Genotoxicity Of Soil Research Articles

Evaluating the Effects of Bioremediation on Genotoxicity of Polycyclic Aromatic Hydrocarbon-Contaminated Soil Using Genetically Engineered, Higher Eukaryotic Cell Lines

Bioremediation is one of the commonly applied remediation strategies at sites contaminated with polycyclic aromatic hydrocarbons (PAHs). However, remediation goals are typically based on removal of the target contaminants rather than on broader measures related to health risks. We investigated changes in the toxicity and genotoxicity of PAH-contaminated soil from a former manufactured-gas plant site before and after two simulated bioremediation processes: a sequencing batch bioreactor system and a continuous-flow column system. Toxicity and genotoxicity of the residues from solvent extracts of the soil were determined by the chicken DT40 B-lymphocyte isogenic cell line and its DNA-repair-deficient mutants. Although both bioremediation processes significantly removed PAHs from the contaminated soil (bioreactor 69% removal, column 84% removal), bioreactor treatment resulted in an increase in toxicity and genotoxicity over the course of a treatment cycle, whereas long-term column treatment resulted in a decrease in toxicity and genotoxicity. However, when screening with a battery of DT40 mutants for genotoxicity profiling, we found that column treatment induced DNA damage types that were not observed in untreated soil. Toxicity and genotoxicity bioassays can supplement chemical analysis-based risk assessment for contaminated soil when evaluating the efficacy of bioremediation.

Improvement of Vicia-micronucleus test for assessment of soil quality: A proposal for international standardization

The Vicia faba root tip micronucleus test is one of the most employed plant genotoxicity assays, and has been used on various types of contaminated materials. This test has been standardized by AFNOR, the French member organization of ISO. However, this test is usually performed with a water extraction step but soil genotoxicity assessment would be more relevant when performed directly in the soil itself. In order to harmonize these protocols, an ISO standard for the V. faba micronucleus test in both liquid phase (exposure of plants to different liquid matrix, including soil water extracts) and solid phase (direct exposure of plants to the soil) would be very useful. In this context, we compared two exposure durations in the solid phase (48 h and 5 d) for the V. faba micronucleus test with two different well-known genotoxicants, maleic hydrazide and copper sulfate. We concluded that these two durations induced equivalent sensitivity: the micronucleus frequency was significantly increased with 5 μmol maleic hydrazide per kg dry soil and with 2 mmol copper sulfate per kg dry soil with both exposure durations. However, exposing roots to soil during 48 h is more practical. Moreover, organically and conventionally cultured seeds were employed to determine whether the seed provenance influenced the test sensitivity. Organic seeds were less sensitive to copper, possibly because copper-based treatments are permitted, and often applied, in organic farms. Therefore, in the absence of completely non-treated seeds, organically-cultured seeds did not appear to offer any advantages over conventional seeds.

Screening evaluation of the ecotoxicity and genotoxicity of soils contaminated with organic and inorganic nanoparticles: The role of ageing

This study aimed to evaluate the toxicity and genotoxicity of soils, and corresponding elutriates, contaminated with aqueous suspensions of two organic (vesicles of sodium dodecyl sulphate/didodecyl dimethylammonium bromide and of monoolein and sodium oleate) and five inorganic nanoparticles (NPs) (TiO 2, TiSiO 4, CdSe/ZnS quantum dots, Fe/Co magnetic fluid and gold nanorods) to Vibrio fischeri and Salmonella typhimurium (TA98 and TA100 strains). Soil samples were tested 2 h and 30 days after contamination. Suspensions of NPs were characterized by Dynamic Light Scattering. Soils were highly toxic to V. fischeri, especially after 2 h. After 30 days toxicity was maintained only for soils spiked with suspensions of more stable NPs (zeta potential > 30 mV or <−30 mV). Elutriates were particularly toxic after 2 h, except for soil spiked with Fe/Co magnetic fluid, suggesting that ageing may have contributed for degrading the organic shell of these NPs, increasing the mobility of core elements and the toxicity of elutriates. TA98 was the most sensitive strain to the mutagenic potential of soil elutriates. Only elutriates from soils spiked with gold nanorods, quantum dots (QDs) and TiSiO 4 induced mutations in both strains of S. typhimurium, suggesting more diversified mechanisms of genotoxicity.

