生物硝化池污水中硝化细菌的快速定量研究

实验采用聚合酶链式反应（PCR）技术与最大几率数法（MPN）相结合的MPN－PCR法对生物硝化池污水中的硝化细菌进行快速定量。所用的一对PCR引物是在对硝化细菌的16SrRNA基因进行系统比较的基础上设计合成的，可以扩增出大小为388bp的DNA片段。以从生物硝化池污水中抽提的含硝化细菌DNA的混合DNA为模板，进行PCR扩增并确定合适的扩增条件。运用MPN－PCR法进行定量检测的整个过程可在几小时之内完成。     

彗星试验检测区域水源有机污染致突变量(Study on Mutagenic Threshold of Organic Pollutants in Typical Water Area by Comet Assay)

采用彗星试验检测了区域水源有机污染对人血淋巴细胞的致突变性,探讨了水源风险的评价方法.结果表明:Cr(VI)对人外周血淋巴细胞有强的致突变作用,并具有明显的剂量-效应关系;Cr(VI)剂量的对数与人外周血淋巴细胞DNA损伤的程度呈线性相关.可以用Cr(VI)作为阳性参照物量化表征水中有机污染混合物对人血淋巴细胞的致突变性.现行《地表水环境质量标准》、《污水综合排放标准》中Cr(VI)的限值可以用于评价水体中有机污染对人体健康的风险.     

彗星试验技术在水生与土壤环境中的毒理学应用

彗星试验是一种检测真核细胞脱氧核糖核酸(DNA)损伤的技术,快速、低成本、适用于几乎所有真核细胞,因此常应用于遗传毒理学、辐射生物学、肿瘤细胞学、毒性检测等方面。尤其以灵敏度高、检测联合毒性的特点大量运用于环境研究中。主要介绍彗星试验方法学发展历程,彗星试验的原理及其在环境领域中的应用,并指出应用中的受限因素以及未来的发展前景,为今后进一步推广该试验技术提供理论上的指导与参考。     

乌鲁木齐大气PM2.5对质粒DNA的损伤研究

2012年1月~2012年12月采集乌鲁木齐大气PM2.5样品,使用质粒DNA评价法研究了不同季节PM2.5的氧化能力,并进行氧化性毒性与相应气象因素和质量浓度之间的相关性研究.结果表明,乌鲁木齐大气PM2.5的质量浓度具有冬季最高,春季和秋季次之,夏季最低的季节性变化特征;PM2.5全样和水溶部分氧化能力的季节差异较大,对质粒DNA的氧化性损伤具有冬季最大,春季和夏季之次,秋季最低.冬、春、夏、秋季大气PM2.5全样的TD30(PM2.5对质粒DNA造成破坏达到30%所需要的颗粒物的剂量)平均值分别为440,491,503,515μg/mL,水溶部分分别为474,721,666,600μg/mL.绝大部分PM2.5样品全样的TD30值均小于水溶部分样,表明全样的毒性大于相应的水溶部分样.全样TD30值与平均温度显著(P<0.05)正相关,表明寒冷的天气/季节可能造成PM2.5的高毒性.水溶样TD30值与风速显著(P<0.01)正相关,与相对湿度显著负相关.这表明,高的风速和低的相对湿度可能跟较低和较高的PM2.5的毒性有关.PM2.5氧化性损伤能力的大小与其质量浓度之间的相关性不明显,表明仅以颗粒物的质量浓度来评价大气颗粒物氧化性损伤能力大小的方法并不能真实地反映其对人体健康的危害程度,起决定作用的还是颗粒物的化学组成及其表面吸附的有害成分.     

彗星实验检测渤海主要入海河流遗传毒性

利用彗星实验检测渤海区主要入海河流遗传毒性.以虾虎鱼为受试生物,暂养在河口水样中,染毒48h,取外周血细胞,运用彗星实验检测外周血细胞内DNA损伤程度,以尾相(TM)作为DNA损伤程度指标,并据此评估入海河流邻近海域遗传毒性风险.实验结果表明,入海河流中的特征污染物可导致虾虎鱼外周血细胞的DNA损伤,且损伤程度可以通过彗星实验定量分析,同时该试验方法操作简便、快速、灵敏度高,能够反映出多种污染因子的综合致毒能力.因此,通过彗星实验建立实验室检测入海河流遗传毒性方法具有可行行和创新性.     

