The Molecular Characterization and Immunity Identification of Microneme 3 of Eimeria acervulina

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• 作者：ZhenChao Zhang ; XinChao Liu ; XinChao Yang ; LianRui Liu ; Shuai Wang ; MingMin Lu ; Mohammad Ehsan ; Javaid A. Gadahi ; XiaoKai Song ; LiXin Xu ; RuoFeng Yan and XiangRui Li
• 刊名：Journal of Eukaryotic Microbiology
• 出版年：2016
• 出版时间：November/December 2016
• 年：2016
• 卷：63
• 期：6
• 页码：709-721
• 全文大小：1282K
• ISSN：1550-7408

文摘

The gene of Eimeria acervulina microneme protein 3 (EaMIC3) was cloned and characterized. According to the conserved sequence, the 3′- and 5′-ends of EaMIC3 were amplified by the rapid amplification of cDNA ends (RACE). The full length cDNA of this gene was obtained by overlapping the sequences of 3′- and 5′-extremities and amplification by reverse transcription PCR. The sequence analysis revealed that the opening reading frame (ORF) of EaMIC3 was 2,607 bp and encoded a protein of 868 amino acids with 93.04 kDa. Western blotting assay showed that the recombinant protein was successfully recognized by the sera of chickens experimentally infected with E. acervulina, whereas the native protein in the somatic extract of sporozoites was as well detected by sera from rats immunized with the recombinant protein of EaMIC3. Immunofluorescence analysis indicated that EaMIC3 was expressed in the sporozoites and merozoites stages of E. acervulina. Animal challenge experiments demonstrated that the recombinant protein of EaMIC3 could significantly increase the average body weight gains, decrease the mean lesion scores and the oocyst outputs of the immunized chickens and presented anticoccidial index (ACI) more than 165. Moreover, EaMIC3 protein produced significantly high level of IgG antibody, IFN-γ, IL-10, IL-17 TGF-β, CD4⁺, and CD8⁺.