The mechanism by which indigenous bacteria on the follicle-associated epithelium (FAE) of lymphatic follicles (LFs) accelerate the differentiation of microvillous columnar epithelial cells (MV) into M-cells was immunohistochemically investigated in rat Peyer’s patches. The results showed that the number of Toll-like receptor (TLR) -4⁺ M-cells was greater in the FAE with expansion of bacterial colonies (LFs with bacterial colonies on the FAE: b-LF) than the FAE without expansion of bacterial colonies (nb-LF). TLR-4 was also expressed in the striated borders of MV upstream next to M-cells in the FAE of the b-LF. TLR-4⁺ vesicles were frequently detected in the cytoplasms of MV with TLR-4⁺ striated borders upstream next to TLR-4⁺ M-cells in the FAE of b-LF. These findings suggest that TLR-4⁺ MV take up TLR-4 ligands and differentiate into M-cells in the b-LF. Neither the distribution of RANK nor that of RANKL was coincident with that of M-cells in the b-LF. Moreover, RANK, but not RANKL, was expressed in intestinal villi, whereas cleaved caspase-3 was immunonegative in the MV and M-cells of the FAE, unlike in villous epithelial cells. Therefore, RANK/RANKL signaling in the LF might contribute to the down-regulation of epithelial apoptosis to facilitate the differentiation of MV into M-cells in rat Peyer’s patches.

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We aimed to determine the resistance mechanisms of 27 T. pyogenes isolates from swine in the Jilin province of China. Drug sensitivity analysis indicated that most of the isolated strains were resistant to aminoglycosides. We investigated the genes involved in target alteration, drug inactivation, and increased efflux as potential resistance mechanisms. Two known aminoglycoside resistance genes (aphA1 and strB) were not found in the genomic DNA of any isolate. A 3-bp (CCC) deletion in one 16S rRNA operon was detected in all isolates, and efflux pumps were not active in the resistant group. Ultimately, genes encoding aminoglycoside-modifying enzymes carried by class 1 integrons were identified as the main cause of resistance to aminoglycosides in T. pyogenes.

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The objective of the study was to evaluate the effects of body weight (BW), breed, and sex on two-dimensional (2D) echocardiographic measures, reference ranges, and prediction intervals using allometrically-scaled data of left atrial (LA) and left ventricular (LV) size and LV wall thickness in healthy cats. Study type was retrospective, observational, and clinical cohort. 150 healthy cats were enrolled and 2D echocardiograms analyzed. LA diameter, LV wall thickness, and LV dimension were quantified using three different imaging views. The effect of BW, breed, sex, age, and interaction (BW*sex) on echocardiographic variables was assessed using univariate and multivariate regression and linear mixed model analysis. Standard (using raw data) and allometrically scaled (Y=a × M^b) reference intervals and prediction intervals were determined. BW had a significant (P<0.05) independent effect on 2D variables whereas breed, sex, and age did not. There were clinically relevant differences between reference intervals using mean ± 2SD of raw data and mean and 95% prediction interval of allometrically-scaled variables, most prominent in larger (>6 kg) and smaller (<3 kg) cats. A clinically relevant difference between thickness of the interventricular septum (IVS) and dimension of the LV posterior wall (LVPW) was identified. In conclusion, allometric scaling and BW-based 95% prediction intervals should be preferred over conventional 2D echocardiographic reference intervals in cats, in particular in small and large cats. These results are particularly relevant to screening examinations for feline hypertrophic cardiomyopathy.

