基于代谢组学指标的土壤亚致死剂量汞对蚯蚓的毒性研究

The toxicity of Hg in soil with sublethal doses to earthworms based on the metabolomics

将赤子爱胜蚓(Eisenia fetida)暴露于亚致死剂量(1.0、5.0、25.0 mg·kg^(-1))汞污染土壤中4周,以蚯蚓个体(致死率、体重增长率)及小分子代谢物(代谢组)为指标研究其对汞的动态毒性响应,并采用最小二乘判别分析(OPLSA-DA)对暴露组及对照组的代谢物进行分类,进而识别潜在的标记物.结果表明,蚯蚓对汞的吸收尚未达到稳态,蚯蚓体内代谢物的响应依赖于暴露剂量及暴露时间.暴露1周时,蚯蚓体重略有增长但不显著;最低暴露剂量(1.0 mg·kg^(-1))导致蚯蚓体内亮氨酸、异亮氨酸、色氨酸、组氨酸、酪氨酸、5-氧脯氨酸、2-脱氧肌苷显著低于对照水平,柠檬酸、肌苷酸与腺苷在5.0、25.0 mg·kg^(-1)剂量下显著高于对照水平;暴露4周时,最高暴露剂量(25 mg·kg^(-1))显著抑制了蚯蚓的生长;汞添加组的蚯蚓体内谷氨酸、酪氨酸、马来酸、2-脱氧肌苷水平显著低于对照.上述代谢物对汞的动态变化表明它们可作为潜在生物标记物,用于诊断土壤汞污染.对代谢途径分析发现,1.0～25.0 mg·kg^(-1)汞即可破坏蚯蚓正常氨基酸代谢、三羧酸循环,扰乱能量代谢,对蚯蚓产生氧化损伤.本研究结果表明,相比个体水平的受试终点,代谢组学指标比个体水平指标能更敏感地响应较低剂量汞,是土壤汞污染生态毒性效应诊断的有效指标.另外,本研究结果可为土壤汞污染的风险评估及相关环境标准的修订提供大量基础数据.

In this study,earthworms(Eisenia fetida)were exposed to soil with sublethal doses of Hg(with concentration of 1.0,5.0 and 25.0 mg·kg-1 soil)for 4 weeks.Dynamic responses of the individuals(mortality and weight growth rate)and the small molecular metabolites(the metabolomics)were studied.The supervised orthogonal partial lease squares discriminant method(OPLS-DA)was used to analyze the metabolites in the exposed groups and the control group,by which the potential biomarkers can be identified.Experimental results indicate that Hg accumulation in earthworms has not reached a steady state for 4 weeks exposure,and the responses of the metabolites depend on the exposure dose and the exposure time.After 1 week,the weight of earthworms increased insignificantly.For the lowest exposure dose(1.0 mg·kg-1),the concentration of leucine,isoleucine,tryptophan,histidine,tyrosine,5-oxyproline and 2-deoxyinosine in the exposure group were significantly lower than those in the control group.While,for 5.0,25.0 mg·kg-1 dose,the concentration of citrate,inosine and adenosine in the exposure group were significantly higher than those in the control group.After 4 weeks,the worms'growth was significantly inhibited at the highest exposure dose(25.0 mg·kg-1).In addition,the concentration of glutamic acid,tyrosine,maleic acid and 2-deoxyinosine in the exposure group were significantly lower than those of the control.The dynamic responses of above metabolites suggest that they can be used as potential biomarkers for the diagnosis of Hg-contaminated soil.Analysis on metabolic pathways revealed that 1.0~25.0 mg·kg-1 Hg could disrupt the normal amino acid metabolism,the tricarboxylic acid cycle and the energy metabolism,thus leading to oxidative damage to earthworms.The results of this study suggest that metabolomics are more sensitive and effective than individual endpoints in response to lower-dose Hg.In addition,our results provide a large amount of basic data for the risk assessment of Hg-contaminated soil and the revision of environmental standards.