Under pathological conditions such as autoimmune encephalomyelitis or autoimmune uveoretinitis, many T cells infiltrate the central nervous system (CNS) and retina. Even in normal condition, a small number of T cells are detected in the CNS. However the characteristics of the T cells are not defined. To investigate the T cell characteristics in a healthy retina, the chicken and the embryo were observed by morphological and immunohistochemical methods. In the chicken retina, T cells were regularly detected, and the main subset was CD-8+/ γδ cells. Developmentally, CD positive cells appeared on embryonic day 13, and the constituent T cell repertoires became the same as in the chicken by embryonic day 17. Many T cell repertoires were detected on embryonic day 15 and 16. The present results confirm that the retina receives an immunological surveillance by T cells. The composition of T cells in retina is constructed after embryonic day 17. Many ganglion cells die in embryonic days 15 and 16. So the T cell subsets in these periods may involve in autoimmune diseases.
The efficacy of Mycoplasma hyopneumoniae oral vaccine was investigated in microsphere dosage form. A co-spray drying process was used to apply an encapsulating material, Eudragit L30 D-55, to microspheres containing Mycoplasma hyopneumoniae antigens. The microspheres were generally effective (>93%) with protein release at pH 7.4, but almost none were released at pH 1.2, for 3 hr in an in vitro dissolution test. An SPF-swine model was used to evaluate the effectiveness of the microspheres as an oral vaccine, and the related immune responses. The serum's systemic IgG against M. hyopneumoniae was evoked by ELISA analysis, after a 2nd immunization of all pigs. The vaccinated groups' mean lesion score was significantly lower after the Mycoplasma hyopneumoniae challenge than that of the nonvaccinated/challenged groups (P<0.05). This study strongly suggests that the oral microspheres vaccine prepared by a co-spray drying method can provide effective protection against M. hyopneumoniae infection in pigs.
To isolate Campylobacter spp., the feces of healthy cattle, pigs, and broilers were examined between June 1999 and January 2000. Campylobacter lanienae strains were isolated from the feces of healthy pigs, but not from the feces of cattle or broilers. In six C. lanienae isolates, there was only 21-38% DNA-DNA homology to Campylobacter hyointestinalis subsp. lawsonii strain NCTC 12901. Thus, the primary host of C. lanienae is likely to be the pig and C. lanienae appears to be a species distinct from C. hyointestinalis subsp. lawsonii. In addition, an intervening sequence of 226 bp in the 16S rRNA gene was found in four isolates.
To characterize amino acid polymorphisms in sheep prion protein (PrP), we analyzed the PrP genes from 271 sheep of 4 breeds (Khalkh, Yeroo, Orkhon and Khangai) raised in central Mongolia (Tuv, Uvurkhangai and Selenge prefectures). A total of 16 genotypes and 8 allelic variants of the PrP gene at codons 112, 136, 154 and 171 were found. At codon 171, 1.8% of the sheep had arginine/arginine (R/R) (resistant to scrapie) and 66.8% had glutamine/glutamine (Q/Q) (susceptible to scrapie). Several Yeroo and Orkhon sheep raised in Selenge prefecture had valine at codon 136 (136V) (highly susceptible to scrapie). Several Yeroo, Orkhon and Khangai sheep raised in Selenge prefecture had histidine at codon 154 (154H). Novel polymorphisms of valine (V) and serine (S) at codon 127, lysine (K) at codon 171, and leucine (L) and arginine (R) at codon 189 were also found in Khalkh, Yeroo and Orkhon sheep. It is not known whether these novel polymorphisms affect scrapie susceptibility.
An epidemiological investigation of an outbreak of intestinal atresia in Israeli Holstein-Friesian newborn calves showed a linkage with rectal palpation for early pregnancy diagnosis, performed less than 42 days after insemination. The odds of an exposed calf, i.e., one born to a dam that was diagnosed by early palpation as having intestinal atresia were 119.7 times higher than one born in normal control herds (95% CI; 7.4-1946.3). A total of 682 calves-at-risk was recorded from mid-1998 to mid-2000 and a total of 47 calves (6.9%) were born with intestinal atresia during this period. Two forms of intestinal atresia were recognized at post-mortem: atresia coli and atresia ilei.
