Evaluation of bovine spongiform encephalopathy (BSE) infection risk of cattle via sewage sludge from wastewater treatment facilities in slaughterhouses in Japan

Scattered SRM residues from BSE-infected cattle are possible to contaminate sewage during the slaughtering process in slaughterhouses. A proportion of the sludge discharged from wastewater treatment facilities at slaughterhouses has historically been processed into fertilizer. We therefore investigated the associated risk of BSE infection to cattle via sludge-derived fertilizer. Each stage of the process associated with BSE exposure was qualitatively evaluated and quantitative evaluations were subsequently performed using infectious dose as a unit of concern. Results of these qualitative evaluations indicated that installation of filter(s) at the drains to the wastewater treatment facilities has been undertaken by many slaughterhouses and has decreased the likelihood of SRM contamination of sewage. The level of sludge-derived fertilizer ingested by cattle was considered to be very low since the fertilizer is mixed with the ground soil, and the amount of soil ingested by cattle is likely to be small. Results from the quantitative analysis indicated the total infectious dose ingested by cattle in Japan from an infected cow has been estimated to be 5.5 x 10(-3) ID(50). Preventing scattering of SRM during the slaughtering process, installing filters to the drains with the removal of residues from the drain water and preventing the application of sludge-derived fertilizer to pasturelands would be effective to reduce the risk. Although the limited extent of available information, this study should provide useful indication for the development of an inclusive risk assessment for slaughterhouse sludge in the future.     

Normal calves produced after transfer of embryos cultured in a chemically defined medium supplemented with epidermal growth factor and insulin-like growth factor I following ovum pick up and in vitro fertilization in Japanese black cows

The objective of this study was to examine whether high concentrations of epidermal growth factor (EGF) and/or insulin-like growth factor I (IGF-I) would have a beneficial effect on bovine embryo development in vitro and to obtain normal calves by using an ovum pick up method and embryo culture in a chemically defined medium. When compared with controls, EGF (100 or 200 ng/ml) or IGF-I (50 or 100 ng/ml) significantly increased the rate of embryos that developed into blastocysts during an 8-day culture after the in vitro fertilization of oocytes obtained from ovaries from a slaughterhouse. IGF-I induced a dose-dependent increase in cell number in both the inner cell mass and the trophectoderm, whereas EGF stimulated proliferation only in the inner cell mass. A combination of EGF (100 ng/ml) and IGF-I (50 ng/ml) produced an additive effect, and embryos developed into blastocysts at a comparatively high rate (27.9%) compared with controls (12.0%). A similar rate of development was achieved using a combination of EGF and IGF-I in the culture of embryos following ovum pick up by ultrasound-guided transvaginal follicular aspiration and in vitro fertilization, and 5 blastocysts that developed after the culture were transferred into uteri; two embryos implanted, and normal calves were born. These results suggest that the combined use of EGF and IGF-I makes bovine embryo culture in a chemically defined medium a practical and useful procedure for producing blastocysts, and its application to embryo culture following ovum pick up and in vitro fertilization could be useful for producing normal calves.     

Estimation of glomerular filtration rate in calves using the contrast medium iodixanol

To develop a simple procedure for estimating glomerular filtration rate (GFR) in calves, a three-sample method using iodixanol was first compared to that using the standard agent inulin. Iodixanol and inulin were co-administered intravenously to calves at 40 mg I/kg and 40 mg/kg, respectively, and blood was collected 30, 60, 120, and 180 min later. Serum iodixanol and inulin concentrations were separately determined by high performance liquid chromatography and colorimetry. Serum urea nitrogen (UN) and creatinine concentrations were also measured. GFR estimated by iodixanol was consistent with that using inulin in clinically healthy calves. Based on GFR estimations in healthy calves and those renal-loaded with iodixanol, it was found that the serum creatinine concentrations became elevated when GFR decreased to 60% of the reference value. In contrast, serum UN concentrations fluctuated widely, presumably due to extra-renal factors. When GFR was estimated using the three-sample method and compared with the single-blood-sample method, 62/69 (90%) of samples tested were within the agreement plots. The results demonstrated that the single-blood-sample method using iodixanol may be useful in monitoring GFR in calves.     

