Toxin Biosynthesis Research Articles

First report of cyanobacterial paralytic shellfish toxin biosynthesis genes and paralytic shellfish toxin production in Polish freshwater lakes

<p>In central and southern Europe, <em>Aphanizomenon</em> spp., <em>A. gracile</em> Lemmermann in particular, have been associated with paralytic shellfish toxin (PST) production. In western Poland, <em>A. gracile</em> is very common, and <em>Cylindrospermopsis raciborskii</em> (Woloszyńska) Seenayya & Subba Raju, another potentially PST-producing species, is often found as well. To date it is, however, unknown if the cyanobacterial populations in this area harbour the genetic capability to produce PSTs, and to what extent toxin biosynthesis occurs. The objective of this study was to survey the prevalence of potentially PST-producing cyanobacteria by measuring paralytic shellfish toxin biosynthesis gene <em>sxtB</em> copy numbers, <em>sxtA</em>, <em>sxtG</em> and <em>sxtS</em> gene presence, and PST concentrations in Polish freshwater lakes. In total, 34 lakes in western Poland were sampled twice during summer 2010. The presence of PST biosynthesis genes <em>sxtA</em>, <em>sxtG</em> and <em>sxtS</em> was determined using conventional qualitative PCR. Quantitative PCR (qPCR) was used to measure <em>sxtB </em>copy numbers, and the samples were analysed for PSTs using ion-pair high performance liquid chromatography with post-column oxidation and fluorescence detection (HPLC-FLD). Cyanobacteria carrying the <em>sxtB</em> gene were present in 23.5% of all samples (n=16) and in 14 lakes of the studied 34. Gene copy numbers ranged from 8.2×10<sup>4</sup> to 5.1×10<sup>7</sup> <em>sxtB</em> copies L<sup>-1</sup> (mean 3.8×10<sup>6</sup>). The median was 4.5×10<sup>5</sup> <em>sxtB</em> gene copies L<sup>-1</sup> and the majority of results clustered at the lower end of the <em>sxtB</em> qPCR linear range. In 12 out of the 16 samples positive for <em>sxtB</em> the gene co-occurred with the other three targeted PST biosynthesis genes <em>sxtA</em>, <em>sxtG</em> and <em>sxtS</em>. However, five additional samples lacked one or two of the targeted four genes. Thirteen samples contained PSTs, of which 12 samples at levels <0.072 µg L<sup>-1</sup>, <em>i.e.</em>, close to or below the quantitative detection limit of the HPLC-FLD method (0.01 µg L<sup>-1</sup>). One sample contained 0.57 µg L<sup>-1</sup> saxitoxin, co-occurring with all four <em>sxt</em> genes studied. No correlation between PST and <em>sxt</em> gene occurrence or copy numbers was observed. <em>A. gracile</em> and <em>C. raciborskii</em> occurred in 92% and 50% of samples, respectively, containing PSTs, <em>sxt</em> genes or both. In conclusion, the results confirm that potential PST producers constitute an established subpopulation of cyanobacteria in Polish freshwater lakes. However, none of the <em>sxt</em> genes targeted in this study could serve as a reliable marker for active PST biosynthesis.</p>

Lipidomics of human umbilical cord serum: identification of unique sterol sulfates.

Aim:There are currently limited lipidomics data for human umbilical cord blood. Therefore, the lipidomes of cord sera from six newborns and sera from six nonpregnant females were compared.Materials & methods:Sera lipidomics analyses were conducted using a high-resolution mass spectrometry analytical platform.Results:Cord serum contained a diverse array of glycerophospholipids, albeit generally at lower concentrations than monitored in adult serum. The unexpected observations were that cord serum contained several neurosteroid sulfates and bile acid sulfates that were not detectable in adult serum.Conclusion:Our data are the first to demonstrate that cord serum contains bile acid sulfates that are synthesized early in the hydroxylase, neutral and acidic pathways of primary bile acid biosynthesis and support previous publications of cord blood perfluoralkyl toxins in newborns.

