水牛精子胞质内显微受精技术的初步研究

本研究探讨了水牛精子胞质内显微受精技术的可行性及操作条件的改善对水牛ICSI结果的影响。结果表明 :1.将精子操作液中PVP的浓度从 10 %降低到 5 % ,注射后卵母细胞的存活率无显著差异 (10 0 %vs .97.8% ;P >0 .0 5 ) ,分裂率明显提高 (79.4 %vs.88.6 % ;P <0 .0 5 ) ,囊胚率有所提高 ,但无显著差异 (2 3.5 %vs .2 4 .2 % ;P >0 .0 5 )。 2 .显微注射时卵母细胞第一极体置于相当于时钟 6点的位置与 12点位置相比 ,分裂率无显著差异 (89.5 %vs.86 .2 % ;P >0 .0 5 ) ,但囊胚发育率明显提高 (43.4 %vs.2 7.5 % ;P <0 .0 5 )。 3.离心和不离心的卵母细胞注射后的分裂率分别是 94 .3%和 91.3% ;囊胚发育率分别是 2 3.4 %和 2 7.5 % ,差异均不显著 (P>0 .0 5 )。结论 :1.适当降低PVP的浓度可以促进水牛卵母细胞获得胚胎发育的能力 ,并且不会增加精子显微注射的难度。 2 .注射时水牛卵母细胞的极体位于 6点的位置 ,卵母细胞获得胚胎发育的能力高于 12点位置。3.水牛卵母细胞的低透明度不会影响显微受精的效果。     

水牛卵母细胞去核方法的摸索

本研究对水牛卵母细胞的去核方法进行了摸索。首先通过盲吸法去核,摸索水牛卵母细胞在体外成熟不同时间的成熟率和去核率;其次采用spindle-View系统去核,摸索用这种系统去核的最佳时间。水牛卵母细胞IVM16-18h与IVM 19-21h的成熟率之间有显著差别(43.0％vs58.2％,P<0.05),与22-24h组的成熟率之间有极显著差别(43.0％vs 68.8％,P<0.01).而去核率的情况刚好相反。随着卵母细胞成熟时间的延长,卵细胞的核与PB1的相对距离增大。在体外成熟18h,20h,22h,有较大比例(分别为72.6％、67.9％、64.0％)的卵母细胞中的核与PB1的位置较近,但当成熟时间增加到24h时,这一比例下降到57.6％(72.6％vs 57.6％,P<0.05),如采用Spindle-View系统去核则可消除成熟时间对去核率的影响,这些结果表明:1.水牛卵母细胞的成熟时间的延长不断提高,而去核率呈下降趋势;2.水牛卵母细胞成熟时间的延长会增加核与PB1的相对距离;利用Spindle-View系统可提高成熟时间较长的卵母细胞的去核率。     

日本血吸虫重组GST抗原对水牛的保护性研究

在日本血吸虫病非流行区，选无疫水接触史，８个月龄左右的水牛，用日本血吸虫重组ＧＳＴ抗原进行免疫观察对水牛的保护性。结果显示，攻击感染血吸虫尾蚴后，实验组比对照组平均虫荷减少２２．３％，粪卵ＥＰＧ差异有显著意义，毛蚴孵出率降低近４０％，肝、肠组织血吸虫卵沉积量减少４７．９４％～５６．８３％。     

Evaluation of whole fresh blood and dried blood on filter paper discs in serological tests for Trypanosoma evansi in experimentally infected water buffaloes.

In this study we investigated if whole blood could substitute for serum in the direct card agglutination test (CATT/Trypansosoma evansi) and the indirect card agglutination test (LATEX/T. evansi) for the sero-diagnosis of T. evansi in buffaloes. Likewise blood spots on filter paper were compared with sera for use in the indirect enzyme-linked immunosorbent assay/T. evansi (ELISA) and immunotrypanolysis test (T.L./T. evansi). Samples were collected weekly from experimentally T. evansi infected- and non-infected water buffaloes. To estimate test agreement between serum and respectively whole fresh blood and dried blood spots on filterpaper of the tests, kappa values with 95% confidence intervals were calculated, 0.75+/-0.11 for the CATT/T. evansi; 0.80+/-0.11 for the ELISA/T. evansi; 0.84+/-0.11 for the LATEX/T. evansi and 0.93+/-0.11 for the T.L./T. evansi. In addition kappa values with 95% confidence intervals were computed to assess agreement between results obtained in the reference T.L./T. evansi test and those obtained in the other assays; 0.70+/-0.10 for the CATT-Serum; 0.75+/-0.11 for the LATEX-Blood; 0.77+/-0.11 for the LATEX-Serum; 0.81+/-0.10 for the CATT-Blood; 0.81+/-0.11 for the ELISA-Serum and 0.84+/-0.11 for the ELISA-Confetti. Based on the high kappa values as calculated, we conclude that serum can be replaced by fresh whole blood for the agglutination assays or blood on filter paper for the ELISA/T. evansi and T.L./T. evansi.     

Proper Quality Control of Formulated Foot‐and‐Mouth Disease Vaccines in Countries with Prophylactic Vaccination is Necessary

Vaccination is considered as an important tool to control foot‐and‐mouth disease (FMD). A good quality vaccine containing relevant serotypes and matching strains is a pre‐requisite for vaccination to be effective. The present study investigated the quality of different brands of FMD vaccine available in Pakistan, including three locally produced and two imported products. All the vaccines were found free of bacterial or fungal contamination. No adverse effects were noted in suckling mice and buffalo calves inoculated with the vaccines, showing that the vaccines were sterile and safe. The humoral immune response to the FMD vaccines was determined in buffalo calves for 234 days post‐vaccination. Very low humoral immune responses against FMD serotypes O, A and Asia 1 viruses were detected to the locally produced vaccines. The imported vaccines, however, elicited a higher antibody response which persisted for a long period in one of the 2 vaccines. The present study highlights the need of assessing an independent vaccine quality control of finished FMD vaccine products.     