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Although alteration of commensal microbiota is associated with chronic gastrointestinal (GI) diseases such as inflammatory bowel disease (IBD) in dogs, the microbiota composition in intestinal lymphoma, an important differential diagnosis of canine IBD, has not been investigated. The objective of this study was to compare the fecal microbiota in dogs with IBD, dogs with intestinal lymphoma, and healthy dogs. Eight dogs with IBD, eight dogs with intestinal lymphoma, and fifteen healthy dogs were included in the study. Fecal samples were analyzed by 16S rRNA gene next-generation sequencing. Rarefaction analysis failed to reveal any difference in bacterial diversity among healthy dogs and diseased dogs. Based on PCoA plots of unweighted UniFrac distances, the bacterial composition in dogs with intestinal lymphoma was different from those observed in dogs with IBD and healthy dogs. When compared with healthy dogs, intestinal lymphoma subjects showed significant increases in organisms belonging to the Eubacteriaceae family. The proportion of the family Paraprevotellaceae and the genus Porphyromonas was significantly higher in dogs with IBD compared to healthy dogs. These observations suggest that dysbiosis is associated with intestinal lymphoma as well as IBD in dogs.

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An 8-year-old neutered female Yorkshire terrier with mediastinal neoplasm and subsequent cranial vena cava invasion developed multiple venous collaterals from the brachiocephalic venous trunks to the caudal vena cava. Collateral venous pathways have been described in dogs with obstruction or increased blood flow resistance of the caudal vena cava but cranial vena cava collaterals have not been reported until now in veterinary patients. In this report, the CTA characteristics of such peculiar vascular routes are described and compared to similar findings reported in human medical literature. The recognition of such ancillary CT finding could help radiologists to reach a more accurate diagnosis of superior vena cava syndrome.

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In this study, two commercially available oral electrolyte solutions (OES) with high sodium (CF) or with high glucose and glycine (SL), and two prototype OES were evaluated in terms of rehydration and preventing catabolism. Prototype OES based on CF were prepared by doubling the glucose amount (CFG) or by doubling both glucose and glycine (CFGG). Thirty-two diarrheic calves were randomly assigned four groups with eight calves in each group. Blood volume increased with CF and CFGG compared with that of other OES. The catabolic preventive effect was excellent in CFGG and SL. Our results suggest that both the amount of sodium, glucose, and glycine, and ratio of these factors aid dehydration and provide energy.

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The safety and biological effects of a long-term dose of D-allulose were evaluated in healthy dogs. For 12 weeks, the dogs were administered D-allulose (0.2 g/kg) or placebo daily. Plasma total cholesterol concentrations in the D-allulose group were significantly lower than those in the control group at and after week 2 (P<0.05). D-Allulose administration did not cause clinical signs or changes in hematological and biochemical levels, except for lipids. D-Allulose administration also did not influence body weight. Plasma glucose and insulin concentrations in the glucose tolerance test, performed one day after the termination of D-allulose administration, were not different between groups, suggesting no cumulative effects of D-allulose on glucose metabolism in healthy dogs. In conclusion, long-term administration of D-allulose caused no harmful effects in dogs.

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A five-year-old rabbit was evaluated for a 7 to 8 month history of regurgitation, weight loss, and hyporexia. Previously performed whole body radiographs, plasma biochemistry results and complete blood count revealed had no significant abnormalities. A computed tomography (CT) scan revealed a circumferential caudal esophageal thickening. The animal received supportive care until euthanasia was performed 6 weeks later. Caudal esophageal smooth muscle hypertrophy was diagnosed on necropsy. This case indicates that regurgitation can occur in rabbits and advanced imaging can investigate the underlying cause.

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On Amami Oshima Island, free-ranging and feral cats are harmful to wildlife populations. In this study, the seroprevalence of Toxoplasma gondii in these cats was examined using a newly developed Gaussia luciferase immunoprecipitation system assay. Of 1,363 cats, 123 cats (9.0%) was positive for T. gondii. The prevalence was significantly different in different areas; among cats in the rural area, where many wild animals live, including endangered species, T. gondii infection was more prevalent than in the urban area of the island. This finding indicates a possible risk to wildlife of infection from free-ranging and feral cats. Therefore, management of cats is important for wildlife conservation.

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Natural infection with larval Echinococcus multilocularis was recognized in one of eight Norway rats, Rattus norvegicus, caught indoors in 2009 in Ebetsu, Hokkaido, northern Japan. Cystic lesions were found in the right median and lateral lobes of the liver, with numerous alveolar cysts in the periphery of the lesions. Protoscolices were formed within large cysts. The laminated layers of the cysts were positive for PAS staining. Nested PCR using the primers specific for Taenia mitochondrial 12S rDNA yielded a 250-bp product, and the sequence of the PCR product matched that of E. multilocularis isolates from Hokkaido and Germany. This is the third natural alveolar hydatidosis in R. norvegicus in Japan.

