Canine necrotizing meningoencephalitis (NME) is characterized by autoantibodies against glial fibrillary acidic protein (GFAP) in cerebrospinal fluids (CSFs). To clarify the time-course changes in autoantibodies, serial examinations were conducted in three dogs with NME (two Pugs and a Pomeranian) that were treated by immunosuppressive therapy. The Pugs retained high autoantibody titers throughout the observation periods (146 and 813 days) and died with neurological signs. On the other hand, the Pomeranian switched from being positive for autoantibody to negative after day 580, and its NME seemed to be in clinical remission until death on day 1238. Therefore, the anti-GFAP autoantibodies can be detected over time in canine NME even during immunosuppressive therapies. However, the autoantibodies can also disappear within a certain period after onset.

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Both ehrlichioses and anaplasmoses are zoonotic, fatal infectious diseases that caused by ticks. White-tailed deer (Odocoileus virginianus) are important hosts for Ehrlichia chaffeensis, Anaplasma phagocytophilum and Anaplasma-like organisms. In the present study, an evaluation of infection with tick-borne pathogens was conducted using a PCR assay on the blood of a deer that expressed anorexia and decreased activity. The results of the PCR assay revealed natural co-infection of E. chaffeensis and A. bovis in the deer. This indicates that deer may be a natural reservoir of both E. chaffeensis and A. bovis in South Korea.

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Theileria parva (T. parva) causes a highly serious bovine disease called East Coast fever (ECF), which is characterized by pyrexia, dyspnea and cachexia and is of great economic importance in African countries. We hypothesize that the clinical symptoms of ECF could be explained by a cytokine dysregulation. In this study, we investigated the relationship between T. parva DNA load and expression levels of cytokine mRNAs in leukocytes from experimentally infected calves by quantitative PCR. The p104 gene, which encodes the T. parva 104 kDa microneme-rhoptry protein, was detected in cattle blood from day 10 after T. parva-infected tick infestation, and the protozoan DNA load was increased together with severity of disease. The mRNA expressions of pro-inflammatory cytokines, such as interleukin (IL)-1β and IL-6, were up-regulated with protozoan DNA load increasing. In addition, the level of a type-2 cytokine (IL-10) transcript was also increased during the acute phase. In contrast, the down-regulation or no detectable levels of the expression of type-1 cytokines, such as IL-2 and interferon (IFN)-γ were observed in T. parva-infected animals. Thus, our observations indicated that high protozoan load and resulting intense inflammatory responses might be involved in the severity of clinical signs observed in T. parva-infection.

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The objective of the present study was to evaluate the usefulness of the vertebral heart score (VHS) in coughing dogs with chronic degenerative mitral valve disease (MVD). Survey thoracic radiographs of 90 dogs with a history of cough and clinical and echocardiographic evidence of MVD were evaluated by 2 independent observers. The observers were asked to first determine the origin of the cough as cardiac, non-cardiac or mixed and then to measure the VHS. Agreement regarding diagnosis of the origin of cough was obtained (kappa=0.64) in 69 dogs. Of these 69 dogs, 28 (41%), 32 (46%) and 9 (13%) had a cough of cardiac, non-cardiac and mixed origin, respectively. The dogs with a cough of non-cardiac origin had a significantly lower VHS (mean ± SD, 11 ± 0.9) compared with those of dogs with a cough of cardiac or mixed origin (12.8 ± 1 and 12.9 ± 0.9, respectively). Receiver operating characteristic curve analysis showed that a VHS ≤ 11.4 is fairly accurate for exclusion of a cough of cardiac origin in dogs with MVD. The results indicate that the VHS may be an additional tool for differentiating the origin of cough in dogs with MVD.

