聚合嵌合法制作黄牛和水牛种间嵌合胚的初步研究

通过聚合嵌合法初步建立一套黄牛和水牛种间嵌合的程序与方法。聚合嵌合法采用链酶蛋白酶消化透明带或用机械剥离法去除透明带，然后在含有100 μg/ml PHA的培养液中聚合形成嵌合胚。结果发现， 8-细胞黄牛胚胎聚合水牛桑椹胚与黄牛囊胚聚合水牛桑椹胚相比，聚合胚存活率和囊胚发育率均无显著差异（P>0.05）。采用显微手术法分离黄牛和水牛8-细胞胚胎卵裂球进行聚合，聚合率为92.3%，囊胚发育率为58.3%，与用0.25%链酶蛋白酶分离胚胎卵裂球进行胚胎聚合的聚合率（86.7%）和囊胚发育率（46.2%）均无显著差异（P>0.05）。以上结果表明：①水牛和黄牛胚胎通过卵裂球聚合获得的种间嵌合胚胎能继续发育；②胚胎聚合前的发育阶段对其聚合成功率和随后的胚胎发育无明显影响。③胚胎卵裂球的分离方法（显微手术法和酶消化法）对其聚合率和囊胚发育率无明显影响。     

猪ICM注入囊胚获得嵌合胚的研究

以湖北白猪囊胚为ＩＣＭ供体，杜洛克猪囊胚为受体囊胚，进行嵌合胚的研究，采用免疫外科和显微外科两种方法，分离１８４枚供体囊胚的ＩＣＭ。用酶将ＩＣＭ分散，而后注入到受体囊胚的囊胚腔中，共获６７枚嵌合胚，经体外短期培养，其中３５枚嵌合胚恢复，恢复率为５２．２％，将恢复的嵌合胚分别移入５头与供体囊胚猪同步的受体母猪子宫内，两头受孕，其中一头维持到分娩，产仔４头，但未见毛色嵌合现象。     

猪ICM注人囊胚获得嵌合胚的研究

以湖北白猪囊胚为ICM供体,杜洛克猪囊胚为受体囊胚,进行嵌合胚的研究.采用免疫外科和显微外科两种方法,分离184枚供体囊胚的ICM.用酶将ICM分散,而后注入到受体囊胚的囊胚腔中,共获67枚嵌合胚,经体外短期培养,其中35枚嵌合胚恢复,恢复率为52.2%.将恢复的嵌合胚分别移入5头与供体囊胚猪同步的受体母猪子宫内,两头受孕,其中一头维持到分娩.产仔4头,但未见毛色嵌合现象.     

砷污染区黄牛和水牛姐妹染色单体交换和细胞周期动力学的研究

本文以外周血淋巴细胞姐妹染色单体互换(SCE)频率和细胞周期动力学为指标,以较长期(4～13年)生活在砷污染区的黄牛和水牛为对象,对砷在动物体内慢性蓄积所诱发细胞遗传学效应进行了研究。其结果表明,砷污染区黄牛和水牛SCE频率均显著高于对照组,参照有关评价标准,可判为弱阳性,提示在当地污染水平下,黄牛和水牛都诱发了一定程度的细胞遗传学效应。对细胞周期动力学的观察结果表明,砷对黄牛和水牛没有明显的细胞毒性。本试验还报告了正常黄牛和水牛外周血液淋巴细胞SCE的自发频率以及培养72小时各期细胞的正常比率。     