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β-catenin, E-cadherin and N-cadherin are adhesion molecules that play important roles in organogenesis, tissue homeostasis, renal epithelial integrity and polarity. The present study demonstrated their immunolocalization in adult and neonate rat kidney. Membranous or cytoplasmic expression of β-catenin, E-cadherin and N-cadherin were seen in adult and developing renal tubular epithelial cells. Particularly, in adult kidney, E-cadherin and β-catenin were intensively expressed in distal renal tubules, whereas N-cadherin was expressed in proximal renal tubules. In neonate rat kidney on 1 and 4 days old, developing renal tubular epithelial cells were mainly reacted with E-cadherin and very weakly expressed N-cadherin; β-catenin was expressed in developing renal tubules and mesenchymal blastemal cells. Interestingly, β-catenin-positive renal tubular epithelial cells simultaneously expressed E-cadherin in the kidney of adult and developing rats. Collectively, the adhesion molecules were differentially distributed in the renal tubules of adult rats and β-catenin and E-cadherin are predominant adhesion molecules in developing kidney. The present findings would provide the basic information of evaluating renal tubular toxicity using rats, in addition to renal genesis, in terms of adhesion molecules.

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Macrophage polarization is a process by which macrophages acquire a distinct phenotypic and functional profile in response to microenvironmental signals. The classically and alternatively activated (M1 and M2, respectively) macrophage phenotypes are defined by the specific molecular characteristics induced in response to prototypic pro- and anti-inflammatory cues. In this study, we used LPS/IFN-γ and IL-4 to stimulate porcine alveolar macrophages (PAMs) in vitro and investigated the expression changes of several novel markers during macrophage polarization. Notably, we found that LPS/IFN-γ-stimulated PAMs express prototypical M1 molecules, whereas IL-4-stimulated PAMs express M2 molecules. We also demonstrated that replication of the highly pathogenic porcine reproductive and respiratory syndrome virus (PRRSV) strain HuN4 was effectively suppressed in LPS/IFN-γ-stimulated M1 PAMs (M1 type), but not IL-4 stimulated M2 PAMs. However, this was not observed with the classic, less pathogenic CH-1a strain. Moreover, we found that M2 marker expression gradually increased after PAM infection with PRRSV, whereas no significant changes were found with M1 marker expression, suggesting that PRRSV infection may skew macrophage polarization towards an M2 phenotype. Finally, we found that anti-viral cytokine expression was significantly higher in M1 macrophages than in M2 macrophages or nonpolarized controls. In summary, our results show that PRRSV replication was significantly impaired in M1 PAMs, which may serve as a foundation for further understanding of the dynamic phenotypic changes during macrophage polarization and their effects on viral infection.

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Two cases of extralobar pulmonary sequestrations from a walrus (Odobenus rosmarus) and a Steller sea lion (Eumetopias jubatus) are described in the present study. Grossly, an independent, soft unilocular cystic mass was found within the abdominal cavities of both animals, adherent to the diaphragm in O. rosmarus and attached to the cardia of the stomach in E. jubatus. Histopathologically, the cysts were lined by pseudostratified ciliated columnar epithelium with abundant goblet cells, while the wall comprised of glands, hyaline cartilage, bronchiole- and alveolus-like structures, smooth muscles, and large, well-developed elastic and muscular arteries. The pinniped cases presented are exceptionally rare and to the best of the authors’ knowledge, marks the first descriptions of this congenital anomaly in wildlife.

