Steroid-producing tissues such as ovary, placenta, testis and adrenal gland and non-steroid-producing cells including lymphocytes are known to contain 20α-hydroxysteroid dehydrogenase (20α-HSD). In the present study, we measured and partially characterized the 20α-HSD in 11 neoplastic tissues surgically removed from dogs and a cat. The tissues were pathologically classified as 4 benign and 7 malignant. The enzyme activities in the cytoplasmic fraction were positive in 6 of the 7 malignant tissues and 2 of the 4 benign ones. Following DEAE chromatography analysis of 2 malignant tumour samples, multiple forms of enzyme activities with different electric charges were detected. Furthermore, 20α-HSD activity in the cytosol fraction from malignant tumours was increased dramatically by passage through the DEAE column, suggesting that the enzyme is present in the cytosol as an inactive or Sequestered form.

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The prevalence of antibodies against 5 zoonosis agents was determined in serum samples of 443 breeding monkeys. Of the monkeys, 296 were bred or kept for a long time at R institute, and the remaining 147 were newly imported from the Philippines and kept at S institute for quarantine. Antibodies to simian virus 40 were highly prevalent at 89.1% among monkeys of R institute, whereas no antibody could be detected in those of S institute. Antibodies to Chlamydia psittaci and Yersinia pseudotuberculosis were detected in 14.4 and 11.6% at R institute, and in 9.0 and 3.5% at S institute, respectively, evidencing a significant difference (P<0.05) between those of the two institutes for both agents. Antibodies to Toxoplasma gondii and Leptospira interrogans were found in 3.6 and 2.9% of the overall, respectively, showing no difference in positive rates in relation to breeding place. Even in cases positive to the latter 4 zoonosis agents, the antibody titers were low. The results obtained suggest that all zoonoses tested do not seem to be so serious diseases among monkeys at the present time except simian virus 40 infection, which is highly prevalent among breeding monkeys in Japan.

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In the pig, cattle and horse, the right and left atria and ventricles were examined by immunohistochemistry, and the right atrial and auricular cardiocytes were studied by transmission electron microscopy. Moreover, ANP-granules in the cardiocytes were analyzed by ultrastructural morphometry. Immunohistochemically, the most intensely ANP-reacted cardiocytes were localized in the right auricle, particularly more prominent in the pig and cattle than in the horse. Ultrastructurally, ANP-granules were located principally in the perinuclear region associated with the Golgi apparatus and throughout the sarcoplasmic layers. They were classified into 2 types, A and B. A-granule had a conspicuous electron-dense core enveloped by a membrane. B-granule had a fibrillogranular core enveloped by an indistinct membrane. By TEM equipped with a goniometer stage, a part of the limiting membrane of B-granules became visible as the stage tilt progressed. By ultrastructural morphometry, the number of each type of granules was significantly greater in the pig and cattle than in the horse. In the pig and cattle, the total number of both types of granules in the auricular cardiocytes was significantly greater than that in the atrial ones. The diameter of each type of granules was significantly larger in the pig and cattle than in the horse. The diameter of A-granules was significantly larger than that of B-granules.

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Total serum creatine kinase (CK) and its isozyme activities were determined in dogs with dirofilariasis. Before heartworm removal, total CK and isozyme activities in dogs of the mild group were not different from those in dogs of the heartworm-free group. BB activity was higher in dogs of the hemoptysis group. Dogs of the ascites group displayed a mild increase in MM activity. In dogs of the caval syndrome (CS) group, total CK and MM activities were highest among the heartworm-free and heartworm-infected dogs, and MM isozyme accounted for most (75%) of total CK activity. MB and BB activities were also higher. However, there were no significant differences in CK activities between the surviving and non-surviving cases. In dogs with pulmonary heartworm disease (mild and ascites groups), MM activity correlated significantly with the number of heartworms (r=0.45), hematocrit value (Ht, r=-0.40), serum alanine aminotransferase (ALT, r=0.42) and lactate dehydrogenase (LDH, r=0.46) activities, mean pulmonary arterial pressure (r=0.64) and total pulmonary resistance (r=0.50). In dogs with CS, MM activity did not correlate with any parameter, but BB activity correlated with the number of heartworms at the right atrium (r=0.61), Ht (r=-0.53), ALT (r=0.80), LDH (r=0.73) and serum urea nitrogen (r=0.47). At 1 week after heartworm removal, BB and MM activity decreased in dogs of the hemoptysis and ascites groups, respectively. In dogs of the CS group, total CK and MM isozyme activities decreased markedly (P<0.01) regardless of their prognosis.