Effects of option mitigating ammonia volatilization on CH4 and N2O emissions from a paddy field fertilized with anaerobically digested cattle slurry

A lysimeter experiment was carried out to evaluate the effects of the NH₃ volatilization mitigation by adding anaerobically digested cattle slurry (ADCS) alone, with wood vinegar (WV) or with a higher level of floodwater (HFW), on emissions of CH₄ and N₂O from a paddy soil planted with fodder rice. We have carried out the following treatments: (1) chemical fertilizer, (2) ADCS, (3) ADCS + WV, and (4) ADCS + HFW; the height of floodwater was 10 cm in the latter treatment, and it was 3 to 4 cm in the other treatments just before fertilizer applications. Nitrogen fertilizer rate added to soil in each treatment was 30 g NH₄⁺–N m⁻² (split in one basal and two top-dressing additions). Ammonia volatilization in the ADCS treatment was 2.7 g NH₃–N m⁻² throughout the growing season, and it was significantly reduced by 79% and 55% in the ADCS + WV and ADCS + HFW treatments, respectively. The total amount of CH₄ emitted in the ADCS treatment in the growing season was not significantly enhanced by the mitigation of NH₃ volatilization either by adding wood vinegar or by increasing the height of the floodwater. Negligible N₂O emissions were observed in all treatments during the growing period.     

Effect of trehalose supplementation in milk replacer on the incidence of diarrhea and fecal microbiota in preweaned calves

Trehalose, a nonreducing disaccharide consisting of d-glucose with α,α-1,1 linkage, was evaluated as a functional material to improve the gut environment in preweaned calves. In experiment 1, 173 calves were divided into two groups; the trehalose group was fed trehalose at 30 g/animal/d with milk replacer during the suckling period, and the control group was fed nonsupplemented milk replacer. Medication frequency was lower in the trehalose group (P < 0.05). In experiment 2, calves (n = 20) were divided into two groups (control group [n = 10] and trehalose group [n = 10]) based on their body weight and reared under the same feeding regimens as in experiment 1. Fresh feces were collected from individual animals at the beginning of the trial (average age 11 d), 3 wk after trehalose feeding (experimental day 22), and 1 d before weaning, and the fecal score was recorded daily. Fecal samples were analyzed for fermentation parameters and microbiota. The fecal score was significantly lower in the trehalose group than in the control group in the early stage (at an age of 14 to 18 d; P < 0.05) of the suckling period. Calves fed trehalose tended to have a higher proportion of fecal butyrate on day 22 than calves in the control group (P = 0.08). Population sizes of Clostridium spp. were significantly lower (P = 0.036), whereas those of Dialister spp. and Eubacterium spp. tended to be higher in the feces of calves in the trehalose group on day 22 (P = 0.060 and P = 0.083). These observations indicate that trehalose feeding modulated the gut environment and partially contributed to the reduction in medication frequency observed in experiment 1.     

Salsolinol is present in the bovine posterior pituitary gland and stimulates the release of prolactin both in vivo and in vitro in ruminants

