The porcine coccidium Cystoisospora suis is characterized by a complex life-cycle during which asexual multiplication is followed by sexual development with two morphologically distinct cell types, the micro- and macrogametes. Genes related to the sexual stages and cell cycle progression were previously identified in related Apicomplexa. Dynein light chain type 1 and male gamete fusion factor HAP2 are restricted to microgametes. Tyrosine-rich proteins and oocyst wall proteins are a part of the oocyst wall. The Rad51/Dmc1-like protein and Nima-related protein kinases are associated with the cell cycle and fertilization process. Here, the sexual stages of C. suis were characterized in vitro morphologically and for temporal expression changes of the mentioned genes to gain insight into this poorly known phase of coccidian development. Sexual stages of C. suis developing in vitro in porcine intestinal epithelial cells were examined by light and electron microscopy. The transcriptional levels of genes related to merozoite multiplication and sexual development were evaluated by quantitative real-time PCR at different time points of cultivation. Transcription levels were compared for parasites in culture supernatants at 6–9 days of cultivation (doc) and intracellular parasites at 6–15 doc. Sexual stage of C. suis was detected during 8–11 doc in vitro. Microgamonts (16.8 ± 0.9 µm) and macrogamonts (16.6 ± 1.1 µm) are very similar in shape and size. Microgametes had a round body (3.5 ± 0.5 µm) and two flagella (11.2 ± 0.5 µm). Macrogametes were spherical with a diameter of 12.1 ± 0.5 µm. Merozoite gene transcription peaked on 10 doc and then declined. Genes related to the sexual stages and cell cycle showed an upregulation with a peak on 13 doc, after which they declined. The present study linked gene expression changes to the detailed morphological description of C. suis sexual development in vitro, including fertilization, meiosis and oocyst formation in this unique model for coccidian parasites. Following this process at the cellular and molecular level will elucidate details on potential bottlenecks of C. suis development (applicable for coccidian parasites in general) which could be exploited as a novel target for control.

中文翻译：

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表征Cystoisospora猪链球菌性阶段，在体外

猪球孢梭状芽胞杆菌的特征在于复杂的生命周期，在此过程中，无性繁殖继之以具有两种形态上不同的细胞类型（微型配子和大型配子）的有性发育。与性阶段和细胞周期进程有关的基因先前已在相关的蚜虫复合体中鉴定出来。Dynein轻链1型和雄配子融合因子HAP2仅限于微型配子。富含酪氨酸的蛋白和卵囊壁蛋白是卵囊壁的一部分。Rad51 / Dmc1样蛋白和Nima相关蛋白激酶与细胞周期和受精过程有关。在这里，猪C.suis的有性阶段在体外进行了形态学表征，并提到了上述基因的时间表达变化，从而洞悉了这一球虫发育的鲜为人知的阶段。通过光镜和电子显微镜检查了猪肠上皮细胞在体外发育的猪链球菌的有性阶段。通过定量实时PCR在培养的不同时间点评估与裂殖子繁殖和性发育有关的基因的转录水平。比较培养6–9天（doc）培养上清液中寄生虫的转录水平（doc）和6–15 doc时细胞内寄生虫的转录水平。在8-11 doc体外检测到猪链球菌的性阶段。微型（16.8±0.9 µm）和大型（16.6±1.1 µm）的形状和大小非常相似。小配子有一个圆形的身体（3.5±0.5 µm）和两个鞭毛（11.2±0.5 µm）。大型配子为球形，直径为12.1±0.5 µm。裂殖子基因转录在10 doc达到峰值，然后下降。与性阶段和细胞周期有关的基因显示上调，在13 doc时达到峰值，然后下降。本研究将基因表达的变化与体外念珠菌性发育的详细形态描述联系起来，包括在这种针对球虫的独特模型中的受精，减数分裂和卵囊形成。在细胞和分子水平上遵循此过程，将阐明可能存在的猪链球菌发育瓶颈（一般适用于球虫寄生虫）的详细信息，可将其用作控制的新靶标。在这种针对球虫寄生虫的独特模型中，减数分裂和卵囊形成。在细胞和分子水平上遵循此过程，将阐明可能存在的猪链球菌发育瓶颈（一般适用于球虫寄生虫）的详细信息，可将其用作控制的新靶标。在这种针对球虫寄生虫的独特模型中，减数分裂和卵囊形成。在细胞和分子水平上遵循此过程，将阐明可能存在的猪链球菌发育瓶颈（一般适用于球虫寄生虫）的详细信息，可将其用作控制的新靶标。