It has been suggested that the sympathetic nervous system communicates with lymphocytes expressing cell surface receptors for neurotransmitters such as norepinephrine (NE), on the basis of the finding that neurotransmitters modify immune responses in mammalian species. We confirmed that chicken lymphocytes in the brusa of Fabricius, thymus and spleen expressed β-adrenergic receptor (β-AR) mRNA from embryonic day (E) 10 and that intracellular cAMP level was elevated by NE, suggesting that lymphocytes express functional β-AR on their surface at an early embryonal stage. To clarify whether the nervous system is involved in the development of the immune system, the effects of 6-hydroxydopamine (6-OHDA), one of sympathectomizing agents, on chicken lymphocytes was investigated. A single injection of 6-OHDA at a dose of 400 μg into a chicken embryo was carried out at E7 or 14 (as referred to E7 group and E14 group, respectively). NE level and the relative proportion of Bu-1a+, CD4+ and CD8+ cells in the spleen of 3-week-old chickens were not altered by 6-OHDA treatment. However, the proliferative responses and expression of IL-2 mRNA in spleen cells cultured with pokeweed mitogen were reduced in E7 group compared with those of control. Furthermore, in CD8+ spleen cells of E14 group of 3-week-old chickens, the expression of β-AR mRNA and the relative increase of intracellular cAMP stimulated with NE were significantly decreased. These results suggest that the sympathetic nervous system affects the development of the immune system.
The effects of sugar cane extracts (SCE) on immune responses in chickens were studied. Two- or 10-month-old chickens orally administered SCE (500 mg/kg/day), for 3 consecutive days before immunized with sheep red blood cells, Brucella abortus and Salmonella Enteritidis organisms, showed significantly increased and prolonged antibody responses to these antigens, compared to control chickens without SCE. Furthermore, chickens orally administered SCE also revealed enhanced delayed type hypersensitivity responses to human gamma globulin. These results indicated that SCE has immunostimulating and adjuvant effects in chickens.
To evaluate the hemostatic effects of desmopressin (DDAVP) in dogs with aspirin-induced platelet dysfunction and hemostatic impairment in chronic liver diseases, 3 μg/kg DDAVP was administrated subcutaneously. In aspirin-induced platelet dysfunction dogs (n=5), prolonged BMBT (buccal mucosal bleeding time) was shortened significantly after DDAVP injection (2.2 ± 1.2 min, P<0.05). In dogs with chronic liver diseases (n=4), activated partial thromboplastin time (APTT) tended to shorten by 0.9 to 3.0 sec, and prolonged BMBT was shortened in two cases for 4.2 and 1.7 min after DDAVP injection. Therefore, the present results indicated that DDAVP shortened the prolonged BMBT in dogs with aspirin-induced platelet dysfunction and chronic liver disease. DDAVP might be helpful in hemostasis under invasive procedures such as biopsy or surgery for dogs with hemostatic impairment.
The canine Bcl-xL gene was cloned and sequenced. Canine Bcl-xL cDNA clone was 1252 bp in length, and encoded 233 deduced amino acides. The predicted canine Bcl-xL amino acid sequence shared 99.6%, 97.0%, 97.9%, 98.7% and 98.3% homology with that of human, mouse, rat, sheep and pig Bcl-xL, respectively. RT-PCR analysis revealed that canine Bcl-xL mRNA was constitutively expressed in CL-1 (canine lymphoma) and GL-1 (canine B cell leukemia) cell lines.