Genotoxic effects of metals on human salivary gland tissue and lymphocytes as detected by the Comet assay

The etiology of salivary gland malignancies still remains unclear. Metal compounds are of special interest since they show ubiquitous presence in the environment, are present in many working places, and are accepted (co-)carcinogens in some other malignancies. Metals enter the body as xenobiotics by inhalation or ingestion. This study investigated the genotoxic potential of sodium dichromate (Na₂Cr₂O₇), nickel sulfate (NiSO₄), cadmium sulfate (CdSO₄) and zinc chloride (ZnCl₂) on human salivary gland cells and lymphocytes. Macroscopically healthy tissue of salivary glands was harvested from 46 patients during surgery and isolated to single cells by enzymatic digestion. The cells were incubated with Na₂Cr₂O₇, NiSO₄, CdSO₄ or ZnCl₂. Na₂Cr₂O₇ was also incubated in combination with the other metal compounds listed. Carcinogenic and co-carcinogenic effects of cadmium were tested by incubation with Na₂Cr₂O₇ and consecutive repair intervals. DNA damage and repair were evaluated by the Comet assay, determining DNA-strand breaks. The extent of damage was quantified using a digital analysis system. Na₂Cr₂O₇ produced significantly enhanced DNA-strand breaks in human salivary gland tissue and lymphocytes. All other metal compounds exerted no damaging effect on both cell types. Co-incubation of Na₂Cr₂O₇ with the other metals revealed a significant additive effect only for CdSO₄. Specific analysis of the influence of cadmium showed a reduction of DNA-repair after Na₂Cr₂O₇-induced strand breaks in salivary gland cells. This study provides evidence that exposure to distinct metals may significantly contribute to malignant salivary gland tumors. In consequence, further studies as epidemiological and toxicological data are warranted to determine the role of distinct metals as potential (co-) carcinogens.     

Mutagenicity of polyaromatic hydrocarbons by chemical models for cytochrome P450 in Ames assay

DNA damage is an important step in carcinogenesis. The Ames assay is a short-term screening of carcinogens that induce DNA damage. Most carcinogens require enzymatic activation through oxidation by cytochrome P450 (CYP450) in the presence of S9 mix. A combination of iron (Fe)(III) porphyrin and an oxidant is also able to oxidize compounds as an alternative metabolic pathway to CYP450. Previously it was reported that a chemical model containing a water-soluble 5,10,15,20-tetrakis(1-methylpyridinium4-yl)porphyrinatoiron(III) chloride (4-MPy) and tert-butyl hydroperoxide (t-BuOOH) activated aromatic amines and amides. In this study, a chemical model composed of an Fe porphyrin, water-insoluble 5,10,15,20-tetrakis(pentafluorophenyl)porphyrinatoiron(III) chloride (F₅P) or water-soluble 4-MPy was optimized with an oxidant – t-BuOOH, magnesium monoperoxyphthalate (MPPT), or iodosylbenzene (PhIO). Subsequently the mutagenicity of benzo[a]pyrene (B[a]P) and chrysene in Salmonella typhimurium TA strains was compared. B[a]P was activated by a combination of F₅P or 4-MPy plus MPPT or PhIO in S. typhimurium TA1538. The B[a]P-induced mutagenicity with F₅P plus oxidant was higher than 4-MPy plus oxidant. Mutagenicity of chrysene, a tetracyclic aromatic hydrocarbon, was not detected in the presence of F₅P/PhIO in S. typhimurium TA98, but was activated in the presence of F₅P/MPPT. The F₅P/MPPT activated other polyaromatic hydrocarbons (PAH) in the S. typhimurium TA98 assay including dibenz[a,c]anthracene, dibenz[a,h]anthracene, 3-methylcholanthrene, and benzo[a]anthracene. The results indicated that the F₅P/MPPT was the most efficient model for detecting PAH-induced mutagenicity in the Ames assay.     

