toxins Review The Diversity of Cyanobacterial Toxins on Structural Characterization, Distribution and Identification: A Systematic Review Xingde Du 1, Haohao Liu 1, Le Yuan 1, Yueqin Wang 1, Ya Ma 1, Rui Wang 1, Xinghai Chen 2, Michael D. Losiewicz 2, Hongxiang Guo 3,* and Huizhen Zhang 1,* 1 College of Public Health, Zhengzhou University, Zhengzhou 450001, China; [email protected] (X.D.); [email protected] (H.L.); [email protected] (L.Y.); [email protected] (Y.W.); [email protected] (Y.M.); [email protected] (R.W.) 2 Department of Chemistry and Biochemistry, St Mary’s University, San Antonio, TX 78228, USA; [email protected] (X.C.); [email protected] (M.D.L.) 3 College of Life Sciences, Henan Agricultural University, Zhengzhou 450002, China * Correspondence: [email protected] (H.Z.); [email protected] (H.G.); Tel.: +86-151-8835-7252 (H.Z.); +86-136-4386-7952 (H.G.) Received: 27 August 2019; Accepted: 9 September 2019; Published: 12 September 2019 Abstract: The widespread distribution of cyanobacteria in the aquatic environment is increasing the risk of water pollution caused by cyanotoxins, which poses a serious threat to human health. However, the structural characterization, distribution and identification techniques of cyanotoxins have not been comprehensively reviewed in previous studies. This paper aims to elaborate the existing information systematically on the diversity of cyanotoxins to identify valuable research avenues. According to the chemical structure, cyanotoxins are mainly classified into cyclic peptides, alkaloids, lipopeptides, nonprotein amino acids and lipoglycans. In terms of global distribution, the amount of cyanotoxins are unbalanced in different areas. The diversity of cyanotoxins is more obviously found in many developed countries than that in undeveloped countries. Moreover, the threat of cyanotoxins has promoted the development of identification and detection technology. Many emerging methods have been developed to detect cyanotoxins in the environment. This communication provides a comprehensive review of the diversity of cyanotoxins, and the detection and identification technology was discussed. This detailed information will be a valuable resource for identifying the various types of cyanotoxins which threaten the environment of different areas. The ability to accurately identify specific cyanotoxins is an obvious and essential aspect of cyanobacterial research. Keywords: cyanobacterial toxins; diversity; structural characterization; distribution; identification Key Contribution: The existing information on the diversity of cyanotoxins was reviewed and the routine and advanced detection methods of cyanotoxins were also discussed. 1. Introduction Cyanobacteria, which have existed for about 3.5 billion years, are believed to be the oldest creatures on Earth. Even though over 2600 cyanobacterial species have been described so far, it is believed that there are still many unknown species [1]. Cyanotoxins are toxic secondary metabolites produced by cyanobacteria, and pose a major threat to the ecological environment and human health. Cyanotoxins are diverse in functional properties and chemical structures. Based on functional properties, cyanotoxins can be mainly classified into hepatotoxins, neurotoxins, dermatotoxins and cytotoxins [2]. According to the chemical structure, cyanotoxins are mainly divided into cyclic peptides, alkaloids, lipopeptides, Toxins 2019, 11, 530; doi:10.3390/toxins11090530 www.mdpi.com/journal/toxins Toxins 2019, 11, 530 2 of 34 non-protein amino acids and lipoglycans. Different chemical structures of cyanotoxins contribute to the diversity of their functional properties. A number of lipopeptide cyanotoxins have been found to haveToxins peculiar 2019, 11, 530 biological activities [3,4], such as anticancer, antifungal and molluscicidal2 activities.of 36 However, there are still many cyanotoxins that are believed to have special activities that have not yet beenpeptides, identified. alkaloids, lipopeptides, non‐protein amino acids and lipoglycans. Different chemical structures of cyanotoxins contribute to the diversity of their functional properties. A number of Cyanobacterial toxins are widely distributed in fresh and saltwater, and they have been reported lipopeptide cyanotoxins have been found to have peculiar biological activities [3,4], such as hundredsanticancer, of times antifungal in various and molluscicidal countries activities. and regions However, of the there world, are still even many incyanotoxins the Arctic that and are arid desertbelieved areas to [5 ,have6]. The special extensiveness activities that of have the not distribution yet been identified. of cyanotoxins is self-evident, however, the characteristicsCyanobacterial