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Homeobox (Hox), Sonic hedgehog (SHH), and Wingless-type MMTV integration site family (Wnt) are known to modulate the self-renewal and expansion of hematopoietic progenitor/stem cells in humans and mice. Frizzled (Fzd) and Patched1 (PTCH1) represent the receptors of Wnt and SHH, respectively. In this study, the amounts of mRNA transcripts of the genes associated with the self-renewal of hematopoietic stem cells, HoxB3, HoxB4, HoxA10, Wnt5a, Wnt2b, Fzd1, Fzd6, SHH, and PTCH1, were measured in canine unfractionated bone marrow cells, CD34-enriched cells, and various colony-forming units in culture (CFU-C). Partial cDNA sequences of these 9 canine genes were determined in this study. Quantitative real-time polymerase chain reaction was employed to indicate their relative amounts of mRNA transcripts. Amounts of mRNA transcripts of HoxB3, HoxA10, PTCH1, and Wnt5a genes in canine CD34-enriched cell fraction were significantly larger than those in the CD34-depleted cell fraction. Amounts of mRNA transcripts of HoxB3, HoxA10, PTCH1, Wnt5a, and Wnt2b genes in various CFU-C cells were significantly smaller than those in the seeded CD34-enriched cell fraction. These results suggested important roles of the products of these genes in self-renewal, expansion, and survival of hematopoietic progenitor cells in dogs as shown in humans and rodents.

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Using 24 diarrheic dairy calves under 8 weeks old, multiple fecal samples (4-12) were collected individually during the clinical advancing (max. 10 days) to evaluate the importance of fecal ammonia, lactate and volatile fatty acid (VFA) levels. Removing 3 calves not recovered during the sampling, 21 calves were grouped into under 3 weeks (< 3 wk; n=11) and 3-8 weeks old (3-8 wk; n=10). Data were divided into diarrheic and recovered feces with averaging in individuals. Diarrheic feces showed lower VFA and n-butyrate, and higher acetate proportions than recovered feces at <3 wk, but not at 3-8 wk. Diarrhea showed higher lactate, and lower ammonia and minor VFA (i-butyrate, valerate), which might reflect insufficiency in gut flora and fermentation.

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Generally, a conventional culture-based examination procedure (detection by egg-yolk salt agar and subsequent identification by phenotypic tests) for confirmation of the presence of S. aureus (SA) in laboratory mice and rats requires approximately 4 days. To improve the culture-based examination procedure for SA in terms of rapidity and reliability, combined use of chromogenic X-SA agar (XSA) and PCR using newly designed specific primers for SA (XSA-PCR) that can shorten the examination time (25.5 hr) was compared with the conventional procedure for SA. In 425 samples from mice and rats, 193 suspected isolates were detected by egg-yolk salt agar (EYSA), and 216 suspected isolates were detected by XSA. In the subsequent identification, 189 of 193 suspected isolates detected by EYSA were identified as SA by phenotypic tests (97.9%), and all 216 suspected isolates detected by XSA were identified as SA by PCR (100%). All SA-positive samples by the conventional procedure were included in the SA-positive samples by XSA-PCR. As a result, XSA-PCR was superior to the conventional procedure in detection rate and identification rate of SA. Therefore, XSA-PCR appears to be an effective tool for examination of SA in laboratory mice and rats that improves precision and shortens the examination time.

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Bruceine A, a natural quassinoid compound extracted from the dried fruits of Brucea javanica (L.) Merr., was evaluated for its antibabesial activity in vitro and in vivo. Bruceine A inhibited the in vitro growth of Babesia gibsoni in canine erythrocytes at lower concentration compared with the standard antibabesial drug diminazene aceturate and killed the parasites within 24 hr at a concentration of 25 nM. Oral administration of bruceine A at a dosage of 6.4 mg/kg/day for 5 days resulted in no clinical findings in a dog with normal ranges of hematological and biochemical values in the blood. Three dogs were infected with B. gibsoni and two of them were treated with bruceine A at a dosage of 6.4 mg/kg/day for 6 days from day 5 post-infection. An untreated dog developed typical acute babesiosis symptoms including severe anemia, high fever, and complete loss of appetite and movement. However, the two bruceine A-treated dogs maintained their healthy conditions throughout the experimental period of 4 weeks although complete elimination of parasites from the peripheral blood was not achieved and decreases in the packed cell volume and the erythrocyte and platelet counts were observed. Since natural quassinoid compounds have been used as traditional medicines for the treatment of various ailments including cancer and malaria, the present results suggest that bruceine A or other related compounds are potential candidates for the treatment of canine babesiosis.

