State Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural University. Wuhan, Hubei, 430070, China.
State Key Laboratory of Agricultural Microbiology, College of Veterinary Medicine, Huazhong Agricultural University. Wuhan, Hubei, 430070, China; Key Laboratory of Preventive Veterinary Medicine in Hubei Province, The Cooperative Innovation Center for Sustainable Pig Production, Wuhan, Hubei, 430070, China; Key Laboratory of Development of Veterinary Diagnostic Products, Ministry of Agriculture of the People's Republic of China, Wuhan, Hubei, 430070, China.
Microb Pathog. 2019 Nov;136:103706. doi: 10.1016/j.micpath.2019.103706. Epub 2019 Sep 3.
Riemerellosis, a Riemerella anatipestifer infection, can cause meningitis, pericarditis, parahepatitis, and airsacculitis in ducks, leading to serious economic losses in the duck meat industry. However, the molecular mechanism of the pathogenesis and virulence factors of this infection are poorly understood. In the present study, we created a mutant strain RA-YMΔCas9 using trans-conjugation. Bacterial virulence tests indicated that the median lethal dose (LD) of RA-YMΔCas9 was 5.01 × 10 CFU, significantly lower than that of the RA-YM strain, which was 1.58 × 10 CFU. The distribution and blood bacterial load from the infection groups showed no significant difference in the brain between the RA-YMΔCas9 mutant and the wild-type RA-YM strains, however, the number of mutant strains were significantly reduced in the liver, heart, and blood. Animal immunization experiments demonstrated that the intranasal administration of RA-YMΔCas9 in ducklings provided 80% protection after challenge with the wild-type strain, showing potential use as a live mucosal vaccine. RNAseq analysis indicated that Cas9 protein played a regulatory role in gene expression. This study is the first to report on the involvement of Cas9 in the regulation and pathogenesis of R. anatipestifer, and provides a theoretical basis for the development of relevant genetic engineering vaccines.
里默氏菌病是由鸭疫里默氏杆菌引起的一种疾病，可导致鸭的脑膜炎、心包炎、肝炎和气囊炎，给鸭肉产业造成严重的经济损失。然而，这种感染的发病机制和毒力因子的分子机制还了解甚少。在本研究中，我们通过转导法创建了一个突变株 RA-YMΔCas9。细菌毒力试验表明，RA-YMΔCas9 的半数致死剂量 (LD) 为 5.01×10 CFU，明显低于 RA-YM 株的 1.58×10 CFU。感染组的分布和血液细菌载量显示，突变株在脑中与野生型 RA-YM 株无明显差异，但在肝脏、心脏和血液中的数量明显减少。动物免疫实验表明，RA-YMΔCas9 滴鼻免疫雏鸭后，对野生型菌株的攻毒保护率达到 80%，显示出作为活黏膜疫苗的潜在用途。RNAseq 分析表明 Cas9 蛋白在基因表达调控中发挥作用。本研究首次报道了 Cas9 参与鸭疫里默氏杆菌的调控和发病机制，为相关基因工程疫苗的开发提供了理论依据。
