Kaffashi Amir, Eshratabadi Fatemeh, Shoushtari Abdelhamed
Razi Vaccine and Serum Research Institute, Hesarak, Karaj, Iran.
Virus Genes. 2017 Apr;53(2):312-316. doi: 10.1007/s11262-016-1417-8. Epub 2016 Dec 8.
An Iranian field strain of chicken anemia virus (CAV), designated IR CAV, was isolated in the Marek's disease virus-transformed lymphoblastoid cell line MDCC-MSB1 (MSB1) culture for the first time. The full-length CAV DNA of this strain was cloned in the bacterial plasmid pTZ57R/T to create the molecular clone pTZ-CAV. The nucleotide and deduced amino acid sequences of viral proteins of IR CAV were compared with those of representative CAV sequences including reference and commercial vaccine strains. IR CAV was not related to vaccine strains and also found to have glutamine at positions 139 and 144 confirming previous studies in which such mutations were associated with a slow rate of virus spread in cell culture. pTZ-CAV was digested with PstI to release IR CAV DNA and then transfected into MSB1 cell by electroporation. The transfected cells showed cytopathic effect similar to virion-initiated infection. One-day old specific pathogen-free chicks were inoculated with the regenerated virus, which had been obtained from transfected MSB1 cells, and compared with the chicks inoculated with IR CAV. Gross lesions in the birds inoculated with the regenerated virus illustrated the infectious nature of the regenerated virus from the cloned IR CAV DNA.
一种伊朗鸡贫血病毒（CAV）田间毒株，命名为IR CAV，首次在马立克氏病病毒转化的淋巴母细胞系MDCC - MSB1（MSB1）培养物中分离得到。该毒株的CAV全长DNA被克隆到细菌质粒pTZ57R/T中，构建了分子克隆pTZ - CAV。将IR CAV病毒蛋白的核苷酸序列和推导的氨基酸序列与包括参考毒株和商业疫苗株在内的代表性CAV序列进行了比较。IR CAV与疫苗株无关，并且在第139和144位发现有谷氨酰胺，这证实了先前的研究，即此类突变与病毒在细胞培养中的缓慢传播速度有关。用PstI消化pTZ - CAV以释放IR CAV DNA，然后通过电穿孔将其转染到MSB1细胞中。转染后的细胞表现出与病毒粒子引发的感染相似的细胞病变效应。用从转染的MSB1细胞中获得的再生病毒接种1日龄无特定病原体雏鸡，并与接种IR CAV的雏鸡进行比较。接种再生病毒的鸡的肉眼病变说明了从克隆的IR CAV DNA中获得的再生病毒具有感染性。
