Between November 1997 and February 2000, winter migratory waterfowls of several species staying in San-in district, western Japan were surveyed for influenza A virus and paramyxovirus at four stations. A total of 18 influenza A viruses was isolated from 1,404 fecal samples of whistling swans, pintails, mallards, and white-fronted geese. Five different hemagglutinins and eight neuraminidases were identified in the viruses isolated, in 11 different combinations, including H7N8 related to a subtype of a highly pathogenic chicken virus. In 2000, five lentogenic (non-pathogenic) Newcastle disease viruses were also isolated from white-fronted geese. These results suggested that possible precursor viruses for highly pathogenic avian myxoviruses are still brought into Japan by migratory waterfowls. The results also support the contention that continued surveillance of wild waterfowl population should be an integral part of control policies for these serious poultry diseases.

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The effectiveness of the polymerase chain reaction (PCR) as a field application test for the eradication of contagious equine metritis (CEM) was evaluated. Seven-thousands five-hundred and thirty-four genital swabs were collected from 4,026 Thoroughbred broodmares and stallions in Japan to test "high risk" horses as well as for general surveillance testing from 1998 to 2001. Bacterial isolation as well as PCR testing of original specimens and cultured specimens was performed for detection of Taylorella equigenitalis from genital swabs. As a result, T. equigenitalis was detected in 12 mares and 1 stallion by PCR, although the bacteria were isolated from only 2 of the PCR-positive mares. CEM-infected and carrier horses were treated by a combination of chemotherapy and surgery. Subsequent follow-up testing over a 3-year period did not detect T. equigenitalis. It was demonstrated that PCR testing was more sensitive than isolation as a method for the detection of T. equigenitalis from genital swabs of horses in the field. It was therefore suggested that a combination of PCR testing and treatment were useful measures in the eradication of CEM from Japan.

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The role of birds as sources of Shiga toxin-and intimin-producing Escherichia coli was studied. Fecal samples from live gulls (n=86), pigeons (n=33) and broiler chickens (n=199) from 23 flocks were analyzed for stx and eae by PCR. No stx positive samples were detected. In contrast, eae E. coli were highly prevalent among gulls (40%), and was also found in pigeons (7%) and chickens (57% of the flocks contaminated). The eae positive isolates were analyzed genetically and O-serogrouped. One isolate from a pigeon was found to have stx _2f. The isolates of gulls differed from those of pigeons and chickens, and all eae E. coli isolates from birds differed from human pathogenic strains by the lack of EHEC-hlyA and bfp/EAF as well as distribution of O-serogroups. Thus, birds cannot be regarded as important carriers of zoonotic stx or eae E. coli in Finland.

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Serum leptin concentrations were measured in antenatal and postnatal cows housed at two different locations. The mean serum leptin concentration was 9.2 ± 0.6 ng/m l (n=22) in one group, and was slightly lower in the other (7.4 ± 0.4 ng/ml, n=54), probably because of the different nutritional conditions between the two groups. There was no consistent variation in relation to the menstrual cycle and the periparturient period in both groups. Moreover, serum leptin concentrations during the periparturient period were independent of the number of delivery and the incidence of mastitis and milk fever. These results are quite different from those in rodents and human, suggesting the different regulatory mechanism of circulating leptin concentration in cows.

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Activities of malate dehydrogenase (MDH) and aspartate aminotransferase in the malate-aspartate shuttle were significantly increased in the cytosolic fractions of livers with early neoplastic symptoms such as swelling and discoloration in transgenic mice carrying the prototype human c-Ha-ras gene, Tg-rasH2 mice, which were administered with diethylnitrosamine (DEN) as a carcinogenic chemical at a dose of 200 mg/kg body weight. Cytosolic MDH/LDH (lactate dehydrogenase) (ML) ratio increased significantly and was considered to be a useful marker to characterize the energy metabolism at early neoplastic stage in livers of the Tg-rasH2 mice.

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To detect the anti-P53 antibodies of dogs with tumors, a GST-recombinant canine (rc) P53 fusion protein was expressed and purified. Immunoblot analysis was performed using this GST-rcP53 fusion protein as an antigen and serum samples from dogs suffering from tumors as primary antibodies. Out of 16 serum samples obtained from various tumor cases, four samples showed reaction with GST-rcP53. In contrast, serum from other 12 dogs with tumors, four dogs with non-neoplastic diseases and two control healthy dogs (as controls) did not show any reaction with GST-rcP53 in immunoblotting. The p53 gene mutation and the P53 protein expression were examined, using the tumor tissues to explore the relationship between the existence of the GST-rcP53 bands, gene mutations of p53 and the accumulation of P53 protein. One case, which showed a clear GST-rcP53 band, had a point mutation of the p53 cDNA and showed nuclear accumulation of P53 protein. These results suggest that the anti-P53 antibodies are also produced in tumor dogs with p53 gene mutations.

