Analytical Methods for Anatoxin-a Determination: A Review.

Anatoxin-a (ATX-a) is a potent neurotoxin produced by several species of cyanobacteria whose exposure can have direct consequences, including neurological disorders and death. The increasing prevalence of harmful cyanobacterial blooms makes the detection and reliable assessment of ATX-a levels essential to prevent the risk associated with public health. Therefore, the aim of this review is to compile the analytical methods developed to date for the detection and quantification of ATX-a levels alone and in mixtures with other cyanotoxins and their suitability. A classification of the analytical methods available is fundamental to make an appropriate choice according to the type of sample, the equipment available, and the required sensitivity and specificity for each specific purpose. The most widely used detection technique for the quantification of this toxin is liquid chromatography-tandem mass spectrometry (LC-MS/MS). The analytical methods reviewed herein focus mainly on water and cyanobacterial samples, so the need for validated analytical methods in more complex matrices (vegetables and fish) for the determination of ATX-a to assess dietary exposure to this toxin is evidenced. There is currently a trend towards the validation of multitoxin methods as opposed to single-ATX-a determination methods, which corresponds to the real situation of cyanotoxins' confluence in nature.

In Vitro Mutagenic and Genotoxic Assessment of Anatoxin-a Alone and in Combination with Cylindrospermopsin.

Anatoxin-a (ATX-a) is a cyanobacterial toxin whose occurrence has been reported worldwide and has attracted increasing scientific interest due to its toxicity. Moreover, in nature, ATX-a usually appears together with other cyanotoxins, such as cylindrospermopsin (CYN), so possible interaction phenomena could happen and should be considered for risk assessment purposes. For this reason, the aim of this work was to explore the potential mutagenicity and genotoxicity of pure ATX-a and an ATX-a/CYN mixture using a battery of in vitro assays, including the bacterial reverse-mutation assay in Salmonella typhimurium (OECD 471) and the micronucleus test (MN) (OECD 487) on L5178Y Tk+/- cells. The results showed that ATX-a was not mutagenic either alone or in combination with CYN under the conditions tested. Nevertheless, genotoxic effects were observed for both ATX-a and its mixture with CYN following the in vitro MN assay. The genotoxicity exhibited by ATX-a was only observed in the absence of S9 mix, whereas in the cyanotoxin mixture the concentration-dependent genotoxicity of ATX-a/CYN in vitro was observed only in the presence of S9. Thus, the toxicity induced by cyanotoxin mixtures may vary from that produced by toxins alone, and consequently more studies are necessary in order to perform more realistic risk assessments.

Toxic Effects Produced by Anatoxin-a under Laboratory Conditions: A Review.

The presence of cyanotoxins and its bioaccumulation in the food chain is an increasingly common problem worldwide. Despite the toxic effects produced by Anatoxin-a (ATX-a), this neurotoxin has been less studied compared to microcystins (MCs) and cylindrospermopsin (CYN). Studies conducted under laboratory conditions are of particular interest because these provide information which are directly related to the effects produced by the toxin. Currently, the World Health Organization (WHO) considers the ATX-a toxicological database inadequate to support the publication of a formal guideline reference value. Therefore, the aim of the present work is to compile all of the in vitro and in vivo toxicological studies performed so far and to identify potential data gaps. Results show that the number of reports is increasing in recent years. However, more in vitro studies are needed, mainly in standardized neuronal cell lines. Regarding in vivo studies, very few of them reflect conditions occurring in nature and further studies with longer periods of oral exposure would be of interest. Moreover, additional toxicological aspects of great interest such as mutagenicity, genotoxicity, immunotoxicity and alteration of hormonal balance need to be studied in depth.