The existence of apathogenic strains of Yersinia pseudotuberculosis (Yp) has not so far been reported. Recently, the authors characterized new serogroups and a new subgroup in Yp, that is, O9, O10, O12, O13, O14 and O1c, and the pathogenicity of these new strains was of interest. A total of 137 strains of serogroups O1c, O6, O7, O9, O10, O11, O12, O13 and O14 of Yp were investigated for their pathogenicity in vivo and in vitro. Although catalase activity and the inv gene were detected in all strains except those of groups O13 and O14, only a few strains, from serogroups O6 and O10 caused severe infection in mice. The remaining strains caused no mortality or severe infection even when they grew in limited tissues of infected mice. All the strains of Yp not possessing the virulence plasmid pYV caused no severe infection in mice. It is evident that less pathogenic Yp exists and that not only pathogenic but also less pathogenic Yp organisms exist in the same serogroup.
An allelic variation of the genes encoding the protective outer membrane lipoprotein (omlA) in Actinobacillus pleuropneumoniae was investigated with polymerase chain reaction (PCR)-restriction fragment length polymorphisms. Primers for PCR were selected from a conserved sequence compared between the omlA genes of A. pleuropneumoniae serotypes 1 and 5a. A DNA fragment of 970 bp was amplified from the genomic DNA of all 12 serotypes of A. pleuropneumoniae. The amplified DNA sequence specifically hybridized under a low stringent condition to the cloned omlA gene of A. pleuropneumoniae. Digestion of the amplified DNA with the enzymes either HinfI or VspI yielded specific polymorphic patterns, allowing discrimination of all serotypes into five distinct groups.
One-day-old broiler chickens were given commercial chicken starters during first 4 weeks of growth. The chickens were then divided into two groups: one group was fed commercial intensive growers (control feed), and the other group was fed commercial broiler finishers (high-nutrient feed). Body weights were calculated weekly. At eight weeks old, blood samples were collected and heart tissues were examined histopathologically. Significant increases in body weight as well as pathological findings in the heart muscle and an elevation of serum creation kinase (CK) were observed in chickens fed with broiler finisher. Histopathological changes in heart fiber resembled that of sudden death syndrome, and this heart damage was thought to be related to the increase of CK levels.
The atrioventricular (AV) conductive functions were investigated before and after the pharmacological autonomic nervous blockade (PAB) through the electrophysiological study on six horses (AV block group) in which the dropped beats occurred more frequently (over 200 beats/24 hr) and on five horses (control group) in which the dropped beats occurred sporadically (under 200 beats/24 hr) or in which the arrhythmias were not recognized at all on the long-term electrocardiogram. As a result, AV conductivity, conduction time and refractoriness in the AV block group were lower, longer and higher, respectively, than those in the control group even after PAB. These results indicate the intrinsic AV conductive functions in the AV block group were withdrawing than in the control group.
Bitches were examined to see whether canine deciduoma could be induced at some reproductive stages with the different conditions of corpora lutea by inserting a silk suture into the uterine lumen. The bitches stimulated in the early and middle stages of diestrus or in unilateral pregnancy corresponding to these diestrous stages formed deciduoma at a high induction rate, however, no difference in the strength of decidual reaction between the pregnant and diestrous stages was recognized. On the other hand, no reaction could be seen in bitches in late diestrus, the late stage of unilateral pregnancy or the post partum repair phase in which stromal decidual cells similar to those of the rodentia can be seen. In already implanted uteri, however, no deciduoma was formed in the interplacental areas. Even though the corpora lutea were functional, new additional stimulations were not accepted at the interplacental area in which the uterine horn had already been influenced by fertilized ova. From these results, it was suggested that in the dog as well as the rodentia, the endometrium has to be under the influence of functional corpora lutea in order to form deciduoma.
The effect of ovariectomy in the early first half of the diestrus was examined on the induction or maintenance of suture-induced canine deciduoma. Ovariectomy immediately, or some days, after the insertion of suture had no effect on the induction or maintenance of deciduoma. Even when ovariectomy was performed within 4 days before insertion, deciduoma could be induced in spite of there being no ovary. However, when ovariectomy was done 4 or more days before suture insertion, the rate of deciduoma was decreased or no deciduoma was induced. These results indicate that the influence of the ovary on the endometrium may persist for at least 4 days after ovariectomy. Ovariectomy after the suture insertion had few effects. It is suggested that canine uterine glands in the early first half of the diestrus maintain a certain degree of self-proliferative ability even after ovariectomy, and thus canine deciduoma is not as dependent on the ovary that of the rodentia.
We cloned the part of the bovine PrP gene which contains the 5'-flanking region, exon 1, exon 2 and intron 1 to analyze its promoter region. The 5' non-coding region of the bovine PrP gene consisted of three exons and two introns, and its organization was similar to that of the mouse, rat and sheep PrP genes. The 5'-flanking region of the bovine PrP gene from the transcription start site to nucleotide position -88 was (G+C)-rich (78%) and contained three potential binding sites for the transcription factor Sp1, but no CCAAT-box or TATA-box. This region showed high homology (89%) with that of the sheep PrP gene, but relatively low homology (approximately 46-62%) with the same region of the mouse, rat, hamster and human PrP genes. The position from -88 to -30 within the 5'-flanking region of the bovine PrP gene showed major promoter activity. However, this region was able to function properly only in collaboration with the region at +123 to +891 of intron 1 of the bovine PrP gene.