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A total of 95 rodents and shrews including 82 Rattus norvegeicus, 7 Rattus rattus, and 6 Suncus murinus were trapped from different localities of Taichung, Taiwan. The overall prevalence of parasites was 93.7%. The infection rates for R. norvegeicus, R. rattus, and S. murinus were 93.9%, 85.7%, and 100%, respectively. The rats were infected with four cestodes, Taenia taeniaeformis (48.4%), Hymenolepis diminuta (38.9%), Hymenolepis nana (5.3%), and Raillietina celebensis (45.3%); ten nematodes, Angiostrongylus cantonensis (16.8%), Capillaria hepatica (49.5%), Gongylonema neoplasticum (1.1%), Heterakis spumosa (35.8%), Nippostrongylus brasiliensis (57.9%), Physaloptera sp. (1.1%), Strongyloides ratti (81.1%), Syphacia muris (2.1%), Trichosomoides crassicauda (29.5%), and Trichurus sp. (1.1%), and one protozoan, Sarcocystis spp. (33.7%). Physaloptera sp. from S. murinus and Sarcocystis spp. from both R. norvegicus and R. rattus were reported for the first time in Taiwan. The importances of zoonotic species were discussed.

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A questionnaire survey was conducted by giving 14 statements about echinococcosis to 2,070 dog owners residing in Hokkaido in order to evaluate their understanding about the biology and epidemiology of Echinococcus multilocularis. Analysis of the answers revealed that dog owners understood the disease superficially, and there were several points of confusion in their understanding, especially regarding differences in the modes of transmission and disease development in dogs and humans. The results suggest the need for the proper education of dog owners to perform proper prophylactic measures against the disease.

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Neospora caninum has been detected in several wild mammalian species, i.e., deers, coyotes, dingoes, and foxes. Farm-bred foxes were rarely reported to be affected by the parasite. In this study, we detected for the first time the infection of N.caninum in the farm-bred young blue foxes (Alopex lagopus) in China. Neospora-like tissue cysts were observed in brains and kidneys of the foxes by histopathological and immunohistochemical examinations. One hundred and three sera from the clinically normal vixens were tested for the presence of N.caninum and Toxoplasma gondii antibodies by two commercial ELISA test kits. Twenty-eight of 103 (27.2%) sera were positive for N. caninum and 1 serum (0.97%) was positive for T. gondii. A portion of the Nc5 gene of N. caninum was amplified from the DNA extracted from the fox brains by semi-nested PCR, further confirmed the existence of N. caninum among the farm-bred fox herd in China.

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Pseudomonas aeruginosa Exotoxin A (PEA) has been generally used to induce liver injury in mice for experimental study. No PEA-induced hepatotoxicity study has ever been conducted in rats, although rats are the most common rodents used in toxicologic bioassay and pharmacological evaluation. The present study was conducted in male Wistar rats that were injected (i.v.) with PEA at doses of 0, 10, 20, 30 or 40 μg/kg body weight and evaluated at 12, 24, 36, 48 and 60 hr post-exposure (HPE). Rats exposed to PEA at 40 μg/kg died before 36 HPE, and the mortality was dose and time dependent. Liver injury was noted as increases in serum enzymes, along with alterations of liver histology in the 40 μg/kg group at 12 HPE. TUNEL-positive staining indicative of hepatocyte apoptosis was observed in the 20 μg/kg group at 12 HPE. Significant levels of DNA fragmentation ladder were observed in the 30 μg/kg group starting at 24 HPE. Serum levels of TNF-α was increased in the 30 and 40 μg/kg groups at 48 and 24 HPE, respectively. Other cytokines, such as IL-2, IL-6, and IL-10 were also increased at various doses and times. Furthermore, the elevated serum heaptic index levels decreased significantly by dexamethasone pretreatment. In contrast, these markers were exacerbated by co-administration of a non-toxic dose LPS. In overall evaluation, the PEA-induced liver injury can be used as a model for study of hyperimmune-mediated hepatotoxicity.