Role of Modular Polyketide Synthases in the Production of Polyether Ladder Compounds in Ciguatoxin-Producing Gambierdiscus polynesiensis and G. excentricus (Dinophyceae).

Gambierdiscus, a benthic dinoflagellate, produces ciguatoxins that cause the human illness Ciguatera. Ciguatoxins are polyether ladder compounds that have a polyketide origin, indicating that polyketide synthases (PKS) are involved in their production. We sequenced transcriptomes of Gambierdiscus excentricus and Gambierdiscus polynesiensis and found 264 contigs encoding single domain ketoacyl synthases (KS; G.excentricus: 106, G.polynesiensis: 143) and ketoreductases (KR; G.excentricus: 7, G.polynesiensis: 8) with sequence similarity to type I PKSs, as reported in other dinoflagellates. In addition, 24 contigs (G.excentricus: 3, G.polynesiensis: 21) encoding multiple PKS domains (forming typical type I PKSs modules) were found. The proposed structure produced by one of these megasynthases resembles a partial carbon backbone of a polyether ladder compound. Seventeen contigs encoding single domain KS, KR, s-malonyltransacylase, dehydratase and enoyl reductase with sequence similarity to type II fatty acid synthases (FAS) in plants were found. Type I PKS and type II FAS genes were distinguished based on the arrangement of domains on the contigs and their sequence similarity and phylogenetic clustering with known PKS/FAS genes in other organisms. This differentiation of PKS and FAS pathways in Gambierdiscus is important, as it will facilitate approaches to investigating toxin biosynthesis pathways in dinoflagellates.

Predator cues reduce intraspecific trait variability in a marine dinoflagellate

BackgroundPhenotypic plasticity is commonplace and enables an organism to respond to variations in the environment. Plastic responses often modify a suite of traits and can be triggered by both abiotic and biotic changes. Here we analysed the plastic response towards a grazer of two genotypes of the marine dinoflagellate Alexandrium fundyense, evaluated the similarity of this response and discuss potential strain-specific trade-offs. We compared the expression of the known inducible defensive traits paralytic shellfish toxin content, and chain length. The effectiveness of the induced defense was assessed by monitoring grazing rates in both strains.ResultsOur results show that the grazer cues diminish phenotypic variability in a population by driving the phenotype towards a common defended morphotype. We further showed that the expression of the sxtA gene that initiates the paralytic shellfish toxin biosynthesis pathway does not correlate with an observed increase in the paralytic shellfish toxin analogue saxitoxin, and that toxin induction differs in its physiological characteristics in both strains.ConclusionInduced defense response in Alexandrium thus can directly affect further species interactions by reducing phenotypic variation and can result in genotype-dependent ecological trade-offs.

Involvement of FvSet1 in Fumonisin B1 Biosynthesis, Vegetative Growth, Fungal Virulence, and Environmental Stress Responses in Fusarium verticillioides.

Fusarium verticillioides (teleomorph, Gibberella moniliformis) is an important plant pathogen that causes seedling blight, stalk rot, and ear rot in maize (Zea mays). During infection, F. verticillioides produce fumonsins B1 (FB1) that pose a serious threat to human and animal health. Recent studies showed that Set1, a methyltransferase of H3K4, was responsible for toxin biosynthesis in filamentous fungi. However, to date, the regulation of FvSet1 on FB1 biosynthesis remains unclear. In the current study, we identified only one Set1 ortholog in F. verticillioides (FvSet1) and found that the deletion of FvSET1 led to various defects in fungal growth and pathogenicity. More interestingly, the FvSET1 deletion mutant (ΔFvSet1) showed a significant defect in FB1 biosynthesis and lower expression levels of FUM genes. FvSet1 was also found to play an important role in the responses of F. verticillioides to multiple environmental stresses via regulating the phosphorylation of FvMgv1 and FvHog1. Taken together, these results indicate that FvSet1 plays essential roles in the regulation of FB1 biosynthesis, fungal growth and virulence, as well as various stress responses in F. verticillioides.