Protective effect of Kappaphycus alvarezii (Rhodophyta) extract against DNA damage induced by mercury chloride in marine fish

Marine organisms are continuously exposed to agents, both exogenous and endogenous, that damage DNA. Consequently, it is important to determine the ability of compounds to provide protection against damaging chemicals. The aim of this study was to evaluate the anti-genotoxic activity of crude aqueous extracts of Kappaphycus alvarezii (Rhodophyceae), collected from the Southeast coast of India. This study focused on possible anti-genotoxic potential of aqueous extract of K. alverazii to interfere with clastogenicity induced by mercury chloride (HgCl₂) in marine fish, Therapon jarbua as measured by cytogenetic endpoints such as cell viability and comet assay. In the first set of experiments, fish were exposed to a single treatment of Hg at 0.125, 0.25, 0.5, 1, or 2?ppm along with controls. Mercury exposure produced significant DNA damage in all comet classes, maximum as >79% (Class 4) at 0.5, 1, and 2?ppm exposure in a time dependent manner. Algal extract did not induce genotoxicity when given alone and prevented Hg-induced genotoxicity. The algal extract reduction in genotoxicity was significant but not time- and concentration-dependent. Results suggested that under present experimental conditions, K. alvarezii extract exhibit potent anti-genotoxicity effects in this fish model; and thus these extracts may be recommended as a supplement in fish meal and may benefit humans ingesting Hg-contaminated fish.     

Genotoxic effects of two different classes of insecticide in developing chick embryos

Pesticides provide considerable protection against pest population; however, rampant accumulation of these chemicals into varied habitats across the globe necessitates the need for a careful screening of each chemical due to toxic manifestations. In the current study, the genotoxic potential of two different classes of commercial insecticides – chlorpyrifos and cypermethrin combination and Spinosad, a naturalyte were compared. Rhode Island Red chick embryos were exposed to different doses of either of these insecticides individually, by in ovo treatment. Genotoxicity was then evaluated through micronucleus (MN) test and Comet assay. The combination insecticide exposure at low doses of 0.05 and 0.1 μg/egg induced DNA damage as evidenced by an increased tail moment in the Comet assay. Further, the presence of micronucleated erythrocytes and also various abnormal cells including dacryocytes, microcytes, erythroplastids, squashed/notched nuclei, and spindle-shaped erythrocytes in the blood smear consolidates indicate the presence of insecticide-induced genotoxicity. Spinosad, however, was found only mildly genotoxic but at a high dose of 1.5 mg/egg. The results indicate that usage of naturalyte insecticide may be a better option to minimize the harmful effects of chemical insecticides.     

Oxidative stress and genotoxicity of 1-methyl-3-octylimidazolium chloride on loach (Misgurnus anguillicaudatus)

This study was a preliminary step to evaluate the acute toxicity of 1-methyl-3-octylimidazolium chloride ([C₈mim]Cl) on loach (Misgurnus anguillicaudatus) by determining the effects on hepatic antioxidant enzyme activities and by the comet assay. The results showed that [C₈mim]Cl had acute toxicity at concentrations above 20 mg L^?1, inducing oxidative stress and genotoxicity on fish liver cells. In respect to enzyme activities, [C₈mim]Cl induced changes in the activities of superoxide dismutase, catalase, and glutathione content the livers of fish exposed at 20–80 mg L^?1. [C₈mim]Cl at the same exposure level caused a remarkable increase in malondialdehyde level. The comet assay indicated that [C₈mim]Cl at 20–80 mg L^?1 induced genotoxicity in liver cells. With increased exposure concentration and time, the two comet parameters trailing rate and tail moment were significantly increased, with significant differences (P < 0.05) observed between control group and each treatment group. The present study shows that ionic liquids can be a threat to the health of aquatic organism when accidentally released to aquatic ecosystems.