Zou Ronghua, Wu Xiaoni, Chen Qiwei, Gong Xiaowei, Chu Yuefeng, Zheng Fuying
State Key Laboratory of Veterinary Etiological Biology, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, No. 1 Xujiaping, Yanchangbao, Lanzhou 730046, China.
State Key Laboratory of Veterinary Etiological Biology, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, No. 1 Xujiaping, Yanchangbao, Lanzhou 730046, China.
Vet Microbiol. 2023 May;280:109692. doi: 10.1016/j.vetmic.2023.109692. Epub 2023 Feb 16.
Bacterial meningitis is a major cause of morbidity and mortality. Despite advances in antimicrobial chemotherapy, the disease remains detrimental to humans, livestock, and poultry. Riemerella anatipestifer is a gram-negative bacterium causing duckling serositis and meningitis. However, the virulence factors contributing to its binding and invasion of duck brain microvascular endothelial cells (DBMECs) and penetration of the blood-brain barrier (BBB) have never been reported. In this study, immortalized DBMECs were successfully generated and used as an in vitro-model of duck BBB. Furthermore, ompA gene deletion mutant of the pathogen and multiple complemented strains carrying the complete ompA gene and its truncated forms were constructed. Bacterial growth, invasion, and adhesion assays and animal experiments were performed. The results show that the OmpA protein of R. anatipestifer had no effect on bacterial growth and adhesion ability to DBMECs. The role of OmpA in the invasion of R. anatipestifer into DBMECs and duckling BBB was confirmed. The amino acids 230-242 of OmpA represents a key domain involved in R. anatipestifer invasion. In addition, another OmpA1164 protein constituted by the amino acids 102-488 within OmpA could function as a complete OmpA. The signal peptide sequence from amino acids 1-21 had no significant effect on OmpA functions. In conclusion, this study illustrated that OmpA is an important virulence factor mediating R. anatipestifer invasion of DBMECs and penetration of the duckling BBB.
细菌性脑膜炎是发病和死亡的主要原因。尽管抗微生物化疗取得了进展，但该疾病对人类、家畜和家禽仍然有害。鸭疫里默氏杆菌是一种革兰氏阴性菌，可引起雏鸭浆膜炎和脑膜炎。然而，其导致与鸭脑微血管内皮细胞（DBMECs）结合和侵入以及穿越血脑屏障（BBB）的毒力因子从未被报道过。在本研究中，成功构建了永生化的DBMECs并将其用作鸭血脑屏障的体外模型。此外，构建了该病原体的ompA基因缺失突变体以及携带完整ompA基因及其截短形式的多个互补菌株。进行了细菌生长、侵入和黏附试验以及动物实验。结果表明，鸭疫里默氏杆菌的OmpA蛋白对细菌生长以及与DBMECs的黏附能力没有影响。证实了OmpA在鸭疫里默氏杆菌侵入DBMECs和雏鸭血脑屏障中的作用。OmpA的第230 - 242位氨基酸代表了参与鸭疫里默氏杆菌侵入的关键结构域。此外，由OmpA内第102 - 488位氨基酸构成的另一种OmpA1164蛋白可发挥完整OmpA的功能。第1 - 21位氨基酸的信号肽序列对OmpA功能没有显著影响。总之，本研究表明OmpA是介导鸭疫里默氏杆菌侵入DBMECs和穿越雏鸭血脑屏障的重要毒力因子。