DEVELOPMENT OF THE ALGORITHM FOR IDENTIFICATION OF THE LEVEL OF VIBRIO CHOLERAE CTXA AND TOXR GENE EXPRESSION USING RT-PCR WITH REAL-TIME HYBRIDIZATION-FLUORESCENT REGISTRATION OF RESULTS

Objective of the study is to design the algorithm for assessment of expression of the structural and regulatory virulence Vibrio cholerae genes by the model of ctxA and toxR genes encoding and controlling biosynthesis of cholera toxin. Materials and methods . Utilized were 10 strains of Vibrio cholerae, classical and El Tor biovars. Cloning of gene fragments was carried out through transformation and ligation. RT-PCR was done in “BIS M112” and “Rotor-GeneQ” amplifiers. Processing of the results was performed by means of the software package in set with Rotor-GeneQ (Software 1.8.17.5). Results and conclusions . Developed has been the algorithm for evaluation of the level of expression of V. cholerae ctxA and toxR genes applying RT-PCR with real-time hybridization-fluorescent registration of results. The stated algorithm allows for rapid and effective statistically significant specification of the expression of structural and regulatory virulence V. cholerae genes and can be used for evaluation of newly discovered cholera vibrio strains.

Penicillium roqueforti PR toxin gene cluster characterization.

PR toxin is a well-known isoprenoid mycotoxin almost solely produced by Penicillium roqueforti after growth on food or animal feed. This mycotoxin has been described as the most toxic produced by this species. In this study, an in silico analysis allowed identifying for the first time a 22.4-kb biosynthetic gene cluster involved in PR toxin biosynthesis in P. roqueforti. The pathway contains 11 open reading frames encoding for ten putative proteins including the major fungal terpene cyclase, aristolochene synthase, involved in the first farnesyl-diphosphate cyclization step as well as an oxidoreductase, an oxidase, two P450 monooxygenases, a transferase, and two dehydrogenase enzymes. Gene silencing was used to study three genes (ORF5, ORF6, and ORF8 encoding for an acetyltransferase and two P450 monooxygenases, respectively) and resulted in 20 to 40% PR toxin production reductions in all transformants proving the involvement of these genes and the corresponding enzyme activities in PR toxin biosynthesis. According to the considered silenced gene target, eremofortin A and B productions were also affected suggesting their involvement as biosynthetic intermediates in this pathway. A PR toxin biosynthesis pathway is proposed based on the most recent and available data.

MAP kinase gene STK1 is required for hyphal, conidial, and appressorial development, toxin biosynthesis, pathogenicity, and hypertonic stress response in the plant pathogenic fungus Setosphaeria turcica

The mitogen-activated protein kinase (MAPK), a key signal transduction component in the MAPK cascade pathway, regulates a variety of physiological activities in eukaryotes. However, little is known of the role MAPK plays in phytopathogenic fungi. In this research, we cloned the MAPK gene STK1 from the northern corn leaf blight pathogen Setosphaeria turcica and found that the gene shared high homology with the high osmolality glycerol (HOG) MAPK gene HOG1 of Saccharomyces cerevisiae. In addition, gene knockout technology was employed to investigate the function of STK1. Gene knockout mutants (KOs) were found to have altered hyphae morphology and no conidiogenesis, though they did show similar radial growth rate compared to the wild-type strain (WT). Furthermore, microscope observations indicated that STK1 KOs did not form normal appressoria at 48 h post-inoculation on a hydrophobic surface. STK1 KOs had reduced virulence, a significantly altered Helminthosporium turcicum (HT)-toxin composition, and diminished pathogenicity on the leaves of susceptible inbred corn OH43. Mycelium morphology appeared to be significantly swollen and the radial growth rates of STK1 KOs declined in comparison with WT under high osmotic stress. These results suggested that STK1 affects the hyphae development, conidiogenesis, and pathogenicity of S. turcica by regulating appressorium development and HT-toxin biosynthesis. Moreover, the gene appears to be involved in the hypertonic stress response in S. turcica.

