Tradescantia bioassays for the determination of genotoxicity of water in the Panlong River, Kunming, People's Republic of China.

The Panlong river passes through Kunming City and receives a large quantity of municipal sewage and wastewater from industrial effluent. Along the river, 20 sites were selected to collect water samples to assess the genotoxicity using two Tradescantia assays, the micronucleus (Trad-MCN) and the stamen-hair-mutation (Trad-SHM) assays. The lowest frequencies in the Trad-MCN assay and the Trad-SHM assay are 3.19 MCN/100 tetrads and 1.32 M/1000 stamen hairs, respectively. In the water samples obtained from the Songhua Reservoir, the MCN frequencies and mutation rates are not statistically significantly different from the data found for the negative control (2.49 MCN/100 tetrads and 0.71 M/1000 stamen hairs). Among the other water samples, 19 in Trad-MCN assay and 17 in Trad-SHM assay show significantly higher genotoxicity than the control. The highest genotoxicity was in samples No. 19 for the MCN assay (8.73 MCN/100 cells), over three times higher than the negative control, and in sample No. 11 for the SHM assay (4.30 M/1000), six times higher than the negative control, and were about the same as for the positive control (10.0 mg/l NaN3, 9.28 MCN/100 tetrads and 7.44 SHM/1000 stamen hairs), respectively. The peak frequencies for the Trad-MCN assays were observed in the water samples obtained from the sites that were near industrial and municipal wastewater that ran into the river as effluent. In general, the frequency of MCN and SHM mutations increased where the river passed through Kunming. The Trad-MCN assay seemed more sensitive than that of the Trad-SHM assay in detecting genotoxicity of the river water pollution.     

Genotoxicity of water samples from Dianchi lake detected by the Vicia faba micronucleus test.

Dianchi Lake covers about a 300 km2 area. Kunming city on the edge of the lake is surrounded by industrial establishments. Farm land surrounds the remaining areas of the lake. The lake water is polluted by the Kunming city municipal sewage from 3 million inhabitants, the industrial effluent and farm runoff. Water samples were collected from 12 sites along the shore of the lake during the dry (May) and rainy (August) seasons for genotoxicity testing with the Vicia micronucleus assay during the year 1995. Genotoxicity in terms of micronuclei (MCN) frequencies in the root tip cells of Vicia showed a consistent elevated frequency of MCN over the control sample at the 0.05-0.01 levels of significance in both seasons. The MCN frequencies of sites A (Daguanhe), B (Gaoqiao) and C (Xiyuan) were 3.5-4 times as high as the control values (5.25/1000 cells) in the dry season. In the rainy season, the MCN frequencies of water samples from most of the 12 sites were relatively lower than those of the dry season except sites J (Haikou), and K (Kunyang). The average MCN frequency of the dry season samples was 14.97 per 1000 cells and that of the rainy season samples was 12.24 per 1000 cells while the average control value was around 5.00/1000 cells.     

Proficiency of the Tradescantia-micronucleus image analysis system for scoring micronucleus frequencies and data analysis.

The Tradescantia-micronucleus (Trad-MCN) bioassay is an efficient short-term test for genotoxicity of pollutants. In order to increase the efficiency and to standardize the micronucleus (MCN) scoring process, an automated scoring system was developed using the principle of image analysis in computer science. This assemblage is called the Tradescantia-micronucleus image analysis (Trad-MCNIA) system. The MCN frequencies scored by this system were compared with those scored by human observation for its proficiency. A set of low MCN frequency (around 5 MCN/100 tetrads) slides prepared from a control group, a set of medium MCN frequency (around 20 MCN/100 tetrads) slides prepared from sodium azide treated plant cuttings and a set of high MCN frequency (around 50 MCN/100 tetrads) slides prepared from X-ray treated materials were used for this study. In the low MCN frequency slides, the Trad-MCNIA system scored about the same value as human observation. In the medium and high frequency slides, MCN frequencies scored by the system were lower than those scored by human observers. This discrepancy was corrected by increasing the power of the objective of the microscope in the system. The MCN frequencies scored by the system attained 90% congruity with those scored by human observers after the correction. The scoring speed of the system was about 3.5 times as fast as that by human observers, and the data could be statistically analyzed immediately after the data scores were recorded. Further improvements can be made by upgrading the video camera and the computer speed.     

