检测猪流行性腹泻病毒的R-PCR方法的建立

根据猪流行性腹泻病毒 (PEDV)的N基因自行设计和合成了一对可扩增长度为 641bp目的片段的引物 ,成功地建立了检测的猪流行性腹泻病毒的RT PCR方法。对猪轮状病毒 (PRV)、猪传染性胃肠炎病毒 (TGEV)的RT PCR检测结果均呈阴性。对PEDV JS株的RT PCR产物的序列分析表明 ,与CV777株的同源性为 97 3 %。     

重组鸡IL-2增强鸡四联灭活苗免疫效果的研究

使用在原核表达系统中表达的重组鸡白细胞介素 2 (IL 2 )蛋白 ,经过初步的分离纯化 ,与鸡新城疫 禽流感 法氏囊 传染性支气管炎四联油乳剂灭活苗同时使用 ,检测其对疫苗的免疫增强效果。试验共分 5组 ,每组 1 0只鸡。其中对照组只注射四联油乳剂灭活苗 ;4个试验组 ,在注射疫苗的同时分别注射重组鸡IL 2蛋白 0 0 5mg、 0 0 1mg、 0 1 5mg、 0 2 0mg。试验组与对照组同时各注射新城疫等多联油苗 0 5mL。采用HA、HI试验测定鸡新城疫、禽流感抗体滴度。结果表明 :重组鸡IL 2对该四联油乳剂灭活苗有较强的免疫增强作用。以第 3、 4组的增强效果较为明显 ,其中对鸡新城疫病毒和禽流感抗原的抗体效价 ,与对照组相比分别提高 2 42 8和 2 2 3 0个滴度。重组鸡IL 2在使用的过程中 ,没有发现任何毒副作用 ,证明是一种安全高效的免疫增强剂。     

圆点印迹法检测葡萄斑点病毒技术研究

采用圆点印迹法检测GFkV，结果表明：Tris-HCl和PBS在提取GFkV时效果相同，在带毒葡萄植株中韧皮部的GFkV含量较高。     

Factors affecting the development of disease symptoms in potatoes infected by <Emphasis Type="Italic">Tobacco rattle virus</Emphasis>

Infection of potato plants with Tobacco rattle virus by its nematode vector can have different outcomes, including the development of spraing symptoms in progeny tubers. A novel syndrome described here comprises a generalized mottle in leaves on all stems, together with virtually total failure to produce daughter tubers. The outcome of infection depends partly on the potato genotype, but field trials with 15 varieties showed that the incidence and severity of spraing symptoms at three sites differed. There was no evidence that this was due to differences in virulence among the virus isolates at the sites, but it was probably the result of environmental differences that influenced the numbers and activity of the vector nematodes. At the most severely affected site, spraing symptoms were found in all varieties tested, except Record, including several that were not affected at the other two sites. Taking both severity and incidence of symptoms into account, the ranking of varieties was similar at each site. The incidence of spraing symptoms was greater in larger than in smaller tubers, and increased with later harvest dates, but did not increase when early harvested tubers were stored. Tobacco rattle viruswas detected in the roots of many of the weeds growing at one of the sites and in the roots of a few plants of the subsequent barley crop.     

Validation of the specificity and sensitivity of species-specific primers that provide a reliable molecular diagnostic for <Emphasis Type="Italic">Xiphinema diversicaudatum</Emphasis>, <Emphasis Type="Italic">X. index</Emphasis> and <Emphasis Type="Italic">X. vuittenezi</Emphasis>

Xiphinema diversicaudatum and X. index are vector nematode species of economic importance in viticulture regions as they can transmit Arabis Mosaic, Grapevine Fanleaf and Strawberry Latent Ringspot viruses to grapevine. Wang et al. (2003) designed species-specific diagnostic primers from ribosomal genes for both these vector species as well as a vector and a non-vector species X. italiae and X. vuittenezi, respectively. Our study aimed to confirm the specificity and determine the sensitivity and reliability of the primers for the two vector species, X. diversicaudatumand X. indexwhen challenged with closely related longidorid species and general nematode communities typical of vineyard soil. With one exception, no PCR product was observed when the primers were tested against six Longidorus, one Paralongidorus and one Xiphinema non-target species. Occasionally (three out of eight replicate PCR reactions) a weak PCR product was noted when primers for X. index were tested with L. elongatus. Furthermore, when challenged with a range of non-target nematode species comprising the nematode community typical of viticulture soil, no PCR product was amplified. An experimental dilution series of extracted DNA rigorously demonstrated that DNA from an equivalent single specimen of the target virus-vector species, X. diversicaudatum and/or X. index, could be detected amongst 1000 equivalent non-targetX. vuittenezi. Also, extracted DNA from an equivalent single target specimen was detected when added to DNA extracted from the overall soil nematode community. The primers were assessed further by using serial mixtures of actual nematodes rather than extracted DNA to simulate field soil. Using this method, a single target nematode could be detected amongst 200 non-target specimens. Given their specificity, sensitivity and reliability, it appears that these diagnostic primers will be of great benefit to phytosanitary/quarantine services related to the viticulture industry.     

