Safety assessment of surfactin-producing Bacillus strains and their lipopeptides extracts in vitro and in vivo.

Beneficial Bacillus strains can be administered to livestock as probiotics to improve animal health. Cyclic lipopeptides produced by Bacillus such as surfactins may be responsible for some of the beneficial effects due to their anti-inflammatory and immunomodulatory activity. The aim of the present study was to isolate and evaluate the biocompatibility of native Bacillus spp. strains and their surfactin-like lipopeptides in vitro and in vivo to determine their potential to be used on animals. Biocompatibility of endospore suspensions (108 UFC/mL), and different dilutions (1:10; 1:50; 1:100; 1:500, and 1:1000) of Bacillus lipopeptide extracts containing surfactin was tested on Caco-2 cells by microculture tetrazolium-based colorimetric assay. Genotoxicity was tested on BALB/c mice (n = 6) administered 0.2 mL of endospore suspensions by the bone marrow erythrocyte micronuclei assay. All the isolates tested produced between 26.96 and 239.97 µg mL- 1 of surfactin. The lipopeptide extract (LPE) from isolate MFF1.11 demonstrated significant cytotoxicity in vitro. In contrast, LPE from MFF 2.2; MFF 2.7, TL1.11, TL 2.5, and TC12 had no cytotoxic effect (V% > 70%) on Caco-2 cells, not affecting cell viability signifficantly in most treatments. Similarly, none of the endospore suspensions affected cell viability (V% > 80%). Likewise, endospores did not cause genotoxicity on BALB/c mice. This study was elementary as a first step for a new line of research, since it allowed us to choose the safest isolates to keep working on the search of new potentially probiotic strains destined to production animals to improve their performance and health.

Towards the Therapeutic Use of Thrombospondin 1/CD47 Targeting TAX2 Peptide as an Antithrombotic Agent.

OBJECTIVE: TSP-1 (thrombospondin 1) is one of the most expressed proteins in platelet α-granules and plays an important role in the regulation of hemostasis and thrombosis. Interaction of released TSP-1 with CD47 membrane receptor has been shown to regulate major events leading to thrombus formation, such as, platelet adhesion to vascular endothelium, nitric oxide/cGMP (cyclic guanosine monophosphate) signaling, platelet activation as well as aggregation. Therefore, targeting TSP-1:CD47 axis may represent a promising antithrombotic strategy. Approach and Results: A CD47-derived cyclic peptide was engineered, namely TAX2, that targets TSP-1 and selectively prevents TSP-1:CD47 interaction. Here, we demonstrate for the first time that TAX2 peptide strongly decreases platelet aggregation and interaction with collagen under arterial shear conditions. TAX2 also delays time for complete thrombotic occlusion in 2 mouse models of arterial thrombosis following chemical injury, while Thbs1-/- mice recapitulate TAX2 effects. Importantly, TAX2 administration is not associated with increased bleeding risk or modification of hematologic parameters. CONCLUSIONS: Overall, this study sheds light on the major contribution of TSP-1:CD47 interaction in platelet activation and thrombus formation while putting forward TAX2 as an innovative antithrombotic agent with high added-value.

Cytotoxic and immunological responses of fish leukocytes to nodularin exposure in vitro.

Nodularin (NOD) is a cyclic peptide released by bloom-forming toxic cyanobacteria Nodularia spumigena commonly occurring in brackish waters throughout the world. Although its hepatotoxic effects are well known, other negative effects of NOD have not yet been completely elucidated. The present study aims were to evaluate and compare the cytotoxic and immunotoxic effects of the toxin on primary leukocytes (from head kidney [HK]) and stable fish leukocytes (carp leucocyte cell line [CLC] cells). The cells were incubated with the cyanotoxin at concentrations of 0.001, 0.01, 0.05, or 0.1 µg/ml. After 24 h of exposure, the concentrations ≥0.05 µg/ml of toxin resulted in cytotoxicity in the primary cells, while in CLC cells, the toxic effect was obtained only with the highest concentration. Similarly, depending on the concentration, exposure to NOD causes a significant inhibition of chemotaxis of the phagocytic abilities of primary leukocytes and a significant reduction in the proliferation of lymphocytes isolated from the HKs. Moreover, CLC cells and HK leukocytes incubated with this toxin at all the mentioned concentrations showed an increased production of reactive oxygen and nitrogen species. NOD also evidently influenced the expression of genes of cytokine TNF-α and IL-10 and, to a minor extent, IL-1ß and TGF-ß. Notably, the observed changes in the mRNA levels of cytokines in NOD-exposed cells were evident, but not clearly dose-dependent. Interestingly, NOD did not affect the production and release of IL-1ß of the CLC cells. This study provides evidence that NOD may exert cytotoxicity and immune-toxicity effects depending on cell type and toxin concentration.