Phenotypes of lymphocytes from laparoscopically biopsied liver tissues of eleven healthy beagle dogs were analyzed. The proportion of CD3+ lymphocytes (T cells), CD3 -CD21+ lymphocytes (B cells) and CD3 -CD21- lymphocytes (non-T non-B lymphocytes), and the CD4+/CD8+ ratio in the canine hepatic lymphocytes were 54.8 ± 11.9%, 4.7 ± 3.1%, 40.7 ± 13.2%, and 0.33 ± 0.12, respectively, while those in peripheral blood lymphocytes were 85.4 ± 6.5%, 9.3 ± 6.1%, 5.3 ± 1.8%, and 1.64 ± 0.36, respectively. These results indicated that the constitution of hepatic lymphocytes quite differed from that of peripheral blood lymphocytes in dogs, and suggested that the regional immunity in canine liver might be specific.
Giardia has been detected in domestic dogs in Japan, but the genotype of isolates has remained unclear because identification has relied on conventional microscopy. Here we tried to identify the genotypes of four isolates from dogs in Japan by direct sequencing of the PCR amplified Giardia glutamate dehydrogenase (GDH) gene. The primer pair GDHF3 and GDHB5, targeting the GDH gene, was designed to prime a region of the GDH gene sequence conserved in the strains found to have the dog-specific genotype. The specific PCR product (approximately 220 bp), amplified with this primer pair, was only observed when Giardia DNA was used as the template. The sequences of the diagnostic fragments were identical among the isolates from dogs, and were differed by 15 bp or 1 bp from the strains, which were found to be the dog-specific genotypes, Assemblage C or D respectively. To verify the identity of the amplified DNA, a phylogenetic analysis was performed. Consequently, the sequence of the isolates from dogs clearly clustered with the strain found to be Assemblage D with neighbor-joining analyses. Therefore, all the isolates from dogs examined were identified as the dog-specific genotype, Assemblage D. In the present study, we revealed the genotype of Giardia isolates in Japan, and showed that direct sequencing of the PCR product amplified with the primer pair GDHF3 and GDHB5 was a useful tool for distinguishing between the zoonotic and dog-specific genotypes.
An epidemiological study was carried out in natural water supplies of Hokkaido, one of the largest dairy prefectures in Japan. To investigate the prevalence of Cryptosporidium parvum (C. parvum) oocysts water samples were collected from three rivers in the eastern area of Hokkaido from August 1999 to October 2001, and C. parvum oocysts were collected and purified by the ferric sulfate flocculation method. The oocysts were detected using the immunofluorescent assay test (IFAT) and 4', 6-diamidino-2-phenylindole (DAPI) staining. The seasonal change in the number of oocysts detected was observed. Oocysts increased in numbers from the late summer to the early autumn (from August to November), thereafter, they exhibited a trend to decrease until December, when no oocysts could be detected. The maximum number of oocysts detected in the three rivers was 3.50, 5.00 and 3.33 oocysts/l, respectively. The oocyst density in river water changed in relation to the season in 1999, 2000 and 2001. This report first cleared up the seasonal changes in C. parvum oocysts number in river water.
The prevalence of Cryptosporidium infection was examined in 480 healthy cattle (0-39 months old) in the Tokachi district in Hokkaido during the period from June to September in 2000 and from June to July in 2001. C. parvum oocysts were detected in 6 of 50 cattle (0-2 months old) in 2001; while C. muris was detected in 2 of 56 cattle (6-8 months old) in 2001, in 1 of 15 cattle (9-11 months old) in 2001, in 1 of 88 cattle (15-17 months old) in 2000, in 4 of 89 cattle (18-21 months old) in 2000 and in 2 of 53 cattle (21-23 months old) in 2000.
