Isolation and Identification of a Variant Porcine Epidemic Diarrhea Virus and the Immune Efficacy of Its Inactivated Vaccine

This study was aimed to isolate a mutant strain of porcine epidemic diarrhea virus and prepare a PEDV inactivated vaccine with high valence by suspension culture process for immunizing against PEDV effectively in China.200 small intestines and theirs contents of diarrhea piglets died of diarrhea,collected from many large-scale pig farms in China,were detected by RT-PCR and sequenced,a mutant strain of porcine epidemic diarrhea virus was selected and put on the suspension-cultured Vero cells in a 2 L reactor for virus isolation and continuous cell culture,the harvested virus suspension,which was identified and determined TCID₅₀,was inactivated by formaldehyde and mixed with aluminum hydroxide gel adjuvant to prepare PEDV inactivated vaccine.After its physical behavior,stability viscosity,sterility test were checked out,the safety and immune efficacy were studied by immunizing the pregnant pigs and theirs piglets.The results were as follows:86 samples were detected positive in 200 samples,cytopathy occurred after the mutant strain samples screened were passaged to 5th generation,the virus suspension was harvested in 10th generation and identified as a mutant strain of PEDV,named PEDV-GF10 strain.The virus titer of harvested virus suspension was measured up to 1×10^8.0 TCID₅₀/mL after concentrated.After the vaccine was checked out,the sows,40 and 25 days before delivery in experimental groups,were injected into Xuehai acupoint with 4 mL vaccine and the pigs in blank group were free of immunifications.The results showed that there were no obvious differences in the production status of the sows in experimental groups and blank group and the temperature of theirs 3-day-old healthy piglets injected different doses of vaccine,and the vaccine was safe to the sows and piglets.Forty 3-day-old piglets producted by pregnant sows in experimental groups and blank group were randomly selected and taken 4 mL 1×10^8.0TCID₅₀/mL F10 virus culture.The PEDV morbidity of piglets in blank group was 100% after injection and the antibodies were negative;10% piglets in blank group had mild diarrhea symptoms,the protection rate was up to 90%,antibody of passive immunity in piglets lasted for more than 35 days.Virus titer of mutant strain of PEDV-GF10 improved a lot by suspension cell culture,the PEDV-GF10 inactivated vaccine was safe,and could effectively prevent and control the variation strain of PEDV in China.     

Effects of Lactobacillus and Bacillus subtilis Mixed Fermentation Feed on Growth Performance,Slaughter Performance and Intestinal Morphology of Broilers

The purpose of this experiment was to study the usage and proportion of fermented feed in broiler feed.By adding 0,10% and 20% fermented complete formula feed to the broiler feed,and granulating normally,the influence of the usage mode of the fermented feed on the production performance,slaughter performance,intestinal tract and other related indexes of the broiler feed were studied,so as to provide reference for the application of the fermented feed in the broiler feed.180 1-day-old healthy AA male broilers were randomly divided into 3 groups with 6 replicates in each group and 10 chickens in each replicate.The three treatment groups were fed with 0,10% and 20% of the total fermented feed for 42 days.The results showed that:From the whole period (1-42 days old),the 10% fermentation feed substitution group significantly reduced the feed consumption and weight gain ratio (P<0.05),and the average daily weight gain increased (P=0.05).At 42 days old,there was no significant effect of fermented feed on slaughter performance (P>0.05).The ileal crypt depth of broilers in the 20% fermentation feed replacement group was significantly higher than that in the control group and the 10% replacement group (P<0.05).The length of duodenal villi and V/C in 20% and 10% fermentative feed substitution group was significantly higher than that in control group (P<0.05).The results showed that the replacement of 10% fermented feed in broiler feed could reduce the ratio of feed to weight gain,increase the average daily gain,and have a certain impact on intestinal development.     

