NDV La Sota株三个病毒克隆株间生物学特性的比较研究

为比较新城疫病毒(NDV)La Sota株不同克隆株之间的生物学特性,本研究通过挑取细胞病变区对LaSota株病毒进行纯化,并获得了3个病毒克隆株。通过血凝素热稳定性试验、病毒复制动力学试验和免疫原性试验,对3个病毒克隆株进行分析比较。结果显示,不同的病毒克隆株间在这些生物学特性上存在着不同程度的差异,表明为了获得性质均一、稳定的病毒,有必要进行病毒的纯化,并根据实验目的进行比较取舍。     

不同鸡传染性支气管炎疫苗毒株对新城疫La Sota株的增殖干扰试验

为观察IBV不同疫苗毒株H120、H52、4/91、28/86、Ma5及W93株对NDV增殖的干扰作用,采用不同浓度的IBV疫苗毒株与NDV La Sot a毒株分别按不同顺序接种,同胚增殖。利用血凝试验(HA)测定NDV效价,以判断IBV对NDV的干扰作用,为同胚增殖两种病毒提供参考数据。试验结果表明,IBV不同疫苗毒株能干扰NDV La Sot a增殖,干扰作用与其接种顺序、接种浓度及收毒时间有关。     

鸡新城疫、传染性支气管炎病毒同胚接种试验

通过对五个批次的无特定病原体(SPF)鸡胚进行鸡新城疫病毒（NDV）、鸡传染性支气管炎病毒（IBV）同胚接种,收获72～120 h死胚及120 h活胚尿囊液,分别测定NDV和IBV的病毒含量,结果表明NDV病毒含量达到了108.3～108.7EID50/0.1mL, IBV的病毒含量达到了106.3～106.5EID50/0.1mL。两种病毒含量均达到《中华人民共和国兽用生物制品制造及检验规程》（2000版）中鸡新城疫、传染性支气管炎二联活疫苗病毒含量要求的标准。表明该方法即可降低成本,又简化了生产工艺。     

Ultrastructural changes of the tracheal epithelium after vaccination of day-old chickens with the La Sota strain of Newcastle disease virus

The progression of tracheal lesions induced by vaccination of day-old specific pathogen-free chicks with the La Sota strain of Newcastle disease virus (NDV) was examined by relating surface changes as observed by scanning electron microscopy with subcellular changes seen by transmission electron microscopy. NDV infection resulted in hypertrophy of goblet cells, their rupture, and the formation of excess mucus. Activation of goblet cells peaked within 4 days postvaccination. Afterward, the activation levels gradually decreased. At the level of the ciliated cells, a marked increase in the proportion of nonciliated to ciliated cells and later an almost complete deciliation of the tracheal surface were observed because a simple squamous to cuboidal epithelium replaced the original pseudostratified epithelium. Fifteen days postvaccination, all epithelial damage was restored. Because the observed vaccination-induced lesions are detrimental to epithelial integrity and function as a barrier against invading microorganisms, they might explain at the ultrastructural level the secondary complications of vaccination with the La Sota strain against NDV.     

Tracheal mucus transport rate and bacterial clearance in turkeys exposed by aerosol to La Sota strain of Newcastle disease virus

Tracheal mucus transport rate (TMTR) and quantitative clearance of aerosolized Escherichia coli from the trachea, lung, and air sac were measured in healthy unanesthetized turkeys and in turkeys exposed by aerosol to a La Sota vaccine strain of Newcastle disease virus (NDV). The TMTR of uninfected turkeys was 42.4 +/- 14.7 cm/min. The TMTR of NDV-infected turkeys was depressed on days 3 through 7 postexposure (PE); depression was significant (P less than or equal to 0.05) on day 7 PE. Tracheal E. coli clearance in NDV-infected turkeys was reduced on days 4 through 9 PE, significantly so on day 5 PE (P less than or equal to 0.01). Depression of TMTR and tracheal E. coli clearance were associated histologically with replacement of normal pseudostratified columnar epithelium by 3 to 8 layers of immature nonciliated cells. E. coli clearance by the lung and air sac of NDV-infected turkeys was depressed on days 5 through 9 PE.     