During a year from 1999 to 2000, a total of 247 blood samples were collected from 214 cattle and 33 water buffaloes in 16 distinct geographical locations of Thailand and analyzed by allele-specific PCR amplification of major piroplasm surface protein (MPSP) genes of benign Theileria parasites. Four allelic MPSP gene types were determined namely C-type, I-type, B-type and Thai-type, which were originally designated from Japanese Theileriaorientalis (Chitose, Ikeda), Australian T. buffeli (Warwick) and Thai T. sp. (Kamphaeng Saen), respectively. Only two allelic MPSP gene types were successively amplified from 204 (82.6%) blood samples. Among positive cases, 138 (67.6%) and 17 (8.3%) samples contained either Thai-type or C-type parasites, respectively, while 49 (24%) samples contained both types. However, nucleotide sequences of MPSP genes of Thai T. sp. amplified by C-type specific primers revealed higher (96.3%) similarity to Indonesian T. sp. rather than (87.8% similarity) to Japanese T. orientalis (Chitose) designated as C-type.
The effects of salivary gland extract (SGE) from R. sanguineus were examined on the production of IgG1 and IgG2 and the mRNA expression of IFN-γ, IL-2, IL-4, IL-5 and IL-10 in the mononuclear cells from canine peripheral blood, treated with concanavalin A (ConA) in vitro. SGE suppressed the ConA-induced production of IgG2. It also inhibited the expression of IFN-γ, IL-2 and IL-5 mRNA in a dose-dependent manner. No dose-dependent suppression was observed of IL-10 mRNA expression although a significant effect was observed at a SGE protein concentration of 25 μg/ml. SGE had no effect on the mRNA expression of IL-4. These results suggest that the suppression of IgG2 production by SGE from R. sanguineus was caused by the suppression of IFN-γ production.
A solitary brain mass of a 4-month-old miniature dachshund showing seizure-like neurological signs was examined histopathologically. At necropsy a white tumor mass, replacing the thalamus, approximately 1.5 cm in diameter, was found. There was cystic space filled with yellowish pale fluid in the central area of the tumor mass. Histopathological examination revealed that the mass consisted of irregularly arranged well-differentiated neuronal and glial cells, and multifocal mineral deposits. The neuronal cells had a large clear nucleus and various amount of Nissl substances in the cytoplasm. Some neural cells were bi-nucleated. Neither mitotic figures nor proliferating cell nuclear antigen (PCNA)-positive nuclei was found in the neuronal cells. Immunostaining for glial fibrillary acidic protein (GFAP) revealed diffuse proliferation of GFAP-positive glial cells and their processes, while these glial cells did not show apparent cellular atypism, mitotic activity, or PCNA-immunoreactivity. Accordingly, the present tumor was diagnosed as ganglioglioma, and hamartomatous histogenesis might be possible.
The role of rhoA/rho-associated kinase (ROK) signaling pathways in agonist-induced contraction of the rat myometrium was investigated. We measured the [Ca2+]i-force relationship, phosphorylation of myosin regulatory light chains (MLC20) in intact tissue and the Ca2+-sensitization of force in permeabilized myometrial cells of rat. In measurements of the relationship between [Ca2+]i and tension in intact tissue, Y-27632, a ROK inhibitor, significantly attenuated the carbachol-induced contraction without changing [Ca 2+]i. Phosphorylation of MLC20 was increased by carbachol and this increased phosphorylation was blocked by treatment of tissue with Y-27632. In tension measurements of single hyperpermeable cells, carbachol evoked sustained contraction at constant pCa 6.7 and these agonist-induced contractions were decreased by treatment with Y-27632. These results suggest that activation of a ROK-mediated signaling pathway(s) plays an important role in agonist-induced alterations in MLC20 phosphorylation and force of rat myometrium.
Calponin (h1 or basic) is an actin-binding protein that is expressed abundantly in smooth muscle. Our previous study using h1 calponin-null mutant mice demonstrated that h1 calponin inhibits the shortening velocity of smooth muscle contraction without significantly affecting the amplitude of force production. Furthermore, early onset of osteogenesis and increased bone formation have been reported in mutated mice. In the present study, we examined the effect of h1 calponin depletion on the metabolism and behavior of mice and found that the mutated mice showed increased locomotor activity, as well as increased intake of food and water, associated with the decreased number of neurons in the paraventricular nucleus of the hypothalamus (PVN).