昆明小鼠内细胞团和滋胚层细胞注入去核2-细胞胚的核移植研究

采用核移植技术将单个细胞注入去核2-细胞卵裂球中,比较来自于囊胚的内细胞团(ICM)和滋胚层细胞(TE)产生孕体或嵌合体的发育潜力。用TE和ICM重构胚胎的发育潜力,主要取决于注射hCG后从输卵管收集到的2-细胞胚胎的时间。在注射hCG后38～42 h和43～46 h收集得到的2-细胞胚胎,其重构胚胎仅仅能够发育到4-细胞期。注射hCG后48～51 h收集得到的2-细胞胚胎,重构后发生卵裂的胚胎比率显著提高,有少部分会发育到囊胚期。用诺考达唑(Nocodazole)处理这些重构胚,发生卵裂的胚胎比率会显著提高,并且有部分重构胚发育到囊胚阶段。将这些囊胚移植到受体鼠中,在其妊娠中期未检测到供体核的出现。用ICM和TE进行重构得到的嵌合2-细胞胚胎,其体外发育潜力有限,本试验中也未得到嵌合孕体。     

水牛和黄牛同种及种间显微授精(ICSI)的初步研究

试验探讨了水牛和黄牛同种及种间显微授精的可行性。体外成熟22~24 h的水牛和黄牛卵母细胞分别注入冷冻/解冻的尼里－拉菲水牛和利木赞黄牛精子,进行同种或种间的显微授精操作,构建的ICSI胚胎一部分培养到16~18 h固定染色检查原核形成情况,另外一部分胚胎培养2~9 d分别记录胚胎的分裂情况及囊胚发育情况。结果发现,水牛同种(水牛+水牛)和水牛异种(水牛+黄牛)显微授精胚胎的双原核率(47.73%和36.67%)、分裂率(68.00%和72.64%)和囊胚发育率(16.44%和22.39%)均无显著差异(P＞0.05);黄牛同种(黄牛+黄牛)和黄牛异种(黄牛+水牛)显微授精胚胎的双原核率(60.00%和45.28%)、分裂率(73.33%和71.31%)和囊胚发育率(28.57%和33.70%)亦无显著差异(P＞0.05)。以上结果表明:①水牛精子注射到黄牛卵子和黄牛精子注射到水牛卵子的种间授精胚胎均可以体外发育到囊胚阶段;②黄牛卵子无论同种和种间显微授精的效果均优于水牛卵子。     

百色市屠宰场待宰水牛和黄牛蜱感染的初步调查

牛蜱叮咬不仅可造成牛体的机械性损伤,而且由于它可携带多种病毒、细菌和寄生虫,可导致被叮咬动物的染病。为了调查百色市水牛和黄牛牛蜱的感染情况,从5月份到7月份之间我们共调查了202头水牛和81头黄牛的牛蜱感染情况。调查中水牛有9头(4.46%),黄牛有14头(17.28%)观察到牛蜱感染,捕捉的牛蜱经鉴定均为微小牛蜱。在牛蜱感染强度上,水牛的感染强度比较低,一般为每头牛2只~5只,而黄牛的感染强度有明显的差异,感染最多的每头牛可检查到82只。说明我市待宰牛中,水牛、黄牛都有牛蜱感染,以黄牛的感染情况比较严重。     

水牛徒手克隆的初步研究

徒手克隆是近年发展起来的一种手工化的核移植技术。本试验探讨了不同的去核方法、融合方法和供体细胞类型对水牛HMC效率的影响。试验结果表明,CB处理法去核与荧光染色法去核相比,去核效率及之后的重组胚发育能力均无显著差异(P＞0.05);一步法的融合率极显著高于两步法(P＜0.01),而两种方法构建的重组胚发育能力则无显著差异(P＞0.05);利用卵丘细胞所构建的HMC胚胎的融合率极显著高于耳皮肤成纤维细胞(P＜0.01),但重组胚的发育效果差异不显著(P＞0.05)。说明利用卵丘细胞作为HMC的供体细胞,采用CB处理法去核和一步融合的方法进行徒手克隆,其生产核移植胚胎效果较好。     

