腺胃病变型鸡传染性支气管炎病毒变异株的比较研究

Coronavirus IBV Strains with H 95 Variant Isolated from Chickens Suffered from the Avian Infectious Proventriculus Disease

腺胃病变型鸡传染性支气管炎病毒（ＩＢＶ）变异株Ｈ９５提纯样品经ＳＤＳ－ＰＡＧＥ和Ｗｅｓｔｅｒｎｂｌｏｔｉｎｇ，证明Ｈ９５株单抗ＤＥ７和Ｍ４１株单抗６ＤＨ８均能识别Ｈ９５株５４０００蛋白多肽。采用单抗、多抗介导间接ＥＬＩＳＡ试验表明，Ｈ９５毒株能与抗ＩＢＶＭ４１Ｎ蛋白单抗６ＤＨ８和ＩＢＶＭ蛋白单抗ＭＣ发生反应，也能与抗Ｍ４１、Ｇｒａｙ、Ｈｏｌｔｅ、Ｔ、Ｈ５２、Ｎ１１５和分离株Ｃ９００１、ＤＬＺ９１１１的多抗血清反应，同时抗Ｈ９５株的４株单抗、多抗血清也能与上述毒株反应。卵磷脂酶Ｃ处理的Ｈ９５株的血凝活性能被相应的Ｈ９５株的单克隆抗体和高免血清所抑制，也能被Ｍ４１单抗和高免血清所抑制。通过ＲＴ－ＰＣＲ获得了Ｈ９５毒株的免疫原基因Ｓ１，经Ｓｏｕｔｈｅｒｎｂｌｏｔｉｎｇ和ＩＢＶ的Ｓ探针检测呈阳性。

H 95 strain virus isolated from chickens with infectious proventriculus disease has been identified. By using SDS PAGE and Western blotting, both monoclonal antibody (Mab) DE 7 and Mab 6DH 8 against nucleoprotein of H 95 strain and that of M 41 strain could react respectively with the 54 000 peptide of H 95 strain virus. Mab based or polyclonal antibody based indirect ELISA indicated that H 95 strain could react not only with Mab 6DH 8 and Mab Mc (to membrane Proteim of M 41 strain), but also with polyclonal sera against M 41 , Gray, Holte, T, N 115 , H 52 and their isolates (C 9001 and DLZ 9111 ). On the other hand, all above mentioned IBV strains could also be recognized by the Mabs of four hybridoma cell lines or Polyclonal antibody against H 95 strain. The H 95 virus could agglutinate chicken erythrocytes when treated with phospholipase C. The agglutination activity could be inhibited by the specific antibody (Mabs and polyclonal antibody) against H 95 strain and the Mabs and polyclonal antibody against M 41 . Viral RNA was extracted from the allantoic fluid of chicken embryos infected with H 95 strain, then converted to cDNA. The PCR was performed with two primers which spaned S 1 protein spike region and the amplified produsts of 1 700 was identified by agarose gel electrophoresis, Southern blot with IBV S probe.