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An enzyme-linked immunosorbent assay (ELISA) was evaluated and compared in parallel with the standard complement fixation test (CFT) for the diagnosis of bovine subclinical paratuberculosis. Bovine sera preabsorbed with the mixture of Mycobacterium phlei and kaolin suspension were assayed for antibody activities to the crude protoplasmic antigen of Mycobacterium paratuberculosis in the ELISA. ELISA antibody titer was expressed as ELISA antibody index (EAI) value: EAI = (At - An) / (Ap - An), where At, Ap and An are the absorbance values of a 1:200 dilution of unknown test sera, a 1:400 dilution of positive control serum, and a 1:200 dilution of negative control serum. An EAI of 0.6 or greater was established as a reasonable cutoff point for a positive antibody titer by ELISA. Of the 156 sera from cattle with subclinical M. paratuberculosis-infection, 106 (67.9%) were positive by ELISA and 41 (26.3%) by CFT. Of the 3, 880 sera from cattle in the herds which had no history or evidence of paratuberculosis, 3, 875 (99.9%) were negative by ELISA, and 3, 787 (97.6%) by CFr. Positive ELISA titers were detectable l to 5 months earlier than positive CFT titers in experimentally infected cattle, and 7 to 10 months earlier in naturally infected cattle. These results indicate that the ELISA should replace the CFT as the routine test of choice for the diagnosis of bovine paratuberculosis.

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A transplantable mononuclear cell leukemia (MCL) was established from spontaneous MCL in an F344 rat. In this work, we paid special attention to a nodular tumor, named MCL-YSK, developed at the subcutaneous transplant site. MCL-YSK was serially passaged in subcutaneous tissue of syngeneic rats up to the 19th generation. Transplants from MCL-YSK grew into nodules 3cm in diameter and 11.3g in weight 9weeks after subcutaneous implantation. Neoplastic cells forming the nodules had azurophilic cytoplasmic granules, which ultrastructurally appeared to be lysosomes. The cells reacted positively for acid phosphatase and nonspecific esterase, but not for alkaline phosphatase, alpha-1antitrypsin and lysozyme, nor reacted with anti-rat monocyte/macrophage monoclonal antibody and anti-rat CD-8 monoclonal antibody. They possessed Fc-receptor. Leukemic cells first appeared in the peripheral blood 6weeks after transplantation when subcutaneous nodules reached an average diameter of one cm. Subsequently, leukemic changes progressed in recipients as MCL-YSK grew larger. The recipients died during the period from 8 to 12 weeks after transplantation, showing anemia, depression, splenohepatomegaly and lymph node enlargement. Diffuse or focal proliferation of sprinkled tumor cells was present in many organs. Hematologic changes suggestive of hemolytic anemia, elevated plasma enzymes and decreased drug-metabolizing enzymes, indicative of hepatic malfunction, were seen in transplant recipients. MCL-YSK was easily transplanted into athymic nude mice. The transplanted mice showed leukemic changes similar to those observed in rats with transplanted MCL-YSK.

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The fine structure of acrosomic abnormalities such as fragmentation and vacuolization of boar spermatids was studied in 4 different breeds (Duroc, Hampshire, Landrace and Yorkshire). These abnormalitids initially appeared from the cap phase of spermiogenesis. The acrosome did not consist of serial sac covering the hemispherical nucleus, but only of a large acrosomic vesicle containing a dense granule and several pieces of cleaved proacrosomic visicles attached to the anterior half of the round nucleus. The abnormality was frequently observed in the sister cells connected with the intercellular bridge. In the same seminiferous tubules, there was another abnormality on the acrosomic vacuolization of limited membrane system. Multiple vacuoles or vesicles were seen in the apical part of the nucleus in the cap phase, and also in the lower part of the equatorial segment of the acrosome phase.