of the toxins distribution are widely and distributed the severity in fresh of cyanobacterialand saltwater, and toxins they have vary been in di reportedfferent areas. For example,hundreds in of the times Baltic in various region, countries cyanobacterial and regions blooms of the covering world, even an area in the of 100,000Arctic and square arid desert kilometers can beareas formed [5,6]. in The the summerextensiveness [7], and of variousthe distribution cyanotoxins of cyanotoxins produced byis thisself‐ cyanobacteriaevident, however, are seriouslythe threateningcharacteristics the surrounding of the distribution countries. and the severity of cyanobacterial toxins vary in different areas. For Theexample, harm in of the cyanotoxins Baltic region, is obvious,cyanobacterial and itblooms has been covering regarded an area as of a new 100,000 public square health kilometers concern by can be formed in the summer [7], and various cyanotoxins produced by this cyanobacteria are the World Health Organization (WHO) [5]. Common methods such as bioassays, biochemical analysis, seriously threatening the surrounding countries. chemical assays and molecular assays have been used to detect cyanotoxins. However, because of the The harm of cyanotoxins is obvious, and it has been regarded as a new public health concern by diversitythe World of cyanotoxin Health Organization species and chemical(WHO) [5]. structure, Common the methods current such routine as methodsbioassays, cannot biochemical be used to detectanalysis, all the typeschemical and assays variants and ofmolecular cyanotoxins. assays Therefore, have been theused development to detect cyanotoxins. of valuable However, techniques and methodsbecause of of the detecting diversity cyanotoxins of cyanotoxin are species prerequisite and chemical for the structure, accurate the identification current routine of cyanobacterial methods toxinscannot in the be environment. used to detect all the types and variants of cyanotoxins. Therefore, the development of Forvaluable these techniques reasons, data and on methods the structural of detecting characterization cyanotoxins and are regional prerequisite distribution for the of accurate cyanotoxins over theidentification past twenty of cyanobacterial years were collected toxins in for the this environment. paper to provide a review of the cyanotoxin diversity in the world.For these We alsoreasons, summarize data on and the discuss structural the characterization protocols and advancedand regional detection distribution methods of for cyanotoxins over the past twenty years were collected for this paper to provide a review of the cyanotoxins reported in recent years. cyanotoxin diversity in the world. We also summarize and discuss the protocols and advanced 2. Structuraldetection Characterizationmethods for cyanotoxins reported in recent years. 2.1. Cyclic2. Structural Peptides Characterization A2.1. cyclic Cyclic peptide Peptides is a cyclic polypeptide chain formed by a covalent linkage between the amino termini andA cyclic the carboxyl peptide is termini, a cyclic polypeptide the amino termini chain formed and the by sidea covalent chain, linkage the carboxyl between termini the amino and the side chain,termini or and the the side carboxyl chain termini, with another the amino side termini chain. and Microcystins the side chain, (MCs) the carboxyl and nodularins termini and (NODs) the are two classicalside chain, cyclic or the peptide side chain cyanobacterial with another side toxins chain. (Figure Microcystins1). (MCs) and nodularins (NODs) are two classical cyclic peptide cyanobacterial toxins (Figure 1). FigureFigure 1. Structure 1. Structure of cyclicof cyclic peptide peptide cyanotoxins. cyanotoxins. ( (AA)) Microcystin Microcystin-LR,‐LR, YR YR and and RR, RR, (B) Nodularin. (B) Nodularin. Toxins 2019, 11, 530 3 of 34 2.1.1. Microcystins MCs are hepatotoxic cyclic heptapeptides. The general structure of a MC is cyclo (-D-Ala-L-X-D-erythro-β-methylAsp-L-Y-Adda-D-Glu-N-methyldehydro-Ala), in which Adda is the unusual C20 amino acid (3-amino-9-methoxy-2,6,8-trimethyl-10-phenyl-4,6-dienoic acid) [8]. Nuclear magnetic resonance (NMR) was used for the structure determination of MCs in the 1990s [9]. In recent years, more sensitive mass spectrometry (MS) techniques have enabled the detection of MCs in trace amounts in cyanobacterial samples. To date, 246 MC congeners have been found [10]. The most common congeners are MC-LR, MC-YR and MC-RR, resulting from the presence of the L-forms of leucine (L), tyrosine (Y) or arginine (R) in positions 2 and 4 (Figure1A). The structure diversity of MCs results in significant differences in toxicity, and the toxicity of MCs has been demonstrated in 79 countries [11]. As