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An adult, Japanese domestic cat presented with bilateral swelling, distortion, and erythema of ears and deformation of the limbs. Biopsy of the pinnae confirmed auricular chondritis. These lesions and the cat's general condition subsequently deteriorated, and the cat died. At necropsy, chondral changes were present in the pinnae, costae, larynx, trachea, and limbs. Histopathologically, these chondral tissues showed marked deformation with lymphocytic inflammation. The limb joint inflammation was associated with chondral erosion, deformation of subchondral bones with pannus, and thinning of cancellous bones. These lesions were consistent with the diagnostic criteria for human relapsing polychondritis. However, the cat had erosive arthritis, which appeared to be different from human relapsing polychondritis.

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A 14-year-old, spayed female Shih-tzu dog presented with masses in the dorsal aspect of cervical region and digit of the right anterior limb. Extensive necrosis was seen in the dermal tissue overlying the tumor, and diffuse round cell proliferation and infiltration were seen histologically from the superficial dermis to the deep dermis. Two types of proliferating cells were present: lymphoblast-like cells with round-oval, vesicular nuclei and moderate-large nucleoli; and smaller cells with characteristic irregularly shaped nuclei. Electron microscopy of these smaller cells showed cerebriform, pleomorphic nuclei with a chromatin pattern characteristic of lymphoid cells, as seen in lymphoblast-like tumor cells. Immunohistochemically, both types of tumor cells were positive for CD3. Most vessel walls had been invaded by tumor cells, resulting in extensive dermal necrosis and hemorrhage. Based on these histopathological findings, the tumor was diagnosed as vasotropic and vasoinvasive nonepitheliotropic lymphoma, characterized by a notable presence of unusual tumor cells with irregularly shaped nuclei and extensive dermal necrosis.

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The prevalence of hepatitis E virus (HEV) infection in wild boars and pigs in Gunma Prefecture, Japan, was serologically and genetically examined. The positive detection rates of anti-HEV IgG and HEV RNA in the wild boars were 4.5% (4/89) and 1.1% (1/89), whereas those in the pigs were 74.6% (126/169) and 1.8% (3/169), respectively. The positive rates of anti-HEV IgG and HEV RNA on the 17 pig farms in the present study ranged from 20% to 100%, respectively. One male wild boar approximately 5 years of age was positive for HEV RNA but was negative for anti-HEV IgG. Three pigs from 2 farms were positive for HEV RNA; 2 of these pigs were negative for HEV IgG, and the other was positive. A phylogenetic analysis revealed that all of the HEV ORF1 genes detected in the present study belonged to genotype III. In Gunma Prefecture, HEV is highly prevalent and widespread, and uncooked wild boar and pig meat may have the potential to transmit HEV to humans.

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Babesia microti, the erythroparasitic cause of human babesiosis, has long been taken to be a single species because classification by parasite morphology and host spectrum blurred distinctions between the parasites. Phylogenetic analyses of the 18S ribosomal RNA gene (18S rDNA) and, more recently, the β-tubulin gene have suggested inter-group heterogeneity. Intra-group relationships, however, remain unknown. This study was conducted to clarify the intra- and inter-group phylogenetic features of the B. microti-group parasites with the η subunit of the chaperonin-containing t-complex polypeptide l (CCTη) gene as a candidate genetic marker for defining the B. microti group. We prepared complete sequences of the CCTη gene from 36 piroplasms and compared the phylogenetic trees. The B. microti-group parasites clustered in a monophyletic assemblage separate from the Babesia sensu stricto and Theileria genera and subdivided predominantly into 4 clades (U.S., Kobe, Hobetsu, Munich) with highly significant evolutionary distances between the clades. B. rodhaini branched at the base of the B. microti-group parasites. In addition, a unique intron presence/absence matrix not observable in 18S rDNA or β-tubulin set the B. microti group entirely apart from either Babesia sensu stricto or Theileria. These results have strong implications for public health, suggesting that the B. microti-group parasites are a full-fledged genus comprising, for now, four core species, i.e., U.S., Kobe, Hobetsu, and Munich species nova. Furthermore, the CCTη gene is an instructive and definitive genetic marker for analyzing B. microti and related parasites.