The genome of the emerging barley pathogen Ramularia collo-cygni.

BackgroundRamularia collo-cygni is a newly important, foliar fungal pathogen of barley that causes the disease Ramularia leaf spot. The fungus exhibits a prolonged endophytic growth stage before switching life habit to become an aggressive, necrotrophic pathogen that causes significant losses to green leaf area and hence grain yield and quality.ResultsThe R. collo-cygni genome was sequenced using a combination of Illumina and Roche 454 technologies. The draft assembly of 30.3 Mb contained 11,617 predicted gene models. Our phylogenomic analysis confirmed the classification of this ascomycete fungus within the family Mycosphaerellaceae, order Capnodiales of the class Dothideomycetes. A predicted secretome comprising 1053 proteins included redox-related enzymes and carbohydrate-modifying enzymes and proteases. The relative paucity of plant cell wall degrading enzyme genes may be associated with the stealth pathogenesis characteristic of plant pathogens from the Mycosphaerellaceae. A large number of genes associated with secondary metabolite production, including homologs of toxin biosynthesis genes found in other Dothideomycete plant pathogens, were identified.ConclusionsThe genome sequence of R. collo-cygni provides a framework for understanding the genetic basis of pathogenesis in this important emerging pathogen. The reduced complement of carbohydrate-degrading enzyme genes is likely to reflect a strategy to avoid detection by host defences during its prolonged asymptomatic growth. Of particular interest will be the analysis of R. collo-cygni gene expression during interactions with the host barley, to understand what triggers this fungus to switch from being a benign endophyte to an aggressive necrotroph.Electronic supplementary materialThe online version of this article (doi:10.1186/s12864-016-2928-3) contains supplementary material, which is available to authorized users.

Antioxidant Secondary Metabolites in Cereals: Potential Involvement in Resistance to Fusarium and Mycotoxin Accumulation.

Gibberella and Fusarium Ear Rot and Fusarium Head Blight are major diseases affecting European cereals. These diseases are mainly caused by fungi of the Fusarium genus, primarily Fusarium graminearum and Fusarium verticillioides. These Fusarium species pose a serious threat to food safety because of their ability to produce a wide range of mycotoxins, including type B trichothecenes and fumonisins. Many factors such as environmental, agronomic or genetic ones may contribute to high levels of accumulation of mycotoxins in the grain and there is an urgent need to implement efficient and sustainable management strategies to reduce mycotoxin contamination. Actually, fungicides are not fully efficient to control the mycotoxin risk. In addition, because of harmful effects on human health and environment, their use should be seriously restricted in the near future. To durably solve the problem of mycotoxin accumulation, the breeding of tolerant genotypes is one of the most promising strategies for cereals. A deeper understanding of the molecular mechanisms of plant resistance to both Fusarium and mycotoxin contamination will shed light on plant-pathogen interactions and provide relevant information for improving breeding programs. Resistance to Fusarium depends on the plant ability in preventing initial infection and containing the development of the toxigenic fungi while resistance to mycotoxin contamination is also related to the capacity of plant tissues in reducing mycotoxin accumulation. This capacity can result from two mechanisms: metabolic transformation of the toxin into less toxic compounds and inhibition of toxin biosynthesis. This last mechanism involves host metabolites able to interfere with mycotoxin biosynthesis. This review aims at gathering the latest scientific advances that support the contribution of grain antioxidant secondary metabolites to the mechanisms of plant resistance to Fusarium and mycotoxin accumulation.