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Regulation of inflammation in intestinal mesothelial cells in the abdominal cavity is important for the pathogeny of clinical conditions, such as postoperative ileus, peritonitis and encapsulating peritoneal sclerosis. Here we have examined the inflammatory effect of lipopolysaccharide (LPS) and the anti-inflammatory effect of nicotinic acetylcholine receptor stimulation in rat intestinal mesothelial cells. LPS upregulated mRNA expression of interleukin-1β (IL-1β), tumor necrosis factor-α (TNF-α), monocyte chemotactic protein-1 (MCP-1) and inducible nitric oxide synthase (iNOS). The α7, α9 and α10 subunits of nicotinic acetylcholine receptor were detected in intestinal mesothelial cells. Nicotine (10 nM) significantly inhibited LPS-induced mRNA expression of IL-1β and iNOS, but not TNF-α and MCP-1. In addition, the α7 nicotinic acetylcholine receptor selective agonist, PNU-282987 (10 nM), significantly inhibited LPS-induced mRNA expression of IL-1β but not TNF-α, iNOS and MCP-1. Finally, we found that enteric nerves adhered to intestinal mesothelial cells located under the ileal serosa. In conclusion, intestinal mesothelial cells react to LPS to induce the production of nitric oxide from iNOS. The anti-inflammatory action of intestinal mesothelial cells expressing α7nAChR may be mediated via their connectivity with enteric nerves.

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Klotho protein is recognized as having a renoprotective effect and is used as a biomarker for kidney injury. We investigated the level of Klotho protein in hyperoxaluria-induced kidney injury and the effects of vitamin E (Vit E) and vitamin C (Vit C) supplementation. Hyperoxaluria was induced by feeding 2% (w/v) Hydroxy-L-proline (HLP) in the drinking water for 21 days. Rats were divided into 5 groups; control (Group 1, n=7), HLP treated rats that received nothing else (Group 2, n=7), Vit E (Group 3, n=6), Vit C (Group 4, n=6) and both Vit E and Vit C (Group 5, n=7). Vit E (200 mg/kg) was injected on days 1, 6, 11 and 16, while Vit C (500 mg/kg) was given intravenously on days 1 and 11. The Klotho protein levels and oxidative status were measured. The expression level of kidney Klotho protein expression was significantly reduced by HLP-treatment, while the mRNA expression was higher (P<0.05), the plasma and kidney malondialdehyde and kidney superoxide dismutase activities were increased, and the kidney reduced glutathione and urinary total antioxidant status were decreased (P<0.05). All of these changes were ameliorated by administration of Vit E, Vit C or especially the co-administration of both. In conclusion, HLP-induced hyperoxaluria reduced the kidney Klotho protein level, which could be restored by Vit E and/or Vit C.

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We investigated the antidepressant-like effect of lactoferrin (Lf) in a repeated forced-swim test (FST) stress mouse model. FST was performed on days 1, 2, 7 and 14. Bovine Lf (bLf) or bovine serum albumin (BSA) was supplemented at 1% to the commercial diet after the first FST throughout the experimental period. The FST-control and FST+BSA group showed a marked increase in immobility time on day 2, which remained increased up to the 14th day, while the FST+bLf group showed a significant lower immobility time. Brain-derived neurotrophic factor (BDNF) content in the hippocampus significantly decreased in all of FST treated groups. These results suggest that bLf may improve the depressive-like symptoms induced by repeated FST.

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This study aimed to evaluate whether radiographic abnormalities at yearling sales were associated with the failure to start racing at 2–3 years of age. Radiographic abnormalities in the carpal (n=852), tarsal (n=976), metacarpophalangeal (n=1,055), and metatarsophalangeal joints (n=1,031) from 1,082 horses, recorded at yearling sale, were reviewed. Eighty-two horses (7.6%) failed to start racing. Radiographic abnormalities such as wedged or collapsed tarsal bones, irregular lucency of a sagittal ridge at the distal aspect of the distal third metatarsal bone, and proximal dorsal fragmentation of the first phalanx in metatarsophalangeal joints were associated with failure to start racing in these horses. In the follow-up survey of 12 horses with one or more these radiographic abnormalities, the horses failed to start racing due to reasons unrelated to these radiographic abnormalities such as pelvic fractures (2 horses), fracture of a distal phalanx (1 horse), cervical stenotic myelopathy and proximal sesamoid fracture (1 horse), superficial digital flexor tendonitis (2 horses), laryngeal hemiplegia (1 horse), economic problems (2 horses) and unknown causes (3 horses). Although radiographic abnormalities at yearling sales can be associated with failure to start racing at 2–3 years of age, these radiographically detected abnormalities might not necessarily cause that failure.

