Medical College of Yan'an University, Yan'an, China.
Yan'an Key Laboratory of Zoonotic Parasitology Laboratory, Yan'an, China.
J Basic Microbiol. 2024 Jan;64(1):32-41. doi: 10.1002/jobm.202300308. Epub 2023 Sep 12.
The morphological and structural differences of different types of chlamydospore of Arthrobotrys flagrans, a nematophagous fungus, were studied under light microscope and electron microscope to provide a reference for the biological control of parasitic nematodiasis. In this study, A. flagrans isolate F088 dormant chlamydospore and nondormant chlamydospore were selected as the research objects. The structural differences of these spores were observed by optical microscopy through lactol cotton blue, Trypan blue, and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) staining. FunXite -1, 4',6-diamidino-2-phenylindole, and calcofluor white staining were used to observe the metabolic activity, cell wall, and nucleus differences of the two types of spores under fluorescence microscope. Ultrastructure of the two kinds of spores was observed using scanning electron microscope (SEM) and transmission electron microscope (TEM). Since lacto phenol cotton blue, trypan blue staining cannot distinguish dormant spores from dead spores, MTT assay was performed. Fluorescence microscopy observation showed that the cytoplasmic metabolic activity of nondormant spores was stronger than that of dormant spores. The nucleus of dormant spores was bright blue, and their fluorescence was stronger than that of nondormant spores. The cell wall of nondormant spores produced stronger yellow-green fluorescence than that of dormant spores. Ultrastructural observation showed that there were globular protuberances on the surface of the two types of spores but with no significant difference between them. The inner wall of dormant spore possesses a thick zona pellucida with high electron density which was significantly thicker than that of nondormant spores, and their cytoplasm is also changed. In this study, the microstructure characteristics of dormant and nondormant chlamydospores of A. flagrans fungi were preliminarily clarified, suggesting that the state of cell wall and intracellular materials were changed after spores entered to dormancy.
研究了捕食线虫真菌节丛孢（Arthrobotrys flagrans）不同类型厚垣孢子的形态和结构差异，为寄生线虫病的生物防治提供参考。本研究以节丛孢 F088 休眠厚垣孢子和非休眠厚垣孢子为研究对象，通过乳醇棉蓝、台盼蓝和 3-(4,5-二甲基噻唑-2-基)-2,5-二苯基四氮唑溴盐（MTT）染色，在光学显微镜下观察这些孢子的结构差异。利用 FunXite-1、4',6-二脒基-2-苯基吲哚（DAPI）和钙黄绿素白染色，在荧光显微镜下观察两种类型孢子的代谢活性、细胞壁和细胞核差异。采用扫描电子显微镜（SEM）和透射电子显微镜（TEM）观察两种厚垣孢子的超微结构。由于乳酚棉蓝、台盼蓝染色不能区分休眠孢子和死亡孢子，因此进行了 MTT 测定。荧光显微镜观察表明，非休眠孢子的细胞质代谢活性强于休眠孢子。休眠孢子的核呈亮蓝色，其荧光强度强于非休眠孢子。非休眠孢子的细胞壁产生比休眠孢子更强的黄绿光荧光。超微结构观察表明，两种类型的孢子表面都有球形突起，但无明显差异。休眠孢子的内壁具有一层厚的透明带，电子密度高，明显比非休眠孢子厚，其细胞质也发生了变化。本研究初步阐明了节丛孢休眠和非休眠厚垣孢子的微观结构特征，表明孢子进入休眠状态后，细胞壁和细胞内物质的状态发生了变化。