A monofunctional nonribosomal peptide synthase might be involved in the biosynthesis of marine toxins of the palytoxin family

The aim of this study was to identify in the transcriptome of the dinoflagellate Ostreopsis ovata genes, particularly such encoding nonribosomal peptide synthases (NRPSs), that are possibly involved in the biosynthesis of ovatoxins. Polyketide synthases (PKSs) and NRPSs are multienzyme complexes of mainly bacterial and fungal secondary metabolism. They comprise various enzymatic activities and are often organised in so‐called modules, each of which inserts one building block. The minimal NRPS module consists of an adenylation domain (A domain), which selects and activates the substrate, a condensation domain (C domain), which catalyses the formation of the new peptide bond, and a thiolation domain for binding intermediates. In the last decade, PKS and NRPS genes have also been identified in various protists, including some dinoflagellate species. A strain of the dinoflagellate O. ovata that was sampled in Okinawa, Japan, produces several ovatoxin variants. Ovatoxins belong to the same structural family as the extremely poisonous compound palytoxin. The biosynthetic scheme of palytoxin and its congeners is completely unknown. Sequencing of the O. ovata transcriptome by means of RNA‐Seq revealed a wide range of genes encoding PKSs and NRPSs. Most interestingly, O. ovata possesses a new type of NRPS, which is characterised by an unusual monofunctional architecture. This monofunctional architecture resembles a special type of monofunctional PKSs that have been found exclusively in dinoflagellates. The same type of monofunctional NRPSs found in O. ovata was also identified by screening the published draft genome of the dinoflagellate Symbiodinium minutum. These enzymes form a distinct dinoflagellate‐specific clade in comprehensive phylogenetic reconstructions including NRPSs from bacteria, fungi and other eukaryotes. Expression of those genes on the RNA level was confirmed by RT‐PCR, and the sequences of the transcripts demonstrated that the transcriptome‐based prediction of monofunctional NRPSs was not an artefact caused by assembly issues. Therefore, this study has experimentally demonstrated for the first time the existence of a new type of NRPSs, which at present seems to be limited to dinoflagellates. Structural modelling revealed differences between the active centre of A domains of dinoflagellate origin and those from bacteria and fungi. This new type of NRPS found in O. ovata might be involved in the biosynthesis of ovatoxin and related compounds, as the ovatoxin structure hints to incorporation of an amino acid as part of the biosynthesis pathway. The finding presented here will make it possible to initiate the elucidation of the biosynthesis scheme of palytoxin and related compounds.Support or Funding InformationThis study was supported by internal funds of the Okinawa Institute of Science and Technology Graduate University (OIST).

The global regulator LaeA controls biosynthesis of host-specific toxins, pathogenicity and development of Alternaria alternata pathotypes

The global regulator LaeA is required for the expression of biosynthetic genes for secondary metabolites in filamentous fungi such as Aspergillus nidulans. To learn more about the regulation of these genes in Alternaria alternata, which has at least seven pathogenic variants (pathotypes) that produce host-specific toxins (HSTs) and cause severe diseases in their host plants, we identified LaeA homologs encoding methyltransferase in tomato, strawberry and apple pathotypes of A. alternata, designating them AtLAE1, AsLAE1 and AaLAE1, respectively. In the AtLAE1-deleted mutant of the tomato pathotype (∆AtLAE1), expression of the AAL-toxin biosynthetic gene ALT1 was lower than that in the wild type. Correspondingly, AAL-toxin production and virulence of the mutant were significantly lower as were spore production and hyphal growth. Similarly, the AsLAE1-deleted mutant of the strawberry pathotype (∆AsLAE1) and the AaLAE1-deleted mutant of the apple pathotype (∆AaLAE1) produced less of their host-specific toxin (AF- and AM-toxin, respectively), had reduced virulence on the host plant, and hyphal growth and sporulation were defective. Thus, the global regulator gene LaeA positively regulates HST biosynthesis, pathogenicity, growth and differentiation in A. alternata pathotypes.