Aporrectodea caliginosa, a suitable earthworm species for field based genotoxicity assessment?

There is a growing interest for the application of biomakers to field-collected earthworms. Therefore we have evaluated the usability of native populations of endogeic, widely distributed earthworm Aporrectodea caliginosa in the assessment of soil genotoxicity using the Comet assay. Validation of the Comet assay on earthworm coelomocytes has been established using commercially available Eisenia fetida exposed to copper, cadmium, and pentachlorophenol, along with A. caliginosa exposed to copper in a filter paper contact test. Neutral red retention time (NRRT) assay was conducted on copper exposed and field-collected earthworms. Significant DNA and lysosomal damage was measured using Comet and NRRT assays in native populations of A. caliginosa sampled from the polluted soils in the urban area in comparison to the earthworms from the reference site. The results of this study confirm the employment of A. caliginosa as a suitable species for the in situ soil toxicity and genotoxicity field surveys.

Potential toxicicty and genotoxicity of soils around a high impacted site

Potential toxicicty and genotoxicity of soils around a high impacted site

Genotoxicity of Agricultural Soils after one year of Conversion Period and under Conventional Agriculture

Agricultural soils are often contaminated with genotoxic chemicals. Hence, transition to sustainable agriculture requires a conversion period. We performed a comparative evaluation of the soilgenotoxicity in a field after one year of conversion and in a field under conventional agriculture, not located close to sources of pollution. Soil samples were taken from 0-20 cm and 20-40 cm depth. Allium cepa-test system was used for the cytogenetic analysis. The higher mitotic index and lower frequency of chromosome aberrations in the samples from the field in conversion period compared with the field under conventional agriculture are indicative of a decrease of the soil genotoxicity. A slight increase in the frequency of aberrantcells and a substantial decrease in the mitotic index in 20-40 cm soil layer from both fields were observed. Regarding the field in conversion this might be due to the presence of agrochemicals used in the previous years, and indicates the necessity of longer conversion period. The results showed that Allium cepa-test might be wasused for monitoring of genotoxic pollution of the soils without preliminary extraction of the chemicals they contain.

The influence of thermal desorption on genotoxicity of multipolluted soil

A multipolluted soil sampled from a former coking plant in Lorraine (France) was evaluated for its genotoxic effects on coelomocytes of the Eisenia fetida earthworm using the comet assay. The biological efficiency of thermal desorption of the contaminated soil was also investigated. The untreated polluted soil was shown to be genotoxic to earthworms. Although thermal desorption reduced the concentration of PAHs by 94% (∑ 16 PAHs=1846 and 101 mg/kg before and after thermal desorption, respectively), the treatment did not eliminate the genotoxicity of soil pollutants to earthworms but increased it. The concentration of non-volatile metals did not change after thermal desorption. Among metals found in the treated soil, cadmium, chromium and nickel could explain the genotoxicity of the contaminated soil after thermal desorption. The treatment could increase the bioavailability and genotoxicity of heavy metals, through a modification of the soil’s organic matter, the speciation of heavy metals and their binding to organic matter. This study underlines the importance of measuring biological effects, in order to evaluate the risk associated with formerly contaminated soils and the efficiency of remediation.

New direct contact approach to evaluate soil genotoxicity using the Vicia faba micronucleus test

A method to assess micronucleus (MN) induction in Vicia faba roots by direct contact exposure to a solid matrix was developed. The procedure comprised a 5-d germination period, as in the well-known method using aqueous extracts. However, the seeds were here sown directly into the test soil whereas a culture period is necessary before exposing seedlings to a liquid medium. One soil under forest and two contaminated soils from areas affected by industrial installations and a coke works were used. Three durations of direct exposure were tested: 2, 5 and 7 d. The optimal duration was evaluated at 2 d to observe maximal MN induction without observing toxicity symptoms. The methodology using aqueous extracts was applied to the same three soils: MN frequency was higher than in the direct contact assay but the ratios of MN frequencies from tested soils in comparison to the negative control were lower. However, for each soil, both the direct contact method and the aqueous extract exposure led to the same risk assessment diagnosis. The evaluation of a concentration range of a polycyclic aromatic hydrocarbons (PAH)-contaminated soil showed a dose-dependent MN frequency when the seeds were allowed to germinate before sowing in the soil: the soil genotoxicity was the highest at intermediate doses. The direct contact method was found to be rapid, sensitive and well suited to the evaluation of soil quality.