The prophylactic efficacy of desmopressin acetate (DDAVP) on diabetes insipidus (DI) after hypophysectomy was investigated in the dog. In the control group, hypernatremia with a plasma level of 155 mEq/l or higher persisted for 12 hr from the 4th to the 16th hour after hypophysectomy, and symptoms of DI developed within five days after surgery. In the DDAVP treatment group, these changes were not observed, showing that administration of DDAVP (4 μg, instillation, twice daily) effectively prevented hypernatremia that develops immediately after surgery and DI-like symptoms that persists for about one week after surgery.
Experimental osteotomy model of canine tibia was prepared to investigate the changes in biomechanical characteristics during the healing process. After 16 weeks, although radiographs revealed that fracture healing proceeded, the recovery rates of the mechanical parameters ranged from 12.3 to 47.3%, compared to the intact side. After 32 weeks, those recovered to 68.9-93.2%. These results suggested that the biomechanical characteristics of the healing bone could not be recovered sufficiently even after the passage of the healing period which has been empirically proposed from clinical findings. It was also considered that evaluation of the mechanical parameters using this osteotomy model would be suitable for investigating the effect of osteoinductive growth factors on fracture healing.
The effect of electroacupuncture (EA) on minimum alveolar concentration (MAC) of isoflurane was evaluated in dogs. After determination of baseline MAC, EA was applied at each acupoints (LI-4, SP-6, ST-36 and TH-8) and nonacupoint for 30 min. MAC was determined again. EA at acupoints significantly lowered the MAC of isoflurane in dogs (17.5 ± 3.1%, 21.3 ± 8.0%, 21.2 ± 7.5% and 15.4 ± 3.1%, respectively). In control group and nonacupoint electrical stimulation group MAC were not decreased significantly. From these results, electroacupuncture at each acupoints used in the present study would have an advantage in isoflurane anesthesia with reducing its requirement.
To evaluate whether oocytes excluded from somatic cell nuclear transfer (SCNT) could be utilized for embryo production by parthenogenetic activation (PA), porcine oocytes with poor morphology after maturation culture were excluded from SCNT and subsequently used for PA with different stimuli. In the first set of experiment, either electric pulse of different strengths (1.75, 2.0 or 2.25 kV/cm for 30 μsec each) or chemicals with different treatment durations [7% ethanol for 5 min followed by exposure to 6-dimethylaminopurine (6-DMAP) for 0, 2, 3 or 4 hr] was employed. Development to the 8-cell and morula stages was significantly (P<0.05) improved by electric stimulation of 2.0 kV/cm, while blastocyst formation was enhanced by chemical treatment of ethanol and 6-DMAP for 4 hr. Subsequently, oocytes were parthenogenetically activated by one of four stimuli; 1) optimal electric (2.0 kV/cm for 30 μsec), 2) optimal chemical (ethanol followed by 6-DMAP for 4 hr), 3) electric then chemical and 4) vice versa. On the other hand, oocytes with normal morphology were subjected to the same experimental treatments for the control. Regardless of oocyte type, a combination of electric and chemical stimulations did not further stimulate preimplantation development, compared with electric activation only. However, combinational treatment greatly increased the cell number of blastocysts in SCNT-excluded oocytes (21.9 to 22.9 vs. 16.9 cells/blastocyst), while such effect was not found in normal oocytes (22.2 to 23.3 cells/blastocyst). In conclusion, porcine oocytes excluded from SCNT still have a potential to develop blastocysts after PA and this might contribute to increasing the efficiency of SCNT for various purposes. A combined activation by electricity and chemical yielded the best rate of preimplantation development with increasing the quality of blastocyst.
