In early embryogenesis, the posteroventral foregut endoderm gives rise to the budding endodermal organs including the liver, ventral pancreas and gallbladder during early somitogenesis. Despite the detailed fate maps of the liver and pancreatic progenitors in the mouse foregut endoderm, the exact location of the gallbladder progenitors remains unclear. In this study, we performed a DiI fate-mapping analysis using whole-embryo cultures of mouse early somite-stage embryos. Here, we show that the majority of gallbladder progenitors in 9–11-somite-stage embryos are located in the lateral-most domain of the foregut endoderm at the first intersomite junction level along the anteroposterior axis. This definition of their location highlights a novel entry point to understanding of the molecular mechanisms of initial specification of the gallbladder.

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Brucellosis is a key zoonosis of major public health, animal welfare and economic significance, and is endemic in livestock in Uganda. A cross-sectional epidemiological study was carried out to estimate the sero-prevalence of brucellosis and identify factors associated with sero-positivity in cattle in urban and peri-urban Gulu and Soroti towns of Northern and Eastern Uganda, respectively. A total of 1007 sera and data on biologically plausible risk factors from 166 herds and their spatial locations, were collected from cattle reared in urban and peri-urban Gulu and Soroti towns of Uganda. The sera were analyzed using indirect ELISA and sero-positive reactors confirmed by competitive ELISA. Multivariable models were used to investigate for risk factors. The overall animal-level and herd-level sero-prevalence was 7.5% (76/1007, 95% Confidence Interval (CI): 6.15–9.4%) and 27.1% (45/166, 95% CI: 20.9–34.3%), respectively. Herd-level sero-prevalence was significantly (P<0.001) higher in Soroti than Gulu. In Gulu town, sero-positivity increased with an increase in herd size (P=0.03) and age (P=0.002), and was higher in cattle brought in from western Uganda (P<0.0001). In Soroti town, introduction of new cattle into a herd was significantly (P=0.027) associated with herd sero-positivity. There was a geographically differential risk (clustering) of Brucella sero- positivity in herds in Soroti, while sero-positivity was homogeneously distributed in Gulu. The data highlight brucellosis occurrence and major risk factors for its transmission in cattle in urban and peri-urban areas.

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The genetic organization of the gene involved in the capsular polysaccharide (CPS) biosynthesis of Actinobacillus pleuropneumoniae serotype 14 has been determined. The DNA region for the CPS biosynthesis of serotype 14 (cps14) comprised 9 open reading frames, designated as cps14AB₁B₂B₃CDEFG genes, encoding Cps14A to Cps14G protein, respectively. Cps14A was similar to CpsA of A. pleuropneumoniae serotypes 1, 4 and 12; the Cps14B₁ and Cps14B₂ were similar to CpsB of A. pleuropneumoniae serotypes 1, 4 and 12, suggesting that CPS structure of A. pleuropneumoniae serotype 14 would belong to Group I including A. pleuropneumoniae serotypes 1, 4, 12 and 15. Surprisingly, the overall nucleotide sequence, deduced amino acid sequence, and the genetic organization of the cps14 were nearly identical to those of Actinobacillus suis. This study will provide the molecular basic knowledge for development of diagnostics and vaccine of A. pleuropneumoniae serotype 14.

