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

Abstract

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.

Keywords: Artemia salina; Cyanobacteria; Daphnia magna; Hepatic cell lines; PP1 inhibition assay; Toxicity; Zebrafish embryos.