocyte infiltration. The crystals were colorless or basophilic on the hematoxilin and eosin-stained section and could be visualized with polarized light as doubly fractile crystals. The crystals were identified as calcium oxalate crystals by histochemical examinations using von Kossa stain and alizarin red S stain under different conditions and by ultrastructural examination. Judging from the above-mentioned findings, the present renal lesion detected in an infant cat was diagnosed as renal oxalosis which was suspected to be hereditary in nature.

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Antibiotic beads have become popular for the treatment of local bacterial infections. The preparation of antibiotic beads from commercial pharmaceutical antibiotics is a convenient method in clinic. The elution characteristics of cefazolin from polymethylmethacrylate (PMMA) (SmartSet HV, Depuy I and Cemfix 3) beads and calcium sulfate beads were studied. Commercial cefazolin formulation was incorporated in PMMA or calcium sulfate at 1 g cefazolin /10 g of matrix substances to form beads. The concentrations of eluted cefazolin during 15 days were greater than MIC for Staphylococcus aureus (ATCC 25923). The eluted cefazolin concentrations were in the range of 3.6 ± 1.2 to 4.6 ± 0.4 mg for PMMA beads and 15.4 ± 1.7 mg for calcium sulfate beads. The accumulated eluted cefazolin from PMMA beads and calcium sulfate beads for 15 days were 34.41 ± 3.93 to 38.67 ± 3.04% and 95.94 ± 3.93%, respectively. The various storage conditions; at room temperature or 4°C, with or without light-protection, for 6 months had little effects on the amounts of eluted cefazolin. The results showed both in-housed cefazolin-PMMA beads and cefazolin-calcium sulfate beads could be the effective tools for the treatment of local bacterial infections.

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Quantitative analyses of Listeria monocytogenes in imported ready-to-eat (RTE) foods sold at retail stores in Japan were performed. Of the 77 non-cooked meat products, 6 samples (7.8%) tested positive. The levels of contamination of 4 of the samples were below 100 colony-forming units (CFU)/g, which is the microbiological criterion for L. monocytogenes in RTE foods as determined by Codex. However, Listeria cells at levels of 100 and 400 CFU/g were detected in a salami sample and a raw ham sample, respectively. All of the 70 cheese samples and the 3 samples made from raw ham and cheese showed negative test results. These results suggest that imported RTE foods are potential sources of the causative agent of listeriosis.

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The aim of the present study was to evaluate the anesthetic and cardiorespiratory effects of tiletamine/zolazepam and the effect of flumazenil on the recovery from tiletamine/zolazepam anesthesia in the pig. Six Landrace and Yorkshire cross-bred pigs (three females and three males, 3-4 months old) weighing 35.8 ± 1.7 kg were used in this study. Pigs were given tiletamine/zolazepam intramuscularly at a dose of 4.4 mg kg^-1 (2.2 mg kg^-1 tiletamine and 2.2 mg kg^-1 zolazepam) of body weight. Twenty minutes after the administration of tiletamine/zolazem, the pigs were given saline solution (control, Group TZ) or given flumazenil intravenously at a dose of 0.08 mg kg^-1 of body weight (Group TZF). Anesthesia and recovery times, scores of anesthetic effects and cardiorespiratory variables were recorded for each pig. There was a significant difference between the duration of tiletamine/zolazepam anesthesia with and without the antagonist. Flumazenil significantly shortened the recovery time. A significant difference in blood gas variables was observed between the two groups. The anesthetic effects induced by tiletamine/zolazepam could be reversed successfully and safely by flumazenil alone. Therefore, flumazenil administration could be considered in cases in which quick recovery is required in pigs.

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This study is the first report on circulating testosterone and inhibin levels in a species of whales, the beluga. Circulating testosterone and immunoreactive (ir-) inhibin levels in two captive male belugas ("Nack", originally from Canada and "Duke", from the Okhotsk Sea) were measured every month for 9 years between 1995 and 2003. Assuming that clearly increased testosterone levels in the circulation indicates that the belugas had reached sexual maturity, at the ages of 10 ("Nack") and 11 years old ("Duke"). Their testosterone levels before the significant increase (pre-pubertal) were 0.42 ± 0.07 ng/ml (n=18) and 0.35 ± 0.10 ng/ml (n=18) and, those of after the increase (maturity) were 1.65 ± 0.14 ng/m l (n=74) and 2.06 ± 0.14 ng/ml (n=74). Circulating ir-inhibin levels before sexual maturity were 0.78 ± 0.04 ng/ml (n=18) and 0.64 ± 0.04 ng/ml (n=15) and, after sexual maturity were 0.52 ± 0.02 ng/ml (n=56) and 0.43 ± 0.02 ng/ml (n=67). Seasonal changes were observed in the testosterone levels after sexual maturity and the levels increased during March and April in Canadian origin "Nack", and peaked in February in Okhotsk origin "Duke". Circulating ir-inhibin level gradually decreased as they aged. A negative correlation between the circulating testosterone and ir-inhibin was observed. No seasonal changes were observed in the ir-inhibin levels after sexual maturity. These data will surely correspond to clarification of endocrinology and the successful reproduction of the beluga.

