Polyphasic toxicological screening of Cylindrospermopsis raciborskii and Aphanizomenon gracile isolated in Poland

Aphanizomenon gracile and Cylindrospermopsis raciborskii are extensively studied Nostocales of wide geographical distribution and have potential to produce toxins. However, a number of knowledge gaps regarding their toxicity and related health risks in certain locations, including Europe, exists. The present study applied a polyphasic approach to screen the toxicity of different strains of C. raciborskii (LBY-Cr, LBO-Cr and LKM-Cr) and A. gracile (LBY-Ag, LBN-Ag and LWI-Ag) isolated from five freshwater lakes of Western Poland. The following investigations were carried out: (i) in vitro toxicological studies employing human cells isolated from healthy donors; (ii) analytical screening for the presence of cylindrospermopsin (CYN), guanidinoacetate (GAA; initial CYN precursor and postulated general cyanobacterial metabolite), three microcystin (MC) analogues, β-N-methylamino-l-alanine (BMAA) and its isomer α-γ,-diaminobutyric acid (DAB), anatoxin-a (ATX) and ten saxitoxin (STX) analogues; and (iii) molecular studies of genes involved in CYN, GAA, MCs and ATX biosynthesis. Extracts of C. raciborskii LBY-Cr and A. gracile LBN-Ag caused a significant increase in the intracellular reactive oxygen content in human neutrophils during short-term (1 h) exposure and also led to lipid peroxidation and cell death. No cytotoxic effects were noted for the other tested strains. None of the toxin genes (cyrA, cyrJ, anaF and mcyE) and toxins (CYN, GAA, MCs, BMAA, ATX and STX) were detected. The only exception was DAB found at a concentration below 1.0 μg g− 1 dw in A. gracile LWI-Ag. It is the first time that cyanobacterial DAB producer has been identified in the Central European region. The study points to the production of as yet unknown metabolite(s) that may pose a relevant threat to human health through strains of C. raciborskii and A. gracile isolated from two Polish lakes, and adds to the general understanding of the toxicity of European strains of both species.