1株高致病性血凝性脑脊髓炎病毒的分离与鉴定

利用细胞培养方法从临床表现神经症状的死亡仔猪脑组织内分离获得1株病毒,同时对该病毒进行电镜负染观察、昆明种小鼠致病性试验、RT-PCR检测和部分基因测序分析。电镜观察可见细胞培养物中有大量的直径约115 nm的冠状病毒样粒子;接种病毒的小鼠均出现典型的神经症状,死亡率100%;血凝性脑脊髓炎病毒(HEV)特异性引物RT-PCR扩增结果阳性;其S基因序列与GenBank中登录的HEV-67 N相应基因序列的同源性高达99.6%。结果表明,分离的病毒为猪血凝性脑脊髓炎病毒,分离株暂命名为HEV-CC-2007。     

猪附红细胞体可溶性抗原的分析

为了解附红细胞体可溶性抗原的特性,本试验对解离下的猪附红细胞体,经超声波裂解制备粗抗原,再经SephadexG-200分离提纯后,应用对流免疫电泳(CIEP)定位特异性抗原蛋白峰,再进行十二烷基硫酸钠-聚丙烯酰胺凝胶电泳(SDS-PAGE)及免疫印迹(Westernblot),对猪附红细胞体粗提及纯化的可溶性抗原进行了分析。结果表明,猪附红细胞体可溶性抗原的电泳图谱上有四条蛋白带,其分子量分别为77Ku、62Ku、58Ku、29Ku,其中特异性抗原分子量为58Ku和29Ku。从而为附红细胞体抗原成分的进一步分析及该病的免疫学诊断等研究奠定了基础。     

超声波法和超声波酶法提取灵芝多糖的条件研究

灵芝(Ganoderma lucidum)是我国传统中药,具有滋补强壮,扶正固本的作用[1],至今已有2000多年的使用历史。灵芝子实体多糖是重要的有效成份之一,被广泛用作药品及保健食品的原材料。常用的灵芝多糖提取方法有煎煮法、水蒸馏法等,但这些方法耗时长、提取率低、提取物中多糖含量相     

滩羊尿对典型草原三种草地植物生长的短期作用特性

研究了不同尿液浓度和土壤含水量下，羊尿对典型草原植物茵陈蒿、委陵菜和狗尾草生长的短期作用特性。结果表明，不同施尿量施尿后1个月，狗尾草对羊尿沉积的响应为高尿液浓度负效应，低尿液浓度正效应；茵陈蒿和委陵菜为负效应。土壤含水量越高，羊尿对其分蘖（枝）的负效应越低。不同尿液浓度下，三种植物对羊尿响应的负效应顺序为茵陈蒿〉委陵菜〉狗尾草；且其分蘖（枝）对羊尿和土壤水分互作反应的正效应顺序为狗尾草〉委陵菜〉茵陈蒿。具体放牧过程中，可通过家畜适时饮水降低尿液浓度，以降低畜尿对草地的负效应。     

猪尿液中抗菌药物快速筛选拭子法的建立

为防止抗菌药物在动物性食品中的残留,以枯草芽孢杆菌为受试菌,建立一种检测活体动物尿液中抗菌药物残留的快速筛选拭子法,进行宰前活体检疫.添加试验测定猪尿液中5类抗菌药物最低检测限分别为:β-内酰胺类青霉素和氨苄青霉素均为0.05mg/L;氨基糖苷类庆大霉素0.05mg/L、新霉素0.4mg/L;四环素类金霉素0.1mg/L;大环内酯类红霉素0.05mg/L和氟喹诺酮类恩诺沙星0.2mg/L.各抗菌药物添加回收率范围均在64.0%～107.7%,变异系数均小于15%.假阴性结果显示,除青霉素(5%)和氨苄青霉素(4%)出现假阴性外,红霉素、庆大霉素、新霉素、恩诺沙星和金霉素均未出现假阴性.与国外同类试剂盒比较,结果显示两者对10种抗菌药物的检测限一致.     