Genotoxicity of 2,4- and 2,6-dinitrotoluene as measured by the Tradescantia micronucleus (Trad-MCN) bioassay

The phytogenotoxicity of 2,4-dinitrotoluene (2,4-DNT) and 2,6-dinitrotoluene (2,6-DNT) was assessed using the Tradescantia micronucleus (Trad-MCN) bioassay. Tradescantia cuttings bearing young inflorescences were exposed for 6h to 2,4- or 2,6-DNT amended water solutions up to their respective solubilities. The nominal concentrations were 0, 1.9, 3.8, 7.5, 15, 30, 60, 100, 150, 200mg/l of 2,4-DNT, and 0, 7.5, 15, 30, 60, 90, 120, 180mg/l of 2,6-DNT. Each treatment was repeated three or four times. Chemical concentrations in test solutions were analyzed prior to and after the exposure. Cadmium chloride (0-20mM) was used as the positive control. Micronuclei (MCN) were scored in the tetrad-stage pollen mother cells. The MCN frequency (%), i.e. the number of micronuclei scored in 100 tetrads, was the measurement endpoint. Results indicated that both 2,4-DNT and 2,6-DNT were genotoxic with the minimum effective dose (MED) of 30 and 135mg/l, respectively. Longer exposure (30h) without recovery time at 150mg/l of 2,4-DNT and 180mg/l of 2,6-DNT did not induce significantly higher MCN frequencies.     

Impact of microgravity on radiobiological processes and efficiency of DNA repair

Three major plant bioassays, i.e., the Allium root anaphase aberration (Allium-AA), the Tradescantia-micronucleus (Trad-MCN) and the Tradescantia stamen hair mutation (Trad-SHM) tests, were utilized in soil solutions or shallow well water samples to determine the degree of their genotoxicity. Shallow well water samples were collected from five different farms, and soil solutions were extracted with distilled water or dimethyl sulfoxide (DMSO) from pesticide-contaminated (metolachlor, atrazine, extrazine, and 2, 4-D) and pesticide-free soil samples. Genotoxicity was expressed in terms of anaphase aberration (AA) frequencies in the Allium-AA test, in terms of micronuclei frequencies in the Trad-MCN test, and in terms of pink mutation events in the Trad-SHM test. On average, results of Allium-AA tests showed a 2.78-3.01 fold increase in anaphase aberration frequencies in contaminated soil solution samples and well water samples as compared with the negative control. Results of Trad-MCN tests showed a 1.66-4.75 fold increase of MCN frequencies in contaminated soil solution samples and shallow well water samples as compared with the frequencies of the controls. Results of Trad-SHM tests showed a 2.7-2.86 fold increase of pink mutation events in the contaminated soil solution samples over that of the controls. Control groups of the Allium-AA tests had an average of 0.75/1000 anaphase figures, and control groups of the Trad-MCN tests had an average of 3.2 MCN/100 tetrads, while control groups of the Trad-SHM tests had an average of 1.4 mutation events/1000 hairs. In general, soil solutions of DMSO extracts showed higher genotoxicity than that of distilled water extracts. Among these three plant bioassays, the Trad-MCN test has the highest efficiency. The highest toxicity, based upon the Trad-MCN test results, was found in the pesticide contaminated soil samples from Monroe's farm. Water samples from the Fountain Green/Bushnell area ranked second in genotoxicity.     

Genotoxic effects of volatile organic compounds in a chemical factory as evaluated by the Tradescantia micronucleus assay and by chemical analysis

The clastogenic effects of volatile organic compounds in the workplace air of a chemical factory were studied by means of the Tradescantia micronucleus (Trad-MCN) assay and chemical analysis. Sampling was performed at a chemical factory producing PVC film in Cheong-ju, South Korea. Inflorescences of Tradescantia BNL 4430 were placed for 2, 6, and 9 h at the height of 1.40 m at two locations in the workplace and one outdoor of the chemical industry. Air samplings were conducted in the same places and the collected tube samples were analyzed by automatic thermal desorption/gas chromatography/mass spectrometry (ATD/GC/MS). The frequencies of micronuclei in specimens exposed for 2 h in sites 1-3 were 6.13 +/- 0.47, 5.40 +/- 1.60, and 2.93 +/- 0.43 MCN per 100 tetrads, respectively. GC/MS analysis proved the presence of various volatile organic compounds such as trichloroethylene, toluene, ethyl benzene, (m, p, o)-xylene, styrene, 1,3,5-trimethyl benzene, and 1,2,4-trimethyl benzene. Mean values of toluene measured by 2 h sampling in sites 1-3 were 1946.6, 1368.3, and 340.1 microg/m3, respectively. The toluene concentrations in sites 1 and 2 were at least four to six times higher than that in site 3. The micronucleus frequencies increased with exposure time. In addition, there was a correlation between the micronucleus frequencies and toluene concentration in the air (R2 = 0.96). The results of this in situ monitoring proved the applicability of the Trad-MCN assay combined with chemical analysis for monitoring genotoxic chemicals in the work environment.     