Prohibitin is the protein which has an inhibitory function in cell growth, and its gene is suggested to be one of putative tumor suppressor genes. In this report, we described a partial cloning of prohibitin cDNAs from canine, feline, bovine, equine, and rabbit liver mRNAs by RT-PCR, and their homology analysis. The sequences of these RT-PCR products were compared with each other as well as those reported for human and rat. The homology in this region of prohibitin cDNA was approximately 90%, and the amino acid sequence of each RT-PCR product shared more than 95% identity. Therefore, it is concluded that all the RT-PCR products are a part of prohibitin cDNA of each animal.
Three-day-old, specific-pathogen-free (SPF) chicks were inoculated with the strains of influenza A/whistling swan/Shimane/499/83 (H5N3) via the air sac route. The strains had been passaged through air sacs or air sacs and brains of SPF chicks. Two experiments were undertaken to examine the pathogenicity of these strains and the development of brain lesions based on time-interval changes. In experiment 1, original strain (4e) showed low pathogenicity with mild respiratory signs and zero mortality. Air sac passaged strains (18a and 24a) of 4e demonstrated mortalities of 50% and 67%, respectively, and inoculated chicks showed hemorrhages and necrotic lesions in major organs. Air sac-brain passaged strain (24a5b) of 4e produced 100% mortality and severe nervous signs. Severe circulatory disturbance with multiple foci of necrosis in major organs including the brain was found in chicks inoculated with 24a5b. The 24a5b was analogous to highly pathogenic avian influenza virus in regard to its pathogenicity to chicks. Hence, low pathogenic influenza virus (4e) gradually aggravated its pathogenicity to highly pathogenic virus (24a5b) by air sac and brain passages. In experiment 2, chicks were inoculated with 24a5b, and the earliest histological lesion was the enlargement of the vascular endothelial cells at 18 hr post-inoculation (PI) followed by necrotizing encephalitis at 24 to 48 hr PI. Immunohistological staining revealed avian influenza virus antigen initially in the vascular endothelial cells and then in the astrocytes, neurons and ependyma.
In order to clarify the pathogenesis of porcine serum (PS)-induced rat liver fibrosis, three experiments differing in dose of PS or duration of treatment were performed on male Fischer 344 rats. The rats were given an intraperitoneal injection of PS twice a week for 3 to 16 weeks and euthanized 7 days after the last injection for each treatment group. Liver tissues from these animals were subjected to detailed morphological and immunohistochemical examinations. Biochemical tests on treated rat serum revealed an increase in globulin concentration but no elevation in AST, ALT and ALP activities. There were no relationships among the dose of PS, the extent of fibrosis, and the anti-PS antibody titer. A number of α-smooth muscle actin-positive non-myofibroblastic cells, desmin-positive cells, and lipofuscin-laden Kupffer cells were found around the central veins and in the fibrous septa. In advanced stages of fibrosis, a proliferation of elastic fibers were observed in the septa. These findings were considered to indicate gradually occurred hepatocellular necrosis. The vascular endothelial cells in the fibrous septa expressed factor VIII-related antigen, exhibited fenestration accompanied by basement membrane formation, and were surrounded by Ito cells. Most of the portal vein branches showed hypertrophic thickening of the smooth muscle layer, resulting in narrowing of the lumen. These vascular changes suggested that hemodynamic alterations of the intrahepatic circulation induced hepatocellular necrosis/apoptosis and played an important role in the pathogenesis of porcine serum-induced liver fibrosis in rats.
Male 5-week-old Wistar rats orally (po) administered with fusarenon-X (FX) 1.5 mg/kg and control rats po-treated with distilled water were sacrificed at 0-48 hr after gavage. FX-administered rats showed significant dilatation of the stomach with increased fluid contents at 1-24 hr postadministration (PA). Histopathologically, karyopyknosis of chief cells in the basal region of the gastric glands began to appear at 1 hr, and nuclear fragments were seen in the neck cell region at 1.5 hr PA. At 2-4 hr PA, apoptotic cells appeared diffusely in the neck region and focally in the basal region. Electron microscopy revealed that cells phagocytosing apoptotic bodies were the surface epithelia, undifferentiated neck cells, parietal cells and chief cells. No evidence was detected to show that parietal cells underwent apoptotic changes. The apoptotic lesions peaked at 4-6 hr PA, gradually subsided at 12 hr PA, and became minimal leaving apoptotic remnants in the basal region at 24 hr PA. At 48 hr PA, however, diffuse apoptotic lesions reappeared in the basal region at a level similar to that at 2-3 hr PA. This might be attributable to absorption of FX retaining in the stomach for 24 hr. In situ detection of DNA breaks by the terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) reaction was consistent with the histopathologic findings. Agarose gel electrophoresis of DNA fragments isolated from the gastric mucosae of FX-administered rats showed a ladder pattern after 1.5 hr PA and the pattern became more distinct at 2-4 hr PA.