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The traditional metabolic profile test cannot be applied to peripartum dairy cows, because these cows are in a state of physiological abnormality making it difficult to interpret their blood components. This study aimed at establishing and evaluating the practicability of interpreting a metabolic profile test every 10 days (Ten-day criteria) during the dry and lactation periods in herds with high and no incidence of peripartum diseases. Data from 29,043 cows in 1,130 commercial dairy herds were used to establish standard values every 10 days, mean ± 1.0 standard deviation for the metabolic profile test. The practicability of these criteria was evaluated in herds with peripartum diseases. In the ten-day criteria, the body condition score, albumin, blood urea nitrogen, glucose, total cholesterol, nonesterified fatty acids, γ-glutamyl transpeptidase and aspartate aminotransferase, fluctuated during the dry and early lactation periods and there were very big changes in packed cell volume, blood urea nitrogen, total cholesterol and magnesium just after calving. The ten-day criteria were able to detect overconditioned cows, low levels of albumin, total cholesterol and magnesium, and high nonesterified fatty acids in herds with a high incidence of peripartum diseases. In conclusion, the ten-day criteria can be successfully applied to peripartum cows, and is recommended because it is able to detect metabolic abnormalities not only in the herd, but also in individual cows.

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In order to confirm the in vivo effectiveness of anti- feline herpesvirus type 1 (FHV-1) mouse-cat chimeric antibody (FJH2), and anti-feline calicivirus (FCV) mouse-cat chimeric antibody (F1D7), cats that had been experimentally infected with FHV-1 or FCV were administered intravenously with the chimeric antibodies, and observed for clinical manifestations. The symptoms due to FHV-1 or FCV infection in the cats administered FJH2 or F1D7 were obviously decreased when compared with those of the non-administered control cats. From these results, it was confirmed that both FJH2 and F1D7 were effective at reducing the appearance of symptoms due to FHV-1 and FCV infection, respectively.

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Polymorphonuclear cells of the peripheral blood in the chicken significantly increased their phagocytosis when cultured with sugar cane extract (SCE; 250-1,000 μg/ml) for 24 hr. Chickens orally administered SCE (500 mg/kg/day) for 3 or 6 consecutive days at 1 week of age showed significantly higher body weight and gain in body weight/day and a lower food conversion ratio within the growing period of 6 weeks than physiological saline-administered control chickens. Furthermore, oral administration of SCE also resulted in significantly higher immune responses against sheep red blood cells and Brucella abortus. These results suggest that SCE has immunostimulating and growth promoting effects in chickens.

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The natural resistance associated macrophage protein 1 (Nramp1) has been reported to confer resistance or susceptibility to Mycobacterium bovis, Salmonella typhimurium, and Leishmania donovani in the mouse, Mus musculus. A Gly and Asp substitution at position 169 of the mouse Nramp protein is invariably associated with the resistant and susceptible phenotypes, respectively. The present study aimed to detect polymorphisms in the NRAMP1 gene from different cattle and buffalo breeds. Genomic DNAs from five breeds of cattle and four breeds of buffalo were used in the study. Sequencing showed two nucleotide substitutions found in intron 4, three in exon V, and ten in intron 5. An amino acid substitution was observed at nucleotide position 1202 in exon V of the Japanese black, Angus, Philippine and Bangladesh swamp-type buffaloes which coded for Thr, while the Korean cattle, Holstein, African N'dama, Indonesian swamp-type buffalo and the Bangladesh river-type buffalo had Ile. All the breeds of cattle and buffaloes tested in this study coded for Gly at the position in exon VI which corresponds to the same amino acid of the murine Nramp1-resistant phenotype at position 169. The phylogenetic relationship among the different breeds showed a cluster comprised mainly of cattle and another one mainly of buffaloes.

