Shanghai Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Key Laboratory of Animal Parasitology of Ministry of Agriculture, Minhang, Shanghai, 200241, People's Republic of China.
College of Life Sciences, Shanghai Normal University, Shanghai, 200234, People's Republic of China.
Parasitol Res. 2019 Oct;118(10):2989-2999. doi: 10.1007/s00436-019-06437-0. Epub 2019 Aug 31.
Chicken coccidiosis is caused by the apicomplexan parasite Eimeria spp. At present, drug resistance of Eimeria is common because of the indiscriminate use of anticoccidial drugs. The gene encoding surface antigen 10 of Eimeria tenella (EtSAG10) is differentially expressed between drug-resistant and drug-sensitive strains. RNA-seq analysis indicated that this gene was downregulated in strains resistant to maduramicin and diclazuril compared to susceptible strains. EtSAG10 DNA sequence alignment revealed that they contained one and ten mutations in MRR and DZR, compared with DS, respectively. A full-length EtSAG10 cDNA was successfully cloned and expressed, and the polyclonal antibody was prepared. The transcription and translation levels of EtSAG10 were analyzed by quantitative real-time PCR (qPCR) and Western blotting. The localization of EtSAG10 in Spz, Mrz, and parasites in the first asexual stage was determined by indirect immunofluorescence. The potential association of EtSAG10 with sporozoite invasion of host cells was assessed by invasion inhibition assays. The results showed that EtSAG10 had a predicted transmembrane domain at the C-terminal end and a predicted signal peptide at the N-terminal end. EtSAG10 was downregulated in drug-resistant strains, which is consistent with the RNA-seq results. The EtSAG10 protein was localized to the parasite surface and parasitophorous vacuole membrane. This protein was shown to play a role in the infection of chicken intestine by sporozoites.
鸡球虫病是由艾美耳属顶复门寄生虫引起的。目前，由于抗球虫药物的滥用，艾美耳属对药物的耐药性很常见。柔嫩艾美耳球虫（Eimeria tenella）表面抗原 10 基因（EtSAG10）在耐药株和敏感株之间存在差异表达。RNA-seq 分析表明，与敏感株相比，该基因在对马杜霉素和地克珠利耐药的株系中表达下调。EtSAG10 DNA 序列比对显示，它们在 MRR 和 DZR 中分别比 DS 含有一个和十个突变。成功克隆并表达了全长 EtSAG10 cDNA，并制备了多克隆抗体。通过定量实时 PCR（qPCR）和 Western blot 分析了 EtSAG10 的转录和翻译水平。通过间接免疫荧光法确定了 EtSAG10 在 Spz、Mrz 和第一个无性生殖阶段寄生虫中的定位。通过入侵抑制试验评估了 EtSAG10 与子孢子入侵宿主细胞的潜在关联。结果表明，EtSAG10 在 C 末端具有预测的跨膜结构域，在 N 末端具有预测的信号肽。耐药株中 EtSAG10 表达下调，与 RNA-seq 结果一致。EtSAG10 蛋白定位于寄生虫表面和滋养体空泡膜。该蛋白在子孢子感染鸡肠的过程中发挥作用。