Two-day-old specific-pathogen free chicks were inoculated with type A influenza virus (A/whistling swan/Shimane/499/83 (H5N3)) through the air sac. Inoculated chicks showed mild to severe diarrhea and lesions of pancreatitis and atrophy of the pancreas, thymus and bursa of Fabricius. One chick died on each of days 4, 6 and 14 postinoculation (PI). Reduced weight gain was conspicuous from day 22 PI. Positive immunoreaction to the virus antigen was detected in the pancreas, kidneys, liver, lungs and air sacs, and cecal lamina propria. Virus recovery persisted longer in the pancreas. Some of these findings conformed to those of stunting syndrome.
To diagnose the possibility of early estimation for fertility in bovine heterosexual twin females, we designed a new diagnostic program. The 9 freemartins (FM) and 5 normal females (Normal) were used in this study. All 14 cases, at 4 months of age, were given Pregnant Mare Serum Gonadotrophin (PMSG) and human Chorionic Gonadotrophin (hCG) 1.5-2 days later. Thereafter, the concentration of estradiol-17β (E2) was determined by RIA, and that of progesterone (P) was done by RIA and EIA (Ovcheck EIA Kit). The concentration of E2 in the Group of Normal rapidly increased after administration of PMSG, but in the Group of FM, the concentration of E2 changed in very low levels over 14 days. The concentration of P in the Group of Normal rapidly increased after administration of PMSG, but in the Group of FM, the concentration of P changed in very low levels over 14 days.
Fifty cattle were divided into 3 groups (I, II, III) according to FSH-R dosage used to induce superovulation between days 9 and 11 of the estrous cycle. Ova/embryo recovery rate did not differ significantly between groups I and II administered total dosages of 16 and 24 mg, respectively. However, the average number of recovered ova/embryos and transferable embryos was significantly lower in group III (administered a total dosage of 30 mg) (P<0.05) than groups I and II. These results suggest that intramuscular administration of 16 mg FSH-R over 3 days is sufficient for inducing superovulation in Japanese Black cattle.
We investigated whether cationic liposomes are efficient at delivering the gene for diphtheria toxin A-chain (DT-A) under the control of the long terminal repeat (LTR) of bovine leukemia virus (BLV) (pLTR-DT) into BLV-infected cells and are also suitable for in vivo use. The transfection activity of the cationic liposomes composed of N-(α-trimethylammonioacetyl)-didodecyl-D-glutamate chloride (TMAG), dioleoyl phosphatidylethanolamine (DOPE) and dilauroyl phosphatidylcholine (DLPC) (1:2:2, molar ratio) (TMAG-liposome) and liposomes composed of phosphatidylserine (PS) (PS-liposome) was evaluated by the luciferase assay using a plasmid which contains the coding sequence of firefly luciferase under the control of the SRα promoter (pSRα/L-AΔ5). The TMAG-liposome gave highly efficient transfection in the presence of serum. On the other hand, PS-liposome showed inferior efficiency. When BLV-infected cells were co-transfected with a fixed amount of pSRα/L-AΔ5-entrapped TMAG-liposome and various amount of pLTR-DT-containing TMAG-liposome, the luciferase activity in the BLV-infected cells was inhibited by the addition of pLTR-DT-entrapped TMAG-liposome dose-dependently. The cationic TMAG-liposome containing pLTR-DT was successively added to BLV-infected cells in culture. The number of viable cells was markedly reduced by the cationic TMAG-liposome containing pLTR-DT. On the other hand, TMAG-liposome containing pSRα/L-AΔ5 showed no such effect. pLTR-DT entrapped by the cationic TMAG-liposome was not digested by the treatment with DNase I and with serum. These results suggest that the cationic liposomes, such as TMAG-liposome, may be efficient transfection reagent for the BLV-infected cells and can be utilized for DT-A gene delivery into the BLV-infected cells in vivo.
Feline herpesvirus type 1 (FHV-1) gD-expressing Sf9 cells adhered to two feline cell lines, but not to the porcine, bovine, or canine cell lines tested. In addition, this adhesion activity was inhibited by a monoclonal antibody against FHV-1 gD. These results showed that the FHV-1 gD might bind to a specific-molecule(s) on the surface of feline cell lines. We discussed a possible importance of the FHV-1 gD in host cell restriction to FHV-1 infection.
The diagnosis of porcine epidemic diarrhea virus (PEDV) infection in the laboratory is rather fastidious because of difficulties in virus propagation. The feasibility of virus propagation in vivo is also limited by the handling of a number of samples at the same time. In this study, the detection of PEDV by reverse transcription polymerase chain reaction (RT-PCR) is described. The RT-PCR could detect up to 104 TCID 50/ml of PEDV and did not show any cross reaction with transmissible gastroenteritis virus or porcine rotavirus. Using this method, the detection of PEDV in experimentally inoculated piglets was possible as early as one day after inoculation. These results suggest that the RT-PCR could be applicable for a rapid diagnosis of PEDV infection.