Micronucleus Assay with Tetrad Cells of Tradescantia.

The Tradescantia micronucleus assay has been used since 50 years for the detection of genotoxins (including carcinogens) in the environment. A large database concerning the effects of individual chemicals and complex environmental mixtures (soil, air and waters) has accumulated. In contrast to other mutagenicity test systems, the effects of low concentrations of heavy metals, radionuclides, certain herbicides, pesticides and gaseous mutagens can be detected and it is also possible to conduct in situ biomonitoring studies with plant. The test system has been validated and standardized protocols have been developed for laboratory experiments and for field studies which are described in this chapter.

The Tradescantia micronucleus assay is a highly sensitive tool for the detection of low levels of radioactivity in environmental samples.

Environmental contamination with radioactive materials of geogenic and anthropogenic origin is a global problem. A variety of mutagenicity test procedures has been developed which enable the detection of DNA damage caused by ionizing radiation which plays a key role in the adverse effects caused by radioisotopes. In the present study, we investigated the usefulness of the Tradescantia micronucleus test (the most widely used plant based genotoxicity bioassay) for the detection of genetic damage caused by environmental samples and a human artifact (ceramic plate) which contained radioactive elements. We compared the results obtained with different exposure protocols and found that direct exposure of the inflorescences is more sensitive and that the number of micronuclei can be further increased under "wet" conditions. The lowest dose rate which caused a significant effect was 1.2 µGy/h (10 h). Comparisons with the results obtained with other systems (i.e. with mitotic cells of higher plants, molluscs, insects, fish and human lymphocytes) show that the Tradescantia MN assay is one to three orders of magnitude more sensitive as other models, which are currently available. Taken together, our findings indicate that this method is due to its high sensitivity a unique tool, which can be used for environmental biomonitoring in radiation polluted areas.

Impact of common cytostatic drugs on pollen fertility in higher plants.

Cytostatic drugs are among the most toxic chemicals which are produced. Many of them cause damage of the genetic material which may affect the fertility of higher organisms. To study the impact of the widely used anticancer drugs [cisplatin (CisPt), etoposide (Et), and 5-fluorouracil (5-FU)] on the reproduction of higher plants, pollen abortion experiments were conducted with species which belong to major plant families, namely with Tradescantia paludosa (Commelinaceae), Arabidopsis thaliana (Brassicaceae), Chelidonium majus (Papaveraceae), and Alisma plantago-aquatica (Alismataceae). All compounds increased the frequencies of abortive grains. The lowest effective doses were in general in a narrow range (i.e., 1 and 10 mg/kg of dry soil). The effects of the individual drugs were similar in T. paludosa, A. plantago-aquatica, and Ch. majus, while A. thaliana was consistently less sensitive. The highest abortion rate was obtained in most experiments with CisPt, followed by 5-FU and Et. Comparisons of the doses which caused effects in the present experiments in the different species with the predicted environment concentrations and with the levels of the cytostatics which were detected in hospital wastewaters show that the realistic environmental concentrations of the drugs are 4-6 orders of magnitude lower. Therefore, it is unlikely that these drugs affect the fertility of higher plants in aquatic and terrestrial ecosystems.

Analyses of combined effects of cytostatic drugs on micronucleus formation in the Tradescantia.

Recent experiments showed that 5-fluorouracil (5FU), cisplatin (CDDP), etoposide (ET), and imatinib mesylate (IM), which are currently among the most widely used anticancer drugs, cause damage of the genetic material in higher plants. The aim of the present study was to determine whether mixtures of these drugs cause synergistic or antagonistic effects which may have an impact on their environmental safety. Therefore, the effects of binary mixtures of these anticancer drugs on the induction of micronuclei (MN) which reflect structural and numerical chromosomal aberrations were assessed in Tradescantia tetrads. Synergistic/antagonistic effects were determined by comparison with single exposures that would be equally effective in a reference model of independent action. This comparison was performed at two distinct effect sizes. We found clear evidence for synergisms in combination experiments with IM and antagonism in a high-dose experiment with ET and 5FU. Our findings indicate that IM increases the genotoxic effects of other anticancer drugs. The maximal effects which we found were in the range between 19 and 38 % in the excess of effect sizes predicted under independent action. These effects may have an impact on the overall genotoxic activities of untreated hospital waste waters but not on the environment in general as the predicted environmental concentrations of the studied drugs are several orders of magnitude lower as the levels which are required to cause induction of MN in higher plants.

