The objective of this study was to evaluate the effects of post artificial insemination (AI) treatment with intravaginal progesterone device (P4 device) on conception rate, synchronization of returning estrus and plasma P4 concentration in Japanese Black cows. Nineteen cows were treated with DIB (1.0 g P4) from Day 12 to 19 (Day 0=day of the first AI), 27 cows were treated with a CIDR (1.9 g P4) from Day 12 to 19, and 33 cows were not treated after the first AI (control). Estrous behavior was daily examined between Day 20 and 25, and cows returning to estrus were inseminated (the second AI). On Day 19, plasma P4 concentration was not different among DIB, CIDR and control groups. There was no significant difference in conception rate after the first AI among three groups (DIB: 63.2%, CIDR: 66.7% and control: 72.7%). In non-pregnant cows, there was no significant difference in the proportion of cows showed returning estrus between Day 20 and 25 (DIB: 57.1%, CIDR: 22.2% and control: 44.4%), and day of returning estrus was not synchronized. The overall conception rate after the first and second AI was not different among the groups. In conclusion, post-AI treatment with intravaginal devices containing 1.0 and 1.9 g P4 from Day 12 to 19 neither increased plasma P4 concentration nor improved fertility and synchronization of the returning estrus in Japanese Black cows.
The Sunda pangolin (Manis javanica) population in Southeast Asia faces threats such as poaching and deforestation. Health assessments of rescued individuals including physical examination and blood work are crucial for clinicians to determine the health status of these animals. The establishments of reference intervals of hematology and serum biochemistry are important for identifying clinical abnormalities. The objectives of our study were to establish blood reference intervals for Sunda pangolins, to determine if there are age and sex related differences in hematology and serum biochemistry, and to compare our results with those of a previous study on confiscated Sunda pangolins in Thailand. Fifty-eight Sunda pangolins were rescued between January 2011 and December 2015. The hematology and serum biochemistry results of 51 clinically normal Sunda pangolins were selected for the establishment of the blood reference intervals. No sex related differences were noted in this study. Age-related differences were observed, in which adult Sunda pangolins had a significantly higher mean corpuscular volume than juveniles, and juvenile Sunda pangolins had significantly higher red blood cell counts and hemoglobin levels than those of the adults (P<0.05). Age-related differences were also noted in several serum biochemistry parameters: alkaline phosphatase (ALP) was significantly higher in juveniles, and total protein was significantly higher in adult Sunda pangolins. Compared to a previous study the white blood cell counts, neutrophil counts, and ALP were higher, and the lymphocyte counts were lower in the present study.
The present study describes the association between inflammatory cell types and feline infectious peritonitis virus (FIPV) antigen in the brain of 4 cats diagnosed as feline infectious peritonitis (FIP). Immunohistochemically, FIPV antigens were detected in the inflammatory foci of the leptomeninges, choroid plexus and ventricles in 3 of the 4 cats. In 3 cases, inflammatory foci mainly consisted of CD204- and Iba1-positive macrophages, and the FIPV antigens were found in the macrophages. In the other case which was negative for FIPV antigen, severe inflammation predominantly consisting of CD20-positive B lymphocytes was observed in the leptomeninges and subventricles, accompanied with diffuse proliferation of gemistocytic astrocytes. The difference in histopathology may reflect the inflammatory process or the strain variation of FIP virus.
Recently, we reported that follicle-sinus complexes (FSCs) in the muzzle skin are useful for postmortem diagnosis of rabid dogs. Here, we compared the sensitivity and specificity of detecting the viral antigen in the brain and FSCs of 226 suspected rabid dogs, and assessed whether the FSC harbored the virus genome and particles. The viral antigen was detected in 211 of 226 samples with 100% sensitivity and specificity. Viral RNA and particles were observed in the cytoplasm of Merkel cells (MCs). These results suggest that MCs are targets of virus infection and FSCs are useful material for diagnosing rabies.
Asian and African elephants are frequently afflicted by foot disorders that can be very challenging to manage even with aggressive therapy. Such conditions may have indirect life-threatening effects. Mohs' paste (zinc chloride based escharotic agent) was used to treat a female Indian elephant (Elephas maximus indicus) aged 39 years with foot disorder at Kanazawa Zoological Gardens. Degenerated hyperplastic tissue was observed inside the hoofs of digits 2 and 5. Mohs' paste was applied on the lesions, which coagulated the hyperplastic tissue and restrained its proliferation. Subsequently, the hyperplastic tissue could be trimmed with little pain, and the disorder became manageable. Mohs' paste treatment was effective and is expected to be an alternative treatment for hoof disorder.
Insulin degludec (IDeg) is a new insulin formulation that facilitates long-term control of glucose level in humans. In this study, we investigated the effects of IDeg on glycemic control in dogs. Its time-action profiles were monitored in healthy dogs using an artificial pancreas apparatus under euglycemic conditions. At 9.0–13.5 hr post-IDeg injection, an indistinct peak of glucose level was detected. Moreover, the action of IDeg was persistent for >20 hr. Both IDeg and neutral protamine Hagedorn insulin (NPH) lowered blood glucose concentrations in diabetic dogs, but IDeg caused postprandial hyperglycemia and a somewhat lower preprandial glucose level than that caused by NPH. IDeg might be ineffective in concurrently preventing postprandial hyperglycemia and preprandial hypoglycemia in a single-agent administration.
Canine nasal carcinomas are often treated with radiotherapy. Presence of lysis of the cribriform plate by tumor invasion (stage 4 by modified Adams’s staging system) is a well-known prognostic factor. In this study, dogs with stage 4 disease were divided into two subgroups based on the presence or absence of midline shift of the olfactory or frontal lobes of the brain (Stage 4a: without presence of midline shift. Stage 4b: with midline shift). The median survival time of dogs with midline shift was significantly shorter than that of dogs without midline shift (64 days vs. 208 days). Our results indicate that the finding of a midline shift might have a prognostic significance in dogs with nasal carcinoma treated with radiotherapy.