Malformin C, an algicidal peptide from marine fungus Aspergillus species.

A natural compound with the algicidal effect was isolated from the culture medium of Aspergillus sp. SCSIOW2 and was identified as malformin C, which was based on the data of 1H-NMR, 13C-NMR, and ESI-MS. Malformin C exhibited dose-dependent algicidal activities against two strains of noxious red tide algae, Akashiwo sanguinea and Chattonella marina. The activity against A. sanguinea was stronger than that against C. marina (the algicidal activity of 58 and 36% at 50 µM treatment for 2 h, respectively). Morphology changes including perforation, plasmolysis, and fragmentation of algal cells were observed. Malformin C induced a significant increase in ROS level, caused the damage of SOD activity, and led to the massive generation of MDA contents in algae cells. To our knowledge, this is the first report of the cyclic peptide described as an algicidal compound against HABs.

Maculosin, a non-toxic antioxidant compound isolated from Streptomyces sp. KTM18.

CONTEXT: Streptomyces species are prolific sources of bioactive secondary metabolites known especially for their antimicrobial and anticancer activities. OBJECTIVE: This study sought to isolate and characterize antioxidant molecules biosynthesized by Streptomyces sp. KTM18. The antioxidant potential of an isolated compound and its toxicity were accessed. MATERIALS AND METHODS: The compound was purified using bioassay-guided chromatography techniques. Nuclear magnetic resonance (NMR) experiments were carried out for structure elucidation. The antioxidant potential of the isolated compound was determined using DPPH free radical scavenging assay. The toxicity of the isolated compound was measured using a brine shrimp lethality (BSL) assay. RESULTS: Ethyl acetate extract of Streptomyces sp. KTM18 showed more than 90% inhibition of DPPH free radical at 50 µg/mL of the test concentration. These data were the strongest among 13 Streptomyces isolates (KTM12-KTM24). The active molecule was isolated and characterized as maculosin (molecular formula, C14H16N2O3 as determined by the [M + H]+ peak at 261.1259). The DPPH free radical scavenging activity of pure maculosin was higher (IC50, 2.16 ± 0.05 µg/mL) than that of commercial butylated hydroxyanisole (BHA) (IC50, 4.8 ± 0.05 µg/mL). No toxicity was observed for maculosin (LD50, <128 µg/mL) in brine shrimp lethality assay (BSLA) up to the compound's antioxidant activity (IC50) concentration range. The commercial standard, berberine chloride, showed toxicity in BSLA with an LD50 value of 8.63 ± 0.15 µg/mL. CONCLUSIONS: Maculosin may be a leading drug candidate in various cosmetic and therapeutic applications owing to its strong antioxidant and non-toxic properties.

A Convolutional Neural Network-Based Approach for the Rapid Annotation of Molecularly Diverse Natural Products.

This report describes the first application of the novel NMR-based machine learning tool "Small Molecule Accurate Recognition Technology" (SMART 2.0) for mixture analysis and subsequent accelerated discovery and characterization of new natural products. The concept was applied to the extract of a filamentous marine cyanobacterium known to be a prolific producer of cytotoxic natural products. This environmental Symploca extract was roughly fractionated, and then prioritized and guided by cancer cell cytotoxicity, NMR-based SMART 2.0, and MS2-based molecular networking. This led to the isolation and rapid identification of a new chimeric swinholide-like macrolide, symplocolide A, as well as the annotation of swinholide A, samholides A-I, and several new derivatives. The planar structure of symplocolide A was confirmed to be a structural hybrid between swinholide A and luminaolide B by 1D/2D NMR and LC-MS2 analysis. A second example applies SMART 2.0 to the characterization of structurally novel cyclic peptides, and compares this approach to the recently appearing "atomic sort" method. This study exemplifies the revolutionary potential of combined traditional and deep learning-assisted analytical approaches to overcome longstanding challenges in natural products drug discovery.