Phytotoxicity and genotoxicity of soils from an abandoned uranium mine area

The great number of abandoned uranium mines within the Portuguese territory led to the production of specific legislation establishing the rehabilitation of these areas and the definition of those requiring prior intervention. Cunha Baixa uranium mine (Mangualde, Central Portugal) was one of these areas and a site-specific risk assessment is being carried out for this mine, following European frameworks. In this study the Ames test with soil elutriates, and emergence and growth assays with Lactuca sativa L. and Zea mays L. were performed to assess soil retention and habitat functions. Levels of metals bioaccumulated in plant tissues, grown above soil surface, during the assays, were determined as well. No genotoxic effects were recorded for soil eluates (1:10 and 1:2, m/v) except for site F within the mine area, which is under the effluence of runoffs from the effluent treatment pond. Nevertheless, most of all were highly toxic to Salmonella typhimurium (especially for TA98) strains, used in the assay. The results gathered suggested that during the natural process of soil percolation by rainwater, the leachates yielded are contributing with toxic but no genotoxic metals to underground water resources. Phytotoxicity assays demonstrated that soil habitat function was compromised in almost all the sampling sites selected within the mine area. These results have clearly demonstrated how evaluations based on extractable metal concentrations may underestimate the risks of contaminated soils. Lettuce was the most sensitive species probably due to the great ability of this species in accumulating high concentrations of metals in their aboveground tissues. The highest bioaccumulation factors (BAFs) values were recorded for this species namely for Co (16.5), Cu (28.5), Ba (7.3), Sr (7.2) and Ni (6.1). Copper (9.0) and nickel (3.6) were the main elements bioaccumulated by maize. The dilution of mine soils with a local reference soil has proved to mitigate its toxicity in almost all the sites analysed.

Genotoxicity of agricultural soils in the vicinity of industrial area

Soil samples from agricultural fields (cultivated) in the vicinity of industrial area of Ghaziabad City (India) were collected. In this city, wastewater coming from both industrial and domestic sources and without any treatment is used to irrigate the food crops. This practice has been polluting the soil and pollutants might reach the food chain. Gas chromatographic analysis show the presence of certain organochlorine (DDE, DDT, dieldrin, aldrin and endosulfan) and organophosphorus (dimethoate, malathion, methylparathion and chlorpyrifos) pesticides in soil samples. Samples were extracted using different solvents, i.e. methanol, chloroform, acetonitrile, hexane and acetone (all were HPLC-grade, SRL, India), and the extracts were assayed for genotoxic potential using Ames Salmonella/microsome test, DNA repair defective mutants and bacteriophage λ systems. TA98 and TA100 were found to be the most sensitive strains to all the soil extracts tested. Methanol extracts exhibited a maximum mutagenicity with TA98 strain {540 (−S9) and 638 (+S9) revertants/g of soil} and 938 (−S9) and 1008 (+S9) revertants/g of soil with TA100 strain. The damage in the DNA repair defective mutants was found maximum with methanolic extract followed by acetonitrile, chloroform, hexane and acetone at the dose level of 40 μl/ml culture after 6 h of treatment. The survival was 25, 30, 32, 33 and 35% in polA strain after 6 h of treatment when tested with wastewater irrigated soil extracts of methanol, acetonitrile, chloroform, hexane and acetone, respectively. A significant decrease in the plaque forming units of bacteriophage λ was also observed when treated with 40 μl of test samples. Present results showed that methanolic extracts of soil were more toxic than other soil extracts. The soil is accumulating a large number of pollutants due to wastewater irrigation and this practice of accumulation has an impact on soil health.