Bovine tuberculosis (bTB) control programs generally rely on intradermal tuberculin tests for the antemortem diagnosis of Mycobacterium bovis infection in cattle, but these tests detect only a portion of the infected animals. The aim of the present study was to evaluate the diagnostic coverage of a combination of the bTB antemortem techniques known as the comparative intradermal tuberculin test (CITT) and an ELISA based on a recombinant chimera of ESAT-6/MPB70/MPB83 as the antigen in cattle. The results were compared to postmortem findings based on M. bovis culturing and PCR. Paired comparisons of all data (n=92) demonstrated that ELISA and LST results compared to the culturing results did not present significant differences (P=0.27 on McNemar’s test and P=0.12 on Fisher’s exact test, respectively). Using culturing as the gold standard, the sensitivity and specificity of ELISA were 79.5% (95% CI: 64.5–89.2%) and 75.5% (95% CI: 62.4–85.1%), respectively, whereas LST demonstrated 100% sensitivity (95% CI: 91.03–100%) and 92.5% specificity (95% CI: 82.1–97.0%). The ELISA results did not reveal significant differences in relation to the LST results (P>0.99 on Fisher’s exact test). Using the latter as the gold standard, the sensitivity and specificity of ELISA were 79.1% (95% CI: 64.8–88.6%) and 79.6% (95% CI: 66.4–88.5%), respectively. The use of ELISA with the recombinant chimera of ESAT-6/MPB70/MPB83 as the antigen complements the diagnostic coverage provided by CITT and increases the removal of infected animals from herds.
This study assessed the advantages of dextrose and amino acid mixture solution as parenteral nutrition (PN) therapy for diarrheic calves. Thirty diarrheic calves were randomly assigned to receive PN (PN group, n=15) or only dextrose solution (Dex group, n=15). The treatment period for the PN group (4.0 days; min-max, 2–10 days) was significantly shorter than that for the Dex group (6.0 days; min-max, 3–21 days) (P<0.01). The PN therapy tended to improve plasma diamine oxidase activity compared with traditional therapy. One potential association between PN therapy and shortened treatment period may be the repair of damaged intestinal villi. Although our proposal has limitations, PN therapy suggested the potential for new treatment of diarrheic calves.
Plasma luteinizing hormone (LH) and testosterone concentrations were examined in Japanese Black beef bulls with normal and abnormal semen in response to gonadotropin releasing hormone (GnRH) challenge at the start (10 months) and completion (20 months) of puberty. Bulls with normal semen had higher testosterone concentrations after GnRH treatment at 20 months than they did at 10 months, while LH concentrations did not differ between the two age groups. LH and testosterone concentrations were not different between bulls with normal and abnormal semen at 20 months. Thus, testosterone secretions in response to the GnRH challenge were higher for bulls with normal semen at pubertal completion compared to bulls at the start of puberty, but responsiveness of LH to GnRH and of testosterone to the LH increment was not altered in bulls with abnormal semen.
Muscovy duck parvovirus (MDPV) and goose parvovirus (GPV) have both been found to cause high mortality and morbidity in Muscovy ducklings. Specific detection is often rife with false positives due to high identity at the genomic nucleotide level and antigenic similarity between MDPVs and GPVs. In this study, significantly variable regions were found, via NS comparison, between MDPV and GPV NS genes; however, NS genes were conserved within the MDPV and GPV groups. A polymerase chain reaction (PCR) assay for detecting and differentiating MDPVs and GPVs was developed with more specificity based on the NS gene characterization. The assay detected as low as 103 DNA copies of both the MDPV and GPV strains, along with 549 separate base pairs (bp). No bands of the same size from other duck pathogens, including duck circovirus, duck enteritis virus, egg drop syndrome virus, duck-origin goose hemorrhagic polyomavirus, Escherichia coli, Salmonella, Riemerella anatipestifer and Pasteurella multocida were amplified. This indicates that this method for performing PCR provides a useful and reliable alternative tool for more precise differentiation of MDPV and GPV infection in clinical samples.
The control of inflammation, which arises from complex biological responses to harmful stimuli, is an important determinant of both clinical outcomes and patient comfort. However, the side effects of many current therapies such as non-steroidal anti-inflammatory drugs mean that new safe treatments are required. We previously reported that 12.5 μg/ml hydroxytyrosol (HT) suppressed gene expression of the inducible nitric oxide (NO) synthase (iNOS) isoform and NO production, in mouse peritoneal macrophages treated with lipopolysaccharide (LPS), where nuclear factor-κB (NF-κB) gene expression was not altered. The present study evaluated the anti-inflammatory effects of various concentrations of HT in LPS-induced RAW264.7 mouse macrophages. HT suppressed NF-κB signaling and downregulated LPS-mediated expression of iNOS, cyclooxygenase-2, tumor necrosis factor alpha, and interleukin-1β at 12.5 μg/ml, resulting in reduced production of NO and prostaglandin E2. At lower concentrations, HT seemed to act via another signaling pathway to regulate the inflammatory response. In contrast, HT did not suppress LPS-induced expression of phosphorylated p44/42 mitogen-activated protein kinase. This study showed that HT had anti-inflammatory effects on LPS-stimulated RAW264.7 cells. HT is already available as a nutritional supplement and no toxic effects have been reported. Hence, HT represents a potential novel anti-inflammatory agent.
Toxoplasma gondii is a highly prevalent protozoon that can infect all warm-blooded animals, including humans. It is frequently used as an Apicomplexan parasite model in research. In this review, the invasion mechanism of T. gondii is described as a representative Apicomplexan parasite. The invasion machinery of T. gondii consists of the moving junction and the glideosome, which is a specific motor system for Apicomplexan parasites. I provide details about the moving junction, parasite-secreted proteins and host adhesion receptors, the glideosome, and calcium signaling, which generates the power for the gliding mobility of T. gondii. A detailed understanding of parasite invasion can be useful for the development of new effective drugs to inhibit this event and disrupt the Apicomplexan life cycle.