Modulation of nodularin toxicity in shrimp Litopenaeus vannamei (BOONE, 1931) fed with dietary açai (Euterpe oleracea) inclusion.

This study evaluated the effect of dietary inclusion of lyophilized açaí Euterpe oleracea (LEO) on redox status of shrimp Litopenaeus vannamei (initial weight 1.5 ± 0.39 g) upon exposure to cyanotoxin nodularin (NOD) in bioflocs system. Three hundred juvenile shrimps were randomly divided into two groups and fed twice a day with two diets: one containing 0.00 (control diet) and the other 10.0% LEO (w/w) for 30-days. After the feeding period, both shrimp groups were submitted to three treatments (14 L; 7 shrimp/tank) with different concentrations of cyanotoxin NOD (0.00; 0.25; and 1.00 µg/L) dissolved in water with 96 h of exposure. Then, the shrimps were sampled (n = 15/treatment) for the determination of reduced glutathione (GSH), the activity of glutathione-S-transferase (GST), sulfhydryl groups associated to proteins (P-SH), and lipid peroxidation (TBARS) in the hepatopancreas, gills and muscle. The NOD accumulation was measured in the muscle. The results revealed that dietary LEO significantly increased GSH levels in the hepatopancreas and gills of the shrimps exposed to NOD. Toxin exposure did not modify GST activity in all organs. Muscle TBARS levels were lower in the shrimp fed with the LEO diet and exposed to NOD. The NOD toxin did not accumulate in the muscle but notably was detected in the control groups fed or not with dietary LEO. Açaí was able to induce the antioxidant system of L. vannamei, as well as lowered the oxidative damage in shrimps exposed to NOD, suggesting its use as a chemoprotectant against cyanotoxins.

Nodularin induced oxidative stress contributes to developmental toxicity in zebrafish embryos.

Nodularin (NOD) is a kind of cyanobacterial toxins. It is of concern due to elicit severe genotoxicity in humans and animals. The comprehensive evaluation of NOD-induced adverse effects in living organisms is urgently needed. This study is aimed to report the developmental toxicity and molecular mechanism using zebrafish embryos exposed to NOD. The embryonic toxicity induced by NOD is demonstrated by inhibition of embryo hatching, increase in mortality rate, abnormal heart rate, embryonic malformation as well as defects in angiogenesis and common cardinal vein remodeling. NOD triggered a decreased rate of angiogenesis through inhibiting endothelial cells migration. NOD induced embryonic cell apoptosis and DNA damage, which can be alleviated by antioxidant N-acetyl-L-cysteine. NOD significantly caused oxidative damage as indicated by changes in reactive oxygen species, superoxide dismutase, catalase, glutathione and malondialdehyde. NOD also altered the expression of vascular development-genes (DLL4, CDH5, VEGFA, VEGFC) and apoptosis-related genes (BAX, BCL-2, P53, CASPASE 3). Taken together, NOD induced adverse effect on zebrafish embryos development, which may be associated with oxidative stress and apoptosis through the activation of P53-BAX/BCL-2-CASPASE 3-mediated pathway.

Development of Zwitterionic Polypeptide Nanoformulation with High Doxorubicin Loading Content for Targeted Drug Delivery.

Much attention has been drawn to targeted nanodrug delivery systems due to their high therapeutic efficacy in cancer treatment. In this work, doxorubicin (DOX) was incorporated into a zwitterionic arginyl-glycyl-aspartic acid (RGD)-conjugated polypeptide by an emulsion solvent evaporation technique with high drug loading content (45%) and high drug loading efficiency (95%). This zwitterionic nanoformulation showed excellent colloidal stability at high dilution and in serum. The pH-induced disintegration and enzyme-induced degradation of the nanoformulation were confirmed by dynamic light scattering and gel permeation chromatography. Efficient internalization of DOX in the cells and high antitumor activity in vitro was observed. Compared with the free drug, this nanoformulation showed higher accumulation in tumor and lower systemic toxicity in vivo. The DOX-loaded zwitterionic RGD-conjugated polypeptide vesicles show potential application for targeted drug delivery in the clinic.

Preclinical evidences of safety of a new synthetic adjuvant to formulate with the influenza human vaccine: absence of subchronic toxicity and mutagenicity.

