腺胃病变型鸡传染性支气管炎病原特性与多肽分析

STUDY ON THE PATHOGEN CHARACTERISTICS AND POLYPEPTIDES ANALYSIS OF AVIAN INFECTIOUS BRONCHITIS OF PROVENTRICULUS PATHOTYPE

从患传染性腺胃病的病鸡中分离到4 株病毒,其中H95株在SPF鸡胚上已稳定传代。病料及H95 传代后的鸡胚尿囊液经蔗糖梯度和CsCl密度梯度离心后的纯化物,电镜下为大小80～160 nm 、有囊膜的病毒颗粒,囊膜表面有纤突,呈冠状形态。病毒在CsCl中的浮密度为1.19～1.23 g/cm 3, 对热和氯仿敏感,耐酸,能抵抗1% 胰酶,无直接血凝性,经卵磷脂酶C处理后的病毒则能凝集鸡红细胞,能干扰新城疫病毒在鸡胚上的增殖。应用杂交瘤技术筛选出4 株抗H95株的阳性克隆,经Western blotting, 其中3 株单抗能识别54 kd 蛋白多肽,该条带也能与来自鸡传染性支气管炎病毒(IBV) M41 株的针对N 蛋白的单抗反应。中和试验表明, H95株与其它IBV 毒株在血清型上存在差异。上述研究结果初步表明,鸡传染性腺胃病的病原是冠状病毒科的成员,并且很可能是鸡传染性支气管炎(IB)中以腺胃病变为主要特征的一种新的致病病型。

Four strains viruses were isolated from illed chickens with infectious proventriculus disease. And H 95 strain had steadily passed on SPF chicken embryos. The viruses were purified from pathogenic materials and allantoic fluid of chicken embryos inoculated with strain H 95 by sucrose gradient and CsCl isopycnic gradient centrifugation. Under electron microscope observation, the purified virus particles were 80~160 nm in size, enveloped. Bulb shaped spikes on the envelope surface were observed. The buoyant density of the virus in CsCl was 1.19~1.23 g/cm 3. The virus was sensitive to heat and chloroform, stable at pH 3.0, resistant to 1% trypsin, no direct hemagglutination activity and could interfere in propagatation of NDV in chicken embryos. The virus could agglutinated chicken erythrocyte when treated with phospholipase C. Four hybridoma cell lines secreting MAbs against H 95 strain were screened, and three of them could react with 54 kd peptide by SDS PAGE and Western blotting analysis. MAb 6DH 8, which was identified defining an epitope on N protein of IBVM 41 could react with the same 54 kd peptide of H 95 strain, too. Neutralization test showed that H 95 strain was quite diffence from some IBV strains in serum type. All those results showed that the pathogen of infectious proventriculus disease of chicken was a member of coronaviridae and had close correlation with IBV to some extent.