A rapid risk assessment was conducted using a questionnaire composed of 10 questions asking experts in African swine fever (ASF) to identify and rank the potential risk factors associated with the introduction and spread of ASF in Japan. The experts participating in this risk assessment considered illegal food import, followed by transport routes and foreign workers, to be the most relevant pathway of ASF introduction into Japan. Kanto and Kyushu were identified as the most likely regions for ASF introduction. All experts agreed that China is the most likely source of ASF introduction into Japan. Most Japanese experts were of the view that the risk of ASF spread if introduced into Japan would be low, while foreign experts considered the risk to be moderate or high. Most experts answered that wild boars would play an important role in the persistence of ASF if the disease were to spread in Japan.
Bovine herpesvirus 1 and 5 (BoHV-1 and -5) are antigenically and genetically related and can establish latent infection. We aimed to analyze the applicability of the milk sample to detect latently BoHV-infected cattle. BoHV-1 non-vaccinated clinically healthy cows from five dairy cattle herds (herd 1, n=24; herd 2, n=39; herd 3, n=39; herd 4, n=36; herd 5, n=70) were studied. We confirmed the presence of BoHV-1, and for the first time, BoHV-5 in the milk of naturally infected dairy cattle.
The purpose of this study was to detect porcine epidemic diarrhea virus (PEDV) subclinically infected pigs shipped from non-case farms to slaughterhouses. Systematic sampling was conducted at two slaughterhouses. A total of 1,556 blood samples were collected from 80 case and non-case farms from pigs over 6 months old. Blood samples were centrifuged to obtain sera. Serial serum dilutions were subjected to serological examination for PEDV presence using Neutralization test (NT). The cut-off titer was set at titer of 1:2 dilution and farms with at least one positive sample in duplicate were classified as PED-positive farms. Several non-case farms (9.4%, 6/64) and 100% (16/16) of the case farms were indeed positive for PEDV. The proportion of seropositive animals from case farms was 63.7%, significantly different from that of non-case farms (4.3%, P<0.05). In both case and non-case farms, the proportion of seropositive animals in farrow-to-finish farms was significantly higher than in wean-to-finish farms (P<0.05). Seropositive animals in non-case farms were detected by NT in a sero-survey by sampling at slaughterhouses. Therefore, subclinically infected pigs should be considered prior to shipment.
Glycerol has been recently used to induce muscle adiposity in mice. However, its effects on the rat muscles have not been investigated previously. Therefore, we investigated the regeneration outcomes of rat muscles following glycerol-induced injury at different time points. Glycerol injection induced myofiber degeneration with extensive inflammatory infiltration on day 4 followed by appearance of regenerating myotubes on day 7 after injury without adipocyte infiltration. Meanwhile, a significant collagen deposition at early stage of regeneration that increased together with persistent inflammatory infiltration up to day 14 after injury indicates impaired regeneration. In conclusion, glycerol injury in rats is more suitable as a fibrosis-inducing model than in mice due to earlier and higher accumulation of fibrous tissue with lacking adipogenesis.
We tried measurement of visual acuity in laboratory beagle using pattern stimulus visual evoked potential (P-VEP). We recorded P-VEP in 6 beagles which were corrected refractive power. The stimulus pattern size was 1.22 mm. The testing distance were 0.5, 1.0, and 2.0 m. The visual angles and spatial frequency were calculated from stimulus pattern size and distance. In all subjects, P-VEP was clearly recorded in all testing distance, and this result means that the eye could recognized grid pattern on the stimulus monitor. When stimulus monitor was set up 2.0 m, spatial frequency was 14.29 cpd. From our results, it was founded that the visual acuity in laboratory beagle which was corrected refractive power was 14.29 cpd and more.
Intraocular cholesterol granuloma (CG) associated with synchysis scintillans (SS) was diagnosed in a 5-year-old spayed Shetland sheepdog. During the initial clinical examination, the patient exhibited SS in the anterior chamber. Canine SS is usually found in the vitreous cavity, and SS in the anterior chamber has not been described. Since canine SS has been reported to be a non-progressive condition, and its long-term clinical course has not been adequately documented. The present case report describes the long-term clinical course of a case of canine SS, in which SS occurred in the anterior chamber, leading to intraocular CG formation, and eventually glaucoma.
We previously demonstrated that transmissible gastroenteritis virus (TGEV) could induce apoptosis through caspase signaling. However, apoptosis was not completely prevented by caspases inhibitors, suggesting that there may be a caspase-independent pathway involved in TGEV-induced cell apoptosis. In this study, we investigated the regulation of apoptosis-inducing factor (AIF) on TGEV-induced apoptotic pathway. Results indicated that AIF translocated from the mitochondria to nucleus during TGEV infection, and the AIF inhibitor, N-phenylmaleimide (NP), significantly attenuated the apoptosis. In addition, the translocation of AIF was inhibited by Veliparib (ABT-888), an inhibitor of poly (ADP-ribose) polymerase (PARP). And the reactive oxygen species (ROS) scavenger, pyrrolidinedithiocarbamic (PDTC), redistributed AIF in the mitochondria and nucleus in TGEV-infected cells. Moreover, the protein levels in nucleus and the mRNA levels of AIF were inhibited in the presence of the p53 inhibitor, piﬁthrin-α (PFT-α) or in TGEV-infected p53-/- cells. Furthermore, TGEV-induced apoptosis was blocked by combination of three or more inhibitors, such as pan caspase inhibitor Z-VAD-FMK, NP, ABT-888, PDTC, PFT-α, to treat PK-15 cells. Taken together, these results suggest that the p53- and ROS-mediated AIF pathway and caspase-dependent pathway were involved in TGEV-induced apoptosis.