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Seventy-six Staphylococcus aureus strains isolated from various lesions of horses were characterized. All of the 76 strains were identified as biotypes B (38.2%) and C (61.8%). Of 55strains tested, 42 (76.4%) were differentiated into 7 coagulase types. Coagulase types V and VII were predominant in the metritis strains. Coagulase type II was found most frequently in the strains from phlegmon, dermatitis, sinusitis, empyema sinus, and nasal catarrh. Forty-two (55.3%) of the 76 strains were differentiated into 24 phage patterns. Twenty (58.8%) of 34 typable strains from metritis were lysed by the human group I phage 52, and group II phages 3A, 3C, 55 and 71. Forty-five (59.2%) of the 76 strains were resistant to 1 or more of 6 antibiotics. Strains resistant to penicillin G, irrespective of source, were most frequent (95.6%). Forty (93.0%) of 43 strains resistant to penicillin G alone or in combination with other antibiotics produced beta-lactamase.Only 8 (10.5%) of the 76 strains produced enterotoxins A (n=2), B (n=1) or C (n=5), and they all were isolated from metritis. Only 1 strain isolated from phlegmon and 2 from metritis produced exfoliative toxin (ET) and toxic shock syndrome toxin-1 (TSST-1), respectively. The latter 2 strains also produced enterotoxin C. The results of the present study showed the first evidence of the presence of both ET-and TSST-1-producing S. aureus isolated from horses.

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Eight hundred eight Sprague-Dawley rats were examined for ophthalmic abnormalities during a pretest period in various preclinical safety assessment studies. Persistent pupillary membrane, corneal crystal, healed minor trauma, synechia, coloboma of the iris, lens luxation, cataract, persistent hyperplastic primary vitreous, vitreous hemorrhage, coloboma of the optic disc or choroid, remnant of hyaloid arterial system, retinal hemorrhage, retinal detachment, retinal folding and choroidal defect were observed. The incidences of corneal crystal, synechia, and nuclear cataract in this survey were higher than those reported previously. On the other hand, retinal folding in this survey was less common than that reported previously. These results suggest that background data of eye problems in albino rats should be accumulated in each own laboratory colony. In addition, since spontaneous eye problems are common in young albino rats, elimination of rats with ophthalmic abnormalities from study groups by an ophthalmic examination during a pretest period would facilitate to evaluate toxicity potential of test compounds in safety assessment studies.

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Eight calves were inoculated with 1O⁶ or 1O⁷ sporulated oocysts of Eimeria subspherica, Christensen, 1941. All the calves, except one, were successfully infected. Prepatent period was 10 to 11 days with a mean of 10.6 days and patent period was 1 to 5 days with a mean of 2.9 days. Peak oocyst discharge was found on days 11 and 12 after inoculation. The number of oocysts passed in the feces was very small and varied among calves, and no relationship was detected between the inoculum dose and the number of oocysts passed in the feces. No clinical symptoms were associated with the infection in any calves. Infectivity, reproductive potential and patbogenicity of E. subspherica were suggested to be low in the experimentally infected calves, even though relatively large numbers of oocysts were given. The sporulation process of E. subspherica oocysts was observed under a condition of 29°C with constant aeration. Completely sporulated oocysts increased in percentage following a sigmoid curve, and the expected 50% and 95% sporulation times were determined as 161 and 194 hours, respectively, by regression analysis.

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DBA/2FG-pcy mice developed the chronic renal failure by the progressive polycystic formation at five months old. Their bilaterally enlarged kidneys occupied about 15% of the body weight. It was about 9 times larger than the normal kidney of DBA/2N mice. A large number of various-sized cysts appeared in cortex and medulla of bilateral light-yellow kidneys of sponge-like shape. Blood urea nitrogen and creatinine increased. Red blood cells (746±39×10⁴/mm³), hemoglobin (8.8±0.4 g/dl) and hematocrits (31.1±1.5%) were lower than those of normal control mice. Serum erythropoietin level and reticulocytes did not increase. In addition, the treatment with exogenous erythropoietin improved the anemia in DBA/2FG-pcy mice. It was suggested that the anemia in DBA/2FG-pcy mice was due to the disorder of erythropoietin production caused by the progressive polycystic formation in kidneys.