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The aim of this study was to evaluate the immune responses to intranasal and intrapulmonary vaccinations with the attenuated Mycoplasma hyopneumoniae (Mhp) 168 strain in the local respiratory tract in pigs. Twenty-four pigs were randomly divided into 4 groups: an intranasal immunization group, an intrapulmonary immunization group, an intramuscular immunization group and a control group. The levels of local respiratory tract cellular and humoral immune responses were investigated. The levels of interleukin (IL)-6 in the early stage of immunization (P<0.01), local specific secretory IgA (sIgA) in nasal swab samples (P<0.01); and IgA- and IgG-secreting cells in the nasal mucosa and trachea were higher after intranasal vaccination (P<0.01) than in the control group. Interestingly, intrapulmonary immunization induced much stronger immune responses than intranasal immunization. Intrapulmonary immunization also significantly increased the secretion of IL-6 and local specific sIgA and the numbers of IgA- and IgG-secreting cells. The levels of IL-10 and interferon-γ in the nasal swab samples and the numbers of CD4⁺ and CD8⁺ T lymphocytes in the lung and hilar lymph nodes were significantly increased by intrapulmonary immunization compared with those in the control group (P<0.01). These data suggest that intrapulmonary immunization with attenuated Mhp is effective in evoking local cellular and humoral immune responses in the respiratory tract. Intrapulmonary immunization with Mhp may be a promising route for defense against Mhp in pigs.

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Oncolytic virotherapy is a new strategy for cancer treatment for humans and dogs. Reovirus has been proven to be a potent oncolytic virus in human medicine. Our laboratory has previously reported that canine mast cell tumor and canine lymphoma were susceptible to reovirus. In this study, canine solid tumor cell lines (mammary gland tumor, osteosarcoma and malignant melanoma) were tested to determine their susceptibility towards reovirus. We demonstrated that reovirus induces more than 50% cell death in three canine mammary gland tumors and one canine malignant melanoma cell line. The reovirus-induced cell death occurred via the activation of caspase 3. Ras activation has been shown to be one of the important mechanisms of reovirus-susceptibility in human cancers. However, Ras activation was not related to the reovirus-susceptibility in canine solid tumor cell lines, which was similar to reports in canine mast cell tumor and canine lymphoma. The results of this study highly suggest that canine mammary gland tumor and canine malignant melanoma are also potential candidates for reovirus therapy in veterinary oncology.

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A 9-year-old spayed female cat was examined for cheek skin drainage. The skin lesion did not respond to medical therapy; thereafter, facial deformity developed. A computed tomography revealed an intranasal mass and maxillary osteolysis. The mass was histopathologically diagnosed as suppurative granulomatous inflammation caused by filamentous bacteria. The lesion responded well to radiation therapy. Although actinomycosis was suspected histopathologically, no actinomycetes were detected in the nasal lesion by a bacterial culture conducted at a commercial laboratory. The submandibular lymph node and subcutaneous tissue exhibited swelling. Microbiological examination and genetic analysis based on 16S rDNA gene sequence revealed that Nocardia spp. were isolated from both lesions.

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S100B in cerebrospinal fluid (CSF-S100B) was measured in calves with 20 neurologic and 21 non-neurologic diseases to clarify its utility as a biomarker for neurologic diseases. The median CSF-S100B value in the neurologic disease group (43.0 ng/ml) was significantly higher than that in the non-neurologic disease group (10.2 ng/ml). As CSF-S100B levels in calves with neurologic diseases widely differed, the utility of CSF-S100B as a diagnostic marker for neurologic diseases in cattle remains inconclusive.

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Diabetic patients need particular care in case of infection, digestive disorder or external injury, because external stress often exasperates the glucose metabolism, which is known as “sick day management”. In addition, severe trauma can be a cause of hyperglycemia with insulin resistance. In spite of critical component of the treatment, the precise mechanisms of how trauma develops posttraumatic diabetes remain unknown. Here, we ablated body wall muscles in Drosophila larvae by laser beam and found that the level of trehalose, the principal sugar circulating in the hemolymph or in the tissues of most insects, was increased. The model may provide a helpful tool to understand the relationship between trauma and sugar metabolism.