Appropriate dosages of cilostazol have not been studied in veterinary patients, and the degrees of heart rate (HR) increase have not been studied in dogs administered cilostazol. Therefore, this study aimed to investigate the degrees of HR increase in healthy dogs administered cilostazol. Thirty healthy beagle dogs (15 males and 15 females; age, 5–8 years) were divided into 3 groups of 10 dogs each and orally administered 2.5, 5, or 10 mg/kg cilostazol (twice a day at 8:00 AM and 8:00 PM for 10 days). Higher HR increases were seen in the 5 mg/kg group than in the 2.5 mg/kg group at all time points except 7:00 AM, 9:00 AM, 1:00 PM, and 4:00 PM (P<0.01). Higher HR increases were also observed in the 10 mg/kg group than in the 2.5 mg/kg group at all time points except 4:00 PM (P<0.01). The 10 mg/kg group showed higher HR increases than the 5 mg/kg group at all time points except 6:00 AM, 7:00 AM, 6:00 PM, and 7:00 PM (P<0.05 for 4:00 PM and 5:00 PM; P<0.01 for the other time points). These results together show that the HR of healthy dogs increased in a dose-dependent manner after cilostazol administration twice a day at doses of 5 to 10 mg/kg. These results provide a useful basis for choosing cilostazol in the treatment of bradyarrhythmia in dogs.
A total of 449 samples including 385 seafood and 64 water samples in the Mekong Delta of Vietnam collected in 2015 and 2016 were examined. Of 385 seafood samples, 332 (86.2%) samples were contaminated with Vibrio parahaemolyticus and 25 (6.5%) samples were pathogenic V. parahaemolyticus carrying tdh and/or trh genes. The tdh gene positive V. parahaemolyticus strains were detected in 22 (5.7%) samples and trh gene positive V. parahaemolyticus strains were found in 5 (1.3%) samples. Of 25 pathogenic V. parahaemolyticus strains, two strains harbored both tdh and trh genes and the other 23 strains carried either tdh or trh gene. Of 64 water samples at aquaculture farms, 50 (78.1%) samples were contaminated with V. parahaemolyticus. No tdh genepositive V. parahaemolyticus strains were detected; meanwhile, trh gene positive V. parahaemolyticus strain was detected in 1 (1.6%) sample. Twenty-six pathogenic V. parahaemolyticus strains isolated were classified into 6 types of O antigen, in which the serotype O3:K6 was detected in 4 strains. All pathogenic strains were GS-PCR negative except for 4 O3:K6 strains. The result of antimicrobial susceptibility test indicated that pathogenic strains showed high resistance rates to streptomycin (84.6%), ampicillin (57.7%) and sulfisoxazole (57.7%). These findings can be used for understanding microbiological risk of seafood in the Mekong Delta, Vietnam.
This study was conducted on red foxes to determine the appropriate voltage in electroejaculation for semen collection from stud males, and to confirm whether frozen semen with bovine semen extender can be used for artificial insemination. The proper load voltage for electroejaculation was 3-4 V based on semen collection rates and concentrations of spermatozoa. Frozen semen was prepared according to the known procedure for cows. In frozen-thawed semen, a relatively high conception rate (81.3%) was obtained in vixens, in which the optimum insemination time was detected by vaginal electrical resistance. These findings demonstrated that the restricted condition for semen collection by electroejaculation with cryopreservation of semen using bovine semen extender can be applied to artificial insemination of red foxes.
A 10-week-old miniature dachshund presented with acute onset of weakness. Electrocardiography showed sustained ventricular tachycardia, and thoracic and abdominal radiography revealed pleural and peritoneal effusion. Echocardiography revealed severely hypokinetic left and right ventricles. Thoracocentesis and abdominocentesis and subsequent transfer to an oxygen chamber yielded no clinical improvement, and the dog died about 1 hr after admission. Gross examination of a longitudinal section through the entire heart revealed poorly demarcated focal or patchy areas of grayish-white tissue infiltrating extensively into the myocardium. Histologically, these lesions were consistent with infiltrative proliferation of neoplastic lymphoid cells. Immunohistochemical staining confirmed the diagnosis of primary cardiac lymphoma (PCL) of T-cell origin. There have been no previous reports of such young dogs with PCL.
A 7-year-old Miniature Schnauzer presented with exercise intolerance and easy fatigability. Echocardiography revealed the presence of supravalvular pulmonary stenosis. The peak velocity through the stenosis was 6.4 m/sec, and the interventricular septum was flattened. Cutting balloon angioplasty was designed for the treatment of coronary artery stenosis, which was resistant to conventional balloon angioplasty. Accordingly, the dog underwent cutting balloon angioplasty and conventional balloon dilation. One month after treatment, it showed neither exercise intolerance nor easy fatigability. The ventricular septum flattening disappeared. Five months later, the dog showed an increase in activity. Two years later, the peak velocity through the stenosis decreased to 4.4 m/sec. Neither clinical symptoms nor restenosis was observed. Thus, supravalvular pulmonary stenosis was successfully treated using this combination method. The present case showed that combined cutting balloon and conventional balloon angioplasty is a useful and minimally invasive treatment for supravalvular pulmonary stenosis.
Laparoscopic cholecystectomy (LC) is widely accepted as the standard treatment for benign gall bladder diseases in humans because it has proven to be less invasive and safer than are traditional methods. However, the efficacy of LC in dogs remains unclear. The present study aimed to examine the short-term outcome of LC for benign gall bladder diseases in dogs. We enrolled 76 consecutive dogs that underwent LC for benign gall bladder diseases at our hospital between April 2008 and October 2016. Dogs with jaundice, gall bladder ruptures, abdominal effusion, or extrahepatic biliary obstruction were not excluded from the indication. Factors including age, body weight, sex, clinical sign, disease, operative time, conversion to open surgery, perioperative complications, and postoperative hospital stay were investigated. The median age of the dogs was 11 years, and the median body weight was 5.4 kg. Fifty percent of the dogs exhibited no symptoms at the initial visit. Preoperative elevation of total bilirubin levels was observed in 16 dogs (21%). LC was successfully completed in 71 dogs (93%); the median operative time was 124 min. Although gall bladder ruptures were observed in 2 (2.6%) dogs, the operations were completed successfully. Three dogs (4.1%) had to be converted to open cholecystectomy and 2 (2.6%) underwent reoperation. Two dogs (2.6%) died intraoperatively and 2 (2.6%) died postoperatively. LC was a feasible, safe, and appropriate procedure considering the current operative indications for benign gall bladder diseases in dogs.