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Postnatal development of auditory brainstem response (ABR) in spontaneously epileptic rats (SER; zi/zi, tm/tm), a double mutant, their parent mutants, tremor rats (tm/tm) and zitter rats (zi/zi), and Kyo:Wistar rats (control), was studied by repeated ABR recordings from 2 to 13 weeks of age. In tremor rats and Kyo:Wistar rats at 2 weeks of age, ABR pattern was almost the same and 2 or 3 waves were identified. However, no further development in ABR was observed in tremor rats nd prolongation of latency and lowering of amplitude were marked. Only wave I with prolonged latency and lowered amplitude was recognized in zitter rats, SER and SER-N (zitter phenotype without epilepsy; zi/zi, tm/+ or zi/zi, +/+). ABR pattern was almost the same from 2 to 13 weeks of age in them. Electrocochleography revealed delay of N1 latency of compound action potential in SER. Vacuolation was observed in the cochlear nuclei and the brainstem of tremor rats and SER from 3 weeks of age in histopathological examination. The organ of Corti and cochlear nerve of tremor rats and SER exhibited no remarkable histopathological findings. The deafness in SER and its parent mutants was considered to occur as a result of impairment of inner ear, cochlear nerve and brainstem as well.

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A survey of infectious coryza of chickens was performed in West Jawa of Indonesia between 1987 to 1988 by the detection of hemagglutination-inhibition (HI) antibody against Haemophilus paragallinarum (Hpg) in non-vaccinated healthy laying (12 farms, 196 chickens) and native (8 farms, 197 chickens) chickens. HI antibodies against Hpg were detected not only in the native chickens but also in the laying chickens, regardless of the district, and were observed in 70% (14/20) of farms and 19% (73/393) of chickens. HI antibodies against serotype A were detected from 11 farms (55%) and 11% (45/393) in chickens. Those against serotype C were detected from 5 farms (25%) and 8% (30/393) in chickens. Three Hpg strains were isolated from different chickens affected with infectious coryza. Two of them were identified as type A and the other as type C by the rapid plate agglutination test. These results demonstrated that the outbreaks of infectious coryza caused by serotype A and C strains had occurred in Indonesia.

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Micrencephalic neonatal pups were obtained from pregnant Crj:CD (SD) rats once treated with 5 mg/kg of N-methyl-N-nitrosourea (MNU) or 40 mg/kg of methylazoxymethanol (MAM) on day 12, 13, 14 or 15 of gestation (vaginal plug = day 0). They were reared by their own mothers and were subjected to various neurobehavioral tests during the suckling period, days 0 to 22 after birth. The brain weights in the MNU- and MAM-treated pups on postnatal day 22 were significantly less than those in the control pups. These micrencephalic pups were retarded in neurobehavioral ontogeny. By several tests, each of them showed an impaired performance such as paired limb movement, clumsy locomotion or hyperreflexive reaction. These behavioral disorders appeared different according to the day of treatment, without any substantial difference between the test compounds, MNU and MAM. The findings suggest that the different neurobehavioral characteristics in the micrencephalic pups may reflect their different brain disorders induced by the test compounds given on the different period of the treatment.

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A persistent, spasmic and productive cough known as filarial cough often occurs in dogs with dirofilariosis, and has been considered to be the consequence of an allergic response to Dirofilaria immitis. Twenty-one dogs with filarial cough were subcutaneously injected with worm antigen (200 μg of protein concentration) extracted from adult D. immitis once a day for 5 days. These injections were effective for 17 (81%) of the dogs, resulting in a complete cure for 7 dogs and marked improvement for 10 dogs.