Paralytic shellfish toxin production by the dinoflagellate Alexandrium pacificum (Chinhae Bay, Korea) in axenic, nutrient-limited chemostat cultures and nutrient-enriched batch cultures

Blooms of Alexandrium pacificum (formerly Alexandrium tamarense) are common in Chinhae Bay (Korea), presumably linked to anthropogenic eutrophication. Here we examine PSP toxin content and composition in axenic chemostat and batch cultures of A. pacificum using growth conditions that differed according to dilution rate, nutrient limitations, and enrichments. Phosphate (P)-limited cells in chemostat cultures had higher toxin content and a toxin composition that differed from that of nitrogen (N)-limited cells at the highest growth rates. Therefore, toxin composition changes do occur in axenic cultures of A. pacificum following extended growth under steady state conditions. In nutrient-limited batch cultures that received N and P enrichment, the N-enriched cultures showed a more diverse toxin profile than the P-enriched cells; the toxin content of N-enriched cells was lower than in the P-enriched cultures. We infer the following order for the biosynthesis of individual toxins: C1, C2>GTX3>GTX1>neoSTX.

Isoepoxydon dehydrogenase (idh) gene expression in relation to patulin production by Penicillium expansum under different temperature and atmosphere

Penicillium expansum growth and patulin production occur mainly at post-harvest stage during the long-term storage of apples. Low temperature in combination with reduced oxygen concentrations is commonly applied as a control strategy to extend apple shelf life and supply the market throughout the year. Our in vitro study investigated the effect of temperature and atmosphere on expression of the idh gene in relation to the patulin production by P. expansum. The idh gene encodes the isoepoxydon dehydrogenase enzyme, a key enzyme in the patulin biosynthesis pathway. First, a reverse transcription real-time PCR (RT-qPCR) method was optimized to measure accurately the P. expansum idh mRNA levels relative to the mRNA levels of three reference genes (18S, β-tubulin, calmodulin), taking into account important parameters such as PCR inhibition and multiple reference gene stability. Subsequently, two P. expansum field isolates and one reference strain were grown on apple puree agar medium (APAM) under three conditions of temperature and atmosphere: 20°C — air, 4°C — air and 4°C — controlled atmosphere (CA; 3% O2). When P. expansum strains reached a 0.5 and 2.0cm colony diameter, idh expression and patulin concentrations were determined by means of the developed RT-qPCR and an HPLC-UV method, respectively. The in vitro study showed a clear reduction in patulin production and down-regulation of the idh gene expression when P. expansum was grown under 4°C — CA. The results suggest that stress (low temperature and oxygen level) caused a delay of the fungal metabolism rather than a complete inhibition of toxin biosynthesis. A good correlation was found between the idh expression and patulin production, corroborating that temperature and atmosphere affected patulin production by acting at the transcriptional level of the idh gene. Finally, a reliable RT-qPCR can be considered as an alternative tool to investigate the effect of control strategies on the toxin formation in food.

Gene Discovery for Synthetic Biology: Exploring the Novel Natural Product Biosynthetic Capacity of Eukaryotic Microalgae.

Eukaryotic microalgae are an incredibly diverse group of organisms whose sole unifying feature is their ability to photosynthesize. They are known for producing a range of potent toxins, which can build up during harmful algal blooms causing damage to ecosystems and fisheries. Genome sequencing is lagging behind in these organisms because of their genetic complexity, but transcriptome sequencing is beginning to make up for this deficit. As more sequence data becomes available, it is apparent that eukaryotic microalgae possess a range of complex natural product biosynthesis capabilities. Some of the genes concerned are responsible for the biosynthesis of known toxins, but there are many more for which we do not know the products. Bioinformatic and analytical techniques have been developed for natural product discovery in bacteria and these approaches can be used to extract information about the products synthesized by algae. Recent analyses suggest that eukaryotic microalgae produce many complex natural products that remain to be discovered.