The neoplastic mass developed in the left flank of a Border Collie dog. The tumor was resected surgically and evaluated histologically and immunohistochemically. Histologically a moderate number of spindle cells were proliferated with staghorn, placentoid, and myxoid growth patterns and a lack of perivascular whirling. Immunohistochemically, the tumor cells were positive to vimentin, laminin, S-100 protein, CD34 and CD117 antibodies. They were negative to cytokeratin AE1/3, desmin, α-SMA and calponin antibodies. Endothelial cells of the staghorn channels were positive for vWF antibody. The present case was diagnosed as spindle cell tumor, but it was similar to human classical hemangiopericytoma (HEP) and canine HEP classified by Avallon and others.
The number and distribution of Eurasian otters have declined during twentieth century due to human activity and water pollution. The global conservation status of Eurasian otter is presently 'Near Threatened (NT)' and strictly protected by being listed on the international legislation and conventions. A number of studies using the mitochondrial DNA (mtDNA) control region (CR) have been conducted in order to effectively apply conservation and reintroduction programs, especially in Europe. However, aside from Europe, there have been few studies concerning genetic diversity and phylogeny of Eurasian otters. Therefore, in this study, we sequenced partial mtDNA CR sequences (232 bp) from five South Korean Eurasian otters and analyzed 27 otters originating from parts of northeast Asia (South Korea, China, Japan and Russia (Sakhalin)), and Europe. Out of 232 bp partial mtDNA CR sequences, 13 polymorphic sites (5.6%) were identified and 4 novel mtDNA CR haplotypes (Lut16–19) were discovered from 12 Eurasian otters originating from northeast Asian region. In this study, a comprehensive analysis of genetic diversity and population structure of Eurasian otter between Europe and northeast Asia continents were conducted. Of these, different past demographic histories in Pleistocene period might have largely impacted the genetic structure of each population differently. In addition, low degree of gene flow, isolation by distance (IBD) pattern from geographically wide distanced dataset and analysis of molecular variance (AMOVA) also represented distinct genetic characteristics of Eurasian otter between Europe and northeast Asia.
A new cell line (GS-1) was developed from the spleen tissue of the orange-spotted grouper, Epinephelus coioides applied for viral infection studies of fish ranavirus and megalocytivirus. The cells proficiently multiplied in Leibovitz’s L-15 medium supplemented with 10% fetal bovine serum at temperatures between 20°C and 32°C. Morphologically, the cell line comprised fibroblast-like cells, and this was confirmed by immunostaining with vimentin, fibronectin, and desmin antibodies. The optimal temperature for GIV and ISKNV proliferation in GS-1 cells was 25°C, and the highest titer of grouper iridovirus (GIV) was 108.4 TCID50/ml, and the highest titer of infectious spleen and kidney necrosis virus (ISKNV) was 105.2 TCID50/ml. Electron micrographs showed that the mean diameter of GIV virions was 180−220 nm, which was larger than ISKNV virions (160−200 nm). Negatively stained GIV particles possessed an envelope structure that was assembled by the three-layered structure with an inner electron-dense core surrounded by a lighter coat (mean diameter, 27 ± 3 nm). The highest GIV-induced mortality of groupers occurred at 25°C, whereas the highest ISKNV-induced mortality occurred at 30°C. In summary, GS-1 cell line is a valuable tool for isolating and investigating fish ranavirus and megalocytivirus in the same host system.
The name “Actinomyces suis” was applied to each actinomycete isolate from swine actinomycosis by Grässer in 1962 and Franke in 1973. Nevertheless, this specific species was not included in the “Approved List of Bacterial Name” due to absence of the type cultures. Therefore, “Actinomyces suis” based on the description of Franke 1973 has been considered as “species incertae sedis”. We isolated a number of Actinomyces strains from swine. The representative strains of them was designated as Chiba 101 that was closely similar to the description in “Actinomyces suis” reported by Franke in 1973. Interestingly, it was found that the biological characteristics of these strains were also very similar to those of Actinomycesdenticolens. Furthermore, the average nucleotide identity (ANI) value between strain Chiba 101 and the type-strain of Actinomyces denticolens (=DSM 20671T) was found to be 99.95%. Sequences of the housekeeping genes and 16S rRNA gene showed 100% homology. These results strongly suggested that “Actinomyces suis” Franke 1973 is the same species as Actinomyces denticolens. Since actinomycosis caused by Actinomyces denticolens have been demonstrated in horses recently, it is necessary to recognize that Actinomycesdenticolens is the pathogenic actinomycetes in broader range of animals.
We examined the pathogenesis of the attenuated foot-and-mouth disease virus (FMDV) O/JPN/2000 in pigs. The virus used in this study was passaged three times in primary bovine kidney (BK) cells and once in baby hamster kidney-21 (BHK-21) cells after isolation. A plaque assay demonstrated that this virus exhibited the small plaque (SP) phenotype. There was no clinical or histological evidence of vesicular lesions in pigs intraorally inoculated with 106 50% tissue culture infectious dose (TCID50)/ml of the SP virus (SPV) of FMDV O/JPN/2000. Although fever was detected from 2 or 3 days post inoculation (dpi), there was no other prominent clinical sign up to 6 dpi. Virus shedding from saliva and nasal swab samples was not observed in any pigs inoculated with the SPV of FMDV O/JPN/2000. In the foot, mild lamellar degeneration of prickle cells in the upper layer of the stratum spinosum was histologically observed without development into vesicular or necrotic lesions. Immunohistochemical virus antigen- and terminal deoxynucleotidyl transferase-mediated dUTP-nick end labeling (TUNEL)-positive reactions observed in the foot at 1 dpi seemed to disappear after 3 and 6 dpi. Our findings suggest that the SPV of FMDV O/JPN/2000 had low pathogenicity against pigs by intraoral inoculation.