水牛冷冻胚胎分割效果的初步研究

探讨了胚胎分割前后冷冻和不同时期的冷冻胚胎对水牛胚胎分割效果的影响。结果:冷冻后分割组的分割成功率与分割后冷冻组差异不显著(71.39%vs 71.43%,P>0.05),但囊胚冷冻后分割组的半胚发育率显著高于分割后冷冻组(58.33%vs 47.14%,P<0.05);扩张囊胚的冷冻存活率显著高于孵化囊胚(94.84%vs 79.01,P<0.05),但囊胚与孵化囊胚差异不显著(92.67%vs 79.01%,P>0.05),囊胚、扩张囊胚和孵化囊胚冷冻胚胎的分割成功率(70.76%,73.22%,70.92%)及半胚发育率差异均不显著(53.07%,51.49%,57.99%)(P>0.05)。上述结果说明,水牛囊胚冷冻后分割比分割后冷冻能获得更高的半胚发育率;水牛未扩张囊胚和扩张囊胚的冷冻存活率比孵化囊胚高,且这3个时期的冷冻胚胎均适合分割。     

Comparative pharmacokinetics of fenbendazole in buffalo and cattle

Swamp buffalo (Bubalus bubalis) and Droughtmaster cattle (Bos indicus × B. taurus), fitted with gastrointestinal cannulae, were dosed intraruminally with fenbendazole at 7.5 mg/kg liveweight, together with a chromium oxide capsule and a pulse dose of NaCoEDTA, to estimate the flow dynamics of the digesta in the rumen and duodenum. The concentrations of fenbendazole (FBZ) metabolites were measured in plasma and duodenal fluid collected over 120 h. In plasma, significantly lower peak concentrations and earlier disappearance of FBZ and its sulphoxide (OFZ) metabolite were observed in buffalo, which considerably reduced systemic availability in comparison with cattle. The availability of OFZ in the duodenal fluid of buffalo was significantly lower, whereas FBZ disposition was similar to that in cattle. The turnover rate of fluid in the rumen was higher in buffalo than in cattle, while the flow parameters for other digesta were similar in the two species. It is concluded that the decreased absorption of drug in buffalo was attributable to the shorter residence time of the dose in the rumen, and probably in the entire gastrointestinal tract. This may reduce the efficacy of treatment and indicate the need for higher dose rates for benzimidazole anthelmintics in buffalo than in cattle.Abbreviations AAS atomic absorption spectroscopy - AUC area under the concentration-versus-time curve - C _max maximum concentration - FBZ fenbendazole - FBZ.SO₂ fenbendazole sulphone - HPLC high-performance liquid chromatography - OFZ fenbendazole sulphoxide     

The uptake of fenbendazole by cattle and buffalo following long-term low-level administration in urea-molasses blocks

Fenbendazole (Panacur bolus, Hoechst India Ltd) was incorporated at a rate of 0.5 g/kg into urea-molasses blocks made by two different processes. The concentration of the drug in blocks and its bioavailability were measured using plasma oxfendazole as marker. The recovery of the drug in blocks made by a warm process was 68% and the plasma oxfendazole concentration remained fairly stable at 0.2 and 0.12 µg/ml from day 6 of feeding in cattle and buffalo, respectively. The drug seemed to be inactivated in blocks made by a hot process, with reduced bioavailability. A low and sustained plasma concentration of the active metabolite of the drug could be maintained by self-medication using urea-molasses blocks as fenbendazole carrier.Abbreviations FBZ fenbendazole - HPLC high-performance liquid chromatography - MUMB medicated urea-molasses blocks - OFZ oxfendazole - UMB urea-molasses block     

The disposition kinetics of albendazole following the administration of single and divided doses to cattle and buffalo

Concentrations of albendazole sulphoxide and its sulphone metabolite in plasma in cattle and buffalo were measured by high-performance liquid chromatography after single and divided intraruminal administration of albendazole at the recommended nematocidal and fasciolicidal dose rates of 7.5 and 15.0 mg/kg body weight, respectively. No significant differences in the plasma concentrations of the metabolites or their pharmacokinetic parameters were observed between cattle or buffalo at either dose rate. Pharmacokinetic analysis and the disposition curve of the metabolites indicated increased uptake of the drug in both cattle and buffalo when the same total amount of the drug was given in divided doses compared to a single dose (p<0.05). The divided dose schedules of administration could possibly be exploited to extend the life of the available benzimidazole anthelmintics.     

