Foot-and-mouth disease (FMD) occurred recently for the first time in a decade in Japan. The index case was detected on a beef-breeding farm in Miyazaki Prefecture, Southern Japan, on April 20, 2010. After confirmation of this first case, control measures such as stamping out, movement restriction and disinfection were implemented. However, these strategies proved insufficient to prevent the spread of FMD and emergency vaccination was adopted. Up until the last outbreak on July 4, 2010, a total of 292 outbreaks had been confirmed, with about 290,000 animals having been culled. The epidemic occurred in an area with a high density of cattle and pigs, making disease control difficult. Invasion of the disease into a high-density area aided its rapid spread and led to difficulties in locating suitable burial sites. Epidemiological investigations indicated that the disease was introduced into Japan approximately one month before detection. This delay in initial detection is considered to have allowed an increased number of outbreaks in the early stage of the epidemic. Nevertheless, the epidemic was contained within a localized area in Miyazaki Prefecture and was eradicated within three months because of intensive control efforts including emergency vaccination. Although this epidemic devastated the livestock industry in Japan, many lessons can be learnt for the future prevention and control of infectious diseases in animals.

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The distribution and population of immunocompetent cells in bovine hemal node, mesenteric lymph node and spleen were analyzed comparatively by immunohistochemistry and flow cytometry. Many CD8⁺ cells, CD172a⁺ cells and γδ T cells were found in the lymphatic cord along the sinus of the hemal node and the splenic red pulp. A few CD8⁺ cells and γδ T cells were distributed diffusely in the paracortex and medullary cord of the mesenteric lymph node. Many germinal centers were recognized in the lymphatic regions such as the cortex and white pulp of these lymphoid organs. The populations of CD8⁺ cells and γδ T cells in the hemal node and the spleen were higher than those of the mesenteric lymph node. In addition, the populations of CD21⁺ cells and MHC class II⁺ cells in the hemal node and the mesenteric lymph node were higher than those of the spleen. The results suggest that the hemal node has an important role in both cellular and humoral immunity as well as the lymph node and the spleen in cattle.

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The aim in this study is to elucidate the laterality of chicken spinocerebellar (SC) neurons that originate from the caudal cervical to caudal lumbosacral spinal cord. SC neurons in the spinal segment (SS) 17-20 consisted of a mixture of crossed and uncrossed axons. SC neurons in the more cranial and caudal SS than SS 17-20 (transitional zone) were generally uncrossed and crossed, respectively. In the transitional zone, SC neurons in spinal border cells and ventral border cells of the ventral horn changed dramatically from an uncrossed to a crossed type between SS 17 and SS 18. Chicken SC neurons are markedly different in laterality from mammalian SC neurons.

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Atrial (A-type) natriuretic peptide (ANP) is vasodilative hormone involved in the regulation of blood pressure and volume homeostasis. In this study, we examined the differences of the auricular and plasma ANP distribution by immunohistochemistry, ultrastructural morphometry, and radioimmunoassay in five strains of mice. The ANP-immunoreactivities of the auricle were most intense in ICR, and moderate in C57BL/6J and C3H/HeN, and weakest in BALB/cA and DBA/2Cr. The number of ANP-granules was greatest in ICR followed by C57BL, C3H or BALB/c, and DBA/2 mice, in this order. The auricular ANP content was highest in ICR, but the plasma ANP concentration was comparable in all strains. The present study demonstrates that there are differences in the ANP circulating system between five strains.

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A total of 516 slaughter-age ostrich sera were collected in Japan during 2006-2009. Sixty-one of five hundred and sixteen were positive by virus neutralization (VN) test and the titer of most positive samples was low level. Within the 61 positive sera, 35 sera were collected from unvaccinated ostriches. This result implies that these ostriches might have been infected naturally with low-virulent Newcastle disease virus (NDV). Within the 455 negative samples, 125 samples were from vaccinated ostriches. Since ostrich farmers use live attenuated vaccines, it is reasonable that the titer decreased to below detection level by 1 or 1.5 year-old. The above data indicate that NDV has infiltrated into ostrich farms in Japan, and that the efficacy of ostrich ND vaccination is often time-limited.

