Data_Sheet_1_Assessment of the potential risks in SD rats gavaged with genetically modified yeast containing the cp4-epsps gene.docx

IntroductionDespite the absence of definitive evidence indicating that the cp4-epsps gene and its resultant recombinant proteins have significant harmful effects on either human or animal health, the safety assessment of genetically modified (GM) crops expressing the CP4-EPSPS proteins has been controversial. This study endeavor was aimed at evaluating the potential risks posed by the CP4-EPSPS protein in transgenic crops, thereby contributing to the advancement of risk assessment methodologies in the context of genetically engineered crops.
MethodsTo ascertain the appropriate daily dosages for oral gavage administration, the expression levels of the CP4-EPSPS protein in a recombinant yeast were quantified. Subsequently, physiological and biochemical analysis, metabolomics, and metagenomic analysis were conducted based on a 90-day Sprague-Dawley (SD) rats feeding experiment, respectively, thereby enhancing the depth and precision of our risk assessment framework.
ResultsThe results from the physiological and biochemical analysis, organ pathological, blood metabolism, gut microbiota, and correlation analysis of metabolites and gut microbiota revealed several biomarkers for further risk assessment. These biomarkers include clinical biochemical indexes such as total bilirubin (TBIL), direct bilirubin (DBIL), creatine kinase (CK), and lactate dehydrogenase (LDH); metabolites like Methionine, 2-Oxovaleric acid, and LysoPC (16:0); and gut microbiota including Blautia wexlerae, Holdemanella biformis, Dorea sp. CAG 317, Coriobacteriaceae and Erysipelotrichaceae.
ConclusionIn conclusion, the risk can be significantly reduced by directly consuming inactivated recombinant CP4-EPSPS. Therefore, in everyday life, the risk associated with consuming GM foods containing recombinant CP4-EPSPS is substantially reduced after heat treatment.

引言：尽管目前尚无确凿证据表明cp4-epsps基因及其表达的重组蛋白对人类或动物健康存在显著危害，但表达CP4-EPSPS蛋白的转基因（GM）作物的安全性评估仍存在争议。本研究旨在评估转基因作物中CP4-EPSPS蛋白的潜在风险，以期推动转基因作物风险评估方法学的发展。
材料与方法：为确定经口灌胃给药的适宜日剂量，本研究首先定量检测了重组酵母中CP4-EPSPS蛋白的表达水平。随后基于90天Sprague-Dawley（SD）大鼠喂养实验，分别开展生理生化分析、代谢组学及宏基因组学分析，以深化风险评估框架的深度与精准度。
结果：生理生化分析、器官病理学检测、血液代谢组分析、肠道菌群分析以及代谢物与肠道菌群相关性分析的结果，筛选出若干可用于后续风险评估的生物标志物。这些生物标志物包括：临床生化指标，如总胆红素（TBIL）、直接胆红素（DBIL）、肌酸激酶（CK）及乳酸脱氢酶（LDH）；代谢物，如甲硫氨酸、2-氧代戊酸及溶血磷脂酰胆碱（LysoPC，16:0）；以及肠道菌群，包括韦氏布劳特氏菌（Blautia wexlerae）、两形霍尔德曼氏菌（Holdemanella biformis）、Dorea sp. CAG 317、棒杆菌科（Coriobacteriaceae）及红蝽杆菌科（Erysipelotrichaceae）。
结论：综上，直接食用灭活的重组CP4-EPSPS蛋白可显著降低相关风险。因此，在日常生活中，经热处理后食用含有重组CP4-EPSPS的转基因食品，其相关风险将大幅降低。