Polymerase chain reaction (PCR) amplification of a partial fragment of the sex determining region Y (SRY) gene was used for sexing a young Linne's two-toed sloth (Choloepus didactylus), a species in which gender determination from the external genitalia is difficult. DNA was extracted from hairs of a 5-month-old sloth as well as the dam and sire as external controls. A SRY fragment (216 bases) was PCR-amplified both from the offspring and the sire, but not amplified from the dam. The DNA sequence (166 bases without primers) of the sloth PCR product was detetmined and compared with SRY sequences of other mammals previously reported. High homology of their nucleotide (74.1-86.8%) and deduced amino acid (63.6-85.5%) sequences indicates that the PCR product of the sloth was amplified from a region of the SRY gene, and that SRY sequences are conserved throughout mammalian orders. From the result the sex of the young sloth was determined as a male. The PCR method using hairs for sexing the sloth provides an advantageous tool for captive propagation plan in zoos. To the authors' knowledge, no report regarding SRY sequences in the order Xenarthra (Edentata) has been published.
The present immunohistochemical study deals with dynamic alteration of T-cell subsets in the oviduct in sex-hormone-treated chickens. Monoclonal antibodies (CT3, CT4, and CT8) specific for the chicken homologues of CD3, CD4, and CD8 were used in estrogen- or progesterone-treated chickens. In control animals, no lymphocytes appeared throughout the oviduct until 4 weeks of age. When 7-day-old chickens were injected with either diethylstilbestrol (DES) or DES plus progesterone, T cells immunoreactive for CT3 first infiltrated the oviduct at 12 hr after the hormone treatment. Their frequency of occurrence rose from 48 to 96 hr. Subsequently, CT3+ cells in the magnum declined in number per area coincident with the proliferation of albuminous glands in the lamina propria, while in the vagina no decline of T cells was observed. The population of T-cell subsets in the lamina propria of both the magnum and vagina was significantly higher in the DES-treated chickens than in DES plus progesterone-treated chickens. Among T-cell subsets, CT8+ cells were more numerous than CT4+ cells throughout the study, this relative frequency being shared by normal adults. Depopulation of lymphocytes from the thymus, spleen and cecal tonsil, their mobilization to the circulating blood, and subsequent dynamic infiltration into fhe oviduct suggested that the sex hormones induced the traffic of T cells from the lymphoid organs into the oviduct.
We examined bovine c-myb gene expression in six samples of sporadic bovine lymphomas (two calf, three thymic and one intermediate) and five of enzootic bovine leukosis. Tumor cells of the sporadic bovine lymphomas were of immature cell lineage (one B lymphoma and five T lymphomas). The c-myb mRNA was expressed in almost all the sporadic bovine lymphomas (except for one thymic form) including a BoCD8 single positive T lymphoma. On the contrary, c-myb was not expressed in mature B lymphomas of enzootic bovine leukosis. The results suggest that c-myb expression is closely associated with tumor cell differentiation of bovine lymphomas.
Due to its importance in public health, Salmonella Typhimurium originating from a naturally infected bengalee (Lonchura striata), a common cage bird, was examined for its infectivity and persistence for the same species. Eight birds per group for each experiment were used. When bengalees were inoculated orally with 102, 104 or 105 colony forming units (CFU) of S. Typhimurium and observed for 7 days, all the birds receiving 105 CFU were positive for the organism in the liver, spleen or the intestines, and necrotic foci in the liver were observed in 6 birds. When bengalees were inoculated with 105 CFU of S. Typhimurium and observed for 22 days, the organism was found in fecal samples throughout the experimental period and the maximum S. Typhimurium counts in feces were 3.9×108CFU per gram. S. Typhimurium was recovered from the liver, spleen and intestines in 7 birds and necrotic foci in the liver were also observed in 7 birds. The results indicate that S. Typhimurium originating from a naturally infected bengalee is pathogenic to these birds and the persistence of the pathogen lasts at least for 22 days.
Rabbits were treated with a single intravenous injection of various antibiotics. More than 40 per cent of the animals showed diarrhea after being treated with sulbactam/cefoperazone, cefmetazole, clindamycin, piperacillin or aspoxicillin. Clostridium difficile was isolated from sulbactam/cefoperazone-treated diarrheic rabbits, with their cecal contents showing positive reaction in a latex agglutination test for C. diffcile enterotoxin. However, 27 cefmetazole-induced diarrheic cases were not associated with C. difficile. Other enteropathogenic bacteria, such as Campylobacter spp., Bacillus cereus, enteropathogenic Escherichia coli, coagulase positive Staphylococcus aureus, Salmonella spp., Vibrio spp., Clostridium perfringens and Clostridium spiroforme, were not isolated from either of diarrheic rabbit. However, the counts of clostridia remarkably increased in the intestine of cefmetazole-associated diarrheic rabbits. This was ascribed to the overgrowth of Clostridium innocuum and Clostridium sporogenes. There were no remarkable differences in changes in other bacterial population between diarrheic and non-diarrheic rabbits.
