鹦鹉热衣原体荧光环介导等温扩增（LAMP）检测方法的建立

鹦鹉热衣原体(Chlamydia psittaci)是一种人兽共患病原体,能引起自然疫源性衣原体病,目前广泛分布于世界各地。本研究应用环介导等温扩增技术(LAMP),针对鹦鹉热衣原体23S RNA基因序列设计引物,并进行筛选以及敏感性和特异性试验,建立了鹦鹉热衣原体恒温荧光扩增方法。该方法在63℃恒温条件下1 h内即可显示结果,操作简单、快速,特异性强,灵敏度可达100拷贝/μL,且与流产衣原体、动物布鲁氏菌、牛结核分枝杆菌、沙门氏菌、大肠杆菌、金黄色葡萄球菌无交叉反应;用国产仪器可直接通过检测荧光信号判读结果,既增强了准确性,也避免了开盖污染产生假阳性;应用建立的LAMP方法对实验室保存的临床DNA样品进行检测,发现检测结果与QPCR相同。该方法的建立弥补了传统检测技术的不足,为实现鹦鹉热衣原体的现场快速诊断提供了技术支持。     

畜牧兽医执法办案应注意的问题之我见

结合畜牧兽医执法工作实践,总结了10项执法工作中经常遇到的问题,并有针对性的提出了处理建议,以便为国内畜牧兽医执法人员正确执法办案提供参考。     

Green tea polyphenols supplementation alters immunometabolism and oxidative stress in dairy cows with hyperketonemia

Peripartal cows often experience negative energy balance, and are therefore prone to suffering from metabolic diseases such as hyperketonemia, which causes financial losses in dairy farms. This study aimed to investigate the effect of green tea polyphenol (GTP) supplementation during the periparturient period on production performance, oxidative stress and immunometabolism in dairy cows with hyperketonemia. One hundred Holstein cows were assigned to GTP (0.2 g/kg DM; n = 50) or control (without GTP; n = 50) group based on body weight, previous milk yield, and parity on d 15 before expected parturition. Subsequently, 10 cows with hyperketonemia were selected from each group, according to blood β-hydroxybutyric acid (BHBA) concentration between 1.2 and 2.9 mmol/L from 2 to 3 d postpartum. All cows were fed a close-up diet and a lactation diet with or without GTP supply from 15 d prepartum until 30 d postpartum. Milk and blood samples were obtained from 20 cows selected with hyperketonemia on 10, 20, and 30 d postpartum. Compared with control cows, greater milk yield and lower somatic cell count were observed in GTP cows. The GTP group had lower concentrations of BHBA, free fatty acids, cholesterol, triglyceride, reactive oxygen species, malondialdehyde, and hydrogen peroxide, greater concentrations of glucose, lower activities of aspartate aminotransferase, alanine aminotransferase, and glutamyl transpeptidase, alongside greater activities of superoxide dismutase, glutathione peroxidase, and total antioxidant capacity. Additionally, GTP supplementation up-regulated concentrations of interleukin-6 and interleukin-10, but down-regulated concentrations of tumor necrosis factor-α, interleukin-1β, interleukin-2, interleukin-8, and interferon-γ in plasma. Greater concentrations of plasma immunoglobulin G were also detected in the GTP group. Overall, the data suggested that GTP supplementation from 15 d prepartum to 30 d postpartum improved the milk yield and health status in cows with hyperketonemia during early lactation.     

Analysis of Rumen Microbial Diversity and Functional Prediction of Chinese Simmental Cattle Based on 16S rRNA High-throughput Sequencing Technology

To systematically explore the microbial diversity and function in the rumen of Chinese Simmental cattle,this experiment used 16S rRNA gene high-throughput sequencing technology to detect and analyze the rumen fluid samples of Simmental cattle (about 20 months old,the average weight was 550 kg),and its function was predicted by PICRUSt.The results showed that a total of 66 712 high-quality sequences were obtained from the Illumina Miseq sequencing platform,and 1 797 operational classification units (OTU) were obtained by cluster analysis.The taxonomic identification was divided into 13 phylums,20 classes,22 orders,33 families and 59 genuses;Firmicutes and Bacteroidetes were dominant bacteria,accounting for 62.46% and 22.45%,respectively;Based on the genus composition,[Eubacterium]_coprostanoligenes_group (9.16%),Ruminococcus_2 (7.90%),norank_f__Bacteroidales_RF16_group(6.23%),norank_f__Ruminococcaceae(4.79%),Rikenellaceae_RC9_gut_group(4.72%),norank_f_Bacteroidales_BS11_gut_group(4.49%),Ruminococcus_1(4.43%),etc.The PICRUSt function of the 16S rRNA genome predicted that the rumen flora function was mainly concentrated in carbohydrate transport and metabolism,and might contain a large amount of cellulose and lignin degrading enzyme genes.In summary,the high-throughput sequencing technology based on 16S rRNA gene fully revealed the diversity of rumen bacteria in Simmental cattle,and predicted that it was rich in protein decomposition and lignocellulose degrading enzymes.The basis of microbial cognition provided a reference for further research on rumen microbial functional genes closely related to some important nutritional and physiological functions.     