Genotoxicity of Agricultural Soils after One Year of Conversion Period and Under Conventional Agriculture

ABSTRACTPollution of the agricultural areas as a result of used agrochemicals turns into a global problem for contemporary mankind. Sustainable agriculture provides protection of the environment and requires a conversion period. We performed a comparative evaluation of the soil genotoxicity in a field after one year of conversion and in a field under conventional agriculture, not located close to sources of pollution. Soil samples were taken from 0–20 cm and 20–40 cm depth. Allium cepa-test system was used for the cytogenetic analysis. The higher mitotic index and lower frequency of chromosome aberrations in the samples from the field in conversion period compared with the field under conventional agriculture are indicative of a decrease of the soil genotoxicity. A slight increase in the frequency of aberrant cells and a substantial decrease in the mitotic index in 20–40 cm soil layer from both fields were observed. Regarding the field in conversion this might be due to the presence of agrochemicals used in the previous years, and indicates the necessity of longer conversion period. The results showed that Allium cepa-test might be was used for monitoring of genotoxic pollution of the soils without preliminary extraction of the chemicals they contain.

Genotoxicity of Acqueous Elutions of Industrial Soils

The genotoxicity of industrial soils was evaluated. A single cell electrophoresis assay or comet assay, using eleocyte cells of Eisenia foetida, was performed to assess the genotoxicity of aqueous elutions. These were obtained from industrial soils containing metals. All soil samples meet the environmental quality guidelines for metal concentrations. However, elutions have produced genotoxic effects at dilutions as low as 6%. Total metal concentrations for each aqueous elution could express synergistic effects of these compounds.

Genotoxicity evaluation of water soil leachates by Ames test, comet assay, and preliminary Tradescantia micronucleus assay

Combining genotoxicity/mutagenicity tests and physico-chemical methodologies can be useful for determining the potential genotoxic contaminants in soil samples. The aim of our study was to evaluate the genotoxicity of soil by applying an integrated physico-chemical-biological approach. Soil samples were collected at six sampling points in a Slovenian industrial and agricultural region where contamination by heavy metals and sulphur dioxide (SO(2)) are primarily caused by a nearby power plant. The in vitro alkaline version of the comet assay on water soil leachates was performed with Caco-2 and HepG2 cells. A parallel genotoxicity evaluation of the samples was performed by Ames test using Salmonella typhimurium and the Tradescantia micronucleus test. Pedological analyses, heavy metal content determination, and different physico-chemical analyses, were also performed utilizing standard methodology. Water leachates of soil samples were prepared according to standard methods. Since only a battery of biotests with prokaryotic and eukaryotic organisms or cells can accurately estimate the effects of (geno)toxicants in soil samples and water soil leachates, a combination of three bioassays, with cells or organisms belonging to different trophic levels, was used. Genotoxicity of all six water soil leachates was proven by the comet assay on both human cell lines, however no positive results were detected by bacterial assay, Ames test. The Tradescantia micronucleus assay showed increase in micronuclei formation for three samples. According to these results we can assume that the comet assay was the most sensitive assay, followed by the micronucleus test. The Ames test does not appear to be sensitive enough for water soil leachates genotoxicity evaluations where heavy metal contamination is anticipated.

Genotoxicity assessment of soils from wastewater irrigation areas and bioremediation sites using the Vicia faba root tip micronucleus assay

Genotoxicity potential of soils taken from wastewater irrigation areas and bioremediation sites was assessed using the Vicia faba root tip micronucleus assay. Twenty five soils were tested, of which 8 were uncontaminated soils and taken as the control to examine the influence of soil properties; 6 soils were obtained from paddy rice fields with a history of long-term wastewater irrigation; 6 soils were obtained from bioremediation sites to examine effects of bioremediation; and 5 PAH-contaminated soils were used to examine methodological effects between direct soil exposure and exposure to aqueous soil extracts on micronuclei (MN) frequency ( per thousand) in the V. faba root tips. Results indicate that soil properties had no significant influences on MN frequencies (p > 0.05) when soil pH varied between 3.4 to 7.6 and organic carbon between 0.4% and 18.6%. The MN frequency measured in these control soils ranged from 1.6 per thousand to 5.8 per thousand. MN frequencies in soils from wastewater irrigation areas showed 2- to 48-fold increase as compared with the control. Soils from bioremediation sites showed a mixed picture: MN frequencies in some soils decreased after bioremediation, possibly due to detoxification; whereas in other cases remediated soils induced higher MN frequencies, suggesting that genotoxic substances might be produced during bioremediation. Exposure to aqueous soil extracts gave a higher MN frequency than direct exposure in 3 soils. However, the opposite was observed in the other two soils, suggesting that both exposure routes should be tested in case of negative results from one route. Data obtained from this study indicate that the MN assay is a sensitive assay suitable for evaluating genotoxicity of soils.