鸡新城疫La Sota株、传染性支气管炎W93株二联活疫苗研究 Ⅲ．疫苗免疫期试验

分别用3批鸡新城疫、传染性支气管炎二联活疫苗(La Sota W93)，ND La Sota单苗和IB W93单苗免疫SPF鸡，并于免疫后第3、7、14、30、90、150天分别用NDV和NIBV强毒攻击。结果表明，二联活疫苗(La Sota W93)于免疫后7-150d可获得与相应单苗同样的保护力。     

Protection induced by infectious laryngotracheitis virus vaccines alone and combined with Newcastle disease virus and/or infectious bronchitis virus vaccines

Two types of live attenuated vaccines have been used worldwide for the control of infectious laryngotracheitis virus (ILTV): 1) chicken embryo origin (CEO) vaccines; and 2) tissue culture origin vaccines (TCO). However, the disease persists in spite of extensive use of vaccination, particularly in areas of intense broiler production. Among the factors that may influence the efficiency of ILTV live attenuated vaccines is a possible interference of Newcastle Disease virus (NDV) and infectious bronchitis virus (IBV) vaccines with the protection induced by ILTV vaccines. The protection induced by CEO and TCO vaccines was evaluated when administered at 14 days of age alone or in combination with the B1 type strain of NDV (B1) and/or the Arkansas (ARK) and Massachusetts (MASS) serotypes of IBV vaccines. Two weeks after vaccination (28 days of age), the chickens were challenged with a virulent ILTV field strain (63140 isolate, group V genotype). Protection was evaluated at 5 and 7 days postchallenge by scoring clinical signs and quantifying the challenge virus load in the trachea using real-time PCR (qPCR). In addition, the viral load of the vaccine viruses (ILTV, NDV, and IBV) was quantified 3 and 5 days postvaccination also using qPCR. The results of this study indicate that the NDV (B1) and IBV (ARK) vaccines and a multivalent vaccine constituted by NDV (B1) and IBV (ARK and MASS) did not interfere with the protection induced by the CEO ILTV vaccine. However, the NDV (BI) and the multivalent (B1/MASS/ARK) vaccines interfered with the protection induced by the TCO vaccine (P < 0.05). Either in combination or by themselves, the NDV and IBV vaccines decreased the tracheal replication of the TCO vaccine and the protection induced by this vaccine, since the ILTV-vaccinated and -challenged chickens displayed significantly more severe clinical signs and ILTV load (P < 0.05) than chickens vaccinated with the TCO vaccine alone. Although NDV and IBV challenges were not performed, the antibody responses elicited by NDV and/or the IBV vaccinations were significantly reduced (P < 0.05) when applied in combination with the CEO vaccine.     

1株肾型鸡传染性支气管炎病毒的鉴定

鸡传染性支气管炎病毒(IBV)具有不同致病特性,将IBV XDC-2株接种9日龄SPF鸡胚培养,可引起鸡胚死亡和出现侏儒胚,病毒EID50达5×10-5.33/mL。将IBV XDC-2株接种18日龄SPF鸡,饲养观察14 d,病鸡临床症状表现为:精神沉郁,羽毛凌乱,双翅下垂,轻微腹泻,多数拉白色水样稀粪。病死鸡出现肾肿大、呈花斑状、大量尿酸盐沉积。鸡发病率为100%,死亡率为25%。死亡鸡肺脏、肾组织制作组织切片,发现病理变化明显,主要为:肾小管扩张,上皮细胞呈玻璃样变性,部分管腔内可见坏死脱落之上皮细胞,于肾间质可见大量单核细胞浸润,肾间质有充血、出血现象;肺内动脉、毛细血管充血,淋巴细胞浸润。死亡鸡肺脏、肾组织接种鸡胚分离病毒,RT-PCR检测结果为阳性,表明该分离株为鸡传染性支气管炎病毒,具有很强的嗜肾性。     

Efficacy of vaccination with La Sota strain vaccine to control Newcastle disease in village chickens in Nepal

The efficacy of vaccination with Newcastle disease (ND) La Sota and R₂B (Mukteswar) modified live strain vaccines was determined by experimental challenge and with ND La Sota vaccine under field conditions in Nepal. Booster vaccination with ND La Sota vaccine after a primary vaccination with ND La Sota vaccine, induced a geometric mean titre (GMT) of 5.0 log₂ haemagglutination inhibition (HI) units, compared to a GMT of 6.0 log₂ HI units following booster vaccination with R₂B vaccine 1 month after primary vaccination with ND La Sota vaccine. Both vaccines provided 100% protection against challenge with a local field ND strain. Furthermore, booster vaccination with ND La Sota vaccine induced protective levels of antibody after field use in villages in Jhapa, and no outbreaks of ND occurred during the study period. The ND La Sota modified live vaccine is immunogenic and efficacious and is a suitable vaccine for use in vaccination programmes in village chickens in the rural areas of Nepal.

