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.

Cytotoxic and genotoxic activities of waters and sediments from highway and parking lot runoffs.

The genotoxicity of water and sediment samples from stormwater treatment systems and water from urban highway runoff was tested in the Salmonella/microsome assays with Salmonella typhimurium, micronucleus assay (Trad-MN) with plants and with human-derived liver cells (HepG2), or comet assay with HepG2. Cytotoxicity of water samples was studied using either reactive oxygen species (ROS) generation, cell proliferation or dye exclusion assay in HepG2. Concentrations of several contaminants in the tested samples were also measured. Results suggested that urban highway runoff exposed to severe vehicle traffic emissions caused genotoxic effects in comet assay and in Trad-MN assays. Sediments induced either mutagenic effects in strain YG1024 or genotoxic effects in Trad-MN assay. These effects could be due to the presence of nitro-polycyclic aromatic hydrocarbons (NPAHs) which possess carcinogenic and mutagenic properties. Influent and effluents of stormwater treatment systems did not induce genotoxic activity or effects on HepG2 cell viability; however, the influents were able to induce ROS generation and cell proliferation in HepG2 cells. As the methods require a sterile filtration of the water samples, this could have also removed particulate-associated polycyclic aromatic hydrocarbons (PAHs) and resulted in a less pronounced induction of genotoxicity, as would be expected by PAH contamination.

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.

Acute toxic and genotoxic activities of widely used cytostatic drugs in higher plants: Possible impact on the environment.

Cytostatic drugs are highly toxic pharmaceuticals and it was repeatedly postulated that they may cause adverse effects in ecosystems. The acute toxic and genotoxic properties of these drugs have not been adequately investigated in higher plants so far; therefore, we studied the most widely used drugs (5-flurouracil, 5FU; etoposide, Et; cisplatin, CisPt; carboplatin, CaPt; vincristine sulfate, VinS and cyclophosphamide monohydrate, CP) in micronucleus (MN) assays with meiotic pollen tetrad cells of Tradescantia and with root cells from Allium cepa. MNi are formed as a consequence of chromosome breaks and aneuploidy. We monitored also the acute toxic properties of the drugs, i.e. inhibition of cell division (mitotic indices and retardation of root growth) in the latter species. All compounds caused in both indicator plants genotoxic effects. The order of genotoxic potencies expressed as NOELs in µM was CisPt (0.1)≥ Et (0.5)>CP (1.0)>CaPt (10)>5FU (30)>VinS (100) in Tradescantia. A similar order was seen in Allium MN but Et was less active (5.0µM). Four compounds caused alterations of the mitotic indices under the present conditions namely CisPt (0.5), Et (10.0), 5FU (10.0) and VinS (100). Inhibition of root growth decreased in the order CisPt (0.5)>Et (1.0)≥VinS (1.0)>5FU (5.0)>CaPt (33.0)>CP (>1000). Comparisons of the NOELs with the predicted environmental concentrations (PEC) show that the latter values are at least 5 orders of magnitude lower and indicate that it is unlikely that their release in the environment may cause adverse effects in higher plants. However, it is notable that the levels of both platinum compounds and of 5FU in hospital effluents may reach levels which may induce damage of the genetic material.

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.

Assessment of genotoxicity and acute toxic effect of the imatinib mesylate in plant bioassays.

Imatinib mesylate (IM) is at present one of the most widely used cytostatic drugs in developed countries but information on its ecotoxicological activities is scarce. This article describes the results of the first investigation in which genotoxic and acute toxic properties of the drug were studied in higher plants. IM was tested in two widely used plant bioassays namely in micronucleus (MN) assays with meiotic tetrad cells of Tradescantia (clone #4430) and in mitotic root tip cells of Allium cepa. Additionally, acute toxic effects (inhibition of cell division and growth of roots) were monitored in the onions. Furthermore, we studied the impact of the drug on the fertility of higher plants in pollen abortion experiments with three wildlife species (Chelidonium majus, Tradescantia palludosa and Arabidopsis thaliana). In MN assays with Tradesacantia a significant effect was seen with doses ⩾10µM; the Allium MN assay was even more sensitive (LOEL⩾1.0µM). A significant decrease of the mitotic indices was detected at levels ⩾10µM in the onions and reduction of root growth with ⩾100µM. In the pollen fertility assays clear effects were observed at doses ⩾147.3mgkg(-1). Data concerning the annual use of the drug in European countries (France, Germany, Slovenia) enable the calculation of the predicted environmental concentration (PEC) values which are in the range between 3.3 and 5.0ngL(-1). Although comparisons with the genotoxic potencies of other commonly used cytostatic drugs and with highly active heavy metal compounds show that IM is an extremely potent genotoxin in higher plants, it is evident that the environmental concentrations are ⩾5 orders of magnitude lower as the levels which are required to cause adverse effects.

Micronucleus assay with tetrad cells of Tradescantia.

The Tradescantia micronucleus assay is being used since almost 50 years for the detection of genotoxins (including carcinogens) in the environment. A large database on the effects of individual compounds and of complex environmental mixtures (soil, air and water) has accumulated. In contrast to other mutagenicity test systems, the effects of low concentrations of heavy metals, radionuclides, certain herbicides and pesticides, and gaseous mutagens can be detected and it is also possible to use the test for in situ biomonitoring studies. 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.