The objective of this experiment was to evaluate the influences of Ca and P contents in an anionic diet on the mineral metabolism in plasma, urine and bone in periparturient dairy cows. Fifteen multiparous Holstein-Friesian cows were divided into 3 dietary groups (5 cows/group) by dietary Ca and P contents and dietary cation-anion balance [(Na+K)-(Cl+S) mEq/kg DM]; diet 1 [low Ca (0.46%), low P (0.24%), cationic (+195.8 mEq/kg DM)]; diet 2 [low Ca (0.46%), low P (0.24%), anionic (-32.4 mEq/kg DM)]; and diet 3 [high Ca (0.93%), high P (0.60%), anionic (-41.0 mEq/kg DM)]. Cows were fed one of these 3 diets from approximately 4 weeks before the expected calving date to 5 days after calving. There was no outbreak of milk fever in any cows fed these 3 diets; however, plasma Ca levels at 1 and 2 days after calving tended to be higher in the cows fed diet 3 than those in the cows fed diets 1 or 2. Fractional urinary excretion of Ca in the cows fed diet 2 or 3 was higher than that in the cows fed diet 1. Fractional urinary excretion and plasma level of Pi were higher during the periparturient period in the cows fed diet 3 than those in the cows fed diets 1 or 2. There were no significant differences in plasma parathyroid hormone levels among the 3 groups. In the spongy substance of ilium at 5 days after calving, the Ca and Mg contents, bone volume and trabecular thickness were the lowest, but not significant, in the cows fed diet 2. These data suggest that sufficient Ca and P contents in an anionic diet may be effective in maintaining plasma Ca and Pi levels of periparturient cows and further in preventing of potential bone damage brought about by increased urinary mineral excretion following the feeding of an anionic diet.
The structural arrangement and cellular distribution of endothelial and lining cells of the synovial villi were studied in the equine palmar/plantar recess of the metacarpo- and metatarsophalangeal joints by light microscopy and electron microscopy. The extent and distribution of blood vessels varied with villous shape and length. The majority of vessels formed concentric circles in cross and longitudinal sections and probably are arranged in a convoluted, spiral or helical pattern. The villi do not contain smooth muscle cells or typical capillaries as observed in other organs. Under the electron microscope, the endothelium is surrounded by connective tissue and discontinuous circular cells, presumably fibroblasts. The outermost layer was sometimes surrounded by type A and/or B synovial cells. The lumen of the blood vessels at the top of villus appeared to be constricted in most cases, with a diameter of about 12±3μm. Blood vessels formed by more than six endothelial cells in the middle portion of villus generally were not constricted. Well-developed cytoplasmic processes extended into the lumen of blood vessels. The constriction of blood vessels with no apparent smooth muscle presence and the observation of numerous intermediate filaments in the cytoplasm of the endothelial cells suggests that these villous blood vessels constrict through contraction of their own endothelial cells. Lining cells were distributed unevenly even within a single villus; the villous lining cells seemed to have directional preferences with domination of synovial type A cells. Surprisingly, structures resembling myelinated nerve ends (∼0.2μm) were observed between juxtaposed endothelial cells as well as directly on an endothelial cell, suggesting that these nerve endings may be a sensor detector of either pressure or temperature or have a proprioceptive-like function. Synovial villi have a distinctive structural arrangement of vessels, lining cells, and nerve endings.
Rapid infusion is believed to be harmful to the lung, however, the pathological status of pulmonary edema resulting from excessive fluid therapy in horses has not been clarified because the quantitative diagnosis of pulmonary edema is impossible. To evaluate the precision of the double indicator dilution method using heat and sodium in horses, which allows the quantitative diagnosis of pulmonary edema, we compared extravascular lung water volume measured using a lung water computer based on the theory of the double indicator dilution method with that determined by the direct method. The value of extravascular thermal volume (ETV) determined by the double indicator dilution method was 7.82±0.62 ml/kg and the detection ratio of ETV to the value of pulmonary extravascular water volume (PEWV) by the direct method was 0.996±0.038. There was a significant correlation between ETV and PEWV (P<0.05), and the regression line was Y=1.23 X - 1.73 with a correlation coefficient of 0.953. The value of extravascular lung water determined by the double indicator dilution method was significantly consistent with that obtained by the direct method, indicating the high precision of the double indicator dilution method in normal horse lungs.
