肠炎沙门菌 Z11中 iscA 基因的敲除及功能

Knockout and functional characterization of iscA in Salmonella Enteritidis Z11

摘要:

肠炎沙门菌( Salmonella Enteritidis)是一种重要的食源性病原菌，可引起人和动物的胃肠道感染。 iscA是编码铁-硫(Fe-S)簇组装的关键基因之一，参与铁离子的转运和能量代谢，是一种保守的A型铁结合蛋白。 目的 通过构建肠炎沙门菌Z11 Δ iscA突变株，探究IscA在沙门菌感染过程中的作用。 方法 以实验室保存的肠炎沙门菌Z11菌株为研究对象，采用无标记框内缺失突变法构建 iscA突变株。分析并比较野生株与突变株在运动性、生物被膜形成能力等方面的差异；分别在小鼠单核巨噬细胞RAW264.7和小鼠模型上探究IscA对肠炎沙门菌毒力的影响。 结果 本研究成功构建了Δ iscA缺失株；与野生株相比，Δ iscA在生长能力和生物被膜形成能力方面无明显差异，表明缺失 iscA基因不会影响肠炎沙门菌的正常生长和生物被膜形成；在运动性方面，Δ iscA在6 h时的菌圈直径显著小于野生株，表明 iscA基因的缺失降低了肠炎沙门菌Z11的游动能力。在RAW264.7细胞实验中，与野生株相比，Δ iscA缺失株的黏附和侵袭能力均显著降低，分别下降至约37%和20%；并且，Δ iscA缺失株突破细胞屏障进入细胞内的增殖速率也显著低于野生株。小鼠感染实验结果显示，Δ iscA缺失株在空肠和盲肠中的定殖能力显著低于野生株感染组。 结论 基因 iscA与肠炎沙门菌的毒力密切相关，其缺失会影响该菌株的运动性、黏附侵袭能力、胞内增殖能力，并且降低其在宿主肠道中的定殖能力，从而进一步影响肠炎沙门菌的感染过程。

关键词:肠炎沙门菌;iscA基因;运动性：定殖

Abstract:

Salmonella Enteritidis is a major foodborne pathogen that can cause gastrointestinal infections in both humans and animals. As one of the key genes encoding the iron-sulfur cluster assembly, iscA plays a role in the transport of iron ions and energy metabolism. IscA is a conserved A-type iron-binding protein. Objective To study the role of iscA in the infection process of Salmonella by constructing an iscA-deleted mutant (Δ iscA) of Salmonella Enteritidis Z11. Methods The unmarked in-frame gene deletion method was employed to construct Δ iscA from the laboratory-preserved Salmonella Enteritidis Z11 strain. The wild type (WT) and Δ iscA were compared in terms of motility and biofilm formation. Additionally, the impact of IscA on the virulence of Salmonella Enteritidis was explored in both RAW264.7 cells and a mouse model. Results The deletion mutant Δ iscA was successfully constructed. No significant difference in the growth or biofilm formation was observed between Δ iscA and WT, indicating that the deletion of iscA did not affect the normal growth or biofilm formation of Salmonella Enteritidis. However, Δ iscA exhibited a significantly smaller zone of motility than WT at the time point of 6 h, suggesting that the loss of iscA reduced the motility of Salmonella Enteritidis Z11. In RAW264.7 cells, the adhesion and invasion of Δ iscA significantly decreased to 37% and 20%, respectively, of those of WT. Furthermore, the proliferation rate of Δ iscA in the cells was significantly lower than that of WT. Mouse infection experiments revealed that Δ iscA demonstrated reduced colonization in the jejunum and cecum compared with WT. Conclusion iscA is closely associated with the virulence of Salmonella Enteritidis. Its deletion affects the motility, adhesion, invasion, and proliferation, ultimately reducing the colonization in the host intestine and influencing the infection process of Salmonella Enteritidis.

Key words:Salmonella Enteritidis;iscA;motility;colonization