Development of a method for detecting Coxiella burnetii in cheese samples

Coxiella burnetii is the causative agent of Q fever, and the main route of infection in humans is inhalation of contaminated aerosols. Although oral transmission by contaminated raw milk or dairy products is also a possible route of human infection, there have been few studies investigating the presence of C. burnetii in dairy products. We developed a new method of extracting DNA from cheese and detecting C. burnetii DNA in cheese samples with a nested PCR assay. The limit of detection was 6.0 × 10(2) C. burnetii particles per gram. We subsequently used this method to examine the presence of C. burnetii in cheese at commercial markets in Tokyo from June 2005 to December 2008. Twenty-eight of 147 cheese samples were found to be positive for C. burnetii DNA. However, when we assessed the viability of C. burnetii by inoculating mice with DNA-positive samples, all of the samples were found to be negative. Thus, the viability of C. burnetii appears to have been lost in these cheese samples.     

Geographical variation of skull morphology and its functional significances in the red-cheeked squirrel

Skulls of the red-cheeked squirrel (Dremomys rufigenis) from various geographical locations: Malaysia (peninsular area), Vietnam (south district)-Laos, and Thailand (north district) were osteometrically examined. The skull size of the squirrels in the southern (Malaysia) population was fundamentally larger than that in the northern (Vietnam, Laos and Thailand) populations. The proportion indices indicated that the splanchnocranium was relatively longer in the Malaysia population, and that the interorbital space was narrower in Vietnam-Laos, and Thailand populations. We suggest that the long nose and laterally-oriented orbits in the skull may be better adapted for terrestrial-insectivorous life in the Malaysia population and the binocular sense facilitated by rostrally-oriented eyes contributes to the arboreal-fruit eating behavior in the two northern populations. The Malaysia population was clearly distinguished from the other populations by the principal component analysis. We suggest that the geographical barrier of the Isthmus of Kra influences the morphological variation of the skull among the squirrel populations.     

Changes in sexual behavior and fecal steroid hormone concentrations during the breeding season in female Hokkaido brown bears (Ursus arctos yesoensis) under captive condition

The longitudinal changes in fecal steroid hormone concentrations and sexual behavior in 2 mated/pregnant and 3 non-mated female Hokkaido brown bears were investigated during the breeding season. Behavioral estrus (standing) lasted for 14 and 32 days in the mated females and for 25 to 36 days in the non-mated females. In non-mated females, sexual behavior, such as female-female mounting and masturbation, was observed for several days before and after the estrous period. In mated females, mean fecal estradiol-17beta concentrations were higher in the estrous period than in the post-estrous period, while fecal progesterone concentrations were higher in the post-estrous period than in the estrous period. The similar trends of steroid hormone changes were observed in the non-mated females.     

Transglutaminase 2: a novel autoantigen in canine idiopathic central nervous system inflammatory diseases

Necrotizing meningoencephalitis (NME), necrotizing leukoencephalitis (NLE) and granulomatous meningoencephalomyelitis (GME) are common idiopathic inflammatory central nervous system (CNS) diseases with unknown etiology in dogs. We previously showed that IgG autoantibodies in the cerebrospinal fluid (CSF) of NME cases reacted to unknown brain proteins as well as to glial fibrillary acidic protein (GFAP). In the present report, we evaluated the autoantibodies against transglutaminase2 (TG2) in the canine CNS diseases. CSF samples obtained from dogs with NME (n=19), NLE (n=7), GME (n=11) and miscellaneous CNS diseases (n=12) were subjected. CSFs from 20 healthy dogs were used as controls. Indirect fluorescent antibody test on the canine cerebrum revealed astrocyte-binding IgG in the CSF of NME. After absorption of the CSF with bovine GFAP, the CSF still possessed the reactivity to astrocytes. Double-color staining showed clear colocalization of the autoantibodies and anti-human TG2 rabbit polyclonal IgG. An immunoblot assay against human recombinant TG2 revealed anti-TG2 IgG in the CSF from dogs with NME, NLE and GME. The CSF of canine idiopathic encephalitis cases, notably of NME, tended to show high ELISA OD values against human recombinant TG2 compared to healthy controls. The presence of anti-TG2 autoantibodies in the CSF may contribute to the elucidation of the etiology of canine NME, NLE and GME.     