Red mud a byproduct of aluminum production contains soluble vanadium that causes genotoxic and cytotoxic effects in higher plants.

Red mud (RM) is a byproduct of aluminum production; worldwide between 70 and 120 million tons is produced annually. We analyzed RM which was released in the course of the Kolontar disaster in Hungary into the environment in acute and genotoxicity experiments with plants which are widely used for environmental monitoring. We detected induction of micronuclei which reflect chromosomal damage in tetrads of Tradescantia and in root cells of Allium as well as retardation of root growth with contaminated soils and leachates. Chemical analyses showed that RM contains metals, in particular high concentrations of vanadium. Follow-up experiments indicated that vanadate causes the effects in the plants. This compound causes also in humans DNA damage and positive results were obtained in carcinogenicity studies. Since it was found also in RM from other production sites our findings indicate that its release in the environment is a global problem which should be studied in more detail. CAPSULE ABSTRACT: Our findings indicate that the red mud causes genotoxic effect in plants probably due to the presence of vanadate which is contained at high concentrations in the residue.

In situ biomonitoring of the genotoxic effects of mixed industrial emissions using the Tradescantia micronucleus and pollen abortion tests with wild life plants: demonstration of the efficacy of emission controls in an eastern European city.

Aim of the study was to monitor changes of genotoxic activity of urban air caused by an incinerator and a petrochemical plant in Tradescantia micronucleus (Trad-MCN) and pollen fertility assays with wild plants (Chelidonium majus, Clematis vitalba, Cichorium intybus, Linaria vulgaris, Robinia pseudoacacia). While in the first sampling period (1997-2000) significantly (on average 80%) more MN were found at the polluted site in comparison to controls from a rural area, no significant effects were observed during a later period (between 2003 and 2005). A similar pattern was observed in the pollen abortion assays in which the most pronounced effects were found in chicory and false acacia. The differences of the results obtained in the two periods can be explained by a substantial reduction of air pollution by use of new technologies. In particular the decrease of SO(2) emissions may account for the effects seen in the present study.

In situ monitoring of clastogenicity of ambient air in Bratislava, Slovakia using the Tradescantia micronucleus assay and pollen abortion assays.

Aim of this study was to monitor the genotoxic effects of polluted air in Bratislava (Slovakia) with the Tradescantia micronucleus (Trad-MN) test. In situ monitoring was carried out at five locations during two seasons (years 2003 and 2004). Flower pots with Tradescantia paludosa (clone 03) plants were exposed for 6-8 weeks at the different sites each year. The highest MN levels were observed in the vicinity of an agrochemical factory (3.1 times higher than background level in 2003 and 2.7 times higher in 2004). Lower effects were seen when plants were exposed to urban traffic emissions or in the vicinity of a glass-producing plant (the MN frequencies ranged between 2.8 and 4.4 per 100 tetrads, respectively, while the control frequencies were 2.1-2.6 per 100 tetrads); exposure near a petrochemical plant had no significant effects. In pollen abortion assays, three wild growing species were used, namely, chicory (Cichorium intybus L.), old man's beard (Clematis vitalba L.) and common toadflax (Linaria vulgaris Mill.). Again, the strongest effects were observed close to the agrochemical industry (reduction of fertile pollen by 5.6%, 11.1% and 8.3% in chicory, old mans beard and in toadflax, respectively). Cichorium intybus was the most sensitive species and the number of abortive pollen grains was 5.1 times higher in specimens collected near the agrochemical factory than that seen at the control location. These observations indicate that contaminated urban air has an impact on the fertility of wild plants. Furthermore, it is interesting that the same rank order of effects was seen in pollen abortion assays as in the Trad-MN test (agrochemical industry>technical glass industry≥traffic>city incinerator/petrochemical plant). These results confirm the sensitivity of the Tradescantia MN test and pollen abortion assays for the detection of air pollution, and show that distinct differences exist in genotoxicity of different sources of pollutants.