Mutagenicity (micronucleus test in Vicia faba root tips), polycyclic aromatic hydrocarbons and heavy metal content of sediments collected in Tiber river and its tributaries within the urban area of Rome

Sediments collected in Tiber river and in its main tributary water courses within the urban area of Rome were tested for mutagenicity by means of Vicia faba root tips micronucleus (MN) test. Representative samples were scored for micronucleus generating events (chromosome/chromatid loss and fragments) too. Sediments were assayed for content of the thirteen most important chemicals of polycyclic aromatic hydrocarbon (PAH) group and for some heavy metal ions (Cd, Cr, Cu, Ni, Pb, Zn). Samples were collected in four tributary rivers (Prima Porta, Acqua Traversa, Aniene and Magliana) just before their confluence with Tiber river and at different stations along the Tiber river itself upstream and downstream the sites of confluence of the sampled tributaries. All samples were collected in July 1992. An alarming level of mutagenicity was reached in most of the tested stations, with an effect comparable to an X-rays exposure up to 0.4 Gy. Chemical analysis showed that the total amount of identified PAHs ranged from 4.5 to 625.2 ng/g of dry matrix in the different stations and the total amount of heavy metals ranged from 130 to 570 ppm. Tiber mutagenicity is likely to be mainly due to local factors such as the confluence of a small polluted tributary rather than to large scale effect due to an upstream–downstream relationship.     

Toxicity and genotoxicity of surface water before and after various potabilization steps

Many studies have revealed the presence of compounds with genotoxic activity in drinking water by means of short-term mutagenicity tests. In this study, the influence of the different steps of surface water treatment on the mutagenicity of drinking water was evaluated. Four different types of samples were collected: raw lake water, water after pre-disinfection with chlorine dioxide, water after filtration on granular activated carbon, and tap water. Water extracts underwent a bacterial toxicity test (Microtox test) and different in vitro genotoxicity tests: a test with Salmonella typhimurium strains, a Saccharomyces cerevisiae test, the SOS Chromotest with Escherichia coli and the Mutatox test with Vibrio fischeri. The Microtox test revealed high toxicity in the treated water samples. The disinfection steps increased the toxicity: the Mutatox test confirmed these results and the Salmonella/microsome test at the highest doses showed toxicity that could conceal mutagenicity. The SOS Chromotest was positive in all treated water samples without metabolic activation. In the test with S. cerevisiae both toxicity and genotoxicity generally increased during the water treatment steps, especially in cells without induction of cytochrome P450.     

Genetic effects observed in tetrads of Tradescantia induced by radon.

Inflorescences of Tradescantia clone 4430 were exposed to different concentrations of radon (Rn) gas (0.85, 12.10, 36.50 and 98.16 kBq/m3) from plants placed in an acrylic chamber that received radon from a container with pitchblende (containing uranium mineral). The exposure time was 24 h, afterwards the plants were left for 6 h in water and constantly aerated. Positive control plants were irradiated with gamma rays (0.8 Gy) and negative control plants received ambient air only (the background measurement had a mean of 0.38 kBq/m3). Micronuclei (MCN) in the tetrads induced by alpha particles emitted from Rn were tabulated and a linear concentration response was obtained. The potency of radon to induce MCN from the slope of the regression line was 0.13 MCN/kBq/m3 of alpha-radiation. Radon could reach the anthers by diffusion through the aerial spaces within the buds.     

Genotoxic evaluation of the River Paranaíba hydrographic basin in Monte Carmelo,MG, Brazil,by the Tradescantia micronucleus

Pollutants have adverse effects on human health and on other organisms that inhabit or use water resources. The aim of the present study was to assess the environmental quality of three watercourses in Monte Carmelo, MG, Brazil, using the micronucleus test on Tradescantia. For each treatment, 15 plants were exposed to water samples for 24 h. The control group was exposed to formaldehyde (0.2%) and the negative control to Hoagland solution. Subsequently the plants were placed in Hoagland solution for 24 h to recover. Cells were stained with 2% acetic carmine and examined by light microscopy. Three hundred tetrads were analyzed per slide. The frequency of genotoxic alterations was expressed as the number of micronuclei per 100 tetrads, and the groups were compared by ANOVA. At all sample sites for each watercourse significant genotoxicity indices were observed. The results suggest that in the Mumbuca creek, the current situation of effluent discharge should be reconsidered by the municipal environmental authorities. The increase in micronucleus frequency denoted for water samples of the Mumbuca creek, Lambari river and Perdizes river emphasizes the need to adopt environmental vigilance strategies, such as biological monitoring.     