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Hepatitogenicity of three plaque purified mutant strains of mouse hepatitis virus, designated as MHV-2S, -2M and -2L, isolated from MHV-2 infected SR-CDF1-DBT cells was studied. After intraperitoneal inoculation with 2×10⁵ PFU of parental MHV-2 and its mutants to 4-week-old female ICR mice, 40% of mice inoculated with MHV-2S and 20% of mice with -2M died in one week, whereas with -2L all mice survived. All mice inoculated with MHV-2 died in 3 days postinoculation (p.i.). Virus titer of the liver of mice inoculated with MHV-2, -2S and -2M reached peaks (MHV-2:10⁷ PFU/0.2g, -2S:10⁵ PFU/0.2g and -2M:10⁶ PFU/0.2g) at 96 hr p.i., while with -2L a peak titer (10³ PFU/0.2g) was shown at 48 hr p.i. Immunofluorescence revealed MHV spedific antigen in the liver of MHV-2S infected mice in and around necrotic areas though less extensive than that of parental MHV-2 infected mice. With MHV-2M specific fluorescence was restricted in degenerated hepatocytes in the small necrotic foci. In mice inoculated with MHV-2L only faint fluorescence was detedted. Histopathologically, in the liver of MHV-2S infected mice zonal necrosis and cell infiltration were observed. There were spotty necrosis and focal cell infiltration in the liver of MHV-2M infected mice and only small inflammatory foci were seen in MHV-2L infected mice. Large number of extracellular virions were detectable in MHV-2S but notin -2M and -2L infected mice by electron microscopy.

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The temperature-sensitive (ts), thymidine kinase-deficient (TK^-) mutant designated ZHtsTK^- strain, of Aujeszky's disease virus (ADV) was isolated from a virulent strain with the treatments using 5-bromodeoxyuridine and arabinosylthymine. The ZHtsTK^- strain was easily distinguished from the other virulent ADV strains by plaque size on HmLu-1 and chicken embryo fibroblast cells and by restriction endonuclease analyses using Bam HI, Sal I and Kpn I. The ZHtsTK^- strain was avirulent for mice, guinea pigs and rabbits, and produced neutralizing antibodies to ADV in these animals. The rabbits inoculated with the ZHtsTK^- strain did not shed detectable amounts of virus after dexamethasone treatment. The ZHtsTK^- strain was also avirulent for 5-day-old piglets and did not cause disease. No virus was detected from the piglets inoculated intramuscularly in the nasal swabs or the tissues examined on postinoculation day 9. These findings presented here suggested that there is a significant correlation between pathogenicity and properties such as ts and TK^-, and the combination of ts and TK^- properties plays a much larger role in reducing virulence for animals.

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The safety and efficacy of the cell-associated (C-A) vaccine prepared by chicken embryo fibroblast (CEF) cells infected with the tissue-culture-modified strain of infectious laryngotracheitis (ILT) virus were studied in chickens. Over seventy percent of chickens inoculated with the C-A vaccine by the subcutaneous (S.C.) or intramuscular (I.M.) route at 1 day of age was protected against challenge with a virulent strain of ILT virus without any clinical signs. Chickens vaccinated with the C-A vaccine at 1 day of age acquired immunity within 6 days after vaccination, and the protection rate maintained more than 60% until 10 weeks post-vaccination. The C-A vaccine was invariably effective for chickens at various age. There was no evidence that the development of immunity was hindered by further vaccination with Newcastle disease and infectious bronchitis combined live vaccine. In addition, the C-A vaccine was safe when chickens wefe inoculated with 10 doses. In the field trials of the C-A vaccine, no adverse reaction was obserbed, and over 65% of vaccinated chickens was protected against the challenge of the virulent ILT virus at 8 weeks after vaccination.

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Junctional complexes of the epithelia lining the rete testis, efferent ductules, connecting ductules and epididymal duct in the fowl were examined by trandmission electron microscopy and by a tracer method using lanthanum nitrate. The junctional compldxes were composed of tight junctions, adhering junctions and desmosomes. In the rete testis, one or two points of membrane fusion were observed at the tight junctions. In the efferent and connecting ductules and epididymal duct, the tight junctions consisted of a series of punctate membrane fusions. The adhering junctions and desmosomes showed no rematkable structural differences among these excurrent ducts. Vascularly infused lanthanum nitfate penetrated into the tight junbtions of individual epithelia for variable distances, but was prevented from entering the lumen at the site of membrane fusion. These results suggest that the tight junctions can restrict the diffusion of materials via the paracellular route, and that they play an important role in maintaining a suitable fluid environment within the excurrent ducts.

