All experimental data from this study.

Microsporidia are fungal obligate intracellular pathogens, which infect most animals and cause microsporidiosis. Despite the serious threat that microsporidia pose to humans and agricultural animals, few drugs are available for the treatment and control of microsporidia. To identify novel inhibitors, we took advantage of the model organism Caenorhabditis elegans infected with its natural microsporidian Nematocida parisii. We used this system to screen the Pandemic Response Box, a collection of 400 diverse compounds with known antimicrobial activity. After testing these compounds in a 96-well format at high (100 μM) and low (40 μM) concentrations, we identified four inhibitors that restored the ability of C. elegans to produce progeny in the presence of N. parisii. All four compounds reduced the pathogen load of both N. parisii and Pancytospora epiphaga, a C. elegans-infecting microsporidia related to human-infecting species. One of these compounds, a known inhibitor of a viral protease, MMV1006203, inhibited invasion and prevented the firing of spores. A bis-indole derivative, MMV1593539, decreased spore viability. An albendazole analog, MMV1782387, inhibited proliferation of N. parisii. We tested albendazole as well as 5 other analogs and observed that MMV1782387 was amongst the strongest inhibitors of N. parisii and displayed the least host toxicity. Our study further demonstrates the effectiveness of the C. elegans-N. parisii system for discovering microsporidia inhibitors and the compounds we identified provide potential scaffolds for anti-microsporidia drug development.

微孢子虫（Microsporidia）是一类真菌专性细胞内病原体，可感染绝大多数动物并引发微孢子虫病（microsporidiosis）。尽管微孢子虫对人类与畜牧动物构成严重威胁，但目前可用的治疗与防控药物寥寥无几。为筛选新型抑制剂，本研究利用感染天然宿主巴黎线虫孢虫（Nematocida parisii）的模式生物秀丽隐杆线虫（Caenorhabditis elegans）作为研究体系，对大流行响应化合物库（Pandemic Response Box）进行筛选——该库包含400种具有已知抗菌活性的多样化化合物。我们采用96孔板体系，分别以高浓度（100 μM）与低浓度（40 μM）对这些化合物开展测试，最终筛选得到4种可恢复秀丽隐杆线虫在巴黎线虫孢虫感染条件下产生子代能力的抑制剂。这4种化合物均可降低巴黎线虫孢虫与嗜胞质微孢子虫（Pancytospora epiphaga）的病原体载量，其中嗜胞质微孢子虫是一种与感染人类的微孢子虫亲缘关系较近的秀丽隐杆线虫感染性微孢子虫。其中一种化合物为已知的病毒蛋白酶抑制剂MMV1006203，可抑制病原体入侵并阻止孢子弹出；双吲哚衍生物MMV1593539可降低孢子活力；阿苯达唑类似物MMV1782387则可抑制巴黎线虫孢虫的增殖。我们对阿苯达唑及其余5种类似物进行了测试，结果显示MMV1782387是对巴黎线虫孢虫抑制活性最强的化合物之一，且宿主毒性最低。本研究进一步验证了秀丽隐杆线虫-巴黎线虫孢虫研究体系在筛选微孢子虫抑制剂方面的有效性，本次鉴定得到的化合物可为抗微孢子虫药物开发提供潜在的分子骨架。