基于细菌群体感应的抗生素联合毒性评估  

Evaluation of joint toxic action for antibiotics based on bacterial quorum sensing

作　　者：黄颂 张玉莲 姚静怡 戴佳敏 孙昊宇 刘树深[2] 唐量[1] HUANG Song;ZHANG YuLian;YAO JingYi;DAI JiaMin;SUN HaoYu;LIU ShuShen;TANG Liang(Key Laboratory of Organic Compound Pollution Control Engineering(MOE),School of Environmental and Chemical Engineering,Shanghai University,Shanghai 200444,China;College of Environmental Science and Engineering,Tongji University,Key Laboratory of Yangtze River Water Environment,Shanghai 200092,China)

机构地区：[1]上海大学环境与化学工程学院,有机复合污染控制工程教育部重点实验室,上海200444 [2]同济大学环境科学与工程学院,长江水环境教育部重点实验室,上海200092

出　　处：《中国科学：技术科学》2024年第10期1949-1965,共17页Scientia Sinica(Technologica)

基　　金：国家自然科学基金(批准号:42277270,42377359)资助项目。

摘　　要：抗生素作为一类新污染物能够对环境中以细菌为代表的微生物圈产生胁迫,影响生态系统的稳定性.环境中的抗生素往往以混合物形式存在,因此抗生素联合毒性的精准解析对于其生态风险评估具有重要意义.然而,目前缺乏从细菌群体行为入手对抗生素联合毒性进行深入分析的有关报道.本研究以磺胺甲氧嗪(sulfamethoxypyridazine,SMP)和四环素(tetracylinehydrochloride,TH)作为抗生素代表,选择风险评估中常用的费氏弧菌(Aliivibrio fischeri,A.fischeri)生物发光作为测试终点,利用微孔板法测定受试抗生素对生物发光在24 h内的毒性效应,基于独立作用模型评估SMP和TH的联合毒性作用,并考虑细菌在真实环境中可能遇到的群体感应(quorum sensing,QS)信号分子,利用交互效应评估信号分子与抗生素之间的联合作用.结果表明,SMP,TH及SMP&TH混合物均对A.fischeri生物发光产生了低浓度促进、高浓度抑制的毒物兴奋效应(hormesis),且该双相剂量-效应具有随时间变化特征.同时,SMP&TH的联合作用及其强度也表现出浓度和时间依赖特性.利用实时荧光定量多聚核苷酸链式反应技术、体外生物发光反应模拟实验和分子对接技术进行机制探究发现,SMP和TH分别作用于细菌QS系统中的信号通路蛋白和萤光素酶进而产生hormesis和联合作用的异质性模式,而暴露时间依赖特征则是由细菌QS活性在24 h内由无到弱再到强的变化规律所导致的.外源N-3-氧代己酰-DL-高丝氨酸内酯(N-(β-Ketocaproyl)-DL-homoserine lactone,通常以C6表示)信号分子的加入不仅改变了SMP,TH及SMP&TH混合物的毒性效应,还影响了SMP与TH之间的联合毒性作用.此外,SMP,TH与外源C6的交互效应以协同为主,而SMP&TH与外源C6的交互效应随暴露时间变化.本研究证明了细菌群体行为在调控抗生素单一及联合毒性中的关键作用,并阐释了环境系统中外界信号分子对抗生素诱导�Antibiotics,recognized as emergent pollutants,pose a significant stress on environmental microbiota,especially affecting bacterial communities and potentially disrupting ecosystem stability.Given that antibiotics usually exist as mixtures within natural settings,it is necessary to precisely evaluate their combined toxicity for comprehensive ecological risk assessments.Currently,in-depth analyses into the combined effects of antibiotics on bacterial group behavior remain scarce.This study focuses on sulfamethoxypyridazine(SMP) and tetracycline hydrochloride(TH) as representative antibiotics.We employ the bioluminescence of Aliivibrio fischeri(A.fischeri),a widely used biomarker for evaluating pollutant risk,as the test endpoint.Using the microplate method,we determine the toxic effects of test antibiotics on bioluminescence within 24 h.The analysis incorporates the independent action model to judge the joint toxic action between SMP and TH.Furthermore,we evaluated the interaction between antibiotics and quorum sensing(QS)signaling molecules,recognizing the ability of bacteria to sense and respond to the signaling molecules from real environments.Observations indicate that SMP,TH,and an SMP&TH mixture induce hormetic effects on A.fischeri bioluminescence,characterized by low-concentration stimulation and high-concentration inhibition.Moreover,these biphasic dose responses exhibited time dependency.The joint toxic actions between SMP and TH demonstrated concentration and time dependency.Using quantitative realtime PCR technology,in vitro bioluminescence reaction simulations,and molecular docking analyses,the study suggests that SMP and TH target key proteins in the signaling pathway and luciferase in the QS system of A.fischeri,contributing to the observed hormetic bioluminescence responses and the variable patterns of joint toxicity.The dynamics of QS activity,varying from weak to strong over 24 h,underline the time-dependent feature of hormesis.Furthermore,the introduction of the exogenous signaling molecule N-(β

关 键 词：混合抗生素 联合毒性 群体感应 信号分子 交互效应 生物发光 

分 类 号：X172[环境科学与工程—环境科学]