Abstract
The co-occurrence of various cyanobacterial toxins can potentially induce toxic effects different than those observed for single cyanotoxins, as interaction phenomena cannot be discarded. Moreover, mixtures are a more probable exposure scenario. However, toxicological information on the topic is still scarce. Taking into account the important role of mutagenicity and genotoxicity in the risk evaluation framework, the objective of this study was to assess the mutagenic and genotoxic potential of mixtures of two of the most relevant cyanotoxins, Microcystin-LR (MC-LR) and Cylindrospermopsin (CYN), using the battery of in vitro tests recommended by the European Food Safety Authority (EFSA) for food contaminants. Mixtures of 1:10 CYN/MC-LR (CYN concentration in the range 0.04–2.5 µg/mL) were used to perform the bacterial reverse-mutation assay (Ames test) in Salmonella typhimurium, the mammalian cell micronucleus (MN) test and the mouse lymphoma thymidine-kinase assay (MLA) on L5178YTk± cells, while Caco-2 cells were used for the standard and enzyme-modified comet assays. The exposure periods ranged between 4 and 72 h depending on the assay. The genotoxicity of the mixture was observed only in the MN test with S9 metabolic fraction, similar to the results previously reported for CYN individually. These results indicate that cyanobacterial mixtures require a specific (geno)toxicity evaluation as their effects cannot be extrapolated from those of the individual cyanotoxins.
Highlights
CYN/MC-LR mixtures did not induce changes in any of the S. typhimurium strains without S9 fraction (Table 1)
None of the evaluated CYN/MC-LR mixture concentrations induced a mutagenic response in the absence or presence of S9 fraction, neither after a short treatment (4 h) nor a long treatment (24 h)
Caco-2 cells exposure to CYN/MC-LR mixtures did not result in DNA strand breaks in the standard comet assay after 24 and 48 h (Figure 1a)
Summary
A proliferation of cyanobacterial species can be seen globally because of water eutrophication and climate change, leading to an increasing occurrence of cyanotoxins [1,2,3].Cyanotoxins are toxic secondary metabolites produced by various species of cyanobacteria, which involved an ample variety of compounds with different structural and physicochemical properties [4].Humans may be exposed to cyanotoxins via different routes, but oral exposure by means of contaminated water and foods (fish, crops, vegetables and food supplements) is by far the most important [5,6].Microcystins (MCs) and cylindrospermopsins (CYN) are among the most frequently investigated cyanotoxins due to their toxicity and extensive distribution.MCs are cyclic heptapeptides and 246 variants were identified so far [7], with Microcystin-LR (MC-LR) as the reference congener. A proliferation of cyanobacterial species can be seen globally because of water eutrophication and climate change, leading to an increasing occurrence of cyanotoxins [1,2,3]. Cyanotoxins are toxic secondary metabolites produced by various species of cyanobacteria, which involved an ample variety of compounds with different structural and physicochemical properties [4]. Humans may be exposed to cyanotoxins via different routes, but oral exposure by means of contaminated water and foods (fish, crops, vegetables and food supplements) is by far the most important [5,6]. Microcystins (MCs) and cylindrospermopsins (CYN) are among the most frequently investigated cyanotoxins due to their toxicity and extensive distribution. MCs are cyclic heptapeptides and 246 variants were identified so far [7], with Microcystin-LR (MC-LR) as the reference congener. The liver is the main target organ in MC-LR toxicity because of Toxins 2019, 11, 318; doi:10.3390/toxins11060318 www.mdpi.com/journal/toxins
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