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To clarify heart rate modulation by the sympathetic nervous system, dogs with naturally acquired and experimentally induced heart failure were examined. Heart rate and plasma catecholamine concentrations were measured in clinically healthy dogs (control dogs) and dogs with mitral regurgitation (MR) during a resting period, a standing period, a period of standing in a medical examination room (to which the dogs were unaccustomed), a running period, and a period of recovery after running. The heart rate and plasma catecholamine concentration increased in control dogs during the standing period and the medical examination room period, relative to the resting period. However, dogs with MR did not exhibit any clear increase in heart rate or catecholamine concentration under these light stress conditions. Running stress increased plasma catecholamine levels in control dogs; however, dogs with MR did not exhibit any significant changes. Thirty-two dogs with naturally acquired heart disease were classified as grades I to III on the ISACHC scale. The degree of increase in heart rate and plasma catecholamine levels in dogs with naturally acquired heart failure depended on their degree of heart failure. In conclusion, an increased heart rate and an activated sympathetic nervous system were observed, even in mild heart failure. This chronically activated sympathetic activity is expected to increase myocardial oxygen consumption, myocardial hypertrophy, and fibrosis, and to portend a poorer prognosis in heart failure.

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An intact male Pointer aged 9 year was found to have atrial fibrillation (AF). Thoracic radiography and echocardiography as well as routine blood work revealed no cardiac disease. Thus, the dog was diagnosed as lone AF. The arrhythmia disappeared 6 hr after the initiation of the supportive therapy, and relapse was not observed.

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A dog with Japanese cedar (Cryptomeria japonica, CJ) pollinosis had oral allergy syndrome (OAS) after ingesting fresh tomato. The dog showed specific IgE to both CJ and tomato allergens. As a negative control, twenty dogs without atopic dermatitis that had no exposure to tomato and no specific IgE to CJ allergen were used. They had no specific IgE to tomato allergen. Furthermore, IgE cross-reactivity was observed between CJ and tomato allergens in the dog. We found that OAS induced by tomato exists in the dog and there is a relationship between CJ and tomato allergens.

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To determine how blood values in bottlenose dolphins changed during the year, 504 blood samples were taken from 9 dolphins from 1991 to 1999 and clinical blood examinations were undertaken monthly including 3 hematological and 19 serum chemistry tests. In creatinine, significant seasonal changes were found among three groups of adult males, adult females and juveniles, and the average values in summer were 15-38% higher than those in winter. In two out of three groups the average total cholesterol value were highest in winter, and the lowest of all groups were in summer. In two other groups the peaks of average FFA value were recorded in summer, and the lows were in winter.

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A total of 1,013 feces samples and 8 mesenteric lymphonodus samples obtained from apparently healthy dogs were examined for the incidence of salmonella infection. One strain of S. typhimurium (ST) was isolated from feces of one dog, and S. enteritidis (SE) was isolated from the mesenteric lymphonodus of one dog. Sera obtained from 330 apparently healthy dogs were examined for Salmonella antibodies using an ELISA with heated whole cells of SE and ST. Fifty-one of the 330 serum samples were considered to be positive for salmonella antibodies, including 12 which were SE-positive and 39 which were ST-positive. These results indicate that dogs cause possible environmental problems as Salmonella carriers.

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Immunohistochemical localization of TGF-alpha and cell proliferation kinetics during liver regeneration after two-thirds partial hepatectomy (PH) were investigated. Twenty-four to 72 hr after PH, appreciable increase in the number of TGF-alpha-positive hepatocytes was observed in zones 1 and 2. At the peak at 36 hr, almost all positive cells were stained in their nuclei. Considerable increase in the BrdU labeling index was observed 24-36 hr after PH with a peak at 24 hr in zones 1 and 2. These results indicated an association between TGF-alpha expression and hepatocyte regeneration. It is suggested that immunohistochemical localization of TGF-alpha may be a useful marker of cell proliferation activity in rat liver.

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The protective capacity of the cotton rat, Sigmodon hispidus, against the migratory and intestinal phases of Strongyloides venezuelensis infection was examined. After subcutaneous infection with infective larvae (L₃), adult worm recovery rates from male and female animals on Day 71 were only 0.10% and 0.06% of initial dose, respectively. To determine whether this enhanced protection was expressed during the migratory phase or the intestinal phase, larval recovery from the lungs of cotton rat was evaluated 3 days after subcutaneous L₃ infection. The larval recovery rate was only 0.5% of initial dose and about 40-fold lower than that from control mice. Protection in the intestine was also evaluated after intraduodenal implantation of adult worms. About 30% of implanted worms became established and worm burden remained constant until Day 28. Despite a high worm burden on Day 28, EPG was about 25-fold lower than the peak count. To evaluate expulsive capacity and monitor the cellular responses in the intestine of cotton rats, adult Nippostrongylus brasiliensis worms were implanted in addition to S. venezuelensis. Cotton rats were unable to expel adult S. venezuelensis worms, even after 21 days of observation. Although the number of mucosal mast cells increased significantly, the intraepithelial migration of mast cells was not observed. In contrast, N. brasiliensis was expelled by Day 6 in association with goblet cell hyperplasia. These results suggest that in contton rats, the defective intestinal protection against adult S. venezuelensis worms results from dysfunction of mucosal mast cells.