Suppression of Aflatoxin Biosynthesis in Aspergillus flavus by 2-Phenylethanol Is Associated with Stimulated Growth and Decreased Degradation of Branched-Chain Amino Acids.

The saprophytic soil fungus Aspergillus flavus infects crops and produces aflatoxin. Pichia anomala, which is a biocontrol yeast and produces the major volatile 2-phenylethanol (2-PE), is able to reduce growth of A. flavus and aflatoxin production when applied onto pistachio trees. High levels of 2-PE are lethal to A. flavus and other fungi. However, at low levels, the underlying mechanism of 2-PE to inhibit aflatoxin production remains unclear. In this study, we characterized the temporal transcriptome response of A. flavus to 2-PE at a subinhibitory level (1 µL/mL) using RNA-Seq technology and bioinformatics tools. The treatment during the entire 72 h experimental period resulted in 131 of the total A. flavus 13,485 genes to be significantly impacted, of which 82 genes exhibited decreased expression. They included those encoding conidiation proteins and involved in cyclopiazonic acid biosynthesis. All genes in the aflatoxin gene cluster were also significantly decreased during the first 48 h treatment. Gene Ontology (GO) analyses showed that biological processes with GO terms related to catabolism of propionate and branched-chain amino acids (valine, leucine and isoleucine) were significantly enriched in the down-regulated gene group, while those associated with ribosome biogenesis, translation, and biosynthesis of α-amino acids were over-represented among the up-regulated genes. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis revealed that metabolic pathways negatively impacted among the down-regulated genes parallel to those active at 30 °C, a condition conducive to aflatoxin biosynthesis. In contrast, metabolic pathways positively related to the up-regulated gene group resembled those at 37 °C, which favors rapid fungal growth and is inhibitory to aflatoxin biosynthesis. The results showed that 2-PE at a low level stimulated active growth of A. flavus but concomitantly rendered decreased activities in branched-chain amino acid degradation. Since secondary metabolism occurs after active growth has ceased, this growth stimulation resulted in suppression of expression of aflatoxin biosynthesis genes. On the other hand, increased activities in degradation pathways for branched-chain amino acids probably are required for the activation of the aflatoxin pathway by providing building blocks and energy regeneration. Metabolic flux in primary metabolism apparently has an important role in the expression of genes of secondary metabolism.

Possible way of zearalenone migration in the agricultural environment

The aim of this study was to elucidate the zearalenone (ZEA) migration in the agricultural environment, from the field (wheat, corn), through soil and ending in water. All samples were collected in the agricultural area of Western Poland in period between March 2010 and December 2012 (three-year study). Determination of ZEA in environmental samples, after extraction and purification, was carried out with use of high-performance liquid chromatography. The toxin content in water samples was between 0.5 and 4.9 ng/L and increases in the late summer and autumn (after harvest). The results clearly indicate that ZEA concentration in surface waters depends on many factors, of which the most important include weather conditions, the seasons and intensity of rainfall, and it is possibly related to scab severity of cereals and the toxin biosynthesis.

Paralytic shellfish toxin biosynthesis in cyanobacteria and dinoflagellates: A molecular overview