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Microsomal arylhydrocarbon (benzo[a]pyrene) hydroxylase activity in hyperplastic nodules in the livers of rats was decreased by feeding 2-acetylaminofluorene in their diet to 10% of that without 2-acetylaminofluorene feeding. By immunoblotting analysis with the antibodies raised against P-450/B[a]P, which is a form of cytochrome P-450 catalyzing benzo[a]pyrene hydroxylation in liver microsomes of untreated rats, it was shown that P-450/B[a]P content was decreased in the microsomes prepared from the nodular tissues. Microsomal UDP-glucuronyltransferase activity toward phenolic xenobiotics such as 1-naphthol and 4-nitrophenol was increased by 4.5-5.0-fold in the nodular tissues. This increase was inhibited by the addition of antibodies raised against GT-1, a form of UDP-glucuronyltransferase glucuronizing phenolic xenobiotics. Immunoblotting analysis with the antibodies against GT-1 showed that a protein band corresponding to GT-1 was increased in the microsomes from nodular tissues. The increased UDP-glucuronyltransferase also had about 4, 000 daltons "high mannose" oligosaccharide(s) like GT-1. The decreases of P-450/B[a]P and increases of GT-1 were observed mainly in nodular foci of the liver tissues. These results indicate that in liver nodules, decreased cytochrome P-450-dependent benzo[a]pyrene hydroxylase activity and increased UDP-glucuronyltransferase activity toward phenolic xenobiotics result from the decrease of the P-450 corresponding to P-450/B[a]P and the increase of the GT corresponding to GT-1.

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The kinetics of the antigen production of Chlamydia psittaci strains Izawa-1 and Pigeon-1041 (p-1041) was examined every 6 hr after infection up to 48 hr, by the indirect immunofluorescent antibody technique using monoclonal antibodies (MAbs). All three genus-specific antigenic determinants on lipopolysaccharide (LPS) appeared during the whole growth cycle. Antigenic determinants on proteins were, on the other hand, detected at various time periods from the early to the late stages of infection. However, a cross-reactive antigenic determinant on protein recognized by a MAb 3E9 was also detected during the whole growth cycle, similar to that on LPS. The time of appearance of common antigenic determinants on proteins of Izawa-1 and P-1041 was examined using cross-reactive MAbs, and it varied depending on heterologous and homologous MAbs. From the relationship between the detection of antigenic determinants and the morphological changes of chlamydial particles revealed by electron microscopy during the growth cycle, the antigenic determinants on proteins of Chlamydia psittaci were divided into two groups; one was specific to the elementary body and the other was coexisting in both the elementary body and the reticulate body.

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The complex carbohydrates in the bovine duodenal glands were examined histochemically using 5peroxidase-labeled lectins to achieve a better knowledge of their glycoprotein profiles. The presence of at least 2 different cell types and secretion of neutral mucin were observed in the bovine duodenal glands. The duodenal gland cells located in the central area of the lobules contained neutral glycoproteins with different saccharide residues as β-D-gal-(1-3)-D-galNAc, α-D-galNAc, β-(1-4)-D-glcNAc, NeuNAc and D-galNAc. The other cell types located in the peripheral area of the lobules contained α-L-Fuc, α-D-galNAc, β-(1-4)-D-glcNAc, NeuNAc and D-galNAc. These findings seemed to be characteristic of a unique digestive process of the ruminant.

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Two new experimental models of transplantable mouse sarcoma 180 were developed in ICR mice in order to examine the optimum transplantation sites and methods. The cervicodorsal hypoderm was evaluated as the best transplantation site for mouse sarcoma 180 among seemingly usable transplantation sites such as groin, armpit, cervicodorsal, abdominal and lumbodorsal hypoderms by hypodermic transplantation. In addition, the lung transplantation model was established by monitoring the survival period as a reliable parameter for evaluation of anti-tumor effects.

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