The uptake of fenbendazole by cattle and buffalo following long-term low-level administration in urea-molasses blocks: Further studies on block formulations

Fenbendazole (Hoechst India Ltd.) was incorporated at 0.5 g/kg into urea molasses blocks made by two different processes. The proportion of the drug remaining in the blocks and the plasma concentrations of the parent compound and its metabolites were measured. Recovery of the drug in blocks made by the cold and the modified hot processes was 90% and 96%, respectively. The plasma metabolite profile revealed a plateau between days 4 and 6 of feeding in cattle and buffalo. However, the plasma concentrations of fenbendazole and its metabolites were low in buffalo compared to cattle.Abbreviations HPLC high-performance liquid chromatography - MUMB medicated urea molasses blocks - UMB urea-molasses block     

Eating and rumination behaviour in Brahman grade cattle and crossbred water buffalo fed on high roughage diet

The purpose of this study is to compare feeding and other behavior and nutrient digestibility of tropical grade Brahman (body weight (BW) = 231 kg ± 12.4; n = 3) and crossbred water buffalo (BW = 300 kg ± 13.9; n = 3). This experiment on digestibility and measures of muscles of mastication utilized one‐way, and animal behavior two‐way, analysis of variance, respectively. Two video camcorders were installed in each pair of buffalo and Brahman for 24 h period programmed on the 107th, 109th and 111th days of the digestion trials. Frequency and duration of feeding, meal intake, rumination, bolus, chews, drinking, defecating, standing and lying were recorded daily. Muscle diameter of Digastricus, Masseter and Pterygoid and different regions of the tongue were sampled and measured under light microscope using a standard micrometer. Buffalo obtained significantly higher intake of dry matter, roughage, crude protein, total digestible nutrient and metabolized energy than Brahman. This was supported by longer meal duration (P ≤ 0.05), and shorter meal breaks (P ≤ 0.05) of buffalo than Brahman. The diameter of the muscles for mastication was bigger (P ≤ 0.05) in buffalo than in Brahman, which is indicative of stronger chewing ability. Briefly, lesser and slower chewing action; higher intake of roughage and crude protein; and longer resting behavior of crossbred water buffalo than Brahman are all indicative of better digestive and metabolic performance of the buffalo under high roughage feeding conditions.     

Sequence diversity and molecular evolutionary rates between buffalo and cattle

Identification of genes of importance regarding production traits in buffalo is impaired by a paucity of genomic resources. Choice to fill this gap is to exploit data available for cow. The cross‐species application of comparative genomics tools is potential gear to investigate the buffalo genome. However, this is dependent on nucleotide sequences similarity. In this study, gene diversity between buffalo and cattle was determined using 86 gene orthologues. There was approximately 3% difference in all genes in terms of nucleotide diversity and 0.267 ± 0.134 in amino acids, indicating the possibility for successfully using cross‐species strategies for genomic studies. There were significantly higher non‐synonymous substitutions both in cattle and buffalo; however, there was similar difference in terms of dN– dS (4.414 versus 4.745) in buffalo and cattle, respectively. Higher rate of non‐synonymous substitutions at similar level in buffalo and cattle indicated a similar positive selection pressure. Results for relative rate test were assessed with the chi‐squared test. There was no significance difference on unique mutations between cattle and buffalo lineages at synonymous sites. However, there was a significance difference on unique mutations for non‐synonymous sites, indicating ongoing mutagenic process that generates substitutional mutation at approximately the same rate at silent sites. Moreover, despite of common ancestry, our results indicate a different divergent time among genes of cattle and buffalo. This is the first demonstration that variable rates of molecular evolution may be present within the family Bovidae.     