The aims of the present study were to determine whether salsolinol (SAL), a dopamine-related compound, is present in the bovine posterior pituitary (PP) gland, and to clarify the effect of SAL on the secretion of prolactin (PRL) in ruminants. SAL was detected in extract of bovine PP gland using high-pressure liquid chromatography with electrochemical detection (HPLC-EC). A single intravenous (i.v.) injection of SAL (5 and 10mg/kg body weight) significantly and dose-dependently stimulated the release of PRL in goats (P<0.05). Plasma PRL levels reached a peak 10min after the injection, then gradually returned to basal values in 60-80min. The PRL-releasing pattern was similar to that in response to sulpiride (a dopamine receptor antagonist). The intracerebroventricular (i.c.v.) injection of 1mg of SAL had no significant effect on the release of PRL in calves, however, 5mg significantly stimulated the release (P<0.05) with peak values reached 30-40min after the injection. Moreover, SAL significantly stimulated the release of PRL from cultured bovine anterior pituitary cells at doses of 10(-6) and 10(-5)M, compared to control cells (P<0.05). Taken together, our data clearly show that SAL is present in extract of the PP gland of ruminants, and has PRL-releasing activity both in vivo and in vitro. Therefore, this endogenous compound is a strong candidate for the factor having PRL-releasing activity that has been previously detected in extract of the bovine PP gland.     

Effects of urinary excretion of nitrogen, potassium and sodium on urine volume in dairy cows

Data of 50 balance measurements were collected from dry and lactating Holstein cows to clarify the effects of urinary excretion of nitrogen (N), potassium (K) and sodium (Na) on urine volume in cows. In dry cows, orchardgrass silage, timothy hay, alfalfa silage and corn silage were offered to meet the TDN requirement. Orchardgrass silage or alfalfa silage diets were offered in switch back trials in lactating cows. There were no relationships between urinary excretion and plasma concentrations of K or Na in cows, but urinary N excretion increased with the increase of plasma urea nitrogen. There was positive correlation between urinary excretion and urinary content of Na, but urinary K contents increased rapidly by 1.3% with the increasing urinary K excretion and thereafter remained almost constant. The increasing urinary K and N excretion enhanced urine volume in cows, but urine production in cows was accurately estimated from the regression equation of urine K excretion on urine volume. Urine volume was not affected by urinary Na excretion. These results suggest that the increase of urine volume in cows affected by the increasing urinary excretion of K and N may be due to the maintenance of urine or plasma osmolality.     

Evaluation of coproantigen diagnosis for natural Echinococcus multilocularis infection in red foxes.

The validity of a coproantigen ELISA for Echinococcus multilocularis was evaluated by comparison of three diagnostic methods; autopsy, egg examination and the ELISA. Of 71 foxes, 39 were found to be infected with the cestode at autopsy. The overall mean of worm burdens was 3,451, but the number varied (1-34,522). The ELISA could detect 94.9% (37/39) of the worm positives and there were no false-positives. Two false-negatives were infected with 1 and 4 cestodes, whereas 3 cases with similar worm burdens (2, 4 and 6 worms) were diagnosed as positives. This indicates the detection limit of the assay may be equivalent to less than 10 (in the worm burden). On the other hand, egg examination showed low sensitivity (43.6%, 17/39). These results suggest the ELISA has a potential to replace for the conventional methods.     

Histopathology and physiopathology of gastric mucous hyperplasia in rats heavily infected with Taenia taeniaeformis.

Rats heavily infected with larval Taenia taeniaeformis show hyperplasia of the gastric mucosa accompanied by mucous cell proliferation, increase in the level of intragastric pH and hypergastrinemia. Sixty one rats were divided into 2 groups designed as infected (36 rats) and control (25 rats) group. These rats were examined with time course of the infection histopathologically and physiopathologically, during 14-112 days postinfection (DPI). In the infected rats, gastric mucosal hyperplasia began to be observed at 56 DPI, and the structural disturbance of zymogenic units in the corpus and mucous units in the antrum had increased with time. However, the degree of these changes in the antrum was weaker than those in the corpus. Alcianblue and/or PAS-positive cells increased in their numbers with time, and 4 types of cells other than typical surface mucous cell and mucous neck cell were observed by electron-microscopy. However, zymogenic and parietal cells decreased in their number after 56 DPI. Further, the infected rats showed changes in the serum concentration of alanine aminotransferase, aspartate aminotransferase, blood urea nitrogen, glucose and total protein. Some similarities with Menetrier's disease were discussed.