The length of the small intestine, cecum, and the rest of the large intestine of specific-pathogen-free (SPF) swine was compared with that of conventional swine. The average length of the small intestine of SPF swine was shorter than that of conventional swine. The difference was significant for female SPF swine. There was no difference between SPF and conventional swine in the length of the cecum, colon and rectum.
In the present study, osteoclasts were isolated from hen medullary bones at the formative and resorptive phases. The cells were cultured on glass culture dishes and bone slices. After culturing, the adhesion activity of the isolated osteoclasts with the substrates was estimated with a light microscope, and the surfaces of the bone slices were observed with a scanning electron microscope. The results showed that the adhesion activity of tartrate-resistant acid phosphatase (TRAP)-positive osteoclasts is higher at the bone resorptive phase than at the bone formative phase, and this tendency in isolated osteoclasts was observed more frequently on the bone slices than on the glass culture dishes. Furthermore, scanning electron microscopy showed that the isolated osteoclasts in the bone resorptive phase adhered to the bone surface with developed-cytoplasmic projections and formed broad pits where collagen fibrils were exposed. On the other hand, isolated osteoclasts in the bone formative phase adhered to the bone slice with board-shaped cytoplasmic projections and did not form any pits. These results suggest that isolated osteoclasts in the bone resorptive phase have a high level of adhesion activity and actively resorb the bone, whereas isolated osteoclasts in the bone formative phase have a low level of adhesion activity and cease bone resorption. The procedure reported here is useful for studying the bone-resorptive mechanism of authentic osteoclasts.
We investigated the expression of novel protein kinase C (PKC) δ and θ in the testes of pigs and cattle using Western blot and immunohistochemical analysis. PKC δ and θ are recognized in the testes of pigs and cattle by Western blot analysis. We found in immunohistochemical study that PCK δ was localized in the spermatids of seminiferous tubules, but not in the interstitial cells, while PKC θ was recognized only in the interstitial cells of the testes of in both species. These findings suggest that PKC δ and θ play an important role in the development of spermatozoa and the regulation of androgen in the testicular interstitial cells (probably Leydig cells), respectively.
Diarrhea, sudden death after short duration of diarrhea and sudden death without apparent signs were observed in a herd of breeder pigs. Five pigs that died suddenly with diarrhea (SDD pigs) and 6 pigs that died suddenly without signs (SD pigs) were examined. The average age of the pigs was about 28 days. Twelve pigs of age 10 to 14 days old showing diarrhea (D pigs) were also examined. Eleven of them recovered. Large numbers of Escherichia coli were detected in all organs of every SDD and SD pig and in feces of D pigs. All of the isolates were identified as enterotoxigenic E. coli (ETEC) by the polymerase chain reaction (PCR). Porcine reproductive and respiratory syndrome (PRRS) virus cDNA was also detected from the lung of every SD and SDD pig by the RT-PCR. High and low titers of antibodies to PRRS virus were found in 10-day-old and 1-month-old pigs, respectively. In an experiment, 3 ETEC were isolated from 9 healthy weaning pigs during the quiescent stage in the herd. These data showed that growth of the ETEC was not active in healthy weaning pigs; however, following infection with PRRS virus ETEC infection became systemic and caused peracute death in the weaning pigs. It suggested also that infection with PRRS virus in 10-day-old pigs were protected by the colostral antibodies, and fatal infection by ETEC did not occur as a result.
A 30 kDa immunodominant surface antigen (p30) of Babesia equi has been used as a diagnostic antigen. The B cell epitopes on this molecule recognized by horse sera and monoclonal antibody (MAb) against p30, 36/133.97, were determined. A synthetic peptide of p30 with amino acid sequence of 123FYQEVLFKGFEAV 135 exhibited strong positive reaction with the infected horse sera. In contrast, MAb 36/133.97 recognized different region of p30, as peptide synthesized with amino acid sequence of 27ASGAVVDFQLESI 39 reacted strongly. In competitive inhibition ELISA, the binding of MAb 36/133.97 to recombinant p30 was inhibited by horse antibodies, although they did not recognize same or an overlapping epitope. The data on B cell epitopes in this study may be important in improving serodiagnostic methods of B. equi infection.
Two cats with abdominal effusion and anorexia were diagnosed as feline infectious peritonitis (FIP). We tried to evaluate the effect of thromboxane (Tx) synthetase inhibitor, ozagrel hydrochloride, on the progression of symptoms and clinicopathologic data characteristic to FIP. After administration of Tx synthetase inhibitor, improvement of appetite and activity, decreases of peritoneal effusion, reduction of leukocyte number to normal level, and improvement of hyper γ-globulinemia were found in 2 cats with FIP. These findings suggest that the vasculitis in FIP can be successfully treated with Tx synthetase inhibitor which inhibits platelet aggregation.