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Among 16 maras housed as a colony at a zoo, 2 initially showed generalized dermal lesions on the legs, head and abdomen. Approximately 1 month later, following completion of therapy with amitraz, 6 maras in the same colony, including the 2 previously diseased animals, showed dermal lesions with severe alopecia and crusting. Sarcoptic mange was diagnosed on skin scrapings on the basis of morphological criteria. The mites were highly mobile and abundant in all cases, and no other causative agents were detected. Colony-wide treatment with ivermectin and prednisolone was administered weekly for a total of 4 treatments. After therapy was completed in all cohabitants, follow-up scrapings were negative for Sarcoptes scabiei. This report describes the first known outbreak of sarcoptic mange in captive maras and successful treatment with acaricides.

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Context-sensitive half-times (CSHTs) of fentanyl in dogs were determined using pharmacokinetic models reported by Murphy et al. and Sano et al., and compared with a human model. The CSHT was defined as the time required for a 50% decrease in drug concentration in the central compartment after the termination of infusion. Although CSHTs increased gradually as the infusion time increased, the CSHTs in dogs were shorter than those in humans. The CSHTs at steady-state were 31.3 and 69.2 min in dogs, and 306.5 min in humans. The CSHTs of fentanyl in dogs are apparently shorter than those in humans; therefore, a continuous infusion of fentanyl may be a rational regimen in dogs, even if duration of infusion is extended.

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Analysis of L-type amino acid transport expression of hepatocellular carcinoma cells (HCCs) of the dog was performed. The leucine transport activity of canine HCCs was 0.628 ± 0.018 nmol/mg protein/min. The inhibitor of LAT 2-aminobicyclo[2.2.1]heptane-2-carboxylic acid (BCH) reduced 90% of the activity at 1 mM. The deduced amino acid sequences of canine LAT2, LAT3 and LAT4 were well conserved in mammalians, exhibiting 89, 88 and 77% homology, respectively. RT-PCR revealed distinct LAT1 expression compared with normal hepatocytes. Western blotting analysis confirmed the potent LAT1 expression in canine HCCs but not hepatocytes, and real-time RT-PCR analysis indicated that canine HCCs possessed 28 times higher LAT1 expression than hepatocytes. These results indicated that the leucine transport activity of canine HCCs was due to LAT1.

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Although color Doppler ultrasonography has been used to evaluate testicular blood flow in many species, very little has been done in goat. Eight male Shiba goats were exposed to a single intramuscular injection of either gonadotropin-releasing hormone (GnRH group; 1 µg/kg BW) or human chorionic gonadotropin (hCG group; 25 IU/kg BW). Plasma testosterone (T), estradiol (E2) and inhibin (INH) were measured just before (0 hr) and at different intervals post injection by radioimmunoassay. Testis volume (TV) and Doppler indices, such as resistive index (RI) and pulsatility index (PI) of the supratesticular artery, were measured by B-mode and color Doppler ultrasonography, respectively. The results indicated an increase in testicular blood flow in both groups, as RI and PI decreased significantly (P<0.05), but this increase was significant higher and earlier in hCG group (1 hr) than in the GnRH group (2 hr). A high correlation was found for RI and PI with both T (RI, r= −0.862; PI, r= −0.707) and INH in the GnRH group (RI, r=0.661; PI, r=0.701). However, a significant (P<0.05) correlation was found between E2 and both RI (r= −0.610) and PI (r= −0.763) in hCG group. In addition, TV significantly increased and was highly correlated with RI in both groups (GnRH, r= −0.718; hCG, r= −0.779). In conclusion, hCG and GnRH may improve testicular blood flow and TV in Shiba goats.

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To search natural compounds having inhibitory effect on bacterial growth is important, particularly in view of growing multidrug resistant (MDR) strains of bacterial pathogens. Like other bacterial pathogens, MDR Vibrio cholerae, the causative agent of diarrheal disease cholera, is becoming a great concern. As an approach of searching new antimicrobial agents, here, we show that anethole, a well-studied natural component of sweet fennel and star anise seeds, could potentially inhibit the growth of MDR O1 El Tor biotype, the ongoing 7th cholera pandemic variant strains of toxigenic V. cholerae. The minimum inhibitory concentration (MIC) of anethole against diverse O1 El Tor biotype strains is evaluated as 200 µg/ml. Moreover, the effect of anethole is bactericidal and exerts rapid-killing action on V. cholerae cells. This study is the first report which demonstrates that anethole, purified from natural compound, is a potent inhibitor of growth of toxigenic V. cholerae. Our data suggest that anethole could be a potential antimicrobial drug candidate, particularly against MDR V. cholerae mediated infections.