The aims of this study were to investigate whether there are cell-specific distributions of insulin-like growth factor I (IGF-I) and epidermal growth factor (EGF) in the endometrium, and to determine whether or not the expression of these factors varies with the stage of the estrous cycle. Fifty-four endometrial biopsy specimens were collected from normal cycle Holstein-Friesian heifers. The endometrial specimens were divided into nine groups (at least five different animals per group) corresponding to the following days of the cycle: days 20-0, 1-3, 4-5, 6-7, 8-10, 11-13, 14-15, I6-17 and 18-19 (day 0=estrus). IGF-I and EGF were localized by immunohistochemistry in the intact heifer endometrium throughout the estrous cycle. Throughout the estrous cycle, IGF-I was localized in the luminal epithelium and stroma with a little staining in the glandular epithelium (P<0.01). In the luminal epithelium, the number of IGF-I-positive cells increased on days 1-3, 6-7, 11-13 and 16-17. In the stroma, the number of IGF-I-positive cells increased on days 8-10 and 20-3. The number of positively stained cells in the glandular epithelium increased around the estrous period. EGF stained intensely in the stroma but very little in the luminal and glandular epithelia (P<0.01). In the stroma, three peaks in the number of EGF-positve cells were observed: on days 1-3, 6-7 and 11-13. These results demonstrate cyclical changes in the endometrial cell types localization of IGF-I and EGF.
Immunohistochemical detection of rat CAR bacillus antigen in paraffin-embedded experimentally infected rat lungs, using an immunoperoxidase technique based on the labelled streptavidin biotin (LSAB) method and 3-amino-9-ethylcarbazole (AEC) as substrate is described in this paper. The pattern of immunostaining was confined to the ciliated bronchial epithelium and the specificity of this technique was confirmed. The use of AEC as substrate was evaluated more efficient than diaminobenzidine (DAB). The usefulness of this immunoperoxidase technique for the detection of CAR bacillus in rats and its advantages compared to the indirect immunofluorescence (IF) are discussed.
Camelostrongylus mentulatus (Railliet et Henry, 1909) Orloff, 1933 (Nematoda; Trichostrongyloidea) was found from the abomasum of a three-year-old female cape giraffe, Giraffa camelopardalis giraffa, born and died in a zoo park in Yamaguchi prefecture, Japan. This is the new host record from Giraffidae and geographical distribution of C. mentulatus. Present case of C. mentulatus might be infected from other ruminants, e.g., camels, antelopes and goats, kept at a same paddock in the zoo. Risk of imported parasitic diseases by the zoo animals from outside of Japan is discussed.
Bovine lactoferricin(R) (LFcin B) is a strong antimicrobial peptide derived from N-lobe of lactoferrin. To study the immunochemical and structural properties of LFcin B, monoclonal antibody (mAb) was prepared and the amino acid sequence concerning with the binding to mAb has becn identified. Mice injected with LFcin B showed no production of antibody specific to this peptide, whereas those with LFcin B-KLH conjugate produced anti-LFcin B antibodies. None of the mAb reacted with bovine lactoferrin C-lobe, human lactoferrin or LFcin H. By the reactivity of the mAb against the peptides synthesized on cellulose membranes using SPOTsTM and against chemically modified derivatives of LFcin B, the antigenic determinant of LFcin B was identified to be the sequence of "QWR".
T-cell subsets in the chicken bursa of Fabricius were analysed immunohistochemically during postnatal stages. Distinct types of T cells were present both in the surface epithelium and lamina propria of the bursa. TcR1+ and CT8+ cells were predominant in the surface epithelium, while TcR2+ and CT8+ cells were numerous in the lamina propria. These T-cell subsets peaked in frequency of occurrence at 5 weeks. They then decreased in number, and a few T cells remained in the bursa until 15 weeks of age. This result showed that the bursa of chickens is furnished with the distinct types of T-cell subsets at early postnatal stages of development to maintain its local immunity.
Ethylene thiourea (ETU) was administered once orally to pregnant rats on gestation day 12 at a dose of 200 mg/kg, and its concentration-time courses in the maternal plasma, amniotic fluid and embryos were investigated. The ETU concentrations in the maternal plasma and amniotic fluid reached the peak level about 2 hr after dosing, then declined gradually and had disappeared by 48 hr. In embryos, the concentration of ETU peaked at 30 min after dosing and disappeared at 48 hr. The prolonged exposure of the embryos to the high concentration of ETU in the amniotic fluid could be partially responsible for the teratogenic effect of ETU.
The rate of DNA synthesis in the inner cell mass (ICM) of frozen-thawed bovine embryos was examined. Bovine blastocysts derived from in vitro matured/in vitro fertilized oocytes were frozen with 1.8 M ethylene glycol (EG), 1.6 M propylene glycol (PG) and 1.4 M glycerol plus 0.25 M sucrose (GL). Viable embryos, after thawing and culture beyond the blastocyst stage, were examined by immunocytochemical staining for detection of DNA synthesis by ICM cells. The numbers of bromodeoxyuridine-immunoreactive ICM cells of frozen-thawed embryos prepared with EG (10.7) and GL (11.5) were significantly lower than those of unfrozen embryos (17.3). The results suggest that the rates of proliferation of ICM cells of frozen-thawed bovine embryos tend to be lower than those of unfrozen embryos irrespective of the cryoprotectant used.