PCV2感染性分子克隆的制备及体内外感染性试验

将猪圆环病毒2型（Type2 porcine circovirus，PCV2）JXL株cDNA插入到载体pMD18-T中，获得的pMDT-PCV转染猪肾细胞PK15细胞系，盲传4代后，利用PCV2阳性猪血清进行间接免疫荧光抗体试验（IFA），能检测到特异性荧光。将pMDT-PCV质粒DNA腹股沟淋巴结注射40日龄仔猪2头，分别接种2、3次，接种剂量500μg／次，首次注射35d心脏放血致死。在体内感染性试验期间，动物未出现明显临床症状。试验结束，病理切片结果显示，不同淋巴组织中分别存在淋巴细胞缺失或增多的现象；肾小球、扁桃体、空肠淋巴结和股前淋巴结等冰冻组织切片中存在大量PCV2病毒抗原；血清中PCV2特异性的ELISA抗体效价迭1：2560，IFA效价为1：1280；通过聚合酶链式反应（PCR）可以分别从体外感染PK15细胞和体内感染动物及同圈饲养的空白对照猪的淋巴组织中扩增到病毒特异性基因片段。本研究证明含有单拷贝PCV2的重组质粒pMDT—PCV在体内、外均具有感染性，能衍生出病毒，并产生符合自然感染PCV2的大体病变和组织病理学变化，并激发较强的体液免癌应答。     

禽流感病毒分子生物学的研究进展

禽流行性感冒(av ian in fluenza,A I,简称禽流感)是由A型禽流感病毒(av ian in fluenza v irus,A IV)引起的禽类烈性传染病。作为被世界动物卫生组织(O IE)定为A类的传染病,A I不仅给世界养禽业造成了巨大的经济损失,而且对人类健康和生命安全构成了严重威胁。因此,A I已经成为人们关注的焦点,国内外学者也对其进行了大量研究。作者从病原基因组及其编码的蛋白质、致病力、变异性以及对人类感染A IV的分子机制等角度就A IV的分子生物学研究作一综述,为防制A I提供理论基础,并在此基础上探讨了人类禽流感的防治措施,加深人们对A I的认识。     

辣(甜)椒抗黄瓜花叶病毒(CMV)研究进展

黄瓜花叶病毒病(CMV)是为害辣(甜)椒的重要病毒,国内外对其研究较多。通过综述CMV的株系分化、抗性遗传、分子标记、抗CMV基因工程以及抗CMV育种等方面的研究结果和报道,旨在为辣(甜)椒抗CMV育种提供参考依据。     

不同地理种群甘薯茎线虫乙酰胆碱酯酶对抑制剂的敏感性

用改进的Ellman方法测定了河北省昌黎、涿州、卢龙和抚宁四个不同地理种群甘薯茎线虫Ditylenchus destructor的乙酰胆碱酯酶（AChE）的生化特性及其对抑制剂涕灭威和毒扁豆碱的敏感性差异。结果表明：甘薯茎线虫AChE至少存在两种类型，一种类型对毒扁豆碱的IC50值小于10^-6mol／L，另一种类型大于10^-4mol／L。当抑制剂毒扁豆碱浓度为10^-6～10^10mol／L、涕灭威浓度为10^-5～10^10mol／L时，甘薯茎线虫昌黎种群AChE的敏感性抑制率明显高于涿州、抚宁、卢龙三地的甘薯茎线虫种群AChE的抑制率。各地种群AChE对抑制剂敏感性差异是由于甘薯茎线虫AChE的多态性和各分子型综合性改变的结果。     

人参总皂苷及其衍生物对小鼠淋巴细胞和NK细胞免疫活性的影响

为了提高人参总皂苷的生物学活性,对其进行了硫酸化修饰,并探讨总皂苷及其衍生物对小鼠脾T、B淋巴细胞增殖（MTT法）和NK细胞杀伤活性（乳酸脱氢酶释放法）的影响。结果表明,2种衍生物质量浓度在50～500 mg/L时可以促进小鼠脾T、B淋巴细胞的分化增殖和NK细胞对YAC-1的杀伤作用,质量浓度较高时促进作用明显优于总皂苷;衍生物质量浓度达到1 400 mg/L时,仍然可以显著促进T淋巴细胞的增殖。试验结果提示,人参皂苷的这种衍生物有可能成为一种毒副作用较小、免疫活性更强的药物。     

高致病性猪繁殖与呼吸综合征病毒的分离鉴定及其分子流行病学分析

为了弄清2006年在我国南方一些猪场暴发的一种以高热、高发病率和高死亡率为特征的传染病的病因,我们从12个省市48个发病猪场采集临床样品进行了猪繁殖与呼吸综合征病毒(Porcine reproductive and respiratory syndrome virus,PRRSV)的分离鉴定和分子流行病学分析。结果显示从48个猪场样品中均检测出了PRRSV,通过完整的ORF5和部分Nsp2基因序列比较分析发现来自12个省市不同猪场的PRRSV高度同源,而且在Nsp2基因上都有两个相同位点的缺失;从分离的15株PRRSV中选择湖南分离株HuN4人工感染5头60日龄的健康猪,结果5头猪分别在7 d～21 d内全部死亡,感染猪的临床症状和现地所见十分相似。本次试验结果表明新出现的以Nsp2基因部分缺失为特征的PRRSV对猪是高致病性的,可能是2006年大部分猪场暴发"高热综合症"的主要病因。     