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Fifty-one Salmonella enterica serovar 4,[5],12:i:- (S. 4, [5],12:i:-) isolates (14 human strains, 34 animal strains and 3 river water strains) which are assumed to be monophasic variants of S. Typhimurium were analyzed using pulsed-field gel electrophoresis (PFGE) and multiple-locus variable-number tandem repeat analysis (MLVA) in order to investigate their genetic diversities and relationships. PFGE, MLVA and combination of them identified 28, 27 and 34 profiles (Simpson’s diversity indices [DI]=0.94, 0.96 and 0.97), respectively. No correlations were detected between MLVA clustering and PFGE clustering or phage typing. These results suggested that S. 4,[5],12:i:- originated from multiple S. Typhimurium ancestors. Two cattle and one pig isolates showing identical phage types as well as PFGE and MLVA profiles to human isolates S. 4,[5],12:i:- suggested the existence of the links between human infections and animal reservoirs.

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Laparoscopic cryptorchidectomy without insufflation was applied in 10 standing bulls aged 3 to 15 months. Nine bulls were preoperatively pointed out intra-abdominal testes by computed tomography. Preoperative fasting for a minimum of 24 hr provided laparoscopic visualization of intra-abdominal area from the kidney to the inguinal region. Surgical procedure was interrupted by intra-abdominal fat and testis size. It took 0.6 to 1.5 hr in 4 animals weighing 98 to 139 kg, 0.8 to 2.8 hr in 4 animals weighing 170 to 187 kg, and 3 and 4 hr in 2 animals weighing 244 and 300 kg to complete the cryptorchidectomy. In conclusion, standing gasless laparoscopic cryptorchidectomy seems to be most suitable for bulls weighing from 100 to 180 kg.

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Canine epididymal sperm was collected from the cauda epididymis using 2 different methods (flushing and mincing) to compare the qualities (the percentage of progressively motile, viable, morphologically abnormal, immature and intact acrosomes) before and after freezing and thawing. No significant difference was noted in the quality of the cauda epididymal sperm immediately after collection and after freezing-thawing between the collection methods, although the mean levels of sperm quality with the flushing method were slightly better than that of the mincing method. The flushing method is simple and free of blood contamination, although the vas deferens was too small to be perfused in only 1 dog, and our results suggest that the flushing method is preferable to the mincing method for collecting sperm from the canine cauda epididymis.

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The Exaltation of Newcastle disease virus (END) phenomenon is induced by the inhibition of type I interferon in pestivirus-infected cells in vitro, via proteasomal degradation of cellular interferon regulatory factor (IRF)-3 with the property of the viral autoprotease protein N^pro. Reportedly, the amino acid residues in the zinc-binding TRASH motif of N^pro determine the difference in characteristics between END-phenomenon-positive (END⁺) and END-phenomenon-negative (END⁻) classical swine fever viruses (CSFVs). However, the basic mechanism underlying this function in bovine viral diarrhea virus (BVDV) has not been elucidated from the genomic differences between END⁺ and END⁻ viruses using reverse genetics till date. In the present study, comparison of complete genome sequences of a pair of END⁺ and END⁻ viruses isolated from the same virus stock revealed that there were only four amino acid substitutions (D136G, I2623V, D3148G and D3502Y) between two viruses. Based on these differences, viruses with and without mutations at these positions were generated using reverse genetics. The END assay, measurements of induced type I interferon and IRF-3 detection in cells infected with these viruses revealed that the aspartic acid at position 136 in the zinc-binding TRASH motif of N^pro was required to inhibit the production of type I interferon via the degradation of cellular IRF-3, consistently with CSFV.