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The aim of this study was to determine antimicrobial resistance of Aeromonas hydrophila isolated from farmed Nile Tilapia. A total of 50 A. hydrophila isolates from clinical cases were screened for the presence of class 1, 2 and 3 integrons and all the strains resistant to enrofloxacin and/or ciprofloxacin (n=19) examined for mutation in the quinolone resistance-determining regions (QRDRs) of gyrA and parC. The intI1 gene was detected in 23 A. hydrophila strains (46%) but no intl2 and intl3 were detected. Among these, 14 isolates (60.8%) carried gene cassettes inserted in variable regions i.e., partial aadA2, aadA2, dfrA1-orfC and dfrA12-aadA2, of which the most common gene cassette array was dfrA12-aadA2 (26.09%). Conjugal transfer of class 1 integrons with resistance gene array was detected. All the A. hydrophila strains resistant to enrofloxacin and/or ciprofloxacin possessed mutations in the QRDRs of gyrA and parC. Only a Ser-83-Ile substitution was identified in GyrA and only a Ser-80-Ile amino change was found in ParC. The data confirms that A. hydrophila from farm-raised Nile Telapia serve as a reservoir for antimicrobial resistance determinants.

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Contagious equine metritis (CEM), a contagious venereal disease of horses, invaded Japan in 1980 and spread in the Thoroughbred population of the Hidaka-Iburi district of Hokkaido. To eradicate CEM, we ran a program aimed at detecting Taylorella equigenitalis, the causal agent, in carrier horses by using the PCR test, followed by culling or treatment. In 2001, the first year of the program, 12,356 Thoroughbred racing stallions and mares were tested and 11 carriers were found. Four, two, one, and one carrier mares were detected in 2002, 2003, 2004, and 2005, respectively, by application of the program at the same scale as in 2001. No PCR-positive horses were found from 2006 to 2010. These results strongly suggest that CEM was eradicated from Japan by 2010.

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To evaluate the influence of mating behavior on cardiac function, changes in heart rate (HR), electrocardiogram (ECG), hematocrit (Hct) and serum concentration of alpha-atrial natriuretic peptide (alpha-ANP) were evaluated in 10 clinically sound Thoroughbred stallions before and after mating behavior. The stallions were submitted twice to experimental pseudomating in the same month in 2009 and 2010. Measurements and blood samples were collected at a stable before mating (baseline) and at a covering yard before and after mating. ECG was recorded by a Holter-ECG system. Arrhythmias were detected in 5 stallions before or after mating behavior. Minimum HR (HRmin), maximum HR (HRmax) and HR recorded when the stallions entered into yard (HRent) and ejaculated (HRejc) were 34.2 ± 3.7, 168.9 ± 14.2, 141.8 ± 35.3 and 142.6 ± 27.3 beats/min, respectively. Time from entrance into the yard to ejaculation (mating time; MT) ranged from 30 to 2,103 sec and was highly correlated with HRent (r=-0.82) and the time required for attaining HRmax after entrance into the yard (dT HRmax) (r=0.87). Hct and serum alpha-ANP concentration significantly increased after ejaculation (60.0 ± 3.2%, P<0.0001, and 1.54 ± 0.61 ng/ml, P=0.0353) compared with the baselines values (46.9 ± 4.4%, 1.40 ± 0.60 ng/ml). HRent and Hct were significantly higher in the stallions with an MT of less than 5 min (n=5) compared with those (n=5) with an MT of more than 5 min (P=0.0324 and P=0.0082). Mating behavior increases the workload of the heart in Thoroughbred stallions.