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Production of conjugated linoleic acid (CLA) by the intestinal bacteria of dogs and cats was demonstrated by incubating their feces with linoleic acid (LA). CLA accumulated once, and then decreased with time. The numbers of LA-hydrogenating bacteria in the intestines appeared to decrease greatly with the ages of dogs and cats. As a major product of LA biohydrogenation, trans-vaccenic acid (t-VA) was identified. Most CLA and t-VA were readily solubilized by shaking the incubation mixture with bovine serum albumin, which strongly supports the presumption that CLA and t-VA are mostly formed on the outer surface of cell membrane, or excreted to the outer cell surface. This result suggests that CLA and t-VA can readily be absorbed through the large intestines. Triacylglycerol and phospholipid were shown to be hydrolyzed to free fatty acids by fecal bacteria, which is critical for biohydrogenation to occur, because esterified LA is not hydrogenated. However, since the ability of intestinal bacteria to produce CLA is probably low, it is desirable to augment CLA production.

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A total of 267 fecal and serum samples collected from individual pigs reared on a Salmonella-positive farm were subjected to bacteriological and serological examinations of Salmonella. Salmonella was isolated from 47 pigs (17.6%) and prevalence of antibody to lipopolysaccharide (LPS) of S. Typhimurium, which was partly common to S. O4, 12: d: -, was observed in 90 pigs (33.7%). Salmonella was isolated from 26 (28.9%) of 90 antibody-positive pigs and 21 (11.9%) of 177 antibody-negative pigs. Twenty-one of 36 pigs (58.3%) positive for S. O4, 12: d: -, five of 10 pigs (50.0%) positive for S. Havana, and none for S. Anatum had antibodies. Thus, seropositive rates were higher than isolation-positive rates, and antibody prevalence was associated with serovars of the isolates. Then, we analyzed antibody prevalence among pigs on Japanese pig farms. The antibodies to LPS of S. Typhimurium were found in 195 of 1,498 pigs (13.0%) and in at least one serum sample on 35 of 52 farms (67.3%). Our results indicate that Salmonella does not seem to be so prevalent in pigs though it is widely prevalent among pig farms.

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The residue of aflatoxins in the liver, muscle and eggs of laying ducks, hens and quails and in broiler chickens was examined by conducting 7-day feeding experiments with a diet containing 3 ppm Aflatoxin B1 (AFB1). Birds were sacrificed on the 8th or 11th day of AFB1 feeding. AFB1 and its metabolites in the tissues and eggs were determined by HPLC. The tissue levels of AFB1 and its metabolites were higher in quail than in the other birds. The levels of AFB1 and its metabolites, including acid-hydrolyzable metabolites, were more than 10-fold higher in the liver than in the muscle in all the species. The ratio of AFB1 in the feed to the residual level in the liver was 383 in quail, but was ≥ 5769 in the other birds. The corresponding ratios in the egg yolk and albumen of AFB1 were 4615 and 3846, respectively, in chicken hens, and these values were higher than those in the other birds.

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Meat samples and fecal specimens from adult cattle were collected in Changchun, China and were examined for presence of Shiga toxin-producing Escherichia coli (STEC) serogroup O157. STEC O157 strains were isolated from 2 (5%) of 40 beef, 1 (3.3%) of 30 pork, and 3 (1.7%) of 176 adult cattle fecal samples. The strains belonged to phage types (PT) 4, 8, or 47. Two beef strains and a strain previously isolated from a patient in Shandong, China, were PT-4 and showed a similar PFGE pattern, suggesting the possibility of food-borne transmission. It is suggested that cattle are a reservoir of STEC O157:H7 and meat products are contaminated by this pathogen in Changchun, China as well as in other countries.