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We investigated the utility of triple-phase helical computed tomography (CT) in differentiating between benign and malignant splenic masses in dogs. Forty-two dogs with primary splenic masses underwent triple-phase helical CT scanning (before administration of contrast, and in the arterial phase, portal venous phase, and delayed phase) prior to splenectomy. Tissue specimens were sent for pathological diagnosis; these included hematomas (n=14), nodular hyperplasias (n=12), hemangiosarcomas (n=11), and undifferentiated sarcomas (n=5). The CT findings were compared with the histological findings. Nodular hyperplasia significantly displayed a homogeneous normal enhancement pattern in all phases. Hemangiosarcoma displayed 2 significant contrast-enhancement patterns, including a homogeneous pattern of poor enhancement in all phases, and a heterogeneous remarkable enhancement pattern in the arterial and portal venous phases. Hematoma and undifferentiated sarcoma displayed a heterogeneous normal enhancement pattern in all phases. The contrast-enhanced volumetric ratios of hematoma tended to be greater than those of undifferentiated sarcoma. Our study demonstrated that the characteristic findings on triple-phase helical CT could be useful for the preoperative differentiation of hematoma, nodular hyperplasia, hemangiosarcoma, and undifferentiated sarcoma in dogs. Triple-phase helical CT may be a useful diagnostic tool in dogs with splenic masses.

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A major role of the corpus luteum (CL) is to produce progesterone (P4). The CL has immature vasculature shortly after ovulation, suggesting it exists under hypoxic conditions. Hypoxia-inducible factor-1 (HIF1) induces the expression of glucose transporter 1 (GLUT1). To clarify the physiological roles of GLUT1 in bovine CL, we examined GLUT1 mRNA expression in the CL under hypoxic conditions by quantitative RT-PCR. We also measured the effects of glucose (0–25 mM) and GLUT1 inhibitors (cytochalasin B, STF-31) on P4 production in bovine luteal cells. GLUT1 mRNA expression in bovine CL was higher at the early luteal stage compared to the other later stages. Hypoxia (3% O₂) increased GLUT1 mRNA expression in early luteal cells, but not in mid luteal cells. Glucose (0–25 mM) increased P4 production in early luteal cells, but not in mid luteal cells. Both GLUT1 inhibitors decreased P4 production in early and mid luteal cells. Overall, the results suggest that GLUT1 (possibly induced by hypoxic conditions in the early CL) plays a role in the establishment and development of bovine CL, especially in supporting luteal P4 synthesis at the early luteal stage.

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H2N2 influenza virus caused a pandemic starting in 1957 but has not been detected in humans since 1968. Thus, most people are immunologically naive to viruses of the H2 subtype. In contrast, H2 influenza viruses are continually isolated from wild birds, and H2N3 viruses were isolated from pigs in 2006. H2 influenza viruses could cause a pandemic if re-introduced into humans. In the present study, a vaccine against H2 influenza was prepared as an effective control measure against a future human pandemic. A/duck/Hokkaido/162/2013 (H2N1), which showed broad antigenic cross-reactivity, was selected from the candidate H2 influenza viruses recently isolated from wild birds in Asian countries. Sufficient neutralizing antibodies against homologous and heterologous viruses were induced in mice after two subcutaneous injections of the inactivated whole virus particle vaccine. The inactivated vaccine induced protective immunity sufficient to reduce the impact of challenges with A/swine/Missouri/2124514/2006 (H2N3). This study demonstrates that the inactivated whole virus particle vaccine prepared from an influenza virus library would be useful against a future H2 influenza pandemic.