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Dose responses of plasma calcitriol, calcium (Ca), bone metabolic markers and glomerular filtration rate (GFR) were evaluated in four nonpregnant Holstein cows treated subcutaneously with an aqueous formulation of calcitriol at four doses in a 4 × 4 Latin-square design. Calcitriol, Ca, and markers of bone metabolism were analyzed in plasma samples. GFR was measured in predose and day 5 samples. Plasma calcitriol and Ca concentrations increased dose-dependently. The calcitriol dose was positively correlated with the area under the concentration-time curve of plasma calcitriol. Bone formation markers tended to increase from day 3 onward for all doses. No significant changes in GFR were noted. Thus, exogenous calcitriol administered between 0.0625 and 0.5 μg/kg body weight elicited dose-dependent increases in both plasma calcitriol and Ca and elevated bone formation markers without affecting renal function in nonpregnant cows.

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The detection of small amounts of free peritoneal gas in the canine patient can pose a diagnostic dilemma. The objective of this study was to determine how much of this free gas could be detected ultrasonographically. Ultrasound examinations were carried out after increments (0.1 ml) of air were intraperitoneally injected. Via ultrasonography, 0.4 ml of free gas can be reliably detected. The authors concluded that ultrasonography is a very sensitive modality for the detection of small amounts of peritoneal free gas (above 0.4 ml).

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Endoscopic polypectomy and argon plasma coagulation (APC) were performed in a refractory case of inflammatory colorectal polyps in a 7-year-old male Miniature Dachshund. Colonoscopic examination revealed a large sessile polyp and multiple diffuse small polyps, localized to the descending colon and rectum. The case showed a poor therapeutic response to prednisolone and cyclosporine. Under anesthesia, piecemeal resections were performed by polypectomy. APC was carried out to cauterize the polyp remnants. After treatment, reduction of the lesions and the improvement in clinical signs were observed, without recurrence of lesions for at least 10 months. Endoscopic treatment by polypectomy and APC is suggested to be a therapeutic option for refractory cases of inflammatory colorectal polyps in dogs.

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This study was carried out to investigate the presence of Torque teno sus virus types 1 (TTSuV1) and 2 (TTSuV2) in a longitudinally (14 to 150 days of age) collected paired pooled sera (pSE) and peripheral blood mononuclear cells (pPBMCs) using nested polymerase chain reaction. The detection rate of TTSuV1 in pSE increased from 14 to 90 days of age, but a progressive decline was observed from 120 to 150 days of age, while in pPBMC, a high value was maintained till the end of growing-finishing period. On the contrary, except in PBMCs at 30 days of age, high detection rates of TTSuV2 were found in both pSE and pPBMCs in all sampling ages. The detection rate of TTSuVs between pSE and pPBMCs was positively correlated at all sampling ages except for TTSuV1 at 150 days of age. This is the first study showing the presence of TTSuVs in PBMCs from pigs and describing the in vivo infection dynamics of TTSuV in paired sera and PBMCs during the entire growing and finishing periods of pigs reared in conventional farms.

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Circadian pH changes in the fluid of the rumen (bottom and middle) and reticulum were assessed simultaneously using wireless and wired radio-transmission pH-measurement systems in cows fed a control diet (C diet) or rumen-acidosis-inducing diet (RAI diet). The pH in the three sites decreased following the morning and evening feedings. In cows fed the C diet, the bottom-rumen and reticular pH reverted to the basal level by the next morning, while the middle-rumen pH did not recover completely, suggesting that active fermentation occurred in the middle of the rumen. The mean pH at 1 hr intervals was higher in the reticulum than at the bottom and in the middle of the rumen. The relatively stable reticular pH may result from dilution due to salivation. In cows fed the RAI diet, the bottom-rumen pH fell to approximately 5.2 after the evening feeding, but returned to the basal level by the next morning. In contrast, the middle-rumen pH did not return to the basal level (6.5) within 24 hr, presumably owing to continuous, vigorous fermentation. There were positive correlations between the pH at the bottom and in the middle of the rumen and at the bottom of the rumen and in the reticulum. These findings indicate that our radio-transmission pH-measurement system may be suitable tool for simultaneous measurement of pH in the rumen and reticulum fluid.