Biological characteristics of JAK2 transduced CD34+ cells from cord blood during ex vivo expansion

AIM: To explore the feasibility and biological characterization of long-term regulated expansion of JAK2 transduced human CD34⁺ cord blood cells in vitro.METHODS: A retrovirus (RV) vector which contains JAK2 catalytic domain and two binding sites for a chemical inducer,dimerization (AP20187),was cloned (designated MGI-F₂JAK2).CD34⁺cells were enriched from cord blood with a MiniMACS system.The purified CD34⁺ cells were transfected with supernatant from the retrovirus packaging cell line that expressed JAK2.Following transduction,cells were expanded into four groups: AP20187 alone,FL alone,TPO,alone,AP20187+FL+TPO,respectively.The expanded cells were monitored by GFP expression,immunophenotyping,progenitor colony assay,karyotype analysis as well as tumorigenesis in nude mice.RESULTS: The purity of selected CD34⁺ cells was over 91% and gene transfer rate was 49.32%±6.21%.Only the group of AP20187 +FL+ TPO was obtained a significant sustained outgrowth of the transduced CD34⁺ cord blood cells.The percentage of GFP+ cells consistently produced a rise to the 90% peak level by the end of 8th week of culture.Flow cytometry analysis showed that the phenotype of the expanded cells was CD33⁺,CD61⁺ and Gly-A⁺ partial positive;CD38⁺ and HLA-DR⁺ strong positive,while CD2,CD7 and CD19 were almost negative.Colony assays performed in methycelluos,which can give rise to BFU-E,CFU-GM and CFU-Mix,the CFU-GM was predominantly in all colonies.The tumor was not observed in nude mice and the karyotype analysis was normal from expanded cells.CONCLUSION: The results demonstrate that AP20187-mediated activation of JAK2 signaling is capable of stimulating expansion JAK2 transduced CB CD34⁺ cells in combination with FL and TPO.This system may have applications for studies in signaling transduction,hematopoiesis,and for gene and cell therapy.     

Effects of pituitary adenylate cyclase-activating polypeptide on cardiac remodeling in coronary atherosclerotic rabbits

AIM:To study the intervention effects of pituitary adenylate cyclase-activating polypeptide (PACAP) on cardiac remodeling during the development of rabbit coronary atherosclerosis. METHODS:60 male New Zealand rabbits were equally divided into 3 groups randomly: control group (C group), atherosclerosis model group (A group) and PACAP intervention group (P group). At the 4th, 8th and 12th week, 5-6 cases of rabbits in each group were sacrificed, cardiac tissue with coronary arteries were harvested to make paraffin sections. The sections were stained with hematoxylin-eosin and van Gieson separately. The qualitative observation and/or quantitative analysis were made by light microscope. RESULTS:（1）There was no lesion in C group. For A group and P group, there were plaques in large epicardial coronary arteries and small coronary arteries; an impressive accumulation of collagen was also observed in myocardium. In P group, the lesions of small coronary arteries were less serious, and the degrees of perivascular and myocardial fibrosis also appeared to be less.（2）For A group, the wall-to-lumen ratios in small coronary arteries were significantly greater at the 12th week (2.58±1.54) than C group (1.34±0.58) and P group (1.39±0.48) （P<0.05）； and the width of cardiomyocyte (13.85 μm±2.27 μm) was already remarkably narrower than C group (14.68 μm±2.40 μm) at the 8th week (P<0.05) and narrower significantly than C group and P group at the 12th week. （3）There were not difference significantly between the above-related parameters of P group and C group (P>0.05). CONCLUSION:Structure changes exist in coronary arteries and myocardium during the development of rabbit coronary atherosclerosis, PACAP can inhibit the cardiac remodeling.     

我国西南主要干旱河谷生态及其研究进展

在广泛查阅前人工作的基础上, 发现到目前为止, 对干旱河谷生态方面的研究主要集中在成土特征和土壤养分水分分布特征、植被状况及生态恢复上.认为干旱河谷生态研究的进一步工作应该注重人为因素的研究, 对干旱河谷区人口与生态环境间不协调的方面进行干预, 以实现该区域人口与生态环境的良性互动;加强景观生态学研究和多尺度集成, 利用景观生态学原理、方法来探讨干旱河谷景观结构和功能的相应变化, 寻求维持区域生态安全的关键机制.