猪带绦虫TSOL18特异性单克隆抗体的制备及其体外杀虫作用的研究

本研究目的在于制备抗猪带绦虫TSOL18单克隆抗体,并分析单抗对六钩蚴的杀伤作用。采用SephadexG-100纯化在毕赤酵母中表达的猪带绦虫六钩蚴TSOL18蛋白;纯化的蛋白免疫BALB/c小鼠,运用杂交瘤技术建立能分泌抗TSOL18单克隆抗体的细胞株;通过ELISA叠加试验进行mAb的抗原表位分析;采用间接ELISA法测定TSOL18单克隆抗体特异性及腹水效价;采用抗TSOL18单克隆抗体对激活的猪带绦虫六钩蚴进行体外六钩蚴杀伤试验,观察单抗对猪带绦虫六钩蚴活力的影响。结果成功获得12株稳定分泌抗TSOL18单克隆抗体的杂交瘤细胞株,识别2个不同抗原表位,不同抗原表位的2株单克隆抗体腹水效价分别为1×102、1×106。抗体体外六钩蚴杀伤试验结果证明,在有补体的情况下,多抗和单抗作用6 d后,六钩蚴结构模糊,边缘不清晰。表明单抗对六钩蚴有一定的杀伤作用。     

Canine visceral leishmaniasis: comparison of in vitro leishmanicidal activity of marbofloxacin, meglumine antimoniate and sodium stibogluconate

The control of canine leishmaniasis largely depends on the success of treatment. Drugs currently available to treat this disease are toxic and partially effective. The curative effect of marbofloxacin, a third-generation fluoroquinolone developed for veterinarian individual treatment, was evaluated in vitro in the presence of Leishmania infantum promastigotes and dog-monocyte-derived macrophages; meglumine antimoniate and sodium stibogluconate were used as comparative treatments. We observed that the killing of Leishmania promastigotes and intracellular amastigotes by marbofloxacin was dose-dependent. We demonstrated that successful treatment of canine infected macrophages for 48 h was possible with 500 microg/ml of marbofloxacin. Leishmanicidal activity acted through a TNF-alpha and nitric oxide pathway and correlated with the generation of nitric oxide (NO(2)) production by monocytes derived macrophages from infected (23+/-5 microM) or healthy (21+/-6 microM) dogs, in comparison with NO(2) concentration in infected/non-treated macrophages (< 3 microM, P<0.01). This significant induced parasiticidal effect correlated with extensive elimination of amastigotes by macrophages derived from infected (11+/-5) and healthy dogs (6+/-2), when compared to infected/non-treated macrophages (530+/-105 and 472+/-86 amastigotes, respectively, P< 0.01). Marbofloxacin was shown to be non-toxic at 500 microg/ml in vitro and no cell apoptosis was observed. The molecule was able to induce a parasitic process after significant elimination of amastigotes in leishmania-infected dog macrophages. We propose that marbofloxacin, compared to standard chemotherapeutic agents (meglumine antimoniate and sodium stibogluconate), could be an effective and pragmatic oral route alternative to treat canine leishmaniasis.     

Identification and Pathogenicity Analysis of Aeromonas caviae M12 from Fox

We tried to identify the bacteria and explore the mechanism of the bacteria's pathogenicity via housekeeping gene gyrB and in vitro organ culture (IVOC) of ileum and intramuscular injection. Microscope, electron microscope and scanning electron microscope were also used to observe and the structure, pathogen of M12 and changes of infected tissues. The results showed that M12 was short gram-negative bacteria, and both ends of it were obtuse and the size without flagellum was (0.6 to 1.6) μm×(0.6 to 0.7)μm while length of flagellum was about 2 to 3 times of the length of bacteria.It had 100.0% similarity with Aeromonas caviae from GenBank. IVOC test observed M12' adhesion in intestinal epithelial cell, resulting in formulation of biofilm structure and damage in intestinal epithelial cell. Histopathologic examination showed that M12 could cause damage in intestine, liver, lung, kidney, muscle tissue of rabbit.The assay would offer references for researching the pathogenic mechanism of Aeromonas caviae.     

利用体外法研究豆粕型日粮添加烟酸对绵羊瘤胃发酵的影响

本试验培养底物及瘤胃液供体动物日粮为玉米 -豆粕型日粮 ,实验用瘤胃液来自3只安装永久性瘤胃瘘管的绵羊 ,体外实验按不同烟酸添加剂量分为13个试验组 (0、50、150、250、350、400、450、500、600、700、800、900、1000mg/kg) ,每个试验组4个重复。在体外培养24小时后 ,测定培养液中NH3-N、瘤胃细菌N产量、原虫数量、总VFA产量及乙酸、丙酸、丁酸产量和乙酸/丙酸值 ,用以确定在该氮源日粮条件下 ,添加烟酸对瘤胃发酵的影响 ,并对烟酸适宜添加范围进行了研究。实验结果表明 ,在豆粕型日粮来源培养底物中 ,烟酸适宜添加范围为450～600mg/kg。     