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The objectives of this study were to characterize plasma lipid phenotypes and dissect the genetic basis of plasma lipid levels in an obese DDD.Cg-A^y mouse strain. Plasma triglyceride (TG) levels were significantly higher in the DDD.Cg-A^y strain than in the B6.Cg-A^y strain. In contrast, plasma total-cholesterol (CHO) levels did not substantially differ between the two strains. As a rule, the A^y allele significantly increased TG levels, but did not increase CHO levels. Quantitative trait locus (QTL) analyses for plasma TG and CHO levels were performed in two types of F₂ female mice [F₂A^y (F₂ mice carrying the A^y allele) and F₂ non- A^y mice (F₂ mice without the A^y allele)] produced by crossing C57BL/6J females and DDD.Cg-A^y males. Single QTL scan identified one significant QTL for TG levels on chromosome 1, and two significant QTLs for CHO levels on chromosomes 1 and 8. When the marker nearest to the QTL on chromosome 1 was used as covariates, four additional significant QTLs for CHO levels were identified on chromosomes 5, 6, and 17 (two loci). In contrast, consideration of the agouti locus genotype as covariates did not detect additional QTLs. DDD.Cg-A^y showed a low CHO level, although it had Apoa2^b, which was a CHO-increasing allele at the Apoa2 locus. This may have been partly due to the presence of multiple QTLs, which were associated with decreased CHO levels, on chromosome 8.

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Tritrichomonas suis (=T. foetus) has recently been reported to be a causative agent of chronic large-bowel diarrhea in cats. While the disease was previously attributed to Pentatrichomonas hominis, the etiologic agent for feline trichomonal diarrhea was identified as T. suis. Although feline trichomonosis due to T. suis has been reported at prevalences ranging from 14 to 31% in Europe and the U.S., no reports of the pathogen have been published to date in Japan. In 2008, however, we encountered a case of feline trichomonosis at the Veterinary Teaching Hospital of Hokkaido University. The parasite was identified as T. suis by nested PCR amplification of partial internal transcribed spacer region 1 and 5.8S ribosomal RNA gene sequences with T. suis-specific primers and DNA sequencing of the amplified products. We then conducted surveys for feline trichomonosis in three different animal hospitals using either cultivation and/or PCR-based assays. The results revealed that 13 of 147 samples (8.8%) were positive for T. suis, and that 5 of the 13 infected cats, which ranged between 1 month and 7.5 years-old, showed chronic diarrhea. Seven of the infected cats were purebred and 6 were mixed breed. These findings suggested that feline trichomonosis is prevalent in Japan, and that T. suis may play a role as a causative agent of feline chronic diarrhea.

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Curcumin is a constituent phenol compound of turmeric, and has been used as a dietary spice and Indian medicine. Curcumin has been reported to inhibit the formation of amyloid β fibrils and aggregation. In this study, the binding activity of curcumin to various types of canine amyloid was examined. Tissue samples used were lesions of AA, AL, amyloid of canine amyloid-producing odontogenic tumor (Aapot), and senile cardiovascular amyloid (ScA). Curcumin stained all types of amyloid. The binding of curcumin to AA, ScA, and AL was lost by the KMnO₄ treatment, but Aapot maintained the binding. These findings indicate that curcumin binds several types of amyloid, while the binding sites of amyloid molecules might be different from that of Congo red.

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Salivary gland cysts are often concurrent with GM1 gangliosidosis in Shiba dogs. Although the etiology is unknown, these cysts may be misdiagnosed as malignant due to the accumulation of foamy cells. The present study investigated the cytological, histopathological, immunohistochemical and electron microscopic characteristics of salivary gland cysts in a Shiba dog affected with GM1 gangliosidosis. The salivary gland masses were surgically enucleated and examined clinicopathologically and pathologically in a 7-month-old Shiba dog with GM1 gangliosidosis. Many large cells with rich cytoplasm including vacuoles of various sizes, i.e., foamy cells, were observed in stamp smears from the cut-surface of the masses and histopathologically in major parts of the cyst wall. Some of these foamy cells presented features similar to a spider-web appearance. The foamy cells were confirmed to have originated from macrophages based on marked immunohistochemical expression of vimentin, HLA-DR, lysozyme and Iba1. An ultrastructural study demonstrated electron-dense vesicular structures in the vacuolated cells. Therefore, the masses were diagnosed pathologically as benign salivary gland cysts with accumulation of foamy cells. In conclusion, the histopathological features of the salivary gland cysts in this Shiba dog were similar to those of lipoma and/or liposarcoma. In such cases, immunohistochemical and ultrastructural examinations were useful in the differential diagnosis. Practitioners, clinical pathologists and pathologists should take GM1 gangliosidosis into consideration when they encounter salivary gland cysts in Shiba dogs.