Genotoxic Risk Identification of Soil Contamination at a Major Industrialized City in Northeast China by a Combination of in Vitro and in Vivo Bioassays

The present study evaluated the genotoxicity of field soils in the Tianjin area, one of the most industrialized contaminated areas in northeast China. The genotoxicity of organic extracts of 41 soils was assayed by an in vitro SOS/ umu bioassay with Salmonella typhimurium TA 1535/pSK 1002. From the 41 soil samples, 11 samples were selected to confirm the genotoxic effect by in vivo single-cell gel electrophoresis (comet assay) using earthworms (Eisenia fetida). The results obtained demonstrated that, in the in vitro assay, genotoxicity expressed as induction ratios (IR) ranged from 1.00 to 4.60, and in the in vivo assay, the genotoxicity expressed as tail moment (TM) varied from 14.6 to 57.8 microm. All samples with high genotoxicity assessed by the SOS/umu bioassay possessed significantly high genotoxic effects in the comet assay, and there was a correlation (R2 = 0.736, p < 0.05) between IR and TM in both bioassays. It is concluded that soils in the Tianjin area were seriously contaminated by organic genotoxicants and higher levels of genotoxic effects existed in soils in the urban area of Tianjin as well as in areas near the coastal towns in the northeast part of the city. It can be concluded that a combination of in vivo and in vitro bioassays as a powerful and efficient genotoxicity-assessing tool could facilitate the assessment of genotoxic risk at a regional scale.

Evaluation of genotoxicity of combined soil pollution by cadmium and imidacloprid

Cadmium (Cd) is one of the important pollutants of soil and the genotoxicity of Cd-contaminated soil was studied in combination with imidacloprid. The single cell gel electrophoresis or comet assay was used to quantify DNA strand breaks as a measure of DNA damage induced by Cd and imidacloprid contamination in soil. The soil was artificially contaminated by Cd (0.0, 0.2, 0.5, 1.0, 2.0 mg x kg(-1) dry soil) or Cd (0.0, 0.2, 0.5, 1.0, 2.0 mg x kg(-1) dry soil) and imidacloprid (0.5 mg x kg(-1) dry soil). Roots of Vicia faba were exposed to the contaminated soil for 2 h at 25 degrees C and were used in the comet assay. DNA damage was measured as the values of percentage of nuclei with tails, tail length, tail DNA, tail moment (TM), and Olive tail moment (OTM). DNA damages of root tips of Vicia faba increased after Cd treatment and there were dose-related increases in DNA damage measured as these parameters. However, the addition of imidacloprid further increased the DNA damage. These data confirmed the genotoxic effect of Cd to plants, and that the combined pollution with imidacloprid can enhance the genotoxicity of Cd.

Assessment of the contamination and genotoxicity of soil irrigated with wastewater

