College of Veterinary Medicine, Nanjing Agricultural University, Nanjing, China Key Lab of Animal Bacteriology, Ministry of Agriculture, Nanjing, China.
College of Veterinary Medicine, Nanjing Agricultural University, Nanjing, China Key Lab of Animal Bacteriology, Ministry of Agriculture, Nanjing, China
Infect Immun. 2014 Sep;82(9):3867-79. doi: 10.1128/IAI.01769-14. Epub 2014 Jun 30.
Type VI secretion systems (T6SSs) are involved in the pathogenicity of several Gram-negative bacteria. The VgrG protein, a core component and effector of T6SS, has been demonstrated to perform diverse functions. The N-terminal domain of VgrG protein is a homologue of tail fiber protein gp27 of phage T4, which performs a receptor binding function and determines the host specificity. Based on sequence analysis, we found that two putative T6SS loci exist in the genome of the avian pathogenic Escherichia coli (APEC) strain TW-XM. To assess the contribution of these two T6SSs to TW-XM pathogenesis, the crucial clpV clusters of these two T6SS loci and their vgrG genes were deleted to generate a series of mutants. Consequently, T6SS1-associated mutants presented diminished adherence to and invasion of several host cell lines cultured in vitro, decreased pathogenicity in duck and mouse infection models in vivo, and decreased biofilm formation and bacterial competitive advantage. In contrast, T6SS2-associated mutants presented a significant decrease only in the adherence to and invasion of mouse brain microvascular endothelial cell (BMEC) line bEnd.3 and brain tissue of the duck infection model. These results suggested that T6SS1 was involved in the proliferation of APEC in systemic infection, whereas VgrG-T6SS2 was responsible only for cerebral infection. Further study demonstrated that VgrG-T6SS2 was able to bind to the surface of bEnd.3 cells, whereas it did not bind to DF-1 (chicken embryo fibroblast) cells, which further proved the interaction of VgrG-T6SS2 with the surface of BMECs.
VI 型分泌系统（T6SS）参与了几种革兰氏阴性菌的致病性。VgrG 蛋白是 T6SS 的核心成分和效应物，已被证明具有多种功能。VgrG 蛋白的 N 端结构域是噬菌体 T4 的尾纤维蛋白 gp27 的同源物，它具有受体结合功能，并决定宿主特异性。基于序列分析，我们在禽致病性大肠杆菌（APEC）TW-XM 菌株的基因组中发现了两个假定的 T6SS 基因座。为了评估这两个 T6SS 对 TW-XM 发病机制的贡献，我们敲除了这两个 T6SS 基因座的关键 clpV 簇及其 vgrG 基因，以生成一系列突变体。结果，T6SS1 相关突变体在体外培养的几种宿主细胞系中的粘附和侵袭能力降低，在鸭和小鼠感染模型中的致病性降低，生物膜形成和细菌竞争优势降低。相比之下，T6SS2 相关突变体仅在小鼠脑微血管内皮细胞（BMEC）系 bEnd.3 和鸭感染模型脑组织中的粘附和侵袭能力显著降低。这些结果表明 T6SS1 参与了 APEC 在全身感染中的增殖，而 VgrG-T6SS2 仅负责大脑感染。进一步的研究表明，VgrG-T6SS2 能够与 bEnd.3 细胞的表面结合，而不能与 DF-1（鸡胚成纤维细胞）细胞结合，这进一步证明了 VgrG-T6SS2 与 BMEC 表面的相互作用。