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Feline morbillivirus (FmoPV) is an emerging virus that was recently discovered in domestic cats with chronic nephritis. Despite the potential role of FmoPV in chronic nephritis, little is known about its biological characteristics. In this study, we established a quantitative assay of FmoPV by using an indirect immunofluorescence technique. Viral titers of FmoPV were determined in one week. Treatment with polybrene^® or trypsin which was previously used in virus isolation did not augment the virus titers. FmoPV was notably stable at 4°C, retaining high titers for at least 12 days. Heat-treatment at 60°C and 70°C effectively inactivated FmoPV in 10 and 2 min, respectively. The biological characteristics of FmoPV reported here will be beneficial for establishing an efficient virus isolation method and will provide important information to take a measure to reduce the risk of FmoPV infection.

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Pseudorabies virus (PRV) is endemic in some regions of Japan. We investigated the effects of PRV infection status on herd productivity. Serum samples were obtained from 48 swine herds in Japan. Within each herd, three serum samples were obtained from growing pigs at four different ages, as well as from sows in low and high parity groups. Sera were tested for antibodies against wild-type PRV via competitive ELISA. Herds were classified into PRV positive and negative groups based on serological results. Herds infected with PRV exhibited postweaning mortalities (6.84%) that were significantly (P=0.0018) higher than those in unaffected herds (4.73%). Because of the reduced productivity in PRV positive herds, the current PRV eradication program must be strengthened.

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Although the sea otter (Enhydra lutris) is a complete aquatic species, spending its entire life in the ocean, it has been considered morphologically to be a semi-aquatic animal. This study aimed to clarify the unique hindlimb morphology and functional adaptations of E. lutris in comparison to other Mustelidae species. We compared muscle mass and bone measurements of five Mustelidae species: the sea otter, Eurasian river otter (Lutra lutra), American mink (Neovison vison), Japanese weasel (Mustela itatsi) and Siberian weasel (M. sibirica). In comparison with the other 4 species, E. lutris possessed significantly larger gluteus, popliteus and peroneus muscles, but smaller adductor and ischiopubic muscles. The popliteus muscle may act as a medial rotator of the crus, and the peroneus muscle may act as an abductor of the fifth toe and/or the pronator of the foot. The bundles of the gluteus superficialis muscle of E. lutris were fused with those of the tensor fasciae latae muscle and gluteofemoralis muscles, and they may play a role in femur abduction. These results suggest that E. lutris uses the abducted femur, medially rotated crus, eversion of the ankle and abducted fifth digit or extended interdigital web as a powerful propulsion generator. Therefore, we conclude that E. lutris is a complete aquatic animal, possessing differences in the proportions of the hindlimb muscles compared with those in other semi-aquatic and terrestrial mustelids.

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Prevalence of drug-resistant bacteria in wildlife can reveal the actual level of anthropological burden on the wildlife. In this study, we isolated two multiple drug-resistant strains, GG6-2 and GG6-1-1, from 27 fresh feces of wild western lowland gorillas in Moukalaba-Doudou National Park, Gabon. Isolates were identified as Achromobacter xylosoxidans and Providencia sp., respectively. Minimum inhibitory concentrations of the following 12 drugs—ampicillin (ABPC), cefazolin (CEZ), cefotaxime (CTX), streptomycin (SM), gentamicin (GM), kanamycin (KM), tetracycline (TC), nalidixic acid (NA), ciprofloxacin (CPFX), colistin (CL), chloramphenicol (CP) and trimethoprim (TMP)—were determined. Isolate GG6-2 was resistant to all antimicrobials tested and highly resistant to CTX, SM, TC, NA and TMP. Isolate GG6-1-1 was resistant to ABPC, CEZ, TC, CL, CP and TMP.

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