弓形虫Tgcsp2基因敲除株的表型和毒力

Phenotypes and virulence of Tgcsp2 gene knockout Toxoplasma gondii

为研究弓形虫冷休克蛋白2(cold shock domain protein 2 of T.gondii,Tgcsp2)在弓形虫感染宿主过程中的作用,本研究利用CRISPR/Cas9技术构建了弓形虫Tgcsp2基因缺失株(ΔTgcsp2)。通过空斑试验和细胞裂解试验探究ΔTgcsp2虫株在体外的生长能力,结果表明ΔTgcsp2虫株的空斑形成能力和细胞裂解能力相对于RH野生株较弱;弓形虫感染细胞要经过入侵、复制和逸出3个过程。为探究Tgcsp2影响体外感染的具体过程,本研究对ΔTgcsp2虫株和RH野生株进行入侵试验、复制试验和逸出试验,结果表明ΔTgcsp2虫株的入侵能力、复制能力和逸出能力都有不同程度的减弱;将ΔTgcsp2虫株和RH野生株感染BALB/c小鼠进行了体内毒力试验;结果表明,与RH野生株相比,ΔTgcsp2虫株对小鼠的毒力减弱。综上所述,Tgcsp2基因对弓形虫体外生长和体内毒力具有重要的作用。

In order to investigate the role of cold shock domain protein 2 of Toxoplasma gondii(Tgcsp2) in the context of in vitro and in vivo experimental infections,a knockout T.gondii strain deficient in csp2(ΔTgcsp2) was constructed using the CRISPR/Cas9 technology.In vitro growth of ΔTgcsp2 knockout strain was studied by plaque assay and lysis assay,the results indicated that the ability of plaque formation and monolayer lysis in ΔTgcsp2 strain was less than the wild type RH strain.T.gondii infection in host cells involves invasion,replication and egress.To investigate the steps which were affected by Tgcsp2,invasion,replication and egress assays were performed,the results showed that the three processes were all impaired to some extent.The virulence assay in BALB/c mice indicated that the virulence of ΔTgcsp2 strain was attenuated.In conclusion,Tgcsp2 plays an important role in the growth and virulence of T.gondii.