In vitro analyses for predicting standardised ileal digestibility of protein and amino acids in actual batches of feedstuffs and diets for pigs

In vitro digestibility analyses of feedstuffs and diets are integrated in the new Danish feed evaluation system for pigs. This system is based on potential physiological energy and standardised digestible amino acids. For characterisation of the energy value of actual batches of feedstuffs and complete diets for pigs, two different in vitro methods have been developed and proved to analyse precisely the digestibility of organic matter at faecal, as well as at ileal, level in the pigs. For characterisation of the protein value in the diet, table values for standardised digestibility of amino acids from the individual feedstuffs are used for calculation of their contribution to the amino acid recommendations for the specific pig category. The results in the present paper indicate that in vitro analyses for digestibility of crude protein in feedstuffs are a reliable tool for calculation of standardised ileal digestibility of the individual amino acids. Thus, the results may question the need for future experiments with surgically modified pigs for analysing ileal digestibility of amino acids in protein sources. Alternatively, in vitro digestibility analyses appear to have a general potential for a precise evaluation of amino acid digestibility in actual batches of feedstuffs and pig diets.     

山羊卵母细胞体外成熟体系研究进展

卵母细胞体外成熟(IVM)是山羊胚胎体外生产的关键步骤,其对山羊体外生产胚胎的数量和质量均具有非常重要的意义。此外,IVM还可以满足胚胎工程技术如体细胞核移植、转基因动物生产对卵母细胞的大量需求。然而,由于山羊卵母细胞体外成熟研究起步较晚,与牛、猪等家畜相比仍然存在不小的差距,存在诸如体外成熟率低、胚胎质量不佳、培养体系可重复性低等问题。因此,分析山羊卵母细胞体外成熟的主要影响因素,提高山羊卵母细胞体外成熟率,建立山羊卵母细胞体外成熟稳定培养体系,就成为了近年来山羊胚胎体外生产的研究重点。论文综合国内外学者近年的相关研究,对可能影响山羊卵母细胞体外成熟效率的各种因素进行了综述分析,同时对山羊卵母细胞体外成熟现存问题进行了分析,以期为建立山羊卵母细胞体外成熟体系提供理论支持。     

由体外受精囊胚获得ICR小鼠ES细胞

收集鼠卵母细胞和精子，采用体外受精技术，获得早期胚胎。体外培养胚胎至囊胚，分离内细胞团，获得ICR小鼠的ES细胞。结果不但获得了来自体外受精囊胚的ICR小鼠ES细胞，而且表明用体外受精方法获得的小鼠早期胚胎的数量明显大于常规方法，且比较稳定。ICR小鼠ES细胞的获得，为研究小鼠的ES细胞分化提供了条件。     

竹叶提取物对奶牛瘤胃体外发酵参数及产气量的影响

本试验旨在研究竹叶提取物对奶牛瘤胃体外发酵参数及产气量的影响。试验选用健康、体况相近的荷斯坦奶牛用于瘤胃液的采集,以精粗比为4∶6的全混合日粮作为发酵底物。试验组分为6组,各组分别添加0(对照)、0.75、1.50、3.00、4.50、6.00 mg/g的竹叶提取物,每组6个重复,每个重复3个批次。体外发酵24 h后,记录产气量,测定瘤胃发酵参数。结果表明:1)各组的发酵液pH均无显著差异(P>0.05)。试验组的发酵液中氨态氮(NH_3-N)浓度均显著低于对照组(P<0.05),其中3.00和4.50 mg/g竹叶提取物组的发酵液中NH_3-N浓度显著低于其他各组(P<0.05)。试验组的发酵液中干物质消失率均显著高于对照组(P<0.05)。2) 4.50、6.00 mg/g竹叶提取物组的发酵液中总挥发性脂肪酸浓度显著高于其他各组(P <0. 05); 3. 00、4.50、6. 00 mg/g竹叶提取物组的发酵液中乙酸浓度显著高于其他各组(P <0. 05); 4. 50、6.00 mg/g竹叶提取物组的发酵液中丙酸浓度显著高于对照组(P <0. 05); 3. 00、4. 50、6. 00 mg/g竹叶提取物组的发酵液中丁酸浓度显著低于其他各组(P<0.05); 6.00 mg/g竹叶提取物组的发酵液中异戊酸浓度显著低于对照组(P<0.05)。各组的发酵液中异丁酸、戊酸浓度以及乙酸/丙酸均无显著差异(P>0.05)。3)试验组的体外发酵甲烷体积和甲烷含量均显著低于对照组(P<0.05)。各组的体外发酵产气量无显著差异(P> 0.05)。综上所述,体外条件下添加竹叶提取物在不影响瘤胃发酵模式的同时可以显著降低甲烷排放,其中添加4.50 mg/g竹叶提取物效果最佳。