冷冻温度对牛精子复苏率及降温速率变化规律的研究

[目的]为了探讨温度变化对牛精子复苏率的影响,找到降温度变化规律。[方法]利用电脑程控冷冻仪、自动数字测温仪,测定精液在不同温度冷冻精子复苏率变化。[结果]表明,精液在-98℃冷冻精子复苏率最高;在-86- -118℃冷冻精子复苏率较高,精液相变适宜降温速率为35℃/min。[结论]控制精液发生相变阶段的降温速率是冻精制作成败的关键。     

牛卵母细胞体外成熟及体外受精的研究

本研究分析了卵巢不同性周期阶段（卵泡期、黄体期）、外源激素（ＦＳＨ、ＬＨ、雌激素）和不同卵泡液浓度（１０％,２０％,３０％）对卵母细胞体外成熟的影响。结果表明,卵巢的不同性周期阶段对卵母细胞体外成熟的影响不大（６３９％和５８７％）;成熟培养基中添加ＦＳＨ（１０ｍｇ／Ｌ）,ＬＨ（５ｍｇ／Ｌ）和同时添加ＬＨ、雌二醇（１ｍｇ／Ｌ）,可以促进卵母细胞的体外成熟,成熟率分别为６３０％,６４０％,６９１％;卵泡液（１０％,２０％,３０％）代替血清,其成熟率分别为５０７％,５７６％,５１９％,２０％ｂＦＦ效果最好。本研究对精子的两种处理方法进行了比较,发现在相同精子密度条件下（１×１０６～２×１０６个／ｍＬ）,Ｐｅｒｃｏｌ液处理精子的受精率（４７１％）显著低于上浮法处理精子的受精率（５７１％）（Ｐ＜００５）。     

奶牛精子表面电位的群体分布

应用细胞电泳方法,测量了不同电压下奶牛精子的电泳速度,绘制了两类精子在不同电压下的泳动速度分布图,计算了两类精子在磷酸缓冲液中的平均zeta电位,其中X精子为-27.9mV,Y精子为-22.4mV。     

Bovine viral diarrhoea virus in beef cattle herds of Yucatan, Mexico: seroprevalence and risk factors

A survey of bovine viral diarrhoea virus (BVDV) infection was carried out from June 2001 to July 2002 in a non-vaccinated beef cattle population from the livestock region of Yucatan, Mexico, to assess seroprevalence and identify risk factors related to seroprevalence. The aim was also to estimate the intra-herd correlation (r_e) and design effect (D) of BVDV seropositivity. Cattle were selected by a two-stage cluster sampling. Blood samples were collected from 560 animals originating from 40 herds. Sera were tested for antibodies against BVDV using an indirect ELISA test. The sensitivity and specificity of the test was 97.9 and 99.7%, respectively. Risk factors regarding the herd and each animal sampled were recorded through a personal interview at the time of blood sampling. Twenty-four of the 40 herds had at least one seropositive animal. The animal true seroprevalence was estimated as 14%. The marginal logistic regression model used to describe the data found a significant (p < 0.05) association of herd size–cow-origin interaction. The interaction was due to a higher risk of seropositivity in the category of herds with ≤100 animals and purchased cows (OR = 1) as compared to herds with ≤100 animals and cows born in the farm (OR = 0.23). Seropositivity between cows purchased and cows born in the farm was similar for herd sizes of 101–196 and >196 animals. The r_e and D values were 0.17 ± 0.05 and 3.16 ± 0.57, respectively.     

Rat, caprine, equine and bovine erythrocyte ghosts exposed to t-butyl hydroperoxide as a model to study lipid peroxidation using a chemiluminescence assay

The aim of the present study was to analyze the time-course of t-butyl hydroperoxide-induced changes in lipid peroxidation, fatty acid composition and chemiluminescence intensity in rat, caprine, equine and bovine erythrocyte ghosts. A relatively high content of arachidonic acid (C20:4 n6) and docosahexaenoic acid (C22:6 n3) was characteristic of the rat erythrocyte ghosts. The fatty acid composition of native erythrocyte ghosts obtained from caprine, equine and bovine was characterized by a high content of oleic acid (C18:1 n9) and a low content of the peroxidable polyunsaturated fatty acids (C20:4 n6 and C22:6 n3). The proportion of linoleic acid (C18:2 n6) was higher in equine and bovine compared to rat and caprine. Increase in lipid peroxidation in rat erythrocyte ghosts was maximal within 12 min of incubation, t-butyl hydroperoxide concentration dependent and was paralleled by a decrease in C18:2 n6, C20:4 n6 and C22:6 n3 and an increase in chemiluminescence formation. Polyunsaturated fatty acids (PUFAs) present in rat erythrocyte ghosts exhibit the highest sensitivity to oxidative damaged and their sensitivity increases as a power function of the number of double bonds per fatty acid molecule. Light emission in caprine, equine and bovine erythrocyte ghosts was very low, t-butyl hydroperoxide concentration-dependent but changes in fatty acid composition were not observed. The main conclusion of this work is that a low unsaturation degree of fatty acids in erythrocyte ghosts of caprine, equine and bovine prevent the lipid peroxidation on those membranes when they are incubated with t-butyl hydroperoxide.     