重组日本血吸虫26 kDa GST抗原免疫水牛后抗体动态及免疫保护性效果

目的　观察重组日本血吸虫 2 6kDaGST抗原 (reSjc2 6GST)免疫役用放养水牛 (简称水牛 )后抗体动态及免疫保护性的效果。　方法　试验组 96头水牛 ,用reSjc2 6GST免疫 3次 ,每次间隔 2wk ,3次剂量分别为 0 2、 0 2和 0 1mg。对照组 90头水牛不作免疫 ;观察 2组水牛免疫前及免疫后 2、 5、 9、 12、 15和 2 0个月的抗体水平及血吸虫感染率的变化。　结果　试验组机体产生特异性抗reSjc2 6GST抗体 ,其抗体水平呈明显的梯形升高趋势。试验组免疫后 2 0个月血吸虫感染率比免疫前下降了 62 2 % ,比同期对照组低 67 7%。　结论　用reSjc2 6GST免疫水牛能产生特异性抗体 ,在免疫后 2 0个月内维持较高水平 ,有一定的抗血吸虫自然感染的保护力。     

Immunohistochemical studies of the epididymal duct in Egyptian water buffalo (Bubalus bubalis)

Using immunohistochemistry (IHC), this study aimed to evaluate the regional distribution pattern of some biologically active proteins in the epididymis of Egyptian water buffalo and to determine the structural-functional relationships of the different epididymal structures. Wax-embedded sections from different regions of the epididymal duct from adult, clinically healthy, buffalo bulls were used. Primary antibodies against angiotensin converting enzyme (ACE), S-100, galactosyltransferase (GalTase), alpha smooth muscle actin (α-SMA), connexin 43 (Cx43) and vascular endothelial growth factor (VEGF) were used for immunohistochemical studies. The results showed that, in addition to the well-known principal and basal cells, the epididymal epithelium, similar to that of other species, possessed apical cells and intraepithelial leukocytes. IHC showed that, with the exception of VEGF which reacted negatively, all antibodies used displayed variable reactivity in the different epididymal structures. Apical cells expressed a strong reaction with ACE along the entire length of the duct. The principal cells in the caput epididymis exhibited a distinct reactivity with S-100 and GalTase. The peritubular muscular coat displayed a marked immunostaining for α-SMA and for Cx43. In conclusion these findings showed a regional-specific distribution pattern, distinct from that in bovine bulls. Some potential functional capacities, especially absorptive and secretory ones, are discussed in relation to the different epididymal regions.     

Immunosuppressive Potential of Low Fat Buffalo Milk Supplemented with Omega-3 Fatty Acids

The present investigation reports the evaluation of the immunosuppressive potential of low fat buffalo milk supplemented with omega-3 fatty acids in mice. The phagocytic activity of omega-3 milk group was 32% less than that of control group after 4 weeks and 42% less after 8 weeks of dietary regime. The lymphocyte proliferation index was 26% and 61% less than that of control after 4 and 8 weeks, respectively, when lipopolysaccharide was used as mitogen and 17.8% (4 weeks) and 19% (8 weeks) when concanavalin A was used as mitogen. The IgG level in serum reduced after dietary supplementation. IgG in intestine and IgA in intestine did not show any particular trend.     

Transmission of foot-and-mouth disease virus from experimentally infected Indian buffalo (Bubalus bubalis) to in-contact naïve and vaccinated Indian buffalo and cattle

This study investigated the transmission of foot-and-mouth disease virus (FMDV) from experimentally infected Indian buffalo to in-contact naïve and vaccinated cattle and buffalo. In each of six rooms, two donor buffalo that had been inoculated with FMDV were housed for five days with four recipient animals, comprising one vaccinated buffalo, one vaccinated calf, one unvaccinated buffalo and one unvaccinated calf. Vaccination was carried out with current Indian vaccine strain (O/IND/R2/75) and challenged on 28 days post-vaccination with an antigenically similar strain (O/HAS/34/05). All 12 donor buffalo and the six unvaccinated cattle and six unvaccinated calves developed clinical signs of foot-and-mouth disease (FMD). In contrast, all six vaccinated cattle (100%) and four out of six vaccinated buffalo (66.6%) were protected from disease but all became infected with FMDV. This confirms that buffalo have the potential to spread FMD by direct contact and that vaccination can block this spread. The numbers of animals in the study were too small to determine if the differences in clinical protection afforded by vaccination of cattle and buffalo are significant and warrant a different dose regime.     

脱抗原牛角胎骨移植家兔后的特异性免疫反应

目的：研究脱抗原水牛胎移植后的免疫排斥反应和成骨效应。方法：以日本大耳白兔为动物模型，分别植入经30％，H2O2，0．6mol/L HCl脱抗原脱钙，30％H2O2脱抗原不脱钙2种温和方法处理的水牛角胎至兔佐侧肢骨缺损处，体外检测外周血特异性T淋巴细胞转化率和ELISA检测血清特异性抗角胎IgG抗体。结果：脱抗原脱钙处理显著地降低了角胎的抗原性，与对照相比，未见受体对移植物产生显著的特性细胞和体液免疫反应（P＞0．05），尤其是脱抗原不脱钙角胎效果更佳，两者成骨效果良好。结论：脱抗原角胎是一种免疫原性低，成骨效果好，来源方便的骨移植材料。