Genotoxicity of drinking water from three Korean cities

Organic content of drinking tap water from Seoul, Taejon, and Suwon was extracted with an XAD-2 resin column and organic solvents. Four doses of the extract equivalent to 4, 2, 1, and 0.5 l water were tested for mutagenicity in Salmonella typhimurium strains TA98 and TA100 in the presence and absence of S9 mix. The organic extracts of the water from all three cities were mutagenic in TA 98 without S9 mix and in TA 100 with and without S9 mix. The highest number of revertants per plate was found in the absence of S9 mix. Three doses of the extract (equivalent to 22, 11, and 3.7 l water) were also tested in the bone marrow micronucleus test using BDF1 mice. At the highest dose, a significant increase of the micronucleus frequency was observed. The time required to be on the effect, however, varied with the source of the water. Our results indicate that the drinking tap waters from the three cities were genotoxic clearly in the bacterial test and also in the in vivo assay with mice. As we found no genotoxicity of the source water as seen in a previous study, it is likely that the chlorination process leads to the genotoxicity of the tap water.     

太湖蓝藻滤液的遗传毒性研究

蓝藻爆发是环境污染引发的重要事件之一,随之产生的蓝藻毒素又直接危及区域水安全.该论文采用蚕豆和大蒜根尖微核试验研究了太湖蓝藻暴发期间蓝藻滤液的遗传毒性.结果表明,同阴性对照相比,所有试验处理对蚕豆根尖细胞微核发生率的影响显著增加;对大蒜根尖细胞微核发生率而言除蓝藻滤液8倍稀释液的影响不显著外,其它水平效应显著高于阴性对照,而且表现出一定的剂量效应.暴发期蓝藻滤液原液对蚕豆根尖细胞微核发生率影响显著高于阳性对照(0.8mg·mL^-1环磷酰胺)的效应,从而说明蓝藻暴发时期蓝藻滤液具有较强的遗传毒性.通过微核试验效果分析,蚕豆作为植物监测系统的敏感性和稳定性都优于大蒜材料.     

抚仙湖无机污染物化学背景值及动物体内致突变性评定

为了比较抚仙湖不同区域的水质及其致突变性,从抚仙湖周围水域采取７个样点的水样。用电感耦合氩等离子发射光谱法对水样中的无机污染物作定性和定量分析,另外使用蝌蚪红细胞微核试验检测抚仙湖水样的致突变性,结果显示：禄要样点水样带有微核的红细胞率增高,表明这个点水样有致突变活性物质。     

The Salmonella mutagenicity assay in a surface water quality monitoring program based on a 20-year survey

Since 1979, the Environmental Agency of S?o Paulo State in Brazil, CETESB, has been using the Salmonella mutagenicity assay to assess the quality of natural waters. This paper is a compilation of data obtained during the last 20 years from more than a thousand samples. Potencies up to 30,000 revertants/l were observed in 137 positive samples. The Salmonella typhimurium strain TA98 was more sensitive than TA100; 79% of the mutagenicity was detected by this strain, regardless of the presence of S9-mix. A classification of the mutagenic response was proposed to facilitate in the dissemination of the information to the public. The classification was low, moderate, high and extreme for samples with mutagenic potency (revertants/l equivalent) of < 500, 500-2500, 2500-5000 and > 5000, respectively. As a result of this effort to standardize methodologies, compile and classify the mutagenic effect of water pollution, in 1998, the Salmonella mutagenicity assay was officially and systematically included in the S?o Paulo State Water Quality Monitoring Program. This assay has proven to be a useful tool in the identification of important pollution sources. Correction and prevention actions in Water Pollution Control Programs were generated as a result.     

死鱼事件中污染源废水对鱼类急性毒性及诱变微核的研究

为了研究2006年9月广州郊区某鱼塘因化工企业污染源废水进入而导致大量死鱼事件中鱼类死亡原因,用斑马鱼(Brachydanio rerio)和蚕豆(Vicia faba)根尖对污染源废水及鱼塘水分别进行了鱼类急性毒性试验和蚕豆根尖微核试验。污染源废水和鱼塘水的96hLC₅₀分别为1.69%、23.95%,污染源废水和鱼塘水的微核率分别为27.8‰、13.7‰,污染源废水和鱼塘水的COD、BOD、石油类、氨氮、总磷等均出现超标情况,其中氨氮、苯胺、石油类污染物严重超标。监测结果表明,以氨氮污染为主的污染源废水是导致死鱼事件的主要原因,废水对蚕豆根尖微核率的形成具有显著性影响,可能存在潜在的致突变性危险。     