Age Difference in Morphology and Immunohistology inthe Thymus and Spleen in Crl:CD (SD) Rats

We investigated chronological changes in immunohistochemical phenotyping in the thymus and spleen in Crl:CD rats up to the age of about one year. In the thymus, T cells increased markedly from 3 to 4 weeks of age. Proliferating cells also increased markedly at these points. B cells tended towards an increase with age. In the spleen, white pulp increased until 9 weeks of age and remained fairly stable thereafter. In the periarteriolar lymphoid sheath and marginal zone, T cells gradually increased until 9 weeks of age and became almost flat thereafter. In the lymph follicle, T cells increased with age. B cells tended towards an increase with age in all areas of the spleen. It was concluded that development of the thymus was most marked from 3 to 4 weeks of age and that both the thymus and spleen had matured by 9 weeks of age.     

Ovarian mixed germ-cell tumor comprising mature teratoma and embryonal carcinoma in a four-toed hedgehog (Atelerix albiventris)

This report describes the clinical and histopathological characteristics of a rare mixed germ-cell tumor comprising teratoma and embryonal carcinoma in the left ovary of a 10-month-old four-toed hedgehog, with chief complaints of loss of appetite and lethargy. Laparotomy revealed a swollen left ovary with small disseminated peritoneal nodules, and bilateral ovariohysterectomy was performed. The left ovary had a mature teratoma with well-differentiated fat, bone, cartilage, salivary gland, trachea, keratin cyst, and nervous tissues, and an embryonal carcinoma consisting of poorly-differentiated epithelial cells arranged in tubular, alveolar, or solid patterns. Immunohistochemically, the embryonal carcinoma cells were positive for placental alkaline phosphatase and c-KIT. This is the first case of mature teratoma with embryonal carcinoma in the ovary of a hedgehog.     

Effects of whey protein hydrolysate on growth promotion and immunomodulation in mouse pups in artificial rearing system

This study aimed to investigate the effects of whey protein hydrolysate (WPH) on the growth and immunity of mouse pups in artificial rearing (AR) system. Mouse pups were reared in the AR system with artificial milk including 5% WPH (AR with WPH) or not (AR without WPH), and the remaining pups were reared by their mother (dam) for 14 days after birth. The body weight change and body weight gain rates in the AR with WPH group were significantly higher than those observed in the AR without WPH group and similar to those in the dam group. Moreover the feed and protein efficiencies in the AR with WPH group were significantly higher than those of the AR without WPH group. In addition, the supplement of WPH in the AR system was shown to significantly elevate the number of CD3⁺CD8⁺, B220⁺CD19⁺, IA/IE⁺CD11c⁺, and CD11b⁺ in the thymocyte and/or splenocyte, and the thymus weight. Furthermore, MALDI‐TOF/MS analysis identified the amino acid sequences corresponding to some peptides, and indicated that VRTPEVDDE had the highest relative intensity among the peptides from tested WPH. Therefore, WPH would be required to not only promote growth, but also exert immunomodulatory activities in mouse pups in AR system.     

Growth of recombinant equine herpesvirus 1 (EHV-1) replaced with passage-induced mutant gene 1 and gene 71 derived from an attenuated EHV-1 in cell cultures and in the lungs of mice

The relationship of passage-induced mutant genes 1 and 71 of an attenuated equine herpesvirus 1 (EHV-1) with virulence was analysed by constructing nine recombinant EHV-1 viruses by homologous recombination. Gene 1 or/and gene 71 of a virulent EHV-1 strain, HH1, was replaced by a mutant gene 1 or/and 71 of an attenuated HH1 strain, BK343, respectively. The beta-galactosidase gene of Escherichia coli was inserted within the gene 1 or 71 coding sequence of HH1 to inactivate the genes. Virus replications of these recombinant viruses in cell cultures were similar, but release of the gene 71-inactivated virus from infected cells was delayed compared to that of the other viruses. Plaque sizes of the recombinant viruses were similar to those of HH1, but those of BK343 were significantly smaller, indicating an effect of some unknown factor(s) on viral cell-to-cell spread. The growth abilities of the recombinant viruses with a mutant gene 1 or/and 71 in lungs of mice were similar to those of HH1, but those of gene 71-inactivated viruses were reduced to the level of BK343 and the titers were about 100-times lower than those of the other recombinant viruses. These results indicate that the mutant genes 1 and 71 of BK343 might not confer an attenuated nature to EHV-1.