Stability of α-amylase (α-A), butyrylcholinesterase (BChE), lipase, adenosine deaminase (ADA) and total esterase activity (TEA) in two pools of porcine saliva was studied after 1 and 4 days at 4º C, and after 30, 90 and 360 days at –20º and –80ºC. At 4ºC, BChE, lipase and TEA were stable less than 1 day, α-A less than 4 days and ADA for up to 4 days. At –20º C, BChE and TEA were stable less than 30 days, α-A and lipase less than 90 days and ADA up to 360 days. At –80º C, TEA was stable less than 30 days, α-A and lipase less than 360 days, and BChE and ADA for up to 360 days.
Arboviruses, belonging to the Simbu serogroup of the genus Orthobunyavirus, often cause congenital malformations and reproductive loss in cattle. The recent occurrences of such reproductive problems suggest that new arboviruses have emerged in Japan. However, there is no information on the presence of these viruses in South Korea. The aim of this study was to determine the presence of antibodies for Akabane, Aino, Peaton, Sathuperi, and Shamonda viruses in four regions, namely Gyeonggi, Jeollabuk, Jeollanam provinces, and Jeju Island of South Korea by a serum neutralization test. Antibody positivity against Akabane, Aino, Peaton, Sathuperi, and Shamonda viruses was detected in the country, with average seropositive rates of 10.4, 4.5, 1.1, 4.9, and 5.6%, respectively.
Hydropericardium syndrome (HPS) is one of the important emerging diseases causing huge losses to the poultry industry. It affects mainly 3- to 6-week-old broiler chickens and rarely occurs in breeding and laying flocks. Recently, an HPS case was recorded with a sudden heavy mortality in a 100-day-old laying flock. A fowl adenovirus serotype 4 (FAdV-4), named as GDMZ strain, was isolated and identified using polymerase chain reaction coupled with electron microscopy. The animal experiment showed that a mortality of 100% was recorded with hydropericardium as a conspicuous lesion throughout the course of infection. Microscopically, vacuolar changes and intranuclear inclusion bodies were observed in the liver and vacuolar changes were observed in the heart. The complete genome sequence of GDMZ strain was determined to investigate the molecular properties of GDMZ strain. The comparative analysis revealed that the novel Chinese FAdV-4 isolate contained open reading frame (ORF) 19, ORF27, and ORF48 genomic deletions. The phylogenetic analysis revealed that FAdV-4 could be divided into two major clades, of which Chinese FAdV-4 were located at a distinct clade.
Severe papillomatosis occasionally causes astasia leading to euthanizing cattle. There are currently a limited number of reports on virologic approach in severe bovine papillomatosis. Here we report a full genome characterization of bovine papillomavirus type 1 (BPV-1) from the case of severe papillomatosis. A calf developed numerous papillomas on the skin and some nodules in the upper gastrointestinal tract at seven months old. The skin lesion was diagnosed as the epithelial papilloma with BPV antigen expression, while the gastrointestinal lesions were diagnosed as the fibropapilloma without BPV antigen. Full genome analysis revealed that BPV-1s detected in all the lesions were exactly the same. Compared with the reference BPV-1 sequence, there was a single nucleotide insertion in the upstream regulatory region.
A 5-month-old crossbred beef steer died after exhibiting astasia. A postmortem examination revealed verrucous endocarditis and numerous renal hemorrhages. Gram-positive bacteria were identified in the necrotic lesions of the verruca and mitral valve via histopathological analysis. Multifocal necrosis and hemorrhage were detected in the renal cortex. Gram-positive cocci isolated from the verruca were identified via biochemical tests and 16S rRNA gene sequence analysis as Streptococcus suis. Serotyping indicated that the S. suis isolates were untypable, following which these isolates were classified as a new sequence type (ST1000) via multi-locus sequence typing. S. suis is an important pathogen of pigs. However, clinical cases in cattle are rare. This report is intended to provide information that may be useful in the diagnosis of streptococcal disease in cattle.
The protective antibody response of Balb/c mice to Bali rabies virus (RABV) in BHK-21 cells was studied. The virus was isolated from a rabid dog and was adapted to replicate in BHK-21 cell culture for seven passages. The BHK-21-adapted Bali RABV (BHK-Bali RABV) was inactivated with binary ethylenimine and 24 mice were immunized twice at 21-days intervals with the inactivated virus and Rabisin® vaccine. Virus replication was detected using indirect immunofluorescence, immunocytochemistry, and western blotting assays. Enzyme-linked immunosorbent assay examination 2 weeks after the first immunization revealed RABV antibody titers that were mostly below the minimum protective level (<0.5 equivalent unit, EU). Antibody titers increased sharply after the second immunization. Antibody titers in serum of mice induced by inactivated BHK-Bali RABV one week after the second immunization were slightly lower (0.8–3.8 EU) than those induced by Rabisin vaccine (0.9–6.3 EU). RABV antibody titers were stable for at least 6 weeks after the second immunization. Both Rabisin vaccine and inactivated BHK-Bali RABV induced neutralizing antibodies with neutralization titers (50% protective dose per ml) of 29.84 for 0.1 ml Rabisin, 211.41 for 0.2 ml Rabisin, 27.41 for 0.1 ml BHK-Bali RABV, and 28.25 for 0.2 ml BHK-Bali RABV. Thus, inactivated BHK-Bali RABV induces a protective immune response in Balb/c mice, but at lower levels compared to induction by Rabisin vaccine.