Immunohistochemical studies of scrapie archival material from Irish ARQ/ARQ sheep for evidence of bovine spongiform encephalopathy-derived disease

Since scrapie and bovine spongiform encephalopathy (BSE) in sheep are clinicopathologically indistinguishable, BSE in sheep may have been misdiagnosed as scrapie. Disease-specific prion protein (PrP(d)) patterns in archival tissues of 38 Irish ARQ/ARQ sheep diagnosed as scrapie-affected were compared to those in four Dutch BSE-challenged sheep. When medulla oblongata was immunolabelled with an antibody directed against amino acids 93-99 of ovine prion protein (ovPrP), intraneuronal PrP(d) was apparent in all 38 Irish sheep but was absent in BSE-challenged sheep. When lymphoid follicles were immunolabelled with antibodies directed against amino acids 93-106 of ovPrP, granule clusters of PrP(d) were seen in 34 of the 38 Irish sheep. Follicles of the remaining four archive sheep contained either no PrP(d) or single PrP(d) granules, similar to follicles from BSE-challenged sheep. Based on the medulla results, none of the archival cases had BSE-derived disease. The identification of some scrapie sheep with little or no intrafollicular PrP(d) suggests that this technique may be limited in discriminating between the two diseases.     

Follicular fluid concentration of transforming growth factor-beta1 is negatively correlated with estradiol and follicle size at the early stage of development of the first-wave cohort of bovine ovarian follicles

The objectives of this study were to characterize the relationship between estradiol and transforming growth factor-β1(TGF-β1) concentrations in follicular fluid of growing bovine ovarian follicles, and to examine the effect of TGF-β1 on FSH-stimulated estradiol secretion in cultured bovine granulosa cells. Follicular fluid was collected from individual follicles >5 mm in diameter by ultrasound-guided transvaginal puncture (n = 12 heifers). Follicles were sampled at four different stages of development of the first post-ovulatory wave during selection of the single dominant follicle. Estradiol, progesterone and total TGF-β1 were measured in follicular fluid of the three or four largest follicles sampled when the largest follicle (F1) had reached either 6.5, 7.5, 8.5 or 9.5 ± 0.5 mm stage of development. There was a significant negative relationship between follicular fluid TGF-β1 and estradiol concentrations (R² = 0.44; p < 0.002), and between TGF-β1 concentrations and follicle diameter (R² = 0.23; p < 0.01) in cohort follicles at the 6.5 mm stage, but not at any later stage of development of the follicle wave. There was no correlation between progesterone and TGF-β1 concentrations at any stage. To assess the causal relationship between TGF-β1 and estradiol, granulosa cells from follicles measuring 2–5 mm at dissection were placed in serum-free culture. TGF-β1 caused a dose-dependent decrease in FSH-stimulated estradiol secretion (P < 0.001). These findings suggest that TGF-β1 has an inhibitory effect on estradiol secretion in FSH-stimulated follicles and that a reduction in TGF-β1 inhibition may be part of the mechanism of selection of a single dominant follicle.     

Duration of immunity of a quadrivalent vaccine against respiratory diseases caused by BHV-1, PI3V, BVDV, and BRSV in experimentally infected calves

Several laboratory studies assessed the duration of immunity of a quadrivalent vaccine (Rispoval™4, Pfizer Animal Health) against bovine respiratory diseases (BRD) caused by bovine herpes-virus type-1 (BHV-1), parainfluenza type-3 virus (PI₃V), bovine viral-diarrhoea virus type 1 (BVDV), or bovine respiratory syncytial virus (BRSV). Calves between 7 weeks and 6 months of age were allocated to treatment and then were injected with two doses of either the vaccine or the placebo 3 weeks apart. Six to 12 months after the second injection, animals were challenged with BHV-1 (n = 16), PI₃V (n = 31), BVDV (n = 16), or BRSV (n = 20) and the course of viral infection was monitored by serological, haematological (in the BVDV study only), clinical, and virological means for ≥2 weeks. Infection induced mild clinical signs of respiratory disease and elevated rectal temperature in both vaccinated and control animals and was followed by a dramatic rise in neutralising antibodies in all treatment groups. Titres reached higher levels in vaccinated calves than in control calves after challenge with BHV-1, BVDV, or BRSV. On day 3 after PI₃V challenge, virus shedding was reduced from 3.64 log₁₀ TCID₅₀ in control animals to 2.59 log₁₀ TCID₅₀ in vaccinated animals. On days 6 and 8 after BRSV challenge, there were fewer vaccinated animals (n = 2/10 and 0/10, respectively) shedding the virus than control animals (n = 8/10 and 3/10, respectively). Moreover, after challenge, the mean duration of virus shedding was reduced from 3.8 days in control animals to 1 day in vaccinated animals in the BVDV study and from 3.4 days in control animals to 1.2 days in vaccinated animals in the BRSV study. The duration of immunity of ≥6 months for PI₃V, BHV-1 and BVDV, and 12 months for BRSV, after vaccination with Rispoval™4, was associated mainly with enhanced post-challenge antibody response to all four viruses and reduction of the amount or duration of virus shedding or both.