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In South Korea, pigs were vaccinated once between 8 and 12 weeks of age because of the injection-site granulomas. Therefore this study was performed to determine the optimal age for single vaccination of growing pigs with the currently used type O FMD vaccine. With 498 pigs divided into four groups, seroprevalence of the antibody was analyzed with enzyme-linked immunosorbent assay. Although double vaccination is necessary to completely protect growing pigs from FMD virus infection with the current vaccine, the age of 8 weeks can be considered as the optimal age for piglet vaccination if the booster injection is unavailable.

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The genetic characterization and actual prevalence of EIAV in Mongolian horse in the disease endemic region is currently unknown. Here, 11 of 776 horse serum samples from four Mongolian provinces tested positive on agar gel immunodiffusion test. Genomic DNA extracted from all seropositive samples was subjected to nested PCR assay. Among these, three samples tested positive with nested PCR assay and were identified by sequencing analysis based on long termination repeat and tat gene of the virus. Two of the three sequences were identical, with 94.0% identity with the third. These two independent Mongolian EIAV sequences were retained functional motifs, with no dramatic changes but some variability in the U5 region; they were clustered with genotypes from European countries but not with those from China, U.S.A., or Japan.

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A vaccine for equine coronavirus (ECoV) is so far unavailable. Bovine coronavirus (BCoV) is antigenically related to ECoV; it is therefore possible that BCoV vaccine will induce antibodies against ECoV in horses. This study investigated antibody response to ECoV in horses inoculated with BCoV vaccine. Virus neutralization tests showed that antibody titers against ECoV increased in all six horses tested at 14 days post inoculation, although the antibody titers were lower against ECoV than against BCoV. This study showed that BCoV vaccine provides horses with antibodies against ECoV to some extent. It is unclear whether antibodies provided by BCoV vaccine are effective against ECoV, and therefore ECoV challenge studies are needed to evaluate efficacy of the vaccine in the future.

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There is currently no available information regarding the veterinary management of Sunda clouded leopards (Neofelis diardi), either in captivity or in the wild. In this study, 12 Sunda clouded leopards were anesthetized between January 2008 and February 2014 for medical exams, and/or GPS-collaring. Seven wild-caught individuals were kept in captivity and 5 free-ranging animals were captured by cage traps. Two anesthesia combinations were used: medetomidine-ketamine (M-K) or tiletamine-zolazepam (T-Z). Atipamezole (0.2 mg/kg im) was used as an antagonist for medetomidine. Medetomidine (range: 0.039–0.054 mg/kg) and ketamine (range: 3–4.39 mg/kg) were administered during 5 immobilizations, resulting in median induction times of 7 min. After a median anesthesia time of 56 min, atipamezole was injected, observing effects of antagonism at a median time of 12 min. T-Z (range: 6.8–10.8 mg/kg) was administered on 7 occasions. Median induction times observed with this combination were shorter than with M-K (4 min vs 7 min; P=0.04), and anesthesia and recovery times were significantly longer (244 and 35 min vs 56 and 16 min, respectively; P=0.02). Lower heart rates were measured in the M-K group, while lower rectal temperatures were found in the T-Z group. Both combinations resulted in safe and reliable immobilizations, although given the favorable anesthesia and recovery times of M-K, we recommend this approach over T-Z for the veterinary handling of Sunda clouded leopards.

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To reveal the reproductive biology in male bottlenose dolphins (Tursiops truncatus), circulating gonadotropins (follicle stimulating hormone [FSH] and luteinizing hormone [LH]) and testicular hormones (testosterone and inhibin) were monitored for 8−12 years in 2 captive bottlenose dolphins (Mars and Regulus). During the study period, Mars was undergoing sexual maturation, whereas Regulus was already mature at the beginning of the study. Assuming that Mars had reached sexual maturity when the significant increase in circulating testosterone levels was observed, serum concentration of inhibin was higher in the sexually immature stage than in the mature stage, whereas the serum concentration of FSH was higher in the sexually mature stage than in the immature stage. No difference was observed in the LH levels between pre- and post-sexual maturation. There was a significant increase in serum concentration of testosterone during spring in both animals. These results suggest that the mechanism responsible for regulating FSH secretion by inhibin functions during the sexually immature stage in this species.

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