Context: Influenza is a severe, life-threatening viral disease that can be prevented by vaccination. However, the anti-influenza human vaccine failed to show the required efficacy both in infants under 5 years old and in the elder population, who are among those with the highest risk of developing severe complications after influenza infection. Therefore, it is of high importance to improve the vaccine efficacy and ensure its safety in these susceptible populations. GK-1, a novel 18-aa peptide adjuvant, has been proved to increase the immunogenicity of the human influenza vaccine in both young and aged mice. Objective: A preclinical study of the toxicity profile of GK-1 following the World Health Organization guidelines to support its use was herein conducted. Material and methods: GK-1 was synthetically produced following Good Manufacturing Practices. The toxicological evaluation of GK-1 peptide was performed in rats after repeated dose-ranging trials by the subcutaneous route. The mutagenic potential of GK-1 was assessed by the micronucleus, chromosomal aberration, and Ames tests, in accordance with OECD Guidelines. Results: GK-1 did not show toxic effects at doses up to 12.5mg/kg, corresponding to 25 times the dose intended for human use. No indications of mutagenic potential were observed. GK-1 after dermal administration was well tolerated locally. Conclusion: The efficacy of GK-1 to improve influenza vaccine protection, along with the absence of toxicity and mutagenicity, as reported herein, support the evaluation of this peptide in a clinical trial as a novel adjuvant for human use.

Cell-Permeable Bicyclic Peptidyl Inhibitors against NEMO-IκB Kinase Interaction Directly from a Combinatorial Library.

Macrocyclic peptides are capable of binding to flat protein surfaces such as the interfaces of protein-protein interactions with antibody-like affinity and specificity, but generally lack cell permeability in order to access intracellular targets. In this work, we designed and synthesized a large combinatorial library of cell-permeable bicyclic peptides, in which the first ring consisted of randomized peptide sequences for potential binding to a target of interest, while the second ring featured a family of different cell-penetrating motifs, for both cell penetration and target binding. The library was screened against the IκB kinase α/ß (IKKα/ß)-binding domain of NF-κB essential modulator (NEMO), resulting in the discovery of several cell-permeable bicyclic peptides, which inhibited the NEMO-IKKß interaction with low µM IC50 values. Further optimization of one of the hits led to a relatively potent and cell-permeable NEMO inhibitor (IC50 = 1.0 µM), which selectively inhibited canonical NF-κB signaling in mammalian cells and the proliferation of cisplatin-resistant ovarian cancer cells. The inhibitor provides a useful tool for investigating the biological functions of NEMO/NF-κB and a potential lead for further development of a novel class of anti-inflammatory and anticancer drugs.

Safety profile after prolonged C3 inhibition.

The central component of the complement cascade, C3, is involved in various biological functions, including opsonization of foreign bodies, clearance of waste material, activation of immune cells, and triggering of pathways controlling development. Given its broad role in immune responses, particularly in phagocytosis and the clearance of microbes, a deficiency in complement C3 in humans is often associated with multiple bacterial infections. Interestingly, an increased susceptibility to infections appears to occur mainly in the first two years of life and then wanes throughout adulthood. In view of the well-established connection between C3 deficiency and infections, therapeutic inhibition of complement at the level of C3 is often considered with caution or disregarded. We therefore set out to investigate the immune and biochemical profile of non-human primates under prolonged treatment with the C3 inhibitor compstatin (Cp40 analog). Cynomolgus monkeys were dosed subcutaneously with Cp40, resulting in systemic inhibition of C3, for 1 week, 2 weeks, or 3 months. Plasma concentrations of both C3 and Cp40 were measured periodically and complete saturation of plasma C3 was confirmed. No differences in hematological, biochemical, or immunological parameters were identified in the blood or tissues of animals treated with Cp40 when compared to those injected with vehicle alone. Further, skin wounds showed no signs of infection in those treated with Cp40. In fact, Cp40 treatment was associated with a trend toward accelerated wound healing when compared with the control group. In addition, a biodistribution study in a rhesus monkey indicated that the distribution of Cp40 in the body is associated with the presence of C3, concentrating in organs that accumulate blood and produce C3. Overall, our data suggest that systemic C3 inhibition in healthy adult non-human primates is not associated with a weakened immune system or susceptibility to infections.

Bacicyclin, a new antibacterial cyclic hexapeptide from Bacillus sp. strain BC028 isolated from Mytilus edulis.