We investigated the therapeutic effects of a progesterone releasing intravaginal device (PRID) on cystic ovarian disease (COD) and reproduction performance of cows. The possible influence of PRID on metabolic and/or health status was also examined. A total of 40 Holstein-Friesian cattle, with ovarian cystic structures, ≥2.5 cm in diameter, persisting for more than 7-14 days, without a corpus luteum (CL) were used for the study. PRID or placebos were inserted into the vagina for 12 days. Five animals lost the intravaginal device before removal and one was culled. Based on plasma progesterone concentration on the day of treatment, 20 (17 PRID and 3 placebos) of the remaining 34 cows had follicular cysts (progesterone ≤1 ng/ml) and 14 (10 PRID and 4 placebos) had luteal cysts (progesterone >1 ng/m l). Fourteen (82%) of the PRID-treated follicular cystic cows responded with formation of a CL within 14 days after treatment, and an overall conception rate of 53.8%. Likewise, 70% of the treated luteal cystic cows responded with CL formation and 71.4% conception rate. No significant differences were observed in hematocrit (Ht), white blood cell count and serum levels of glucose, blood urea nitrogen, aspartate aminotransferase, and alanine aminotransferase, between the day of PRID insertion and removal, in animals with follicular and luteal cysts. PRID treatment resulted in ovulation 2-4 days later and formation of a CL in cows that recovered.
The objective of this study was to compare the number of recovered spermatozoa, in different parts of the uterine horn and oviduct in gilts, after insemination with fractionated (experiment) and non-fractionated (control) liquid stored semen. The number of spermatozoa and volume of backflow was also investigated. Twenty three cross-bred gilts were used in the study. They were divided into 2 groups, a control group (non-fractionated liquid stored semen, n=10) which were inseminated with 100 ml of liquid stored semen containing 3,000 million spermatozoa per dose and an experimental group (fractionated liquid stored semen, n=10) which were inseminated with 50 ml of liquid stored semen, with 3,000 million spermatozoa per dose and followed by another 50 ml of semen dilutor (Beltsville Thawing Solution, BTS). Thereafter, backflow semen was collected and measured every 15 min for a period of 1 hr. Three or 12 hr after insemination, 5 gilts from each group had the uterus, the horn of the uterus, the oviducts and the ovaries removed under general anaesthesia. The horn of uterus and the oviducts were seperated by ligation into 6 segments. All 6 segments were flushed with BTS to collect all spermatozoa within the segment. Recovered spermatozoa were counted, using a haemocytometer and the volume recorded. It was seen that the percentage of spermatozoa in the backflow semen in the experimental group was less than in the control group. The difference was not significant in the gilts that were operated on 3 hr after insemination, the mean number of spermatozoa in the uterine horn and the utero-tubular junction (UTJ) was more in the experimental than in the control group, but less in the isthmus and the ampulla of the oviduct. The gilts which were operated on 12 hr after insemination, had relativity more ovulating gilts in the control group than in the experimental group (3 of 4 gilts compare to 3 of 5 gilts). The control group had more spermatozoa in the oviduct than the experimental group, but less in UTJ and in the horn of the uterus. Again the difference was not significant. It can be concluded that fractionated (experimental) or non-fractionated (control) insemination of semen with the same number of spermatozoa provides no significant difference in the number of spermatozoa either in the horn of the uterus, the UTJ or the oviduct of gilts.
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-17β 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.
The Republic of Korea had been free from foot and mouth disease (FMD) since 1934, until a recent outbreak in 2000. From March to April 2000, a total of 15 FMD outbreaks due to the serotype O virus were recorded. Coincidental outbreaks of FMD in cattle or pigs by the serotype O virus were reported in the region, including Taiwan, China, Japan, Russia and Mongolia. In this report, the results of emergency investigations of FMD cases on a dairy farm located approximately 5-km from the demilitarized zone in Korea are described. The causative agent of the disease was identified as the FMD virus O by reverse transcription-polymerase chain reaction (RT-PCR) assays using primers derived from the 3D polymerase, internal ribosome entry site (IRES), 1D/2B regions, enzyme-linked immunosorbent assay (ELISA) for antigen detection and typing. Sequence data of the partial 1D/2B region obtained from vesicular fluid showed close similarity (98% sequence identity) to the Kinmen isolate of the FMD virus O in Taiwan. The causative virus was isolated using black goat fetal lung cells following propagation in unweaned mice.