     

鸡新城疫La Sota株、鸡传染性支气管炎W93株二联活疫苗研究Ⅱ.接毒剂量选择试验

将鸡新城疫La Sota毒种和嗜肾型鸡传染性支气管炎W93毒种分别用生理盐水做适当稀释后，将两种病毒液等量混合，接种于同一SPF鸡胚尿囊腔内。结果表明，HBV、NDV在同一鸡胚内增殖时无互相干扰现象；收获的鸡胚液(La Sota W93)中NDV、IBV的效价分别达到相应单苗水平。     

表达鸡传染性支气管炎病毒S基因的重组新城疫病毒构建

为构建以新城疫病毒(NDV)为活毒载体表达鸡传染性支气管炎病毒(IBV)S基因的重组病毒,本研究利用RT-PCR技术,以IBV的Massachusetts41株RNA为模板,通过RT-PCR扩增得到IBV S基因(3534bp),将其插入到NDV感染性克隆pBRN-FL中,构建了含有IBV S基因的重组NDV cDNA克隆pBRN-FL-IBVS。利用磷酸钙转染法,在辅助质粒pBS-NP、pBS-P和pBS-L的共同作用下,将pBRN-FL-IBVS转染表达T7聚合酶重组痘病毒感染的BSR细胞,救获重组NDV(rL-IBVS)。采用RT-PCR检测接种重组病毒的鸡胚尿囊液,结果表明rL-IBVS中含有相应外源基因。IFA试验表明,rL-IBVS可与鸡抗IBV的高免血清发生特异性反应,证明S蛋白在感染的BSR细胞中得到表达。其鸡胚平均致死时间、脑内致病指数和静脉内致病指数等指标显示rL-IBVS保持了亲本疫苗株高滴度的鸡胚生长特性和低致病力特性。本研究采用反向遗传操作技术构建了表达IBV S蛋白的重组NDV,为进一步研制IBV和NDV的重组基因工程活载体疫苗奠定了基础。     

山东省传染性支气管炎病毒分离株的电镜观察

从山东省分离到4株病毒株，经病理剖检、鸡胚传代，血凝检测、动物回归、电镜和免疫电镜观察确证为传染性支气管炎病毒。将分离毒分别接种SPF鸡胚，制备各自的阳性血情，经磷钨酸负染，电镜和免疫电镜观察。可见到典型的冠状病毒粒子，病毒的直径界于60－200mm之间，呈多形性，有囊膜，个别有纤突。病毒大量聚集，各株病毒在形态上无差异。     

Serotypes of avian infectious bronchitis virus isolates from field cases in Japan

Eight etiologic agents isolated from field cases in Japan were identified as isolates of infectious bronchitis virus by agar-gel diffusion, buoyant-density determination on sucrose-density centrifugation, and morphological study by electron microscope of the purified viruses. In studies of the antigenic relationships of the eight isolates and six known infectious bronchitis viruses, antigenic diversity of these viruses was recognized from the degrees of relatedness using a plaque reduction in the "constant-virus decreasing-serum" method.     

Tissue tropism of three cloacal isolates and Massachusetts strain of infectious bronchitis virus

Three virus isolates (ECV-1, -2, and -3) recovered from cloacae of chickens in flocks that experienced drops in egg production were identified as infectious bronchitis virus (IBV), based on characteristic embryo lesions, chloroform sensitivity, coronavirus morphology, and serology. Because these isolates were recovered from the cloacae of the hens, their tissue tropism was compared with the prototype strain of IBV, Massachusetts-41 (M-41), in experimentally inoculated chickens. During the 39-day period postinoculation (PI), virus isolation was attempted from digestive and respiratory tracts, kidney, and cloacal swabs. ECV-1, ECV-2, and M-41 were more frequently recovered from the cecal tonsils than from other tissues. ECV-1, ECV-3, and M-41 were also recovered from kidney for up to 39 days PI. ECV-2 and ECV-3 had a limited distribution in respiratory tissues, being isolated only sporadically from trachea, bronchus, and lung. Surprisingly, ECV-2 was isolated from esophagus at 2, 16, 30, and 39 days PI; otherwise, its distribution in other tissues was sporadic. Results confirmed that IBV, including M-41, can infect a variety of tissues and that some isolates may be recovered frequently from digestive tract tissues, particularly from the cecal tonsils.