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A free-living European red deer calf (Cervus elaphus) was euthanized due to bilateral microphthalmia. Lens was missing, replaced by proliferating squamous epithelial cells; hyperplastic squamous cells, sebaceous and mucinous glands were observed within the cornea with the characteristics of inclusion cyst. Findings were consistent with congenital microphthalmia/aphakia, with multiple eye abnormalities.

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Photodynamic hyperthermal therapy (PHT) with indocyanine green (ICG) is a combination of photodynamic therapy (PDT) and hyperthermia (HT). The low toxicity of ICG with an absorption wavelength of 700-800 nm is thought to make it a good candidate as a photosensitizer for PHT. Upon irradiation, ICG produces oxygen radicals and generates heat. The optimal concentration of ICG and the PHT post-irradiation time effects were evaluated by the cytotoxicity of the treatment on B16F10 murine melanoma. The cytotoxicity of PHT was determined based on the morphology of apoptotic and necrotic cells under phase-contrast microscope, confocal laser scanning microscope (CLSM) with DAPI and Annexin V-FITC/PI double staining, and cell surface structure evaluation by scanning electron microscopy (SEM). The use of ICG at a concentration of 150 μM was selected, as cell proliferation was inhibited from 0 to 24 hr post-PHT with a 3-fold decrease in cell viability (P<0.001) compared to the control group. A morphological observation revealed apoptotic and some degree of necrotic features in the PHT-treated cells.

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It is well known that heart rate or arterial blood pressure may increase in response to surgical stimulation despite the absence of a purposeful movement. However, there is limited information regarding anesthetic requirement for blunting adrenergic response in dogs. This study was designed to compare the minimum alveolar concentrations of sevoflurane required to prevent autonomic response (MAC-BAR) and purposeful movement (MAC) in dogs. Sevoflurane MAC-BAR and MAC were determined in 5 beagle dogs by judging dogs' response to a noxious electrical stimulus applied to the gingiva. The sevoflurane MAC-BAR was significantly higher than MAC (3.33 ± 0.48 vs 2.10 ± 0.28%, P=0.005). These results suggested that autonomic responses occurred at sevoflurane anesthetic concentrations at which purposeful movements were absent.

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Cartilage-derived retinoic acid-sensitive protein (CD-RAP)/melanoma inhibitory activity (MIA), which appears abundantly in hypertrophic cartilage at the stage of endochondral ossification, is also detected in cerebrospinal fluid (CSF) following spinal cord injury. In this study, the localization of the CD-RAP/MIA molecule in normal tissues of the spine and brain obtained from mice, rats, dogs, cattle and horses was examined using immunohistochemistry with a specific antibody. The positive signals of CD-RAP/MIA were found at nerve cells in the spinal cords of all species and were especially strong at cerebellar Purkinje cells. The results suggested that CD-RAP/MIA included in normal cerebrospinal tissues could be a biomarker associated with tissue injuries, as the molecules might flow into the CSF.