猪胸膜肺炎放线杆菌Apx毒素的分子生物学与致病机理

猪传染性胸膜肺炎是由胸膜肺炎放线杆菌(App)引起的猪的一种呼吸道传染病.到目前为止,APP已经被分离鉴定出了15种血清型(1～15).15种血清型的APP能产生属于RTX毒素家族的4种Apx毒素,分别为ApxⅠ、ApxⅡ、ApxⅢ和ApxⅣ.ApxⅠ操纵子包含有ApxⅠ C、A、B、D 4个基因;ApxⅠ毒素含有13个富含甘氨酸的重复区.ApxⅡ操纵子仅包括结构基因ApxⅡA和激活基因ApxⅡC,而无分泌基因;ApxⅡ蛋白含有8个甘氨酸的序列重复.ApxⅢ操纵子与ApxⅠ操纵子相同,具有一个完整的操纵子;ApxⅢ在N～末端有3个疏水区,在C末端部分有13个富含甘氨酸的区域.Apx毒素的致病机理为先由没有活性的前体蛋白转变为有活性的ApxA蛋白,再由有活性的ApxA蛋白侵害细胞.     

鸡传染性支气管炎病毒N蛋白研究进展

鸡传染性支气管炎病毒(IBV)是严重危害养鸡业的重要病原之一.近年来国内外学者在分子生物学领域方面的研究不断深入,在病毒基因组的结构与功能及其N蛋白的研究方面取得了重要进展.对N蛋白如何特异性地识别病毒基因组RNA、参与病毒RNA的复制和包装、诱导IBV体液和细胞免疫应答的机制、IBV新型基因工程疫苗研制以及IBV的抗体水平监测等方面的研究,为IBV基因工程疫苗设计及其免疫防控提供了科学依据.     

Microsatellite markers based assessment of genetic diversity in Iranian olive (Olea europaea L.) collections

To study the genetic variation in Iranian olive collections and some foreign olive cultivars, 47 accessions of 18 local cultivars from 6 olive collections of Iran (Roudbar, Zanjan, Ahvaz, Dezful, Kazeroon and Shiraz), were analyzed along with 30 imported cultivars using 16 microsatellite primer pairs. All the used microsatellite loci revealed polymorphism in the studied genotypes, except GAPU14 and GAPU113 markers. Fourteen microsatellite primers amplified 126 polymorphic alleles in the 87 selected olive accessions. The average number of alleles per locus was 9, ranging from 3 to 14. Polymorphic information content (PIC) was 0.85. The genetic similarity based on Jaccard coefficient ranged from 0.15 to 1. The genetic relationships among accessions were investigated using cluster analysis and principal component analysis (PCA). Most of the accessions with the same name were grouped together; some exceptions were also observed. As expected, close relationship was observed among accessions within same cultivar. Most of the Iranian olive accessions were clustered to a main distinct group. Two-dimensional scatter plot of principal component analysis revealed a clear separation of most of the Iranian olives from Syrian and other introduced cultivars. These suggest that Iranian cultivars have different origin related to West Mediterranean basin cultivars and have evolved independently from the others. Between and within Iranian and foreign cultivars (cultivars including three or more accessions) genetic diversity was analyzed using analysis of molecular variance (AMOVA). AMOVA revealed higher within cultivar genetic variation (62.76%) as compare to that between cultivar variations (37.24%). The intra- and inter-cultivar variance tested by permutation test showed significant genetic variation at both levels. The high level within cultivar genetic variance could be due to mislabeling and presence of homonyms in cultivars produced by vegetative propagation from original plants.     

Assessment of genetic purity of tomato (Lycopersicon esculentum L.) hybrid using molecular markers

Three DNA molecular marker systems, RAPD, ISSR and SSR, were used to test seed genetic purity of two commercial hybrid tomato (Lycopersicon esculentum L.) cultivars ‘Hezuo 903’ and ‘Sufen No. 8’. Genomic DNA from the two F₁ hybrid cultivars and their corresponding parental lines was screened with 218 RAPD decamer primers, 54 ISSR primers and 49 SSR primers. Among the 321 primers, 4 primers for ‘Hezuo 903’ and 3 for ‘Sufen No. 8’, which could produce both female and male parent-specific markers, were selected for testing the genetic purity. A total of 210 hybrid individuals of each cultivar were analyzed using the identified primers. The combined results of the marker analysis showed that eight of the 210 F₁ plants in ‘Hezuo 903’ and 13 of 210 in ‘Sufen No. 8’ were false hybrids, and the overall genetic purity of the two F₁ hybrid seed lots was 96.2 and 93.8%, respectively. This study showed that RAPD and SSR markers could provide a practical and efficient tool in quality control of the tomato commercial hybrid seeds.