The genetic relatedness of 7 Korean type O field strains of foot-and-mouth disease virus (FMDV) in clinical specimens collected from 5 different geographic locations in 2000 was investigated. The sequence of 162 nucleotides (nt 478-639) at the 3' end of the 1D (VP1) genes was determined from amplified cDNA fragments, and subjected to the analysis for the sequence identity/divergence and phylogenetic relationship. The overall nucleotide sequence divergence among the 7 field strains was 0 to 3.8%, suggesting that they are closely related to each other. Phylogenetic analysis with the known Middle East-South Asia (ME-SA) topotype strains showed that the 7 Korean field strains formed two distinct clusters within the same lineage of the ME-SA topotype strains. Cluster 1 consisted of the strains of the primary foci of infection (Paju and Hongseong), and closely related to the strains prevailed in the Far East. Cluster 2 comprised those of subsequently affected regions (Boryeong, Yongin, and Chungju), and was further diverged from the Cluster 1. The result of phylogenetic analysis indicated that the Korean strains may have evolved from a common ancestor of the Pan Asia strains, and that at least 2 phylogenetically clustered variants within the same lineage were prevalent during the epidemic. The potential origin and sources of the virus introduction to Korea were discussed.
Three cattle were experimentally infected with bovine herpesvirus type 4 (BoHV-4), strain B11-41, isolated from the spinal cord of a cow, and monitored for clinical symptoms. None of them showed any clinical signs except increases of leukocyte numbers in two of them, and the body temperature remained normal throughout the experiment. Antibody titers against BoHV-4 continuously increased for one month and were maintained at a high level for more than 1 year by enzyme-linked immunosorbent assay (ELISA). The virus was isolated only from serum and peripheral blood leukocytes (PBL) of one cow in the early stage of infection, but the viral genome was detected in PBL continuously by PCR. When they were euthanized, the viral genome was detected in the lymph nodes and nervous tissues such as medulla, spinal cord, and trigeminal ganglion. These results indicate that cattle are infected with the virus latently and persistently, and the latency site would be in the tissues of the central nervous system as well as lymphoid tissues. When a seroepidemiological survey was performed on antibodies to BoHV-4 among cattle in Japan by ELISA, the rate of antibody-positive cattle was 8.9% and they were found irregularly on certain farms.
The hemagglutinating activity and serological properties of three strains of rabbit hemorrhagic disease virus, Chinese, Korean and Shizuoka, which was first isolated in Japan, were examined by hemagglutination (HA) and cross hemagglutination inhibition (HI) test with human erythrocytes. Similar results were observed between the Chinese and Korean strains, both of which gave positive HA at 4°C with O, A, B and AB, and at 22°C with B and AB blood groups. In the Shizuoka strain, positive HA was observed at 4°C with O, A, B and AB, at 22°C with A, B And AB, and at 37 °C with B blood group. In experimentally infected rabbits, HI antibody in these animals showed a titer of 16,384 or 32,768 at 4 weeks after inoculation. No serological difference was observed in three strains by cross HI test.
Cytotoxic T-lymphocyte (CTL) responses to hemagglutinin (H) protein of canine distemper virus (CDV) were evaluated in dogs using the replication-deficient adenovirus protein expression system. Skin fibroblasts were isolated from two dogs and were infected with recombinant adenovirus bearing the CDV-H gene (Ade-CDVH). CTL assay was performed using fibroblasts expressing CDV-H protein as target cells and peripheral blood lymphocytes (PBL) collected from the same dogs one week after immunization of CDV as effector cells. Specific cytotoxic activity was observed against autologous but not heterologous fibroblasts expressing CDV-H protein. These results indicate that the CTL epitope(s) were localized in the H protein.