犬传染性喉气管炎病毒的分离及鉴定

用犬肾细胞 (MDCK)从疑似犬传染性喉气管炎病毒感染犬的急性期血清中分离到 1株病毒 ,经血凝试验、免疫电镜观察、中和试验、免疫荧光抗体试验、细胞培养特征观察和回归动物试验等鉴定 ,证明所分离的病毒为犬传染性喉气管炎病毒 ,命名为 BJ- JB- 3。     

鸡痘病毒通用高效表达载体的构建及其初步应用

利用分子克隆技术对本室构建的高效鸡痘病毒表达载体 p1 1 S进行改造 ,在其人工合成禽痘病毒 ( FPV)强启动子 Ps的下游引入含 7个单一酶切位点的多克隆位点 ( MCS) ,构建了便于外源基因插入的通用性更强的高效单表达载体 p N1 1 S。然后将 FPV早晚期启动子 PE/ L 及其启动的鹅源新城疫病毒 NDV ZJ1株的 F基因一并插入 p N1 1 S中 ,使PE/ L 与 Ps反向串联 ,从而构建出 1个含 NDV ZJ1株 F基因的重组 FPV高效双表达载体 p N1 1 SEF,其 MCS可以用于插入其他外源基因。在此基础上 ,将 NDV ZJ1株的 HN基因插入 p N1 1 SEF的 Ps启动子下游的 MCS中 ,构建了共表达 NDV ZJ1株 F和 HN基因的重组 FPV双表达载体 p N1 1 SEFHN。将 p H1 1 SEFHN质粒 DNA与 FPV2 82 E4株共转染 CEF,得到共表达 NDV ZJ1株和 HN基因的重组 FPV,间接免疫荧光实验初步证明外源基因得到了较好的表达。表明所构建的通用高效 FPV表达载体有利于高效基因工程活载体疫苗的研制 ,具有广阔的应用前景     

抗鸡传染性喉气管炎重组鸡痘病毒基因工程疫苗同居感染试验

疫苗的接触传播是疫苗免疫接种需要考虑的重要因素,为了检测重组鸡痘病毒载体疫苗水平传播的能力,对隔离条件下饲养的SPF鸡用重组鸡痘病毒基因工程疫苗接种,同时设立非免疫对照鸡,饲养期间特意延长清粪时间以增加感染的机会,1个月之后攻击传染性喉气管炎WG株强毒和鸡痘102株强毒,疫苗免疫鸡全部获得保护,而非免疫鸡则全部发病.在试验动物饲养场的自然条件下,将免疫鸡和试验对照两组鸡饲养在同一个鸡舍内,让疫苗毒的传播更接近自然条件.在每个月的攻毒试验中,对照鸡都没有获得对鸡痘和传染性喉气管炎强毒的保护.在疫苗免疫期间进行连续5个月的跟踪检测,同居未免疫鸡没有检测到抗传染性喉气管炎病毒gB抗体.这些实验结果表明抗鸡传染性喉气管炎重组鸡痘病毒基因工程疫苗不能通过接触传播.     

IMC10200株ALV-J实验诱发禽骨髓性白血病的研究

对分离自肉种鸡群亚临床感染的J亚群禽白血病病毒IMC1o200株进行了实验感染诱发禽骨髓性白血病的病理学研究.IMC10200株J亚群禽白血病病毒尿囊腔接种11日龄肉鸡胚和SPF蛋鸡胚,孵出后跟踪观察.9周龄时随机抽检感染鸡,感染肉鸡特异引物PCR检测全部为阳性;组织病理学观察感染鸡无明显病变.至21周龄时感染肉鸡出现第一例典型骨髓性白血病病例;感染SPF蛋鸡未见明显J亚群禽白血病相关病变.     

Wind-mediated spread of Rice yellow mottle virus (RYMV) in irrigated rice crops

A study was carried out to demonstrate that Rice yellow mottle virus (RYMV), a virus known to be transmitted by beetles, can spread between rice plants by direct leaf contact caused by wind. Almost all healthy plants surrounding an infected plant became infected when exposed to a fan blowing for 15 min at a distance of 50 cm. Spread of RYMV by plant contact, mediated by wind, was also demonstrated in field experiments, the extent of spread depending on plant density. Infection was almost 10 times higher in plots with a density of 33 plants m⁻² than in plots with 16 plants m⁻². Less spread was observed in plots protected by 1·5 m high windscreens. It is suggested that wind-mediated spread of RYMV may result from abrasive contact between leaves of plants.