Post-thaw development of in vitro produced buffalo embryos cryopreserved by cytoskeletal stabilization and vitrification

The present study was conducted to examine post-thaw in vitro developmental competence of buffalo embryos cryopreserved by cytoskeletal stabilization and vitrification. In vitro produced embryos were incubated with a medium containing cytochalasin-b (cyto-b) in a CO₂ incubator for 40 min for microfilament stabilization and were cryopreserved by a two-step vitrification method at 24℃ in the presence of cyto-b. Initially, the embryos were exposed to 10% ethylene glycol (EG) and 10% dimethylsulfoxide (DMSO) in a base medium for 4 min. After the initial exposure, the embryos were transferred to a 7 µl drop of 25% EG and 25% DMSO in base medium and 0.3 M sucrose for 45 sec. After warming, the embryos were cultured in vitro for 72 h. The post-thaw in vitro developmental competence of the cyto-b-treated embryos did not differ significantly from those vitrified without cyto-b treatment. The hatching rates of morulae vitrified without cyto-b treatment was significantly lower than the non-vitrified control. However, the hatching rate of cyto-b-treated vitrified morulae did not differ significantly from the non-vitrified control. This study demonstrates that freezing of buffalo embryos by cytoskeletal stabilization and vitrification is a reliable method for long-term preservation.     

Comparison of carcass and meat characteristics of Brahman grade cattle (Bos indicus) and crossbred water buffalo (Bubalus bubalis)

This study was conducted to determine and compare the live traits, carcass yield and the physical, chemical, processing and sensory properties of meat from 20 Brahman grade cattle (crossbred cattle) and crossbred water buffalo (Philippine carabao × Murrah, Bulgarian or Indian) with an average age and weight of 29 months and 434 kg, respectively. The animals were fed a similar diet for 180 days before slaughter. A survey was performed to establish consumer meat preference between the two species. Live weights before slaughter of the two species were found to be different (P < 0.05) with the crossbred water buffalo showing higher bodyweight. However, the crossbred water buffalo had a lower (P < 0.05) dressing yield based on hot carcass weight and chilled carcass weight. The lower dressing yield of crossbred water buffalo can be attributed to its higher (P < 0.05) weights of edible and non‐edible slaughter by‐products. The chilled carcass yield of the forequarters and hindquarters from crossbred cattle and crossbred water buffalo were comparable when expressed as percent of live weight and chilled carcass weight. The estimated lean yield was higher in crossbred cattle than crossbred water buffalo. Crude protein, ash, fat, cholesterol, myofibrillar, sarcoplasmic and insoluble protein contents of the beef and carabao meat were all similar. Water holding capacity, pH, muscle fiber diameter, tenderness, firmness and marbling score in carabeef were all comparable to the beef. Redness was found to be higher (P < 0.05) in carabeef than the beef as measured with a color meter. The consumer meat preference survey showed that 55.9% selected the beef while 44.1% preferred carabeef. The color and amount of fat on the outside of the beef sample were primary in buyer selection.     

Effects of maize (Zea mays L.) silage feeding on dry matter intake and milk production of dairy buffalo and cattle in Tarai, Nepal

To identify the effects of whole crop maize silage (MS) as a substitute for rice straw (RS) on feed intake and milk production of mid-late lactating buffalo and cattle in Tarai, Nepal, eight Murrah and eight Jersey-Hariana were fed the basal diet, RS ( ad libitum ) with concentrate (0.68% of bodyweight [BW] on a dry matter [DM] basis). A 4 × 4 Latin square design experiment was conducted in each animal species with graded levels of MS substitution for RS (0%, T1; 33%, T2; 67%, T3 and 100%, T4). The MS had higher digestibility and total digestible nutrient (TDN) than RS. The DM intake per BW of the both species was highest in T3. The substitution of MS for RS increased the crude protein intake and the TDN intake in the both species. Although the buffalo showed the highest milking performance in T4, the cattle showed no significant differences in their milking performance among the treatments. The substitution of MS for RS improved the feed intake and milk production in the buffalo. On the other hand, the milk yield was not raised in the cattle, though the feed intake was increased by the substitution.