We investigated possible associations of SLA class II haplotypes with serum antibody titers against swine erysipelas vaccine, reproductive and meat production traits using a population of selective breeding Duroc pigs. In the selective breeding Duroc pigs, four SLA class II-DRB1 and -DQB1 alleles were assigned by using PCR- sequence specific primer technique. Low-resolution haplotype (Lr)-0.30 and/or Lr-0.13, were deduced from the SLA class II alleles in the population of SLA-defined Duroc pigs. SLA-homozygous piglets with the Lr-0.30 haplotype had relatively lower serum antibody titers against the vaccine compared to those with Lr-0.13. In contrast, there were no statistically significant differences in reproductive performance between the SLA-defined pigs with two SLA class II haplotypes. Weaning and rearing rates until the body weight of 105 kg was reached in homozygous piglets with Lr-0.30 were significantly lower than those in homozygous piglets with Lr-0.13. The SLA-defined pigs had lower birth and weaning weights, body weights at 60 days of age, and daily weight gains than non-selective breeding Duroc pigs. Furthermore, the SLA-defined pigs had slightly lower back fat thickness compared to the non-selective breeding pigs. The rib eye areas of homozygous or heterozygous pigs with Lr-0.13 were larger than those of homozygous pigs with Lr-0.30 and non-selective breeding pigs. These data suggested that SLA haplotypes had the potential as useful genetic markers for selective breeding in the population ofSLA-defined Duroc pigs.
Although chondroinductive growth factors are considered necessary for chondrogenesis of bone marrow-derived mesenchymal stem cells (BMSC), independent and spontaneous chondrogenesis has been previously demonstrated in adult horses, bovine calves and adult human BMSC. Surprisingly, adult canine BMSC under similar culture conditions previously failed to demonstrate chondrogenesis. The present study evaluated independent chondrogenic potential of BMSC sourced from three young dogs in the absence of known chondroinductive factors. BMSC were culture expanded in 10% DMEM up to third passage (P3). At each passage, the phenotype of BMSC was evaluated by RT-PCR gel electrophoresis and qPCR. BMSC exhibited a chondrogenic phenotype in the absence of dexamethasone and TGF-β1 as verified by the expression of Sox-9, type II collagen and aggrecan. Sox-9 was significantly downregulated (P<0.05) from P1–P3 compared to P0 while type II and X collagen, and aggrecan were significantly downregulated at P3 compared to P0. There was a significant (P<0.01) negative correlation between passaging and Sox-9, type II collagen and aggrecan gene expression. These results indicate that independent chondrogenic potential and phenotype retention of BMSC decreases in a passage-dependent pattern. Therefore, caution should be exercised for future experiments evaluating the chondrogenic potential of BMSC after extensive expansion cultures in 10% DMEM.
Tetrahymena is commonly used as an alternative eukaryotic system for efficiently expressing heterologous genes. In this study, we inserted the non-structural (NS) 1 gene of avian influenza virus (AIV) into the shuttle vector pD5H8 and transformed conjugating T. thermophila with the recombinant plasmid pD5H8-NS1 by particle bombardment. Positive transformants were selected with paromomycin. We demonstrated that the NS1 protein could be expressed steadily following induction with cadmium in this Tetrahymena system. An enzyme-linked immunosorbent assay detection method was preliminary established using the expressed protein as coating antigens for serodiagnosis. This is the first study in which a Tetrahymena expression system was employed for the expression of the AIV NS1 protein, and it provides a good basis for the development of differential diagnostic kits and vaccines for the prevention and control of avian influenza.
A 2.5-year-old male French Bulldog was evaluated for seizures. Cranial magnetic resonance imaging (MRI) suggested a glioma in the left piriform area. Radiation therapy (RT) and continuous chemotherapy were administered. Although the lesion had regressed significantly 2 months after RT, a follow-up MRI revealed meningeal enhancement in the left piriform area, which expanded further, with hydrocephalus, by day 310 (8 months after RT). Because of the poor prognosis, the dog was euthanized on day 356 and necropsy was performed. Histopathological examination confirmed anaplastic oligodendroglioma with leptomeningeal dissemination. This case suggests that the possibility of leptomeningeal dissemination and hydrocephalus should be considered even after RT and chemotherapy for anaplastic oligodendroglioma.
An Okhotsk snailfish (Liparis ochotensis) kept at Nagoya aquarium exhibited sudden death. Microscopically, the fish showed multiple granulomatous foci in the gills, liver and kidney. Multiple yeast-like organisms as well as pseudohyphal elements were observed within granulomatous lesions. Immunohistochemically, the hyphae were negative for both Asperigullus and Mucor spp., and a weak positive for Candida sp. The seminated-PCR product was consistent with Candida parapsilosis and C. tropicalis. This is the first record of disseminated mycotic granulomatous lesion due to C. parapsilosis and C.tropicalis infection in fish.
The prevention and control of infectious diseases transmitted by wildlife are gaining importance. To establish effective management strategies, it is essential to understand the population structure of animals. Raccoon dogs (Nyctereutes procyonoides) in South Korea play a key role in the maintenance of food web stability and possess genetic compositions that are unique compared to those in other areas. However, wild raccoon dogs play another role as the main host of various infectious diseases. To establish long-term strategies for disease management, we investigated the genetic structure and possible geographic barriers that influence the raccoon dog population in South Korea by analyzing 16 microsatellite loci. The present study showed that mountains were the major factors responsible for genetic structuring, along with distance. We proposed potential management units (MUs) for raccoon dogs based on the genetic structuring and gene-flow barrier data obtained in this study. Four MUs were suggested for the Korean raccoon dog population (northern, central, southwestern, and southeastern). The Korean raccoon dog population structure determined in this study and the proposed MUs will be helpful to establish pragmatic strategies for managing Korean raccoon dog population and for preventing the transmission of infectious diseases.