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The objective of this study was to evaluate the effect of electrical stimulation (EST) on pronuclear formation, chromosomal constitution, and developmental capability among in vitro matured pig oocytes following intracytoplasmic sperm injection (ICSI). After ICSI, the oocytes were randomly distributed and cultured into 3 groups: the EST activated ICSI group, non-activation ICSI group, and in vitro fertilization (IVF) group. The proportion of oocytes in which 2 pronuclei were formed in ICSI groups was significantly higher in the former groups than in the IVF group (96.2 and 93.5 vs. 64.5%, respectively, P<0.05). The cleavage rate was significantly higher in EST activated ICSI group (78.6%) than in the IVF and non-activated ICSI groups (51.8 and 46.0%, respectively, P<0.05), as was the proportion of oocytes that developed to the blastocyst stage at day 7 (18.9 vs. 11.6 and 9.1%, respectively, P<0.05). Diploid blastocysts were observed in 52.4, 63.0, and 65.2% of oocytes in the IVF, activated, and non-activated ICSI groups, respectively. Eight out of 23 gilts (34.8%) were confirmed to be pregnant in activated ICSI groups, but none of these pregnancies were carried to term. These results show that oocyte activation after ICSI is effective in elevating the cleavage rate and blastocyst development, while ensuring normal chromosome composition. Further research is needed to determine the pregnancy maintenance requirements for ICSI-embryos in pigs.

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In peripartum dairy cows, insulin resistance (IR) increases to adjust the direction of energy to lactation after calving. To investigate the effect of prepartum IR on postpartum reproductive performance, the insulin tolerance test (ITT) was applied to 15 cows at 3 weeks (Pre21) and 10 days (Pre10) before the predicted calving date. Blood glucose area under the curve (AUC_glu) within 120 min after administration of 0.05 IU/kg-BW insulin was calculated. The occurrence of first ovulation, days to first artificial insemination (AI) and first AI conception rate were recorded. Nutritional status postpartum was evaluated by blood chemical analysis. Based on AUC_glu changes from Pre21 to Pre10, cows were classified into either the AUC-up group (AUC_glu increase, n=5) or the AUC-down group (AUC_glu decrease, n=10). There was no difference in the decrease in blood glucose at 30 min after insulin injection between groups, although glucose recovery from 30 to 60 min during the ITT was slow at Pre10 in the AUC-up group. The AUC-up group had a higher number of days to first AI and high glucose, total protein, globulin, γ-glutamyltransferase, triacylglycerol levels and a low albumin-globulin ratio at the 14th day postpartum. The present study infers that prepartum slow glucose recovery rather than insulin sensitivity might increase the potential for subclinical health problems postpartum and thus suppress reproductive performance. During the prepartum transition period, glucose dynamics in the ITT can be considered as a new indicator for the postpartum metabolic status and reproductive performance of dairy cows.

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We investigated the influence of Borna disease virus (BDV) infection on the clinical state of dairy cows. Sera from 149 cows were examined using enzyme-linked immunosorbent assay and western blotting detect antibodies to the BDV-nucleoprotein antigen. Among 149 investigated cows, 25 (16.8%) showed a positive reaction to BDV antigen. No significant difference existed in milk production or medical history between seropositive and seronegative cows. Although the estrus cycle appeared normal even in the seropositive cows, the frequency of artificial insemination and calving-to-conception intervals significantly increased in seropositive cows. Therefore, fertilization failure was recognized in the BDV-antibody positive cows.

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H9N2 influenza viruses circulate in wild birds and poultry in Eurasian countries, and have been isolated from pigs and humans in China. H9N2 viruses isolated from birds, pigs and humans have been classified into three sublineages based on antigenic and genetic features. Chicken antisera to H9N2 viruses of the Korean sublineage reacted with viruses of different sublineages by the hemagglutination-inhibition test. A test vaccine prepared from a non-pathogenic A/duck/Hokkaido/49/1998 (H9N2) strain of the Korean sublineage, obtained from our influenza virus library, induced immunity in mice to reduce the impact of disease caused by the challenge with A/Hong Kong/1073/1999 (H9N2), which is of a different sublineage. The present results indicate that an inactivated whole virus vaccine prepared from a non-pathogenic influenza virus from the library could be used as an emergency vaccine during the early stage of a pandemic caused by H9N2 infection.

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