Zhao Feng, Wu Yuting, Liu Lu
School of Life Sciences, Qufu Normal University, Qufu, 273165, Shandong Province, China.
Microb Cell Fact. 2025 Jul 16;24(1):166. doi: 10.1186/s12934-025-02793-y.
Pseudomonas aeruginosa is a facultative anaerobic bacterium widely distributed in oil reservoirs. Efficient production of rhamnolipids by P. aeruginosa in anoxic environments of oil reservoirs contributes to oil displacement. How to enhance the synthesis of rhamnolipids by P. aeruginosa under anaerobic conditions has become a new scientific question.
Pseudomonas aeruginosa SG anaerobically synthesized much lower yield of rhamnolipids than that under aerobic conditions. The key genes rmlBDAC (m), rhlABRI (h) and rhlC (c) and their combination genes were overexpressed in P. aeruginosa. Among seven genetically engineered strains, P. aeruginosa SGhm anaerobically synthesized the highest yield of rhamnolipids, 1.34 g/L, which was about 4.5-fold higher than that of the wild-type strain SG (0.24 g/L). The results of Plackett-Burman (PB) design indicated that glycerol and nitrate were the nutrient factors that significantly influenced the anaerobic production of rhamnolipids by strain SGhm. Optimization of culture medium by response surface method improved the rhamnolipids production of strain SGhm to 1.54 g/L under anaerobic conditions. Pseudomonas aeruginosa SGhm emulsified oil in anaerobic tubes through anaerobic synthesis of rhamnolipids. The percentage of oil droplets formed with a diameter of 0-5 μm was 89.4%.
Genes rmlBDAC and rhlABRI were the best overexpression combination for improvement of rhamnolipids production in P. aeruginosa under anaerobic conditions. Overexpression of key genes and optimization of medium synergistically increase rhamnolipids production of P. aeruginosa under anaerobic conditions. Pseudomonas aeruginosa SGhm is promising for emulsification of crude oil through anaerobic synthesis of rhamnolipids. This study provided an excellent microbial strain resource and nutrients for in-situ production of rhamnolipids in anoxic environments of oil reservoirs.
铜绿假单胞菌是一种兼性厌氧菌，广泛分布于油藏中。铜绿假单胞菌在油藏缺氧环境中高效生产鼠李糖脂有助于驱油。如何在厌氧条件下提高铜绿假单胞菌鼠李糖脂的合成能力已成为一个新的科学问题。
铜绿假单胞菌SG在厌氧条件下合成的鼠李糖脂产量远低于好氧条件下的产量。关键基因rmlBDAC（m）、rhlABRI（h）和rhlC（c）及其组合基因在铜绿假单胞菌中过表达。在七个基因工程菌株中，铜绿假单胞菌SGhm在厌氧条件下合成的鼠李糖脂产量最高，为1.34 g/L，约为野生型菌株SG（0.24 g/L）的4.5倍。Plackett-Burman（PB）设计结果表明，甘油和硝酸盐是显著影响菌株SGhm厌氧生产鼠李糖脂的营养因素。通过响应面法优化培养基，在厌氧条件下将菌株SGhm的鼠李糖脂产量提高到1.54 g/L。铜绿假单胞菌SGhm通过厌氧合成鼠李糖脂在厌氧管中乳化油。直径为0-5μm的油滴形成百分比为89.4%。
基因rmlBDAC和rhlABRI是在厌氧条件下提高铜绿假单胞菌鼠李糖脂产量的最佳过表达组合。关键基因的过表达和培养基的优化协同提高了铜绿假单胞菌在厌氧条件下鼠李糖脂的产量。铜绿假单胞菌SGhm有望通过厌氧合成鼠李糖脂乳化原油。本研究为油藏缺氧环境中鼠李糖脂的原位生产提供了优良的微生物菌株资源和营养物质。
