Evaluation of cyanobacterial toxicity using different biotests and protein phosphatase inhibition assay.

Cyanobacteria are prolific producers of numerous toxic compounds, among which microcystins (hepatotoxins) are the most frequently found. Cyanobacterial bloom in freshwaters is an increasing problem, and there is still a need for rapid and reliable methods for the detection of toxic cyanobacterial samples. In the present study, the toxicity of crude extracts of 11 cyanobacterial strains from different genera has been assessed on two cell lines (human hepatocellular carcinoma HepG2 and rainbow trout (Oncorhynchus mykiss) liver-derived RTL-W1 cells), crustaceans (Daphnia magna and Artemia salina), and zebrafish (Danio rerio) embryos, as well as by protein phosphatase 1 (PP1) inhibition assay and ELISA test to determine whether the toxicity could be due to the presence of hepatotoxins/microcystins. All the tested strains exhibited toxicity on HepG2 cell line (IC50 from 35 to 702 µg mL-1), including Arthrospira (Spirulina) strains, while toxicity against the RTL-W1 cells was detected only in the positive reference Microcystis PCC 7806 and Nostoc 2S9B. Tested strains expressed higher toxicity to D. magna and zebrafish embryos in comparison to A. salina, whereby Nostoc LC1B and Nostoc S8 belonged to the most toxic strains. The PP1-inhibiting compounds have been detected by PP1 assay only in four strains (Microcystis PCC 7806, Oscillatoria K3, Nostoc LC1B, and Nostoc S8), indicating that their toxic potency can be attributed to these compounds. On the other hand, very low levels of microcystins, as confirmed by ELISA, were insufficient to explain toxicity and different toxic potencies of tested cyanobacteria. Results presented in this study suggested HepG2 cell line as a particularly suitable model for cyanobacterial toxicity assessment. In addition, they highlight terrestrial cyanobacterial strains as potent producers of toxic compounds.

Effects of the technical ingredient clomazone and its two formulated products on aquatic macrophytes.

One active ingredient can be a component of different types of formulations of pesticides, while the toxicity of its formulations may vary depending on various constituents used in the mixture. The present study focuses on evaluating the effects of the active ingredient clomazone and its formulations (Rampa® EC and GAT Cenit 36 CS, both containing 360 g a.i./l of clomazone) on non-target aquatic macrophytes. The two formulation types differ in their active ingredient release and presumed environmental impact. In order to cover different ecological traits, two species of aquatic macrophytes - the floating monocot Lemna minor and the rooted dicot Myriophyllum aquaticum, were used as test models. The results of this study revealed differences in the sensitivity of tested plants to clomazone. Based on the most sensitive parameters, M. aquaticum proved to be more sensitive than L. minor to the technical ingredient and both formulations. The species sensitivity distribution (SSD) approach that was tried out in an attempt to create a higher tier step of risk assessment of clomazone for primary producers indicates that tests on rooted macrophytes can add value in risk assessment of plant protection products. The capsule formulation of clomazone was less toxic than the emulsion for L. minor, but more toxic for M. aquaticum. The most toxic for L. minor was the emulsifiable concentrate formulation Rampa® EC, followed by technical clomazone (EC50 33.3 and 54.0 mg a.i./l, respectively), while the aqueous capsule suspension formulation GAT Cenit 36 CS did not cause adverse effects. On the other hand, the most toxic for M. aquaticum was the formulation GAT Cenit 36 CS, followed by technical clomazone and the formulation Rampa® EC, demonstrating a greater effect of the capsule formulation.

The impact of humic acid on toxicity of individual herbicides and their mixtures to aquatic macrophytes.

This study investigates the impact of humic acid (HA) on the toxicity of selected herbicides and their binary mixtures to aquatic plants. The focus was on two auxin simulators (2,4-D and dicamba) and two photosynthetic inhibitors (atrazine and isoproturon). The results suggested that the addition of HA to the standard synthetic medium does not affect Lemna minor growth nor the toxicity of atrazine, but increases the toxicity of 2,4-D and the binary mixture of atrazine and 2,4-D. The addition of HA to the standard synthetic medium reversibly decreased the growth (biomass) of Myriophyllum aquaticum and enhanced the toxicity of individually tested herbicides (isoproturon and dicamba) as well as their binary mixture. The results showed delayed toxic effects of auxin simulators, especially 2,4-D in the Lemna test. The recovery after the exposure to individual photosystem II inhibitors (atrazine and isoproturon) is fast in both plant species, regardless of the presence of HA. In the case of selected mixtures (atrazine + 2,4-D and isoproturon + dicamba), recovery of both plant species was noted, while the efficiency depended on the herbicide concentration in the mixture rather than the presence or absence of HA.

Comparative analyses of cellular physiological responses of non-target species to cypermethrin and its formulated product: Contribution to mode of action research.

Physiological responses of bacterial, fish, rat and human hepatoma cells to the technical cypermethrin (AS), cypermethrin-based plant protection product (PPP), and the major co-formulant (solvent) were compared. The endpoints included: bioluminescence, total protein content, activity of mitochondrial dehydrogenase and cytochrome P450 (CYP) enzymes CYP1A and CYP1B, and expression of several genes encoding different CYP enzyme isoforms. Toxicity of PPP was compared with the toxicity predicted using concentration addition model. Cypermethrin disturbs the activity of mitochondrial dehydrogenase. Induction of CYP1A1-, CYP1A2- and CYP1B1-associated activity was more pronounced in PPP than in cypermethrin treatment. The predominant biotransformation pathway of cypermethrin is related to Cyp3a1 induction. Deviations between observed and predicted toxicity of PPP indicate synergistic effects of cypermethrin and a solvent. In vitro cellular assays may serve as rapid pre-screening tool and provide for a good indication of mixture effects and prompt further in vivo testing of PPPs when really needed.