A new cyclic hexapeptide, cyclo-(Gly-Leu-Val-IIe-Ala-Phe), named bacicyclin (1), was isolated from a marine Bacillus sp. strain associated with Mytilus edulis. The sequences of the amino acid building blocks of the cyclic peptide and its structure were determined by 1D- and 2D-NMR techniques. Marfey's analysis showed that the amino acid building blocks had L-configuration in all cases except for alanine and phenylalanine, which had D-configuration. Bacicyclin (1) exhibited antibacterial activity against the clinically relevant strains Enterococcus faecalis and Staphylococcus aureus with minimal inhibitory concentration values of 8 and 12 µM, respectively. These results demonstrate the potential of marine bacteria as a promising source for the discovery of new antibiotics.

Toxicity and applications of surfactin for health and environmental biotechnology.

Characterized as one of the most potent biosurfactants, surfactin is a cyclic lipopeptide synthesized by several strains of Bacillus genus. The aim of this review was to present the physicochemical and structural properties of surfactin and to demonstrate advances and applications of this biosurfactant for health and environmental biotechnology. Further, this review also focused on toxicological effects of surfactin on in vivo and in in vitro systems. The hydrophobic nature of surfactin enables interaction with membrane-bound phospholipids and indicates the ability of the molecule to act as a new weapon with respect to therapeutic and environmental properties. Seeking to avoid environmental contamination produced by widespread use of synthetic surfactants, surfactin emerges as a biological control agent against pathogen species owing to its antibacterial and antiviral properties. In addition, the mosquitocidal activity of surfactin was suggested as new strategy to control disease vectors. The current findings warrant future research to assess the toxicity of surfactin to enable an optimizing anticancer therapy and to seek refined methodologies, including nanotechnology techniques, to allow for an improved delivery of the biogenic molecule on target cells.

Specific Chemical and Genetic Markers Revealed a Thousands-Year Presence of Toxic Nodularia spumigena in the Baltic Sea.

In the Baltic Sea, diazotrophic cyanobacteria have been present for thousands of years, over the whole brackish water phase of the ecosystem. However, our knowledge about the species composition of the cyanobacterial community is limited to the last several decades. In the current study, the presence of species-specific chemical and genetic markers in deep sediments were analyzed to increase the existing knowledge on the history of toxic Nodularia spumigena blooms in the Baltic Sea. As chemical markers, three cyclic nonribosomal peptides were applied: the hepatotoxic nodularin, which in the sea was detected solely in N. spumigena, and two anabaenopeptins (AP827 and AP883a) characteristic of two different chemotypes of this species. From the same sediment samples, DNA was isolated and the gene involved in biosynthesis of nodularin, as well as the phycocyanin intergenic spacer region (PC-IGS), were amplified. The results of chemical and genetic analyses proved for the first time the thousands-year presence of toxic N. spumigena in the Baltic Sea. They also indicated that through all this time, the same two sub-populations of the species co-existed.

A new cyclic lipopeptide isolated from Bacillus amyloliquefaciens HAB-2 and safety evaluation.

Bacillus is the most widely studied biocontrol agent and has been extensively used in the development of biopesticides and fungicides. In this study, a new cyclic lipopeptide was isolated from Bacillus amyloliquefaciens HAB-2 by column chromatography on silica gel and Sephadex LH-20, and its structures was elucidated on the basis of spectroscopic analysis. This compound is a bacillomycin d-like compound, named as bacillomycin DC. The activity of bacillomycin DC was evaluated against C. gloeosporioides Penz. The median inhibitory concentration of bacillomycin DC was 1.21µg/mL. In addition, bacillomycin DC may have low toxicity to aquatic species as indicated by its 96h half maximum lethal concentration of 22.20µg/mL to zebrafish (Danio rerio). Our current study further provides evidence that bacillomycin DC is a potent fungicide against C. gloeosporioides Penz.

Protective effect of Gelofusine against cRGD-siRNA-induced nephrotoxicity in mice.