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The present study determined the effect of different types of sugars (lactose, fructose, glucose and sorbitol) used in egg yolk-based extender on the post-thawed boar semen quality. Twenty-two ejaculates from 6 fertility-proven Yorkshire boars were cryopreserved by liquid nitrogen vapor method. Sperm motility, viability, acrosome integrity and intact functional plasma membrane were determined at 0, 2 and 4 hr after thawing. It was found that the lactose-based extender resulted in a higher percentage of post-thawed sperm motility, viability, intact acrosome and functional plasma membrane than sorbitol-based extender (P<0.05) and fructose-based extender yielded a higher post-thawed sperm motility and viability than sorbitol-based extender (P<0.05). It could be concluded that sorbitol was not an effective sugar for the cryopreservation in boar semen.

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A 13-year old intact female poodle showed persistent signs of estrus. Ultrasonographically, a large cystic mass containing echogenic fluid with a thin wall was observed, and it was 8 cm in diameter. The cystic mass was located at the mesovarium between the left ovary and uterine horn, and it was identified as a giant parovarian cyst. A right ovarian mass was histologically diagnosed as a granulosa cell tumor. Persistent estrogen hormone secretion by the granulosa cell tumor was thought to cause a rapid increase in the size of the parovarian cyst. This rare cystic mass originated from the genital system and was differentiated from the uterus.

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This study was performed to clarify the sow-to-fetus transmission pathway of Torque teno sus virus (TTSuV) types 1 (TTSuV1) and 2 (TTSuV2). For this purpose, detection of TTSuV1 and TTSuV2 (TTSuVs) in sera of 6 sows (Sows 1-6) at parturition and in sera of their newborn piglets immediately after birth without suckling colostrum was performed by nested polymerase chain reaction (nPCR). These sows were bred using semen that had tested negative for TTSuVs. In a TTSuV1- and TTSuV2-positive sow (Sow 1), TTSuV1 and TTSuV2 were detected in 4 and 5 of 12 newborn littermates, respectively. In a TTSuV1-positive sow (Sow 2), TTSuV1 was detected in 1 of 8 newborn littermates. In 4 TTSuV1- and TTSuV2-negative sows (Sows 3-6), TTSuV1 was detected in 6 out of the 25 newborn piglets of 3 sows (Sows 3-5), while TTSuVs were not detected in all 13 piglets of 1 sow (Sow 6). In addition, to investigate the possibility of a sow-to-piglet transmission pathway of TTSuV via colostrum, TTSuV1 and TTSuV2 in sera of 12 newborn piglets from Sows 1-3 were examined by nPCR. Immediately after birth without suckling colostrum, TTSuV1 and TTSuV2 were not detected in 10 and 8 of 12 newborn piglets, respectively; however, at 24 hr after suckling colostrum, TTSuV1 was detected in 6 piglets, while TTSuV2 was not detected in any piglets. These results confirmed the existence of a sow-to-fetus transmission pathway of TTSuV during normal pregnancy and suggested a possibility of sow-to-piglet transmission of TTSuV via colostrum.

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Rift Valley fever virus (RVFV) is one of the important emerging viral diseases of serious impact in public health and animal hygiene both in human and animal industries. In this study, we developed a monoclonal antibody-based competitive ELISA for the detection of antibodies to RVFV in goats and cattle. The recombinant N protein of RVFV was expressed in E. coli with a six-histidine tag, and the purified N protein was used for detecting antigen with a competitive monoclonal antibody against RVFV antibodies. The competitive ELISA (C-ELISA) could detect antibodies at 9-11 days after inoculation in goats and cattle with a sensitivity of 94.7% (virus neutralization titer >32) and specificity of 99.7%, respectively. In addition, the C-ELISA did not show any cross-reactivity with positive sera against arboviruses such as Akabane, Aino, Chuzan, Ibaraki and bovine ephemeral fever virus, which are prevalent viral agents in ruminant animals throughout Southeast Asia. The results of the present study indicate that the C-ELISA is a simple, rapid and convenient serodiagnostic method for RVFV in goats and cattle.

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We recently found that certain canine live attenuated vaccines produced using `non-feline' cell lines were contaminated with an infectious feline endogenous retrovirus, termed RD-114 virus. We suspected that RD-114 virus may have contaminated the seed stock of canine parvovirus (CPV) during the production of the contaminated vaccines. In this study, we collected stock viruses of CPVs propagated in a feline cell line, and checked the presence of infectious RD-114 virus. Consequently, we found that RD-114 viral RNA was present in all stock viruses, and 7 out of 18 stock viruses were contaminated with infectious RD-114 virus. We also found that RD-114 virus was stable physically and is capable of retaining its infectivity for a long period at -80°C.

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