Wastewater has been widely used for agricultural irrigation in China and countries in arid and semi-arid areas. Because of poor control of the wastewater quality and long-term application, pollutants accumulation and the relevant ecotoxicological effects may occur, especially for persistent pollutants and permanently bound chemical residues. The pollutants accumulated in soil may transfer through the food chains and cause negative effects on human health. In the present study, a field survey in farmlands that have been irrigated with effluents from a sewage treatment plant and untreated wastewater was carried out. The soil quality was evaluated using a battery of chemical and biological parameters to describe the integrated situations of the polluted sites. Organochlorine pesticides (OCPs) were quantified by GC–ECD and polycyclic aromatic hydrocarbons (PAHs) were analyzed by GC-MS with internal standard. Polyphenol oxidase and catalase activities were examined to investigate the soil functions. Vicia faba micronucleus test was performed to detect the potential genotoxicity of water extracts of the soils. It has been concluded that both PAHs and OCPs have accumulated in the soils irrigated with wastewater. The main origin of PAHs was from the wastewater discharged from a coal plant. The site close to the pollution discharge point was heavily polluted by PAHs. OCPs were mainly residues of historical agriculture application. The MN frequencies and the activity of polyphenol oxidase were positively correlated with PAHs concentrations but have no correlation with OCPs. Therefore, polyphenol oxidase activity and MN frequencies were suggested as the biological indicators of PAHs pollution.

Soil genotoxicity assessment: a new stategy based on biomolecular tools and plant bioindicators.

The setting up of efficient early warning systems is a challenge to research for preventing environmental alteration and human disease. In this paper, we report the development and the field application of a new biomonitoring methodology for assessing soil genotoxicity. In the first part, the use of amplified fragment length polymorphism and flow cytometry techniques to detect DNA damage induced by soils artificially contaminated with heavy metals as potentially genotoxic compounds is explained. Results show that the combination of the two techniques leads to efficient detection of the sublethal genotoxic effect induced in the plant bioindicator by contaminated soil. By contrast, the classic mortality, root, and shoot growth vegetative endpoints prove inappropriate for assessing soil genotoxicity because, although they cause genotoxic damage, some heavy metals do not affect sentinel plant development negatively. The statistical elaboration of the data obtained led to the development of a statistical predictive model which differentiates four different levels of soil genotoxic pollution and can be used everywhere. The second part deals with the application of the biomonitoring protocol in the genotoxic assessment of two areas surrounding a steelworks in northern Italy and the effectiveness of this methodology. In this particular case, in these areas, the predictive model reveals a pollution level strictly correlated to the heavy metal concentrations revealed by traditional chemical analysis.

Effects of heavy metal contamination of soils on micronucleus induction in Tradescantia and on microbial enzyme activities: a comparative investigation

The aim of this study was to investigate correlation between genotoxic effects and changes of microbial parameters caused by metal contamination in soils. In total, 20 soils from nine locations were examined; metal contents and physicochemical soil parameters were measured with standard methods. In general, a pronounced induction of the frequency of micronuclei (MN) in the Tradescantia micronucleus (Trad-MN) assay was seen with increasing metal concentration in soils from identical locations. However, no correlations were found between metal contents and genotoxicity of soils from different locations. These discrepancies are probably due to differences of the physicochemical characteristics of the samples. Also, the microbial parameters depended on the metal content in soils from identical sampling locations. Inconsistent responses of the individual enzymes were seen in soils from different locations, indicating that it is not possible to define a specific marker enzyme for metal contamination. The most sensitive microbial parameters were dehydrogenase and arylsulfatase activity, biomass C, and biomass N. Statistical analyses showed an overall correlation between genotoxicity in Tradescantia on the one hand and dehydrogenase activity, biomass C, and the metabolic quotient on the other hand. In conclusion, the results of the present study show that the Trad-MN assay is suitable for the detection of genotoxic effects of metal contamination in soils and furthermore, that the DNA-damaging potential of soils from different origin cannot be predicted on the basis of chemical analyses of their metal concentrations.

Tradescantia Micronucleus and Vicia Chromosome Anatelophase Assays in Monitoring of Genotoxicity of Urban Soil

Tradescantia micronucleus assay (Trad MCN assay) and Vicia anatelophase chromosome aberration assay (Vicia assay) were used to evaluate genotoxicity of extracts from soil samples. Soil samples from six different sites in area of Bratislava - capital of the Slovak Republic - have been taken. As a negative control soil samples from Botanical garden of medicinal plants, Faculty of Pharmacy, Comenius University were used. Soil samples for Trad MCN assay were extracted with distilled water or 5% dimethylsulphoxide (DMSO). Similarly genotoxicity of sample extracted with distilled water by Vicia assay was evaluated. Two sites were found as most polluted. The results of the study indicate different level of pollution in selected sites.