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The inotropic, chronotropic, and vasodilatory effects of five commonly used cyclic adenosine 3',5'-monophosphate (cAMP)-modulating agents were evaluated. Hemodynamic functions were measured continuously in isoflurane-anesthetized cats during infusion of the following: dobutamine (DOB; 2.5, 5 and 10 μg/kg/min; n=8), dopamine (DOP; 1.25, 2.5, 5 and 10 μg/kg/min; n=5), milrinone (MIL; 2.5, 5 and 10 μg/kg/min; n=8), 6-(3-dimethyl-aminopropionyl) forskolin hydrochloride (COL; 0.2, 0.4, 0.8, and 1.6 μg/kg/min; n=7), and bucladesine sodium (BUC; 10, 20, and 40 μg/kg/min; n=9). At the highest infusion rate, DOB and DOP produced the greatest positive inotropic (increase in left ventricular (LV) dP/dt = 89 ± 4% and 75 ± 6%, respectively) and chronotropic (increase in heart rate (HR) = 42 ± 4% and 22 ± 6%, respectively) effects. MIL and COL produced similar albeit less pronounced positive inotropic (increase in LV dP/dt = 18 ± 3% and 22 ± 6%, respectively) and chronotropic (increase in HR = 13 ± 4% and 21 ± 4%, respectively) effects. Both also had significant vasodilatory effects (decrease in peripheral resistance (PR) = -30 ± 2% and -35 ± 7%, respectively). In contrast, BUC produced only vasodilatation (decrease in PR = -33 ± 6%). Hence, MIL, COL, and BUC had significant vasodilatory effects and less-pronounced inotropic effects than the catecholamines DOB and DOP. The vasodilatory effects of non-catecholamine drugs for treatment of congestive heart failure should translate into beneficial decreases in both pre-load and after-load. In contrast, the strong inotropic effects of DOB and DOP should be beneficial in the treatment of acute heart failure and anesthetic crisis.

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Three experiments were conducted to determine the effect of Vascular Endothelial Growth Factor (VEGF) on bovine embryonic development in vitro. Human recombinant VEGF₁₆₅ was employed at 5 ng/ml in modified synthetic oviduct fluid. In Exp. 1, bovine cumulus oocyte complexes were matured with or without VEGF for 22 hr, inseminated without VEGF for 6 hr, then cultured with or without VEGF for 42 hr. The cleavage rate and the development rate to 4- to 8-cell were higher (P<0.05) in groups with VEGF during in vitro maturation (IVM, 71.4% and 59.6%), in vitro culture (IVC, 70.3% and 62.3%), and both IVM and IVC (75.9% and 67.8%) than in the group cultured thoroughly without VEGF (49.9% and 38.4%, respectively). In Exp. 2, 4- to 8-cell embryos produced in vitro without VEGF were removed from cumulus cells at 48 hr post-insemination (Pi) and cultured with or without VEGF for 144 hr. The development rates to blastocyst at 96 hr (D6), 120 hr (D7) and 144 hr (D8) were similar (P>0.05) in both groups. In Exp. 3, cumulus cells were removed from presumptive embryos produced by IVM and IVF without VEGF at 10 hr Pi. Denuded embryos were cultured with or without VEGF for 38 hr or 182 hr. The cleavage rate and the development rates to 4- to 8-cell at 48 hr Pi and to blastocyst on D6, D7 and D8 were similar (P>0.05) in all groups. These results suggest that VEGF has a beneficial effect on the initial development of bovine embryo through surrounding cumulus cells.

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A long-acting prostaglandin F_{2 α} analogue, fenprostalene, was subcutaneously administered singly at doses of 5, 10, 25, 50, 100 or 150 μg on day 25 after ovulation during pregnancy in bitches (n=4 for each dosage), and maintenance of pregnancy, changes in plasma progesterone concentration, interval between the treatment and subsequent estrus and conception rate at that estrus were studied. Abortion was associated with a decrease in the plasma progesterone concentration below about 2 ng/ml, and the abortifacient effect was dose-dependent. Administration of 50 μg or more of fenprostalene induced abortion in all dogs 3 to 13 days after the treatment. The interval between administration and subsequent estrus was markedly shorter in the aborted bitches than in the non-aborted ones (P<0.01). The conception rate at the estrus in the aborted dogs was 50%, whereas all of the bitches who had not aborted became pregnant. The results indicate that a single administration of fenprostalene could induce abortion during mid-pregnancy in bitches, and the subsequent estrus may come early with a low conception rate.

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In combination of utilizing a leader specific primer and primers complementary to porcine reproductive and respiratory syndrome virus (PRRSV) genome through RT-PCR, the leader junction sequences of subgenomic mRNA (sg mRNA) was identified from a Taiwanese isolate of PRRSV. Thirty-six cDNA clones derived from sg mRNAs 2, 3, 4, 5, 6 and 7 were determined. The junction sequences analyzed from different sg mRNA were found to contain a similar 5 nucleotide sequence motif, (U/G)(C/A)(A/G)CC. The distance between the junction site and the translation initiation codon of the down stream open reading frame varied from 4 to 226 nucleotides. Minor heterogenecity was observed in the nucleotide sequence surrounding the junctions from all six sg mRNA analyzed. However, for sg mRNA 7, two junction sites approximately 103 nucleotides apart from each other were identified. The additional site is a new junction not previously reported in sg mRNA 7 from other PRRSV strains.

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