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Hazard analysis of Listeria monocytogenes contamination during processing of salted walleye pollock (Theragra chalcogramma) roe was performed for a seafood plant in Japan from December 2005 to February 2006. As a result, L. monocytogenes number was detected on the pallet used for transport of barrels in the salting process and one of the rollers of the roller conveyor, which rotates while in contact with the bottoms of the barrels, but was not detected in any raw materials, interim products or final products. Thus, we believe that the pallet contamination initially occurred because of insufficient washing, that it was passed on to the bottoms of the barrels and that it was then passed on the roller of the roller conveyor by cross-contamination. Therefore, it is possible that interim and final products may become contaminated by processing devices and machinery. In addition, we conducted an inoculation study designed at the 1/20 actual factory scale using interim products with or without artificial color and seeded with L. monocytogenes to observe changes in its growth. In the inoculation study, multiplication of L. monocytogenes during the salting process was not confirmed in the samples with artificial color.

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Pituitary thyrotroph hyperplasia results from prolonged primary hypothyroidism in humans, mice and rats. In dogs with Cushing's disease, many cases have low serum thyroid hormones concentrations due to euthyroid sick syndrome. A 6-year-old castrated male Beagle diagnosed with Cushing's disease had a high serum thyroid stimulating hormone (TSH) concentration that was treated by hypophysectomy. On histological examination, the resected pituitary gland contained both a corticotroph adenoma and thyrotroph hyperplasia. The TSH-positive cell ratio in this case was greater than that of healthy Beagles. In the present case, the pituitary thyrotroph hyperplasia was probably caused by primary hypothyroidism. In conclusion, this Beagle is the first histological confirmation of the coexistence of a corticotroph adenoma and thyrotroph hyperplasia.

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Bovine mammary epithelial stem cells (MESCs) are very important in agricultural production and bioengineering. In the present study, we compared different isolation and culture methods for MESCs and observed their growth and differentiation characteristics. MESCs have an extremely weak proliferation capacity, and it is very difficult to obtain and prolong subculture of a bovine mammary epithelial stem cell line. We obtained some multipotent MESC aggregates that looked like spherical colonies. These colonies were only derived from suspension culture and were induced to differentiate into epithelial-like cells, myoepithelial-like cells and secretory cells and to establish a ductal-like structure. In contrast, MESCs cultured in adherent culture displayed low morphogenetic competence and only differentiated into epithelial-like cells. MESCs are often identified by testing their differentiation in vivo; however, herein, we have demonstrated the in vitro differentiation potential of bovine MESCs. In our study, beta 1-integrin and alpha 6-integrin which are expressed by human epidermal stem cells, were found in bovine, which shows that bovine MESCs share the same molecular signature as human MESCs.

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Molecular analysis of the polymorphic region of the bovine coronavirus (BCoV)-S gene using recent Japanese field isolates and reference strains revealed that the 148 isolates collected from 1999 to 2008 from 13 prefectures, covering all regions of Japan (Hokkaido, Tohoku, Kanto, Chubu, Kinki, Chugoku, Shikoku, and Kyusyu region) and divided into 3 clusters, show distinctive divergence from the prototype enteric BCoV strains. Almost all isolates after 2005 were clustered into group 4, and there was no regional specificity in these clusters. To differentiate the genotypes without sequencing, a simple technique-reverse transcriptase-polymerase chain reaction/restriction fragment length polymorphism analysis (RT-PCR/RFLP)-was developed. The availability of a simple and easy diagnostic assay will enable larger epidemiological studies of BCoV.

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CXC-chemokine receptor 4 (CXCR4) functions as a receptor for feline immunodeficiency virus (FIV). Although we previously found that a CXCR4 antagonist, T140, inhibited the FIV replication in vitro, it was not effective in cats infected with FIV because of its low stability in feline serum. To resolve this problem, several T140 derivatives have been developed. Here, we examined the efficacy of T140 analogs, TF14016 and TF14013, on the inhibition of FIV infection. These compounds were shown to significantly inhibit the syncytia formation in CXCR4-expressing cells after co-cultivation with FIV-infected cells and the replication of FIV in a feline lymphoid cultured cell line. These results indicated that TF14016 and TF14013 could be useful as antiviral drugs for cats infected with FIV.

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