A serologic survey of Brucella infection was performed in Caspian seals (Pusa caspica, n=71), Baikal seals (P. sibirica, n=7), ringed seals (P. hispida hispida, n=6), and beluga whales(Delphinapterus leucas, n=4) inhabiting Russian waters, by enzyme-linked immunosorbent assay (ELISA) using Brucella abortus and B. canis as antigens. The sera of 4 Caspian seals (4%) tested positive for B. abortus. The same sera samples demonstrated weaker yet detectable affinity for B. canis antigens. Several discrete bands against B. abortus and B. canis antigens were detected on Western blot analysis of the ELISA-positive seal sera; the bands against B. canis were weaker than those against B. abortus. The sera of 3 beluga whales (75%) were positive for B. abortus antigens but showed no binding to B. canis antigens in the ELISA. The positive whale sera showed a strong band appearance only against B. abortus antigens in the Western blot analysis. Many detected bands were discrete, while some of them had a smeared appearance. The present results indicate that Brucella infection occurred in Caspian seals and beluga whales inhabiting Russian waters, and that the Brucella strains infecting the seals and the whales were antigenetically distinct.
A 13-year-old female scimitar-horned oryx (Oryx dammah) died after progressive anorexia, weight loss, and depression. The necropsy showed that the retroperitoneum was compressed by a large white-to-tan uterine mass and on several sections of the mass, the uterine wall was markedly thickened because of ill-defined transmural tumor tissue. Metastatic nodules were detected in the omentum, mesentery, diaphragm, and lung. The genital tract and pulmonary and abdominal nodules exhibited highly pleomorphic sarcoma. The primary and metastatic neoplastic cells showed positive results for vimentin, desmin, and sarcomeric actin and, negative results for smooth muscle actin. Uterine metastatic rhabdomyosarcoma was diagnosed on the basis of the gross, histopathology and immunohistochemistry results.
This study investigated the influence of changing recumbency and mode of ventilation over repeated anesthesias on the alveolar to arterial oxygen tension gradient (PA-aO2) and laboratory analytes in eight horses during a year-long imaging study. Anesthesia was induced with xylazine, diazepam or guaifenesin, and ketamine and maintained with isoflurane. Horses were positioned in right or left lateral recumbency for computed tomography. Ventilation was controlled during 47% of the anesthetics. Blood was sampled from an arterial catheter prior to (30 ± 5 min from connection to anesthetic circuit), within 5 min of changing lateral recumbency, and prior to circuit disconnection (24 ± 6 min after second sample) for measurement of pH, partial pressure of arterial oxygen (PaO2) and partial pressure of arterial carbon dioxide, blood glucose and electrolytes. PA-aO2 was calculated. Data from five anesthetic episodes for each horse were summarized as mean ± standard error and analyzed using a mixed-model ANOVA. t tests were used for pairwise comparisons. P<0.05. PaO2 decreased after turning (198 vs. 347 mmHg), then increased to 291 mmHg prior to disconnection. Correspondingly, PA-aO2 was wider (252 vs.120 mmHg), and improved before disconnection (190 mmHg). Body temperature, ionized-Ca2+ and blood glucose were lower, and Na+ was higher at the last time point. In conclusion, turning anesthetized horses decreases PaO2 andresults in a widening PA-aO2 suggesting a cautious approach in animals with pre-existing hypoxemia.
Bovine foamy virus (BFV) is endemic in many countries, but has not been reported in Japan. A syncytium-forming virus was isolated from peripheral blood leukocytes of clinically healthy cattle on a farm in Kanagawa prefecture during a periodic epidemiological survey of viral diseases. The isolate was propagated in primary fetal bovine muscle cells and subsequently passaged in Madin–Darby bovine kidney cells. Since the isolate appeared to be distinct from the viruses with syncytium-forming ability previously isolated in Japan, we attempted to identify it using genomic analyses and electron microscopy. A phylogenetic analysis revealed that the isolate belongs to the bovine foamy virus cluster and is highly similar to a BFV strain isolated in China. A sero-epidemiological survey was performed using agar gel immunodiffusion test with the isolated virus as the antigen, and five of the 57 cattle tested were found to be seropositive.
This study aimed to characterize two third-generation cephalosporins- and quinolone-resistant Escherichia coli (TGCs- and Q-R-Ec) isolates recovered from the ovaries of a broiler breeder flock and the internal contents of hatching eggs produced by the broiler breeder flock. Clonal relatedness was determined by multilocus sequence typing (MLST). The isolates displayed the same multidrug resistance profile, with resistance to ampicillin, ticarcillin, piperacillin, cefazollin, cephalothin, cefotaxime, nalidixic acid, tetracycline and sulfonamides. Double disk synergy test demonstrated that the two isolates presented an ESBL phenotype. PCR and sequencing results showed that both the isolates harbored the blaCTX-M-1 and qnrS1 genes. MLST revealed a novel allele combination, designated as ST461, in these isolates. This study would contribute to the molecular epidemiological understanding of TGCs- and/or Q-R-Ec.
Stable reference genes are important for gene expression analyses such as quantitative PCR. The stability of 15 candidate reference genes that can be used to developing mouse gonads was thoroughly verified using combinations of multiple algorithms. The expression of these genes fluctuated greatly depending on the analysis period and/or gender. Peptidylprolyl isomerase A (Ppia) and polymerase (RNA) II (DNA directed) polypeptide A (Polr2a) were the reference genes that were used stably for a wide analysis period in developing mouse gonads. Furthermore, the stable reference genes corresponding to the analysis period and/or gender were ranked. These results are useful for the selection of the optimal reference gene required for high-precision measurements.
A large abdominal mass was found in a dog. Histopathologically, the surface of the mass was covered with compressed adrenal gland tissue. The neoplastic cells showed typical features of malignant peripheral nerve sheath tumor (MPNST), including Antoni type A and type B pattern, and nuclear palisading. Immunohistochemically, these cells were positive for S100 protein, nerve growth factor receptor, nestin and claudin-1. The dog was euthanized because of the developing multiple metastatic lesions. The metastatic cells showed quite similar histopathological and immunohistochemical features as those in the original tumor. Although MPNST can develop at many body sites, this is the first report of MPNST originating from the adrenal gland in a dog.