Based on successful targeting to the αvß3 integrin of cyclic arginine-glycine-aspartic acid (cRGD), cRGD-conjugated small interfering RNA (siRNA) exhibits tumor targeting and has become a new treatment strategy for solid tumors. However, the nephrotoxicity caused by its renal retention limits its clinical application. Here, we evaluated the protective effect of Gelofusine against cRGD-conjugated siRNA-induced nephrotoxicity in mice. Male Kunming mice (six per group) were either co-injected with Gelofusine and cRGD-siRNA or injected with cRGD-siRNA alone. After administration of these treatments five times, creatinine and blood urea nitrogen (BUN) levels were determined. Hematoxylin-eosin staining (HE staining) and transferase dUTP nick end labeling (TUNEL) analysis were used to compare the difference in renal damage between the groups. Additionally, fluorescence imaging was used to observe the distribution of cRGD-siRNA in vivo. The group co-injected with Gelofusine and cRGD-siRNA displayed lower creatinine and BUN levels than the cRGD-siRNA-alone group and showed less renal damage upon HE staining and TUNEL analysis. Gelofusine decreased the retention time and accelerated the elimination of cRGD-siRNA from the organs, as observed in the fluorescence images. These data indicate that Gelofusine significantly increased the excretion of cRGD-conjugated siRNA and reduced the associated renal damage.

mZD7349 peptide-conjugated PLGA nanoparticles directed against VCAM-1 for targeted delivery of simvastatin to restore dysfunctional HUVECs.

Endothelial dysfunction is initial and critical step of atherosclerosis. Impaired bioavailability of endothelial nitric oxide synthase (eNOS) is one of the main reasons of endothelial dysfunction. Improving bioavailability of eNOS by increasing its expression or activity using statins is an effective therapeutic strategy in restoring endothelial dysfunction. In this study, simvastatin (SIM) as a poorly water-soluble drug was loaded in poly (lactic-co-glycolic acid) (PLGA) nanoparticles (SIM-PLGA-NPs). NPs were then conjugated with mZD7349 peptide (mZD7349-SIM-PLGA-NPs) and directed against vascular cell adhesion molecule 1 (VCAM-1). In vitro evaluation of the NPs for targeted delivery of SIM was performed on activated Human Umbilical Cord Vascular Endothelial Cells (HUVECs) by tumor necrosis factor alpha (TNF-α). Effect of mZD7349-SIM-PLGA-NPs and SIM-PLGA-NPs was compared on eNOS phosphorylation (ser-1177). Results of western blot showed SIM post-treatment increased significantly phosphor-eNOS (Ser1177) expression but no total eNOS expression. The study showed that mZD7349-SIM-PLGA-NPs have particle size, zeta potential value, polydispersity index (PDI) and encapsulation efficacy % of 233±18nm, -9.6±1.1mV, 0.59±0.066 and 69±17.3%, respectively. Also phosphor-eNOS (Ser1177) expression in activated HUVECs treated with mZD7349-SIM-PLGA-NPs was significantly (p<0.05) better than treated cells with SIM-PLGA-NPs. The results suggest that mZD7349-SIM-PLGA-NPs may be usable as an appropriate drug carrier for restoring endothelial dysfunction.

Synthesis of octapeptin C4 and biological profiling against NDM-1 and polymyxin-resistant bacteria.

The first synthesis of octapeptin C4 was achieved using a combination of solid phase synthesis and off-resin cyclisation. Octapeptin C4 displayed antibiotic activity against multi-drug resistant, NDM-1 and polymyxin-resistant Gram-negative bacteria, with moderate activity against Staphylococcus aureus. The linear analogue of octapeptin C4 was also prepared, which showed reduced activity.

Cyanotoxins: producing organisms, occurrence, toxicity, mechanism of action and human health toxicological risk evaluation.

Cyanobacteria were present on the earth 3.5 billion years ago; since then they have colonized almost all terrestrial and aquatic ecosystems. They produce a high number of bioactive molecules, among which some are cyanotoxins. Cyanobacterial growth at high densities, forming blooms, is increasing in extension and frequency, following anthropogenic activities and climate changes, giving rise to some concern for human health and animal life exposed to cyanotoxins. Numerous cases of lethal poisonings have been associated with cyanotoxins ingestion in wild animal and livestock. In humans few episodes of lethal or severe human poisonings have been recorded after acute or short-term exposure, but the repeated/chronic exposure to low cyanotoxin levels remains a critical issue. The properties of the most frequently detected cyanotoxins (namely, microcystins, nodularins, cylindrospermopsin and neurotoxins) are here critically reviewed, describing for each toxin the available information on producing organisms, biosynthesis/genetic and occurrence, with a focus on the toxicological profile (including kinetics, acute systemic toxicity, mechanism and mode of action, local effects, repeated toxicity, genotoxicity, carcinogenicity, reproductive toxicity; human health effects and epidemiological studies; animal poisoning) with the derivation of health-based values and considerations on the risks for human health.