牛支原体和牛病毒性腹泻病毒二重二温式PCR检测方法的建立

为建立一种牛支原体(Mycoplasma bovis,MB)和牛病毒性腹泻病毒(Bovine viral diarrhea virus,BVDV)的快速鉴别诊断方法,针对MB的uvr C基因和BVDV的5'端非编码区(5'-UTR)保守基因序列,分别设计两对特异引物,并将三温式PCR扩增程序简化为二个温度梯度,建立了鉴别MB和BVDV的二重二温式PCR方法。该方法能同时扩增MB和BVDV,扩增产物大小分别为412和170 bp。特异性试验结果显示,该方法对参试的所有毒株只扩增MB和BVDV基因组,对其它牛病原体无扩增;敏感性试验结果显示,该方法最低能同时检测到104拷贝的两种目的核酸;干扰性试验结果显示,该方法能同时检测两个模板不同浓度的组合,试验结果不受模板影响。综上,本研究所建立的二重二温式PCR方法特异、敏感、快速、简便,可应用于MB和BVDV临床鉴别诊断和流行病学调查。     

Pharmacokinetics of sanguinarine,chelerythrine, and their metabolites in broiler chickens following oral and intravenous administration

Sanguinarine (SA) and chelerythrine (CHE) are the main active components of the phytogenic livestock feed additive, Sangrovit®. However, little information is available on the pharmacokinetics of Sangrovit® in poultry. The goal of this work was to study the pharmacokinetics of SA, CHE, and their metabolites, dihydrosanguinarine (DHSA) and dihydrochelerythrine (DHCHE), in 10 healthy female broiler chickens following oral (p.o.) administration of Sangrovit® and intravenous (i.v.) administration of a mixture of SA and CHE. The plasma samples were processed using two different simple protein precipitation methods because the parent drugs and metabolites are stable under different pH conditions. The absorption and metabolism of SA following p.o. administration were fast, with half‐life (t_1/2) values of 1.05 ± 0.18 hr and 0.83 ± 0.10 hr for SA and DHSA, respectively. The maximum concentration (C_max) of DHSA (2.49 ± 1.4 μg/L) was higher that of SA (1.89 ± 0.8 μg/L). The area under the concentration vs. time curve (AUC) values for SA and DHSA were 9.92 ± 5.4 and 6.08 ± 3.49 ng/ml hr, respectively. Following i.v. administration, the clearance (CL) of SA was 6.79 ± 0.63 (L·h^?1·kg^?1) with a t_1/2 of 0.34 ± 0.13 hr. The AUC values for DHSA and DHCHE were 7.48 ± 1.05 and 0.52 ± 0.09 (ng/ml hr), respectively. These data suggested that Sangrovit® had low absorption and bioavailability in broiler chickens. The work reported here provides useful information on the pharmacokinetic behavior of Sangrovit® after p.o. and i.v. administration in broiler chickens, which is important for the evaluation of its use in poultry.     

宿主域扩大的重组救活昆虫杆状病毒表达载体的构建及外源基因的表达

通过克隆的家蚕核多角体病毒解旋酶基因DNA与重组救活可线性化的苜蓿尺蠖核多角体病毒基因工程载体病毒 (BacPAK6 )DNA在昆虫细胞中发生重组 ,经累代筛选获得了既可感染家蚕又可感染秋粘虫细胞Sf 2 1的宿主域扩大的昆虫杆状病毒表达载体 (HyBacPAK6 )。HyBacPAK6DNA经Bsu36Ⅰ酶切后与含植酸酶基因的转移载体pVL1393 phy在家蚕细胞中重组后 ,通过蓝白斑筛选发现重组率可达 90 %以上。然而以杂交病毒为载体的外源基因表达量仅为以BmNPV为载体病毒的表达量的 2 0 %左右。分析认为HyBacPAK6可用于表达一些对蚕体有害的外源基因。     

有机铬的合成及其产品质量评价

三价铬离子是构成葡萄糖耐量因子的重要组成部分,能够增强胰岛素的作用,是人和动物不可缺少的微量元素之一。有机铬化合物在生物机体中具有多种极其重要的活性功能,如果人体缺铬,将导致糖尿病和其它相关疾病;动物缺铬,则会导致对应激敏感、免疫功能受抑制、繁殖功能下降以及胴体品质下降等。本文综述了有机铬化合物的生物功能及其合成方法,介绍了产品的检测及质量评价方法,并对其发展前景做了一定的展望。     

紫花苜蓿多叶型与三叶型品种同工酶的比较研究

采用聚丙烯酰胺凝胶电泳技术,对国外2种多叶型苜蓿及国内外5种三叶型苜蓿幼苗的5种同工酶进行了测定,将酶谱数量化,应用SPSS统计软件求出各品种间的遗传一致度及欧氏遗传距离,并进行了聚类分析.结果表明:1)测定的5种同工酶中过氧化物酶(PER)、酯酶(EST)、淀粉酶(AMY)具有多态性,综合此3种酶酶谱可以鉴定供试的7个品种;2)2个多叶型品种具有一共同特征酶带PER-4,国内品种的过氧化物酶活性明显高于国外品种;3)品种间的遗传一致度为0.826～0.952,欧氏遗传距离为0.500～1.118,多叶型品种间遗传一致度为0.904,欧氏遗传距离为0.500;4)等级聚类分析结果为,以欧氏遗传距离0.400为分界,供试品种分为3类,2个多叶型品种为一类,其余的5种三叶型品种分为另两类.     

陕西省动物研究所馆藏标本现状及分析

陕西省动物研究所现有馆藏标本5万余份,是秦岭地区野生动物保护及研究的重要参考资料。为加强标本管理,提高标本使用效率,我们在2008年4月至2009年6月对所内馆藏标本进行了系统分类整理,分析总结了馆藏标本的现实状况并就存在的问题进行了探讨,为标本数字化共享平台建设奠定了基础,并为今后科研院校在动物学研究领域的交流与合作奠定基础。