高盐胁迫条件下嗜盐四联球菌生理应答及盐胁迫抗性机制解析

Salt stress is an environmental condition commonly encountered during the fermentation of high-salt foods, which seriously impair the efficiency of food manufacture. In this study, the differences in intracellular micro-environment (activities of key enzymes, intracellular pH, ATP, NAD+/NADH), intracellular metabolites, cell membrane characterization (distribution of fatty acids, membrane fluidity), genes transcription, and proteins expression were investigated based on physiological and omics analysis by using Tetragenococcus halophilus as an initial strain. The physiological responses and regulatory mechanisms of T. halophilu during salt stress were revealed, and based on this, the directed strategies to improve salt tolerance were proposed. The key gene, which was involved in salt tolerance was introduced into the host strain. The mutant, which exhibited high resistance to salt stress was constructed and the tolerance to salt stress was validated. Results presented in this study may contribute to the understanding of salt tolerance mechanisms employed by T. halophilus and provide theoretical basis for the construction of stress-resistant lactic acid bacteria.

盐胁迫是高盐食品发酵过程中广泛存在的一种环境胁迫，严重影响食品微生物制造的效率。本课题以嗜盐四联球菌CGMCC3792为研究对象，从细胞生理特性和组学分析的角度考察细胞在不同盐浓度胁迫条件下胞内微环境(关键酶活性、胞内pH、ATP、NAD+/NADH)、物质代谢、细胞膜生理特性(膜脂肪酸组成、膜流动性)、基因转录水平和蛋白质表达水平上的差异，全面系统地阐释嗜盐四联球菌盐胁迫条件下的生理应答和调控机制，并在此基础上提出定向调控细胞盐胁迫抗性的策略，通过分子生物学手段，导入调控细胞盐胁迫抗性的关键基因，构建具有高盐胁迫抗性的工程菌，并对其耐盐性能进行验证。研究结果将有助于深入理解嗜盐四联球菌耐盐生理机制，并为构建高胁迫抗性的乳酸菌奠定理论基础。

以嗜盐四联球菌为研究对象，选取发酵食品生产中广泛存在的盐胁迫为主要胁迫条件，利用微生物生理学与系统生物学技术考察了细胞在盐胁迫条件下生理特性（胞内微环境、细胞膜水平）、基因转录、蛋白表达以及代谢物水平上的差异，探求嗜盐四联球菌抵御盐胁迫的生理机制。在此基础上，通过分析组学数据以及生理学的研究结果，提出了基于生化工程策略结合发酵优化技术提升细胞耐盐性能，实现了嗜盐四联球菌在高盐胁迫条件下的高密度发酵。本项目的研究结果有助于进一步深入了解嗜盐四联球菌在盐胁迫条件下的生理状态及其抵御盐胁迫的生理应答机制，同时为进一步提高其盐胁迫抗性奠定了理论基础。项目执行期间发表英文论文5篇，其中SCI论文4篇，中文核心期刊4篇，申请国家发明专利4项。