Paralytic shellfish toxins (PSTs) are a group of water soluble neurotoxic alkaloids produced by two different kingdoms of life, prokaryotic cyanobacteria and eukaryotic dinoflagellates. Owing to the wide distribution of these organisms, these toxic secondary metabolites account for paralytic shellfish poisonings around the world. On the other hand, their specific binding to voltage-gated sodium channels makes these toxins potentially useful in pharmacological and toxicological applications. Much effort has been devoted to the biosynthetic mechanism of PSTs, and gene clusters encoding 26 proteins involved in PST biosynthesis have been unveiled in several cyanobacterial species. Functional analysis of toxin genes indicates that PST biosynthesis in cyanobacteria is a complex process including biosynthesis, regulation, modification and export. However, less is known about the toxin biosynthesis in dinoflagellates owing to our poor understanding of the massive genome and unique chromosomal characteristics [1]. So far, few genes involved in PST biosynthesis have been identified from dinoflagellates. Moreover, the proteins involved in PST production are far from being totally explored. Thus, the origin and evolution of PST biosynthesis in these two kingdoms are still controversial. In this review, we summarize the recent progress on the characterization of genes and proteins involved in PST biosynthesis in cyanobacteria and dinoflagellates, and discuss the standing evolutionary hypotheses concerning the origin of toxin biosynthesis as well as future perspectives in PST biosynthesis. Scientific questionParalytic shellfish toxins (PSTs) are a group of potent neurotoxins which specifically block voltage-gated sodium channels in excitable cells and result in paralytic shellfish poisonings (PSPs) around the world. Two different kingdoms of life, cyanobacteria and dinoflagellates are able to produce PSTs. However, in contrast with cyanobacteria, our understanding of PST biosynthesis in dinoflagellates is extremely limited owing to their unique features. The origin and evolution of PST biosynthesis in these two kingdoms are still controversial. Technical significanceHigh-throughput omics technologies, such as genomics, transcriptomics and proteomics provide powerful tools for the study of PST biosynthesis in cyanobacteria and dinoflagellates, and have shown their powerful potential with regard to revealing genes and proteins involved in PST biosynthesis in two kingdoms. Scientific significanceThis review summarizes the recent progress in PST biosynthesis in cyanobacteria and dinoflagellates with focusing on the novel insights from omics technologies, and discusses the evolutionary relationship of toxin biosynthesis genes between these two kingdoms.

Identification of intermediates in the biosynthesis of PR toxin by Penicillium roqueforti.

The sesquiterpenoid 7-epi-neopetasone was synthesized via the Wieland-Miescher ketone. The compound was identical to a previously tentatively identified headspace constituent of Penicillium roqueforti. Feeding experiments with (13) C-labeled mevalonolactone isotopomers demonstrated that oxidation at C12 and an isomerization of the C11C12 to a C7C11 double bond must occur independently and not via a C7-C11-C12 allyl radical in one step. Feeding with (11,12,13-(13) C3 )-7-epi-neopetasone resulted in labelling of the PR toxin, thus establishing this compound as a newly identified pathway intermediate.

Relationship of proteomic variation and toxin synthesis in the dinoflagellate Alexandrium tamarense CI01 under phosphorus and inorganic nitrogen limitation.

Paralytic shellfish toxins (PSTs) are originated from cyanobacteria and dinoflagellates, including Alexandrium tamarense, the common dinoflagellate species. In this study, a toxic dinoflagellate strain of A. tamarense CI01 was selected for studying the PSTs' concentration and the related protein variation during the whole cell cycle under different nutrient conditions. High-performance liquid chromatography, 2-D DIGE and Western blotting were used collectively for protein profiling and identification. Results showed that the toxin content was suppressed under nitrogen limiting condition, but enhanced in phosphorous limiting medium. Based on the results of proteomics analysis, 7 proteins were discovered to be related to the PSTs biosynthesis of A. tamarense CI01, including S-adenosylhomocysteine hydrolase, ornithine cyclodeaminase, argininosuccinate synthase, methyluridine methyltransferase cystine ABC transporter, phosphoserine phosphatase, argininosuccinate synthase and acyl-CoA dehydrogenase, which corresponds to the metabolism of the methionine, cysteine, ornithine, arginine and proline. Moreover, some photosynthesis relating proteins also increased their expression during PST synthesis period in A. tamarense CI01, such as phosphoenolpyruvate carboxylase, chloroplast phosphoglycerate kinase, peridinin-chlorophyll α-binding protein, Mg(2+) transporter protein and chloroplast phosphoglycerate kinase. The above findings are in support of our hypothesis that these proteins are involved in toxin biosynthesis of A. tamarense CI01, but cause-and-effect mechanisms need to be investigated in further studies.