红湖水质对模型动物致突变性的研究

目的:以微核技术及精子畸变率为指示,探讨新疆大学红湖水质对模型动物致突变性作用。方法:采用小鼠嗜多染红细胞(PCE)微核率(MCN)及精子畸变率检测方法,研究红湖水质及其潜在的致突变性。结果:3个水样点的污水都能引起小白鼠嗜多染红细胞及精子细胞不同程度的遗传毒性。与对照组(自来水)喂养的小白鼠相比具有明显的差异(P<0.005)。结论:红湖水质已经被严重污染,对人体及动物具有潜在的危害。     

Tradescantia-micronucleus assay for the assessment of the clastogenicity of Austrian water.

Seven water samples collected from Vienna and Salzburg areas in Austria were tested for their clastogenicity with the Tradescantia-micronucleus (Trad-MCN) assay. There was no indication of clastogenic activity in two drinking water samples; likewise, samples from two major rivers (Danube and Salzburg) and of a river that received effluents from a paper mill also gave negative results. Urban river water as well as ground water samples which were collected near an industrial waste dump site caused a statistically significant and dose dependent increase of the MCN frequencies.     

The effect of gamma-radiation on smoked fish using short-term mutagenicity assays

The effect of gamma-radiation on the mutagenicity potential of wood-smoked fish (Rastrelliger sp.) was investigated. Smoked fish were irradiated with radiation doses of 2.0, 4.0, 6.0 and 8.0 kGy. The DMSO extracts of non-radiated and irradiated smoked fish were tested for mutagenicity using the Ames plate incorporation assay, host-mediated assay, and the micronucleus test. It was observed that gamma-irradiation did not induce any significant increase in the number of revertants of TA98, TA100 and TA104 as compared with the non-radiated smoked fish. Results of the host-mediated assay and the micronucleus test showed no difference in the mutagenic response of non-radiated and irradiated smoked fish. The results indicate that gamma-radiation does not introduce mutagens in smoked fish.     

Quantification of a potent mutagenic 4-amino-3,3'-dichloro-5,4'-dinitrobiphenyl (ADDB) and the related chemicals in water from the Waka River in Wakayama, Japan

4-Amino-3,3'-dichloro-5,4'-dinitrobiphenyl (ADDB) is a novel chemical exerting strong mutagenicity, especially in the absence of metabolic activation. In addition to mutagenicity, ADDB may also disrupt the endocrine system in vitro. ADDB may be discharged from chemical plants near the Waka River and could be unintentionally formed via post-emission modification of drainage water containing 3,3'-dichlorobenzidine (DCB), which is a precursor in the manufacture of polymers and dye intermediates in chemical plants. The main purpose of this study was to make a comprehensive survey of the behaviour and levels of ADDB and suspected starting material or intermediates of ADDB, i.e., DCB, 3,3'-dichloro-4,4'-dinitrobiphenyl (DDB), and 4-amino-3,3'-dichloro-4'-nitrobipheny (ADNB) in Waka River water samples. We also postulated the formation pathway of ADDB. Water samples were collected at five sampling sites from the Waka River four times between March 2003 and December 2004. Samples were passed through Supelpak2 columns, and adsorbed materials were then extracted with methanol. Extracts were used for quantification of ADDB and the related chemicals by HPLC on reverse-phase columns; mutagenicity was evaluated in the Salmonella assay using the O-acetyltransferase-overexpressing strain YG1024. High levels of ADDB, DCB, DDB, and ADNB (12.0, 20,400, 134.8, and 149.4ng/L-equivalent) were detected in the samples collected at the site where wastewater was discharged from chemical plants into the river. These water samples also showed stronger mutagenicity in YG1024 both with and without S9 mix than the other water samples collected from upstream and downstream sites. The results suggest that ADDB is unintentionally formed from DCB via ADNB in the process of wastewater treatment of drainage water containing DCB from chemical plants.     

Mutagenicity of instant coffee and its interaction with dimethylnitrosamine in the micronucleus test

The mutagenicity of instant coffee and its interaction with dimethylnitrosamine (DMN) were examined in mice using the micronucleus test. Although neither a single nor multiple administration of instant coffee by gavage induced a significant rise in micronucleated cells over the dose range tested (100-2500 mg/kg), there was a tendency for the number of micronucleated cells to increase in a dose-related fashion. When coffee was administered with DMN, the difference in the frequency of micronucleated cells was small in comparison to a single treatment with DMN alone, thus indicating a lack of synergism between coffee and DMN.