The location of a specific epitope in the SU proteins (gp70s) of PVC (PVC-111, PVC-211, PVC-321 and PVC-441) murine leukemia viruses (MuLVs) among various MuLVs was determined with chimeric viruses between PVC-211 and F-MuLV and a rat monoclonal antibody to the gp70 of PVC-321. The epitope resided in the N-terminal region from amino acid position 1 to 67 of SU protein and did not correlate to the tropism against brain capillary endothelial cells of the F344 rat or Chinese hamster ovary cells.
Gastrointestinal glutathione peroxidase (GSHPx-GI) is an enzyme expressed in intestinal epithelial cells and may reduce hydroperoxides generated from the ingested diet. We isolated a genomic clone containing the mouse Gpx2 gene encoding 190 amino acids of GSHPx-GI. This gene is composed of two exons and an intron. The nucleotide sequence of the coding region was 89.9% identical with that of the human GPX2 gene. A TGA opal codon predicted to encode a selenocysteine was identified at codon 40. A genomic clone containing a pseudogene for the Gpx2 gene was also isolated. The nucleotide sequence of the pseudogene was 98.3% identical with that of the mouse Gpx2 gene and showed characteristics of a processed pseudogene. Linkage analysis using backcross mouse progeny indicated the mouse Gpx2 gene and its pseudogene to be located on mouse chromosomes 12 and 7, respectively.
Cryptosporidian oocysts were surveyed in rectal stools of adult cattle which were carried into slaughterhouse from April 1995 to July 1996. We morphologically and histologically investigated oocysts, and experimentally infected the isolated oocysts to mice and rats. Cryptosporidian oocysts were detected from 24 of 512 cattle (4.7%). They were spherical or ovoid, and the size was 7.0-7.9 × 5.3-6.1 μm. Mice and rats inoculated orally with 105-107 oocysts became infected and discharged oocysts in the feces for a period of more than two months. Developing parasites were detected from the stomach of mice, and not detected from the other digestive tract. From these findings, present isolates from cattle were identified as Cryptosporidium muris.
Detection of Echinococcus coproantigen using sandwich enzyme-linked immunosorbent assay (sELISA) was performed on fecal samples of red foxes in Hokkaido, Japan. Fecal samples were collected around fox dens in 1990 and 1992. The antibodies used for sELISA recognize heat-resistant antigens, thus all fecal samples were heated to render it safe for handling before examination. Detection of taeniid egg in fox feces collected was considered as an indication of E. multilocularis infection. In fecal samples collected in 1990 and 1992, coproantigen positive results out of taeniid-egg positive cases were 38/40 (95.0%) and 95/97 (97.9%), respectively. In addition, coproantigen was detected regardless of fecal condition when collected from the field, suggesting that the antigens detected by this method are quite stable. These results suggest that detection of coproantigen is useful for field surveys of foxes naturally infected with E. multilocularis.
A 13-month-old female Newfoundland dog suffered from urinary bladder tumor. Histologically the tumor consisted of round or fusiform cells, occasionally having eosinophilic cytoplasms. Apparent mature rhabdomyoblasts possessing elongated eosinophilic cytoplasm and cross striations were infrequently observed. The tumor cells exhibited immuno-positive for anti-myoglobin, desmin and vimentin antibodies. Ultrastructurally, tumor cells have abundant myofibrils in their cytoplasm and Z bands were also detected. The present tumor was diagnosed as a urinary bladder rhabdomyosarcoma in a Newfoundland dog, which has not been frequently reported in dogs.
Five-week-old ddY mice were inoculated intranasally with a low virulent (4e) or highly virulent (24a5b) avian influenza virus strain originated from a water bird. None of mice in the 4e group showed clinical signs and brain lesions. Of the 24a5b group, two mice died and one mouse was killed at a moribund state at day 7 post-inoculation (PI). Four mice of the 24a5b group necropsied at day 5 or 7 PI had mild to severe encephalitis in the brain stem and the cerebellar white matter. Influenza virus antigen was detected in neurons, glial cells and vascular endothelium in the lesions. The distribution of the lesions seems to indicate the transneuronal invasion of the virus via cranial nerve fibers into the brain.
The pharmacokinetics and tissue distribution of two norfloxacin (NFLX) formulations, norfloxacin-glycine acetate (NFLXGA) and norfloxacin nicotinate (NFLXN), were compared after single oral administration with a dose of 5 mg equivalent NFLX base/kg of body weight in twenty rabbits. The pharmacokinetic characteristics of all formulations were fitted by a two-compartment open model. The elimination half-life (T1/2b) of NFLX (3.37 ± 1.37 hr) was not significant as compared with those of NFLXN (3.61 ± 0.65 hr) and NFLXGA (3.93 ± 1.54 hr). The absolute bioavailability (F) of NFLX, NFLXN and NFLXGA was calculated as 29%, 45% and 40%. In addition, tissue distribution of NFLXN and NFLXGA did not show any differences of NFLX concentrations in liver, kidney, serum and muscle. From the present results, it could be suggested that NFLXN and NFLXGA are considered to be bioequivalent when they use oral medication for rabbits.
Jugular vein blood samples were collected from 23 young and sexual mature feral stallions to examine the relationship between plasma testosterone concentration and age, breeding season or harem size. Testosterone concentration increased with the age of the stallions until they formed their own harems, at about 4 to 6 years old. Seasonal variations in testosterone concentrations were observed, and found to be significantly higher (P<0.001) throughout the breeding season than non-breeding season, from 3 years of age. Testosterone levels were correlated with harem size for individual stallions. It can be inferred from these results that there is a relationship between plasma testosterone concentration and age, breeding season and harem size.
The clinical application of MRI of a cat case of traffic accident was examined. On admission, the animal was unconscious and remained so for 2 days. Radiographs disclosed a fracture in the parietal bone. From the temporary unconscious status and the fracture, cerebral damage was suspected and an MRI examination was performed. The contrecoup injury in the cat case of traffic accident which could not be diagnosed by radiography was diagnosed by MRI examination.
We analyzed the expression of mRNAs for growth factor [epidermal growth factor (EGF), insulin-like growth factor-I (IGF-I), basic fibroblast growth factor (bFGF) and platelet-derived growth factor A chain (PDGF-A)] and their receptor (R) by reverse transcription-polymerase chain reaction in bovine ova during oocyte maturation in vitro (0-21 hr) and after fertilization in vitro (6-144 hr: zygotes to blastocysts). Transcripts for EGF were not found before fertilization. Transcripts for IGF-I were present in immature oocytes immediately after collection and in embryos from the 2-cell stage onward. Transcripts for bFGF were present in all stages of oocyte maturation and after fertilization up to the 16-cell stage. Transcripts for PDGF-A were present in all stages of oocyte maturation and after fertilization up to the 2-cell stage. Transcripts for ErbB3 (a member of the EGF-R subfamily), and bFGF-Rα were present in all stages of oocyte maturation, after fertilization up to the 2-cell stage, and the blastocyst stage. Transcripts for IGF-I-R and PDGF-Ra were present in all stages of oocyte maturation and embryo development. The results of this study showed that eight different messages for growth factor and their receptor were detectable in bovine ova during oocyte maturation and/or after fertilization in vitro and their expression patterns were the gene-specific rather than the developmental stage of bovine ova.
Eight male dogs with both asthenozoospermia and teratozoospermia were used in this study. In experiment 1, semen was collected 10 times at intervals of 48 hr, 24 hr and 12 hr in 4 of the 8 dogs, and the semen quality was evaluated. In experiment 2, semen was collected 5 times at 24-hr intervals in the other 4 dogs. The spermatozoa collected on day 1 and day 5 were incubated for 4-6 hr in Canine Capacitation Medium, and the percentages of hyperactivated sperm (%HA), acrosome-reacted sperm (%AR), and the zona pellucida-binding sperm count (ZP sperm count) were assessed. The results of experiment 1 showed that the percentage of motile sperm increased and the percentage of abnormal sperm decreased markedly as the intervals between semen collections became shorter. When semen was collected at 12-hr intervals, the percentage of motile sperm increased from about 65%, the value before frequent collection was started, to about 80%, and the percentage of sperm with abnormal tails decreased from 30% to15%. In experiment 2, the percentages of HA and AR, and the ZP sperm count in specimens collected on day 5 were higher than those in specimens collected on day 1, and the differences in % HA and in ZP sperm count were significant (P<0.05). The results demonstrated that sperm motility, abnormality, and potential fertility in dogs with asthenozoospermia and teratozoospermia can be temporarily improved by frequent semen collection.
Histological variations of canine deciduoma which was induced in the non pregnant horn at several stages of unilateral pregnancy were examined. In the first half of the unilateral pregnancy, deciduoma was characterized by the cystic glandular hyperplasia corresponding to each of the stages in normal early placentation. In the second half, deciduoma could not be induced and few histological reactions were recognized. The endometrium looked normal for late diestrus with no growth of the uterine glands. These differences might reflect the latent strength of the uterine glands to proliferate and dilate in the stimulated periods.
The present study was designed to define whether maximal removal rate of indocyanine green (ICG Rmax), plasma cyclic 3',5'-adenosine monophosphate (cAMP) response to exogenous glucagon (peak to basal ratio of cAMP level: P/B cAMP) and plasma half-life of galactose (tl/2 galactose) can measure the hepatic functional reserve of fatty liver prepared in rats fed choline-deficient (9 weeks), 2% cholesterol (2 weeks) or 0.25% DL-ethionine (2 weeks) diet. Although changes in cholesterol and phospholipid values in serum during feeding periods differed among the models, histopathologic examinations in the liver of almost all animals revealed intermediate to severe fatty liver with or without fibrosis at each termination. ICG Rmax and P/B cAMP were significantly decreased in rats fed choline-deficient or DL-ethionine diet, implying reductions in hepatic functional mass and disturbances in hepatic cAMP production. Meanwhile, tl/2 galactose showed no change in any of the models, suggesting that glucose metabolisms in the models used may be preserved. These findings demonstrate that ICG Rmax and P/B cAMP can apply to measurement of hepatic surviving reserve of fatty liver with fibrosis.
Bovine follicle fluid and oocytes surrounded by follicular epithelial (FE) cells were collected from ovaries of two heifers persistently infected with bovine viral diarrhea virus (BVDV). BVDV was present in the follicle fluid at a higher titer than in serum. The oocytes were matured in vitro under culture conditions of 39°C in humidified air containing 5% CO2. In vitro fertilization was performed after 24 hr in culture (the day of insemination was defined as day l), and culture was continued through day 10. BVDV was present in the culture medium at titers of 102.25 to 103.25 TC ID50/0.l ml. The virus was also detected in FE cells collected on day 10. Viral antigen was demonstrated in the cytoplasm of FE cells by the indirect immunofluorescence technique. However, no BVDV was detected in the embryos on day 10. These findings suggested that the oocytes or embryos were unlikely to be infected with BVDV, but that the FE cells were infected with BVDV and supported virus replication in cattle persistently infected with BVDV.
It was suggested that 3 strains of bovine viral diarrhea virus (BVDV) isolated from persistently infected calves in Tochigi prefecture in Japan belonged to BVDV type II. It was recognized lack of PstI site on the 5'-untranslated region of genome of them as well as BVDV type II reported previously. Inoculated with the 3 strains, the calves showed the mild decrease of platelet counts which was specific clinical sign of BVDV type II. We should report that the 3 strains were the first BVDV type II isolated in Japan. Neutralizing antibody titers of the antisera against the 3 strains using laboratory strains as neutralizing virus were lower than those of them using homologous strains. Therefore, it was indicated that the difference between BVDV type I and BVDV type II in the antigenicity.
The diversity in selected regions of the transmissible gastroenteritis virus (TGEV) and porcine respiratory coronavirus (PRCV) genomes was analyzed among known TGEV and PRCV strains and field isolates. The N-terminal half of the spike (S) glycoprotein gene and open reading frames (ORF) 3, 3-1 and 4 were amplified by reverse transcriptase reaction and polymerase chain reaction (RT/PCR), and analyzed using restriction fragment length polymorphism (RFLP) patterns of the amplified DNA. Reference TGEV strains (Miller and Purdue) and a PRCV strain (ISU-1), and TGEV and PRCV field isolates were analyzed. Based on the size of the ORF 3, 3-1 and 4 RT/PCR products, TGEV and PRCV strains could be quickly and easily differentiated into three groups designated TGEV Miller, Purdue types and PRCV. By RFLP analysis of the N-terminal region of the S glycoprotein gene and ORFs 3, 3-1 and 4, TGEV and PRCV strains were differentiated into five groups using the restriction enzyme Sau3AI. Sequence analysis of a PCR product in the ORFs 3, 3-1 and 4 from virulent and attenuated Miller strains demonstrated additional differences in that region which have been correlated with a change in virulence of TGEV isolates.
The immunogenicity of the bovine leukemia virus (BLV) transactivator protein (tax) was studied by mapping its B-cell and T-cell epitopes. Peptides (18 to 20-mer) overlapping by 10 amino acids, spanning whole amino acid sequence of BLVtax were synthesized. Reconbinant BLV tax protein was used to immunize two different strains of mice, C57BL/6 and BALB/c. B-cell and T-cell epitopes of recombinant BLVtax protein was determined by screening all the 30 synthetic peptides, against immune serum in ELISA for antibody reactivity, and against immune spleen cells in lymphocyte proliferation assay for T-cell stimulation. Peptides with amino acids at position 111-130 and 131-150 were T-cell epitopes for C57BL/6 and BALB/c mice immune cells, respectively. B-cell epitope was mapped to amino acid sequence at 261-280 in both strains of mice. These results imply that BLVtax protein contains some of BLV- immunodominant epitopes and this information may be applied for designing an effective peptide vaccine capable of inducing neutralizing antibodies as well as cellular immunity.
Recombinant vaccinia virus (rVV) was constructed by inserting Rinderpest Virus (RPV) nucleocapsid (N) protein gene. The rVV expressed RPV-N protein in the rVV-